Preface

An early diagnosis, doubled by a comprehensive check-up and a correct choice of treatment, followed by an appropriate follow-up schedule, currently set apart successful management from failure when it comes to dealing with cancer. Although the development of new molecules constantly opens potential pathways to managing malignancies, the overall incidence and mortality rates continue to rise worldwide.

It is important to acknowledge the shifting tendencies, emerging endemic areas, and new at-risk populations in today's rapidly evolving societies. From constant exposure to stress, poor dietary and lifestyle choices, and exposure to new and more insidious polluting factors, many new challenges continue to influence the potential benefits of novel therapies. A precarious balance governs modern oncology, with the patient in the middle of this "silent war" constantly fought by medical professionals, assisted by novel tools provided by researchers from all areas of science.

Novel surgical methods, robotic procedures, and minimally invasive techniques based on newly developed devices will minimize the risk of patients, along with decreased hospitalization times, fewer consumed resources, and better economic impact. After-treatment reintegration of patients will significantly influence the global economy in the long term.

The impact of chemotherapy is further potentiated by novel lines of biological, hormonal, immunotherapeutic, and targeted medications. A more personalized type of approach is supported by recent discoveries in the fields of genetics and molecular biology.

The potential of bioinformatics, with the aid of large-scale data analysis, artificial intelligence, and similar computer-assisted tools, will become increasingly significant in the coming years.

Cancer awareness is important; the rigors of regular check-ups, starting with primary medicine and continuing in specialized centers, are the cornerstones of success stories. The successful collaboration between the poles of excellence in cancer diagnostics that exist in each country and region, their integration with primary and secondary centers, as well as the strong link with the academic world and industry researchers, will unquestionably represent the vector of success against cancer.

All these aspects require constant attention from the academic world, and the collection of interesting and diverse chapters presented within this volume will hopefully raise awareness towards interesting and novel topics. Information is being delivered faster than ever towards both medical professionals and the general

**II**

**Section 4**

*by Richard Lucas Konichi-Dias*

Paraneoplastic Syndromes **125**

**Chapter 7 127**

Paraneoplastic Pemphigus Is a Life-Threatening Disease

population. We feel that the field of oncology is ever-changing, with major breakthroughs always waiting around the corner.

> **Liliana Streba and Michael Schenker** Department of Medical Oncology, University of Medicine and Pharmacy of Craiova, Romania

> **Dan Ionuț Gheonea** Department of Gastroenterology, University of Medicine and Pharmacy of Craiova, Romania

> > **1**

Section 1

Novel Approaches to

Cancer Management

Section 1

## Novel Approaches to Cancer Management

**3**

zebrafish [8].

**Chapter 1**

**Abstract**

early development.

**1. Introduction**

cancer, toxicology, drug discovery

development of nervous system.

Organism

Zebrafish (*Danio rerio*) as a Model

*Farmanur Rahman Khan and Saleh Sulaiman Alhewairini*

Animals as model organisms, the silent sentinels, stand watch over the environmental health of the world. These are non-human animal species which can be used to understand specific biological processes and to obtain informations which can provide an insight into working of other organisms. Among the model organisms, the zebrafish (*Danio rerio*) is one of the best leading models to study developmental biology, cancer, toxicology, drug discovery, and molecular genetics. In addition, the zebrafish is increasingly used as a genetic model organism for aquaculture species and in toxicogenomics and also to generate zebrafish disease models for application in human biomedicines. This tiny fish is a versatile model organism for many fields of research because of its easy maintenance, breeding, and transparent body during

**Keywords:** model organisms, zebrafish, biological process, developmental biology,

Zebrafish (*Danio rerio*) is a prominent model organism in biological researches in recent times. Zebrafish is a tropical freshwater fish, inhabitant of rivers (Ganges mainly) of Himalayan region of South Asia especially India, Nepal, Bhutan, Pakistan, Bangladesh, and Myanmar. It is a bony fish (teleost) that belongs to the

Zebrafish was first used as a biological model by George Streisinger (University of Oregon) in the 1970s because it was simpler over mouse and easy to manipulate genetically. Streisinger's colleagues especially Chuck Kimmel in his university got much impressed by the idea of using zebrafish embryo more attractive to study the

The use of zebrafish as a model organism got impetus from the 1990s when it was used to develop two large genetic mutants, one by Nobel Prize winner Christiane Nusslein-Volhard in Tubingen, Germany, and the other by Wolfgang Driever and Mark Fishman in Boston, USA. The identification of mutants is one of

Zebrafish has a lot of physiological and genetic similarities with humans, including the brain, digestive tract, musculature, vasculature, and innate immune system [1–7]. Also 70% of human disease genes have functional similarities with those of

family Cyprinidae under the class Actinopterygii (ray-finned fishes).

the most important strategies for the study in various areas of biology.

#### **Chapter 1**

## Zebrafish *(Danio rerio)* as a Model Organism

*Farmanur Rahman Khan and Saleh Sulaiman Alhewairini*

#### **Abstract**

Animals as model organisms, the silent sentinels, stand watch over the environmental health of the world. These are non-human animal species which can be used to understand specific biological processes and to obtain informations which can provide an insight into working of other organisms. Among the model organisms, the zebrafish (*Danio rerio*) is one of the best leading models to study developmental biology, cancer, toxicology, drug discovery, and molecular genetics. In addition, the zebrafish is increasingly used as a genetic model organism for aquaculture species and in toxicogenomics and also to generate zebrafish disease models for application in human biomedicines. This tiny fish is a versatile model organism for many fields of research because of its easy maintenance, breeding, and transparent body during early development.

**Keywords:** model organisms, zebrafish, biological process, developmental biology, cancer, toxicology, drug discovery

#### **1. Introduction**

Zebrafish (*Danio rerio*) is a prominent model organism in biological researches in recent times. Zebrafish is a tropical freshwater fish, inhabitant of rivers (Ganges mainly) of Himalayan region of South Asia especially India, Nepal, Bhutan, Pakistan, Bangladesh, and Myanmar. It is a bony fish (teleost) that belongs to the family Cyprinidae under the class Actinopterygii (ray-finned fishes).

Zebrafish was first used as a biological model by George Streisinger (University of Oregon) in the 1970s because it was simpler over mouse and easy to manipulate genetically. Streisinger's colleagues especially Chuck Kimmel in his university got much impressed by the idea of using zebrafish embryo more attractive to study the development of nervous system.

The use of zebrafish as a model organism got impetus from the 1990s when it was used to develop two large genetic mutants, one by Nobel Prize winner Christiane Nusslein-Volhard in Tubingen, Germany, and the other by Wolfgang Driever and Mark Fishman in Boston, USA. The identification of mutants is one of the most important strategies for the study in various areas of biology.

Zebrafish has a lot of physiological and genetic similarities with humans, including the brain, digestive tract, musculature, vasculature, and innate immune system [1–7]. Also 70% of human disease genes have functional similarities with those of zebrafish [8].

#### **1.1 Salient features of zebrafish as a model organism**

*D. Rario* is preferred by scientists because of its variety of features that make it useful as a model organism. The embryo develops rapidly outside mother and optically clear and thus, easily accessible for experimentation and observation. The embryo develops very fast, and the blastula stage lasts only for 3 h, while gastrulation gets completed in 5 h; in an embryo that is about 18 h old, very well developed ears, eyes, segmenting muscles, and brain can be viewed as the embryo is transparent. By 24 h, segmentation gets completed, and most primary organ systems are formed. By 72 h, the embryo hatches out from the eggshell and within the next 2 days starts hunting for food. In a period of just 4 days, the embryo converts rapidly into a small version of adult. The rapid development simplifies development and genetic studies.

The adult zebrafish attains sexual maturity very quickly, having generation time of about 10 weeks, and also this tiny fish has good fecundity rate. When kept under optimal conditions, the zebrafish can lay about 200 eggs per week [9, 10]. Under laboratory conditions the zebrafish can spawn throughout the year that ensures the constant supply of offspring from designated pairs that makes this transparent fish a quintessential choice for large-scale genetic approaches to identify novel genes and to discover their specific functions in vertebrates [11]. The zebrafish is a very hard fish and is very easy to raise.

In addition to the features of zebrafish mentioned above, it requires very low space and maintenance cost. These features make this fish an attractive model organism for developmental, toxicological, and transgenic studies [12].

In this chapter author summarizes some of the recent advances in the area of zebrafish research, viz., developmental biology, toxicology, transgenic studies, human disease, drug discovery, cancer, etc. This review is by no means a comprehensive one but an attempt to provide a flavor to the readers some recent advances about this wonderful creature to use in potential researches.

#### **2. Use of zebrafish in developmental biology**

Much of the pioneer works that established zebrafish as a model organism were done by George Streisinger, Charles Kimmel, and their colleagues [13]. The team of these researchers studied the embryonic axis, cell lineage analysis, embryonic formation, development of central and peripheral nervous systems, muscle development, differential regulation of gene expression, etc. [14–16].

Many of the critical pathways that control development in vertebrates are highly conserved between human and zebrafish. The zebrafish genome shares a lot of similarities with human genome. About 70% of genes associated with disease in humans have functional homologs in zebrafish [8]. Realizing the importance of zebrafish model, Grunwald and Eisen used this developmental model to study the segmental structure of the brain and characterized neurons in zebrafish for the first time in a vertebrate model [17]. Nüsslein-Volhard recognized the importance of zebrafish as a vertebrate model to study developmental biology by identifying developmentally important genes [18]. The zebrafish model has been used to see the development of various systems/processes as follows.

#### **2.1 Development of the enteric nervous system**

Recently advances have been made to study the development of the enteric nervous system (ENS) using the zebrafish model. Like other vertebrates, the zebrafish

**5**

*Zebrafish (Danio rerio) as a Model Organism DOI: http://dx.doi.org/10.5772/intechopen.81517*

resemble with that of amniotes [23].

**2.2 Angiogenesis**

in live specimens [31].

ment also can be tracked.

gastrointestinal tract is a complex organ composed of multiple cell types like epithelial, muscular, vascular, neural, and immune cells. The gut of the zebrafish (teleost) and amniotes have structural similarities, but in teleost it is less complex as compared to amniotes [19]. The zebrafish GI tract has no distinct stomach but an enlarged area of the anterior intestine that is known as the intestinal bulb. This intestinal bulb displays patterns of motility as well as goblet cells that produce acid and neutral mucins like the stomach of mammals [20, 21]. The gut epithelium of zebrafish is simpler than that of amniotes; it lacks crypts and is arranged in an irregular broad fold rather than forming villi [21, 22]. The genes (*sox2*, *barx1*, *gata5*, *and gata6*) which are responsible for the formation of the stomach of zebrafish also

Like that of all vertebrates, the enteric nervous system of zebrafish is also derived from neural crest [24], but it differs potentially from amniotes wherein the enteric nervous system is derived from both the vagal and sacral crests, while in the case of zebrafish, it is derived from the vagal crest only [25–28]. Enteric neural crest cell (ENCC) migration along the gut in zebrafish is also similar with that in amniotes. It takes place in two parallel chains along the length of the developing gut [25, 27, 28]. Afterward the precursors of the ENS voyage circumferentially around the gut and differentiate into the enteric neuron and glia. The final organization of the zebrafish ENS is also very simple as compared to that of amniotes; it is composed of single neuron or small group of neurons rather than more complex

Zebrafish model also has been used in the study of angiogenesis and regeneration. Angiogenesis is the process through which new blood vessels originate from preexisting vascular structures which play essential role in healthy physiological and pathological conditions. It is achieved through interaction between endothelial cells and their niche. Inadequate maintenance leads to the development of many disorders like tissue ischemia, inflammatory disorders, retinopathies, excessive

Being a transparent vertebrate, the zebrafish has emerged as a convenient alternative to study the early development of the cardiovascular system and observe the flow of blood [31]. In zebrafish larvae the vessels and blood flow can easily be visualized by using simple dissecting microscope and also by using fluorescent proteins; the development of the blood vascular system could be examined in great details. By using confocal microscopy and time-lapse imaging, the detailed morphogenetic movements and cell shape changes can be carried out

Vascular anatomy development of zebrafish using molecular tracers during early embryonic stages has high level of similarities with other vertebrates [1, 32, 33]. In one of the experiments, the injected fluorescent microsphere was detected when lumenization and anastomosis of the vascular network were complete [34]. The same approach was adopted to compare the development of blood and lymphatic vasculatures in zebrafish [35]. The individual cell growth during vascular develop-

For vascular development and growth, angiogenesis plays a very important role. During embryonic development, the intersegmental vessels are formed by angiogenic sprouting from the dorsal aorta, and they have been the target of studies using genetic perturbations or drugs [36]. It has been reported that mammalian malignant cells can be xenotransplanted into zebrafish embryos, and they can form tumors [37], and thus models for tumor angiogenesis have been developed [38].

ganglionated myenteric and submucosal plexuses [27, 29].

vascular growth, or abnormal remodeling that promotes cancer [30].

#### *Zebrafish (Danio rerio) as a Model Organism DOI: http://dx.doi.org/10.5772/intechopen.81517*

*Current Trends in Cancer Management*

hard fish and is very easy to raise.

genetic studies.

**1.1 Salient features of zebrafish as a model organism**

*D. Rario* is preferred by scientists because of its variety of features that make it useful as a model organism. The embryo develops rapidly outside mother and optically clear and thus, easily accessible for experimentation and observation. The embryo develops very fast, and the blastula stage lasts only for 3 h, while gastrulation gets completed in 5 h; in an embryo that is about 18 h old, very well developed ears, eyes, segmenting muscles, and brain can be viewed as the embryo is transparent. By 24 h, segmentation gets completed, and most primary organ systems are formed. By 72 h, the embryo hatches out from the eggshell and within the next 2 days starts hunting for food. In a period of just 4 days, the embryo converts rapidly into a small version of adult. The rapid development simplifies development and

The adult zebrafish attains sexual maturity very quickly, having generation time of about 10 weeks, and also this tiny fish has good fecundity rate. When kept under optimal conditions, the zebrafish can lay about 200 eggs per week [9, 10]. Under laboratory conditions the zebrafish can spawn throughout the year that ensures the constant supply of offspring from designated pairs that makes this transparent fish a quintessential choice for large-scale genetic approaches to identify novel genes and to discover their specific functions in vertebrates [11]. The zebrafish is a very

In addition to the features of zebrafish mentioned above, it requires very low space and maintenance cost. These features make this fish an attractive model organism for developmental, toxicological, and transgenic studies [12].

In this chapter author summarizes some of the recent advances in the area of zebrafish research, viz., developmental biology, toxicology, transgenic studies, human disease, drug discovery, cancer, etc. This review is by no means a comprehensive one but an attempt to provide a flavor to the readers some recent advances

Much of the pioneer works that established zebrafish as a model organism were done by George Streisinger, Charles Kimmel, and their colleagues [13]. The team of these researchers studied the embryonic axis, cell lineage analysis, embryonic formation, development of central and peripheral nervous systems, muscle devel-

Many of the critical pathways that control development in vertebrates are highly

Recently advances have been made to study the development of the enteric nervous system (ENS) using the zebrafish model. Like other vertebrates, the zebrafish

conserved between human and zebrafish. The zebrafish genome shares a lot of similarities with human genome. About 70% of genes associated with disease in humans have functional homologs in zebrafish [8]. Realizing the importance of zebrafish model, Grunwald and Eisen used this developmental model to study the segmental structure of the brain and characterized neurons in zebrafish for the first time in a vertebrate model [17]. Nüsslein-Volhard recognized the importance of zebrafish as a vertebrate model to study developmental biology by identifying developmentally important genes [18]. The zebrafish model has been used to see the

about this wonderful creature to use in potential researches.

opment, differential regulation of gene expression, etc. [14–16].

**2. Use of zebrafish in developmental biology**

development of various systems/processes as follows.

**2.1 Development of the enteric nervous system**

**4**

gastrointestinal tract is a complex organ composed of multiple cell types like epithelial, muscular, vascular, neural, and immune cells. The gut of the zebrafish (teleost) and amniotes have structural similarities, but in teleost it is less complex as compared to amniotes [19]. The zebrafish GI tract has no distinct stomach but an enlarged area of the anterior intestine that is known as the intestinal bulb. This intestinal bulb displays patterns of motility as well as goblet cells that produce acid and neutral mucins like the stomach of mammals [20, 21]. The gut epithelium of zebrafish is simpler than that of amniotes; it lacks crypts and is arranged in an irregular broad fold rather than forming villi [21, 22]. The genes (*sox2*, *barx1*, *gata5*, *and gata6*) which are responsible for the formation of the stomach of zebrafish also resemble with that of amniotes [23].

Like that of all vertebrates, the enteric nervous system of zebrafish is also derived from neural crest [24], but it differs potentially from amniotes wherein the enteric nervous system is derived from both the vagal and sacral crests, while in the case of zebrafish, it is derived from the vagal crest only [25–28]. Enteric neural crest cell (ENCC) migration along the gut in zebrafish is also similar with that in amniotes. It takes place in two parallel chains along the length of the developing gut [25, 27, 28]. Afterward the precursors of the ENS voyage circumferentially around the gut and differentiate into the enteric neuron and glia. The final organization of the zebrafish ENS is also very simple as compared to that of amniotes; it is composed of single neuron or small group of neurons rather than more complex ganglionated myenteric and submucosal plexuses [27, 29].

#### **2.2 Angiogenesis**

Zebrafish model also has been used in the study of angiogenesis and regeneration. Angiogenesis is the process through which new blood vessels originate from preexisting vascular structures which play essential role in healthy physiological and pathological conditions. It is achieved through interaction between endothelial cells and their niche. Inadequate maintenance leads to the development of many disorders like tissue ischemia, inflammatory disorders, retinopathies, excessive vascular growth, or abnormal remodeling that promotes cancer [30].

Being a transparent vertebrate, the zebrafish has emerged as a convenient alternative to study the early development of the cardiovascular system and observe the flow of blood [31]. In zebrafish larvae the vessels and blood flow can easily be visualized by using simple dissecting microscope and also by using fluorescent proteins; the development of the blood vascular system could be examined in great details. By using confocal microscopy and time-lapse imaging, the detailed morphogenetic movements and cell shape changes can be carried out in live specimens [31].

Vascular anatomy development of zebrafish using molecular tracers during early embryonic stages has high level of similarities with other vertebrates [1, 32, 33]. In one of the experiments, the injected fluorescent microsphere was detected when lumenization and anastomosis of the vascular network were complete [34]. The same approach was adopted to compare the development of blood and lymphatic vasculatures in zebrafish [35]. The individual cell growth during vascular development also can be tracked.

For vascular development and growth, angiogenesis plays a very important role. During embryonic development, the intersegmental vessels are formed by angiogenic sprouting from the dorsal aorta, and they have been the target of studies using genetic perturbations or drugs [36]. It has been reported that mammalian malignant cells can be xenotransplanted into zebrafish embryos, and they can form tumors [37], and thus models for tumor angiogenesis have been developed [38].

#### **2.3 Regeneration**

The zebrafish exhibits remarkable capacity of regeneration even in adult stages. The caudal fin especially provides an ideal tissue for vascular regeneration studies due to its simple and fine architecture and relative transparency [39]. After successive amputations the full regeneration of the caudal fin used to take place within couple of weeks [40]. The regenerating vessels in the regenerating caudal fin originate from vein-derived cells that have angiogenic potential [41]. These cells migrate individually or in groups and assemble into the vessel in response to chemokine signaling [42].

The zebrafish as an alternative model for angiogenesis and regeneration studies provides the relevance of in vivo assays with simplicity and versatility of in vitro assays. In larvae, access to developing vasculature through fluorophore-tagged strains and small size of zebrafish makes the use of high-throughput strategies possible. In adults, the caudal fin is equally convenient as a model tissue as regenerating vessels can be observed at all stages, and the animals (zebrafish) are suitable for experimental drug manipulations [31].

#### **3. Zebrafish as a cancer model system**

Cancer is a cursed reality for millions of humans worldwide and in fact for all vertebrates. The invertebrates such as flies and nematodes also can develop anomalies in cell proliferation. Clinically and pathologically this dreaded disease is present almost exclusively in all vertebrates, from fish to humans. To understand better the formation, growth, and spread of malignant tumors, vertebrate models are imperative. Being a vertebrate the zebrafish is an ideal model to study cancer, though humans and fishes are separated from their common ancestry but biology of the cancer in both groups of organisms is the same [43]. Because of the variety of benefits to use zebrafish as a model organism which are mentioned in "Introduction," the zebrafish is adroitly exploited to carcinogenic treatment, transplantation of mammalian tumor cells, and transgenic regulations [44].

#### **3.1 Zebrafish as a model for carcinogen effects and development of cancer studies**

Fishes are exposed to many waterborne carcinogens in the wild that lead to the development of a variety of benign and malignant tumors in teleosts, and these tumors have similar histology as in humans [45, 46]. As like humans, cancer is a genetic disease in fishes as shown by melanomas which develop in *Xiphophorus* hybrids [47]. Choosing zebrafish for modeling cancer studies has many advantages. Highly conserved cancer pathways can be screened genetically using zebrafish. Primarily cancer is a disease of adults, but through mutagenesis screens, cell cycle phenotype could be examined in rapidly developing transparent embryos of the zebrafish. The genes regulating cell cycle, cell proliferation, and apoptosis have already been screened in yeast, *Drosophila* and *C. elegans*, in the similar way gene functions for these biological pathways can be screened in zebrafish to understand the events that lead to the development of cancer in any vertebrate species [43].

By inducing different gene mutations or stimulating signaling pathways through chemicals, the tumors can be induced in different organs of the zebrafish like the pancreas, liver, GI tract, vasculature, muscles, skin, and testes [46, 48–51]. It is

**7**

*Zebrafish (Danio rerio) as a Model Organism DOI: http://dx.doi.org/10.5772/intechopen.81517*

**3.2 Tumorigenesis**

cally distant species [66, 67].

been transplanted in the zebrafish [7].

can improve the outcome in cancer patients [69].

**3.3 Xenotransplantation**

**3.4 Angiogenesis**

possible to identify the interacting oncogenes via suppressor and enhancer screens which cause the formation of specific type of tumor. The mammalian tumor cells can be transplanted into the zebrafish, dispensing a novel way to study the interac-

Tumorigenesis is a multistep process induced by chemical carcinogen [52], with accumulation of both epigenetic aberrations and mutations in regulatory regions of genes and disorder of signaling pathways [53, 54]. Methylation of DNA at CpG dinucleotides is an important component of epigenetic gene expression regulation [55] that causes the modulation of protein-DNA interactions [56, 57]. Aberrant methylation of CpG islands (CGT) takes place in the exonic and promoter regions [58, 59] and changes in gene expression associated with tumorigenesis. Hypermethylation of tumorigenetic genes has negative impact (regulation) over tumor suppressor genes (TSGs), DNA repair genes, and antiangiogenic genes, and

A variety of fishes have been used as model to study tumors induced by environmental carcinogens. Among all the zebrafish proved best for investigating embryogenesis, organogenesis, and impact of environmental carcinogen for the development of cancer [64]. Chemically induced tumors in zebrafish and humans are histopathologically similar [43, 65], and orthologous oncogenes and tumor suppressor genes (TSGs) have been identified in fishes and humans [65]. Hepatic gene expression in humans and zebrafish has revealed conservation of gene expression profiles at different stages of tumor aggressiveness between these two phylogeneti-

Xenotransplantation represents another novel way to induce tumor in zebrafish. The most important feature of xenotransplantation is that tumor cells can be stained/marked by fluorescent stain that distinguishes transplanted cells from normal cells and helps in clear observation of developmental process of the tumor [68]. Several other types of tumor, such as pancreatic cancer, lung cancer, ovarian carcinoma, breast cancer, prostate cancer, retinoblastoma, leukemia, etc., have also

Angiogenesis is the most important factor in tumor growth and subsequent metastasis. The importance of angiogenesis has been discussed well in the previous section on development. The vascular network is helpful to transport oxygen and nutrients to the cells; likewise tumor cells also get the supply of all these materials. Because of this reason, the development and the capability of the formation of blood vessels within the tumor determine the malignancy of the cancer as well as influence the therapeutic effects and prognosis. The endothelial cells of the vascular system can be stained by fluorescent dye/protein that helps to visualize the neovascularization of tiny tumor at the earliest stage, and metastasizing tumor cells can be tracked explicitly at cellular level [7]. The vascular system of tumor has always been the target of antitumor therapies; it is evident from research and clinical observations that if angiogenesis inhibitors are used in combination with chemotherapy, it

tions between transplanted tumor cell and vasculature of host.

it is a common quality of neoplastic cells [55, 60–63].

possible to identify the interacting oncogenes via suppressor and enhancer screens which cause the formation of specific type of tumor. The mammalian tumor cells can be transplanted into the zebrafish, dispensing a novel way to study the interactions between transplanted tumor cell and vasculature of host.

#### **3.2 Tumorigenesis**

*Current Trends in Cancer Management*

The zebrafish exhibits remarkable capacity of regeneration even in adult stages. The caudal fin especially provides an ideal tissue for vascular regeneration studies due to its simple and fine architecture and relative transparency [39]. After successive amputations the full regeneration of the caudal fin used to take place within couple of weeks [40]. The regenerating vessels in the regenerating caudal fin originate from vein-derived cells that have angiogenic potential [41]. These cells migrate individually or in groups and assemble into the vessel in response to

The zebrafish as an alternative model for angiogenesis and regeneration studies provides the relevance of in vivo assays with simplicity and versatility of in vitro assays. In larvae, access to developing vasculature through fluorophore-tagged strains and small size of zebrafish makes the use of high-throughput strategies possible. In adults, the caudal fin is equally convenient as a model tissue as regenerating vessels can be observed at all stages, and the animals (zebrafish) are suitable for

Cancer is a cursed reality for millions of humans worldwide and in fact for all vertebrates. The invertebrates such as flies and nematodes also can develop anomalies in cell proliferation. Clinically and pathologically this dreaded disease is present almost exclusively in all vertebrates, from fish to humans. To understand better the formation, growth, and spread of malignant tumors, vertebrate models are imperative. Being a vertebrate the zebrafish is an ideal model to study cancer, though humans and fishes are separated from their common ancestry but biology of the cancer in both groups of organisms is the same [43]. Because of the variety of benefits to use zebrafish as a model organism which are mentioned in "Introduction," the zebrafish is adroitly exploited to carcinogenic treatment, transplantation of

**3.1 Zebrafish as a model for carcinogen effects and development of cancer** 

Fishes are exposed to many waterborne carcinogens in the wild that lead to the development of a variety of benign and malignant tumors in teleosts, and these tumors have similar histology as in humans [45, 46]. As like humans, cancer is a genetic disease in fishes as shown by melanomas which develop in *Xiphophorus* hybrids [47]. Choosing zebrafish for modeling cancer studies has many advantages. Highly conserved cancer pathways can be screened genetically using zebrafish. Primarily cancer is a disease of adults, but through mutagenesis screens, cell cycle phenotype could be examined in rapidly developing transparent embryos of the zebrafish. The genes regulating cell cycle, cell proliferation, and apoptosis have already been screened in yeast, *Drosophila* and *C. elegans*, in the similar way gene functions for these biological pathways can be screened in zebrafish to understand the events that lead to the development of cancer in any

By inducing different gene mutations or stimulating signaling pathways through chemicals, the tumors can be induced in different organs of the zebrafish like the pancreas, liver, GI tract, vasculature, muscles, skin, and testes [46, 48–51]. It is

**2.3 Regeneration**

chemokine signaling [42].

experimental drug manipulations [31].

**3. Zebrafish as a cancer model system**

mammalian tumor cells, and transgenic regulations [44].

**6**

**studies**

vertebrate species [43].

Tumorigenesis is a multistep process induced by chemical carcinogen [52], with accumulation of both epigenetic aberrations and mutations in regulatory regions of genes and disorder of signaling pathways [53, 54]. Methylation of DNA at CpG dinucleotides is an important component of epigenetic gene expression regulation [55] that causes the modulation of protein-DNA interactions [56, 57]. Aberrant methylation of CpG islands (CGT) takes place in the exonic and promoter regions [58, 59] and changes in gene expression associated with tumorigenesis. Hypermethylation of tumorigenetic genes has negative impact (regulation) over tumor suppressor genes (TSGs), DNA repair genes, and antiangiogenic genes, and it is a common quality of neoplastic cells [55, 60–63].

A variety of fishes have been used as model to study tumors induced by environmental carcinogens. Among all the zebrafish proved best for investigating embryogenesis, organogenesis, and impact of environmental carcinogen for the development of cancer [64]. Chemically induced tumors in zebrafish and humans are histopathologically similar [43, 65], and orthologous oncogenes and tumor suppressor genes (TSGs) have been identified in fishes and humans [65]. Hepatic gene expression in humans and zebrafish has revealed conservation of gene expression profiles at different stages of tumor aggressiveness between these two phylogenetically distant species [66, 67].

#### **3.3 Xenotransplantation**

Xenotransplantation represents another novel way to induce tumor in zebrafish. The most important feature of xenotransplantation is that tumor cells can be stained/marked by fluorescent stain that distinguishes transplanted cells from normal cells and helps in clear observation of developmental process of the tumor [68]. Several other types of tumor, such as pancreatic cancer, lung cancer, ovarian carcinoma, breast cancer, prostate cancer, retinoblastoma, leukemia, etc., have also been transplanted in the zebrafish [7].

#### **3.4 Angiogenesis**

Angiogenesis is the most important factor in tumor growth and subsequent metastasis. The importance of angiogenesis has been discussed well in the previous section on development. The vascular network is helpful to transport oxygen and nutrients to the cells; likewise tumor cells also get the supply of all these materials. Because of this reason, the development and the capability of the formation of blood vessels within the tumor determine the malignancy of the cancer as well as influence the therapeutic effects and prognosis. The endothelial cells of the vascular system can be stained by fluorescent dye/protein that helps to visualize the neovascularization of tiny tumor at the earliest stage, and metastasizing tumor cells can be tracked explicitly at cellular level [7]. The vascular system of tumor has always been the target of antitumor therapies; it is evident from research and clinical observations that if angiogenesis inhibitors are used in combination with chemotherapy, it can improve the outcome in cancer patients [69].

#### **3.5 Skin cancer**

Skin or dermal cancers represent the most common type of cutaneous malignancy globally, which includes melanoma and carcinoma of squamous cells [70]. Melanoma is the most pernicious form of skin cancer among all types of skin cancers and has mortality rate over 80% [71, 72]. Melanoma usually develops in the pigmented epidermal cells (melanocytes), which are responsible for the production of melanin. In the beginning of melanoma, it is restricted to the epidermis because of the radial growth phase (RGP) of melanoma, and it can be removed by surgical excision [73]. In later stages of tumor progression, the melanoma cells invade the subcutaneous tissues due to vertical growth phase (VGP) of melanoma and eventually lead toward the metastatic phase. At this stage, very limited therapeutic options are available, and melanoma frequently deteriorates and becomes untreatable [73, 74].

Cutaneous squamous cell carcinoma (cSCC) mostly develops due to UV radiation exposure of epidermal cells, namely, keratinocytes, in which uncontrolled proliferation starts [75]. cSCC accounts for the most frequent type of non-melanoma cutaneous cancer and constitutes about 20% of all skin malignancies [75, 76].

SCCs are curable in situ by surgical excision. Metastatic SCCs are responsible for majority of deaths due to non-melanoma skin cancer [70]. Head and neck squamous cell carcinoma (HNSCC) develops in various places such as the oropharynx and laryngopharynx which is very common worldwide [77]. Especially oral squamous cell carcinoma (OSCC) accounts for about 24% of HNSCC with a mortality rate of 2 million deaths every year [76, 78].

Zebrafish is a powerful in vivo tool to study pathologies and treatment for skin cancer (melanoma and SCC). The zebrafish can be used to study melanoma development, progression, drug screening, and treatment. The zebrafish model has been exploited recently to recognize the key molecules which are responsible for the development of cutaneous squamous cell carcinoma (cSCC) and head and neck squamous cell carcinoma (HNSCC) [72] as well as for SCC target therapies [79].

#### **3.6 Tumor metastasis**

Metastasis is a multistep and complex process in which tumor cells penetrate in the vascular system and spread deep in parenchymatous tissues [80]. For better therapeutic practices like development of antitumor drugs and advancements of clinical treatments, the insight into mechanism of tumor metastasis is very helpful. Because of many significant disadvantages in the previous studies using in vivo mouse model, the metastasis process cannot be abstracted properly, but zebrafish cancer model has overcome the drawback of previous models and has shown exceptional strengths. The adaptive immune system in larvae of zebrafish usually develops after 14 DPF, which provides very conducive environment for survival of transplanted cancer cells and metastasis [81], and the process of tumor metastasis can be observed through the transparent body of zebrafish under microscope. To better understand the process of metastasis, the transplanted tumor cells can be stained/treated by dye like CM-Dil or may be labeled by red fluorescent protein (RFP) [82]. Mammalian tumor cells treated with red fluorescent protein when injected into transgenic zebrafish, the process of tumor cell metastasis and angiogenesis can be viewed well after 48 h of transplantation [83]. By using zebrafish, the suppressing or promoting factors for metastasis can be identified. In RFP treated U87 glioma stem cells (GSCs), when transplanted into the yolk sac of the zebrafish embryo, the various invasive stages of GSCs like approaching, cluster formation, invasion, migration, and transmigration can be observed clearly at 48 h postinjection [83].

**9**

**4.3 Angiogenesis**

*Zebrafish (Danio rerio) as a Model Organism DOI: http://dx.doi.org/10.5772/intechopen.81517*

**4. Toxicology and drug discovery**

MPA, fluorouracil, etc. [85–90].

**4.2 Zebrafish and drug discovery**

**4.1 Drug toxicity**

function assays and assessment of drug effect [84].

As discussed previously in Section 1, because of many advantages, the zebrafish has recently emerged as a prominent model for toxicological studies and drug discovery. The effects of drugs on growth and development can be examined visually through length and shape of the zebrafish body as well as the morphology of internal organs such as the brain, liver, cardiovascular system, pancreas, intestine, kidney, notochord, etc. The zebrafish model also has been used to know the organ

Zebrafish embryos are used as predictive model to assess the toxicity in mammals. The lethal concentration (LC50) of different chemicals has been determined in embryos of zebrafish and has been compared with the mammalian LC50, and it has been found that median lethal dose of zebrafish is lower than mammals [84]. The effects of drugs on specific organs have also been studied, and it has been found that organ toxicity is similar in both zebrafish and mammals. The drugs that were used to evaluate the organ toxicity were gentamicin, cisplatin, vinblastine, quinine, neomycin, doxorubicin, dexamethasone, cyclosporin A, caffeine, camptothecin,

In drug development, the toxicity plays a major role. Due to the toxicity problem, many new drugs have been declined by the FDA. The evaluation of toxicity of drug is very essential to know the end points of toxicity, dose-response relationships, and mechanism of toxicity and also to determine the toxicodynamics of the drug [91]. The zebrafish is acquiring the reputation rapidly as a promising model animal to study drug and chemical toxicology [92, 93]. The toxicity of some of the important drugs has been examined using the zebrafish model, for instance, Amanuma et al. [94] developed a test in which susceptible zebrafish was used to detect small molecule-induced mutagenesis. The embryos of zebrafish were utilized to compare the developmental toxicity resulting from the exposure to ethanol or acetaldehyde [95]. Toxicity of antirheumatic drug like diclofenac was evaluated by using zebrafish. Now, zebrafish has got the status of a successful animal model to study drug

The zebrafish model has been used potentially in drug discovery and to know the effects of neurotoxic, ototoxic, and neuroprotectant drugs. The process of drug discovery is divided into four main components: screening of lead compounds, target identification, target validation, and assay development [96]. The process of target identification involves the recognition of target gene or protein which when modulated by a drug can have positive effects on the progression of disease. After identification of possible target, the validation process of target begins through determination of protein function and assessment of the druggability of the target [84, 97–99]. Zebrafish has great role in each of these areas of drug discovery.

The angiogenesis has already been discussed earlier in detail in previous sections on development and cancer. The impact of various proangiogenic compounds like simvastatine or penicillamine20 or antiangiogenic compounds like vandetanib

toxicity and toxicology caused by environmental contaminants [91].

*Current Trends in Cancer Management*

million deaths every year [76, 78].

can be observed clearly at 48 h postinjection [83].

**3.6 Tumor metastasis**

Skin or dermal cancers represent the most common type of cutaneous malignancy globally, which includes melanoma and carcinoma of squamous cells [70]. Melanoma is the most pernicious form of skin cancer among all types of skin cancers and has mortality rate over 80% [71, 72]. Melanoma usually develops in the pigmented epidermal cells (melanocytes), which are responsible for the production of melanin. In the beginning of melanoma, it is restricted to the epidermis because of the radial growth phase (RGP) of melanoma, and it can be removed by surgical excision [73]. In later stages of tumor progression, the melanoma cells invade the subcutaneous tissues due to vertical growth phase (VGP) of melanoma and eventually lead toward the metastatic phase. At this stage, very limited therapeutic options are available, and

Cutaneous squamous cell carcinoma (cSCC) mostly develops due to UV radiation exposure of epidermal cells, namely, keratinocytes, in which uncontrolled proliferation starts [75]. cSCC accounts for the most frequent type of non-melanoma cutaneous cancer and constitutes about 20% of all skin malignancies [75, 76].

SCCs are curable in situ by surgical excision. Metastatic SCCs are responsible for majority of deaths due to non-melanoma skin cancer [70]. Head and neck squamous cell carcinoma (HNSCC) develops in various places such as the oropharynx and laryngopharynx which is very common worldwide [77]. Especially oral squamous cell carcinoma (OSCC) accounts for about 24% of HNSCC with a mortality rate of 2

Zebrafish is a powerful in vivo tool to study pathologies and treatment for skin cancer (melanoma and SCC). The zebrafish can be used to study melanoma development, progression, drug screening, and treatment. The zebrafish model has been exploited recently to recognize the key molecules which are responsible for the development of cutaneous squamous cell carcinoma (cSCC) and head and neck squamous cell carcinoma (HNSCC) [72] as well as for SCC target therapies [79].

Metastasis is a multistep and complex process in which tumor cells penetrate in the vascular system and spread deep in parenchymatous tissues [80]. For better therapeutic practices like development of antitumor drugs and advancements of clinical treatments, the insight into mechanism of tumor metastasis is very helpful. Because of many significant disadvantages in the previous studies using in vivo mouse model, the metastasis process cannot be abstracted properly, but zebrafish cancer model has overcome the drawback of previous models and has shown exceptional strengths. The adaptive immune system in larvae of zebrafish usually develops after 14 DPF, which provides very conducive environment for survival of transplanted cancer cells and metastasis [81], and the process of tumor metastasis can be observed through the transparent body of zebrafish under microscope. To better understand the process of metastasis, the transplanted tumor cells can be stained/treated by dye like CM-Dil or may be labeled by red fluorescent protein (RFP) [82]. Mammalian tumor cells treated with red fluorescent protein when injected into transgenic zebrafish, the process of tumor cell metastasis and angiogenesis can be viewed well after 48 h of transplantation [83]. By using zebrafish, the suppressing or promoting factors for metastasis can be identified. In RFP treated U87 glioma stem cells (GSCs), when transplanted into the yolk sac of the zebrafish embryo, the various invasive stages of GSCs like approaching, cluster formation, invasion, migration, and transmigration

melanoma frequently deteriorates and becomes untreatable [73, 74].

**3.5 Skin cancer**

**8**

#### **4. Toxicology and drug discovery**

As discussed previously in Section 1, because of many advantages, the zebrafish has recently emerged as a prominent model for toxicological studies and drug discovery. The effects of drugs on growth and development can be examined visually through length and shape of the zebrafish body as well as the morphology of internal organs such as the brain, liver, cardiovascular system, pancreas, intestine, kidney, notochord, etc. The zebrafish model also has been used to know the organ function assays and assessment of drug effect [84].

Zebrafish embryos are used as predictive model to assess the toxicity in mammals. The lethal concentration (LC50) of different chemicals has been determined in embryos of zebrafish and has been compared with the mammalian LC50, and it has been found that median lethal dose of zebrafish is lower than mammals [84]. The effects of drugs on specific organs have also been studied, and it has been found that organ toxicity is similar in both zebrafish and mammals. The drugs that were used to evaluate the organ toxicity were gentamicin, cisplatin, vinblastine, quinine, neomycin, doxorubicin, dexamethasone, cyclosporin A, caffeine, camptothecin, MPA, fluorouracil, etc. [85–90].

#### **4.1 Drug toxicity**

In drug development, the toxicity plays a major role. Due to the toxicity problem, many new drugs have been declined by the FDA. The evaluation of toxicity of drug is very essential to know the end points of toxicity, dose-response relationships, and mechanism of toxicity and also to determine the toxicodynamics of the drug [91].

The zebrafish is acquiring the reputation rapidly as a promising model animal to study drug and chemical toxicology [92, 93]. The toxicity of some of the important drugs has been examined using the zebrafish model, for instance, Amanuma et al. [94] developed a test in which susceptible zebrafish was used to detect small molecule-induced mutagenesis. The embryos of zebrafish were utilized to compare the developmental toxicity resulting from the exposure to ethanol or acetaldehyde [95]. Toxicity of antirheumatic drug like diclofenac was evaluated by using zebrafish. Now, zebrafish has got the status of a successful animal model to study drug toxicity and toxicology caused by environmental contaminants [91].

#### **4.2 Zebrafish and drug discovery**

The zebrafish model has been used potentially in drug discovery and to know the effects of neurotoxic, ototoxic, and neuroprotectant drugs. The process of drug discovery is divided into four main components: screening of lead compounds, target identification, target validation, and assay development [96]. The process of target identification involves the recognition of target gene or protein which when modulated by a drug can have positive effects on the progression of disease. After identification of possible target, the validation process of target begins through determination of protein function and assessment of the druggability of the target [84, 97–99]. Zebrafish has great role in each of these areas of drug discovery.

#### **4.3 Angiogenesis**

The angiogenesis has already been discussed earlier in detail in previous sections on development and cancer. The impact of various proangiogenic compounds like simvastatine or penicillamine20 or antiangiogenic compounds like vandetanib

or PTK787 can be assessed well and visualized through the development of the vascular system in transparent zebrafish embryo [84].

#### **4.4 Cardiotoxicity**

In drug development, cardiotoxicity is one of the major concerns. Through the transparent zebrafish embryo, various cardiac functions like heart rate, rhythm, contraction, circulation, etc. can be assessed directly. It has been demonstrated well that toxic effects of ten cardiotoxic agents in zebrafish embryos have similar impact as in humans [100]. Treatment with terfenadine and clomipramine caused severe impairment of cardiac functions, edema, hemorrhage, arrested heartbeat, and even death. These results in zebrafish exhibit similarities with humans [101]. Another group of researchers proposed to use a transgenic model for high-throughput testing of small molecules that modulate the heart rate of the zebrafish embryo [102]. Thus, zebrafish is a suitable model for preliminary screening of molecules which have potential therapeutic or toxic effects.

#### **5. Human disease and zebrafish**

Most of the tissues and organs found in humans and zebrafish are the same except lungs and prostrate and mammary glands. The cloning of mutated genes screened for specific phenotypes in zebrafish has similarities in humans and thus serves as model for human disease and to study underlying mechanisms. The first human disease identified using zebrafish was a blood disorder involving specific defect in hemoglobin production through ALAS2 mutated gene [103].

Many other mutants which show phenotypic similarities to human disease have been screened and identified. These include neurological disorders [104], hematological disorder [105, 106], cardiovascular diseases [107], muscle disease [108] and cancers [109, 110], Parkinson's disease [111], anxiety, and posttraumatic stress disorder [112].

#### **6. Zebrafish as a model organism for aquaculture species**

Among different fish species of interest to aquaculture, zebrafish is genetically more tractable. The zebrafish model is used commercially in many areas of aquaculture such as in the identification of genes involved in the development of the muscles, bones, and fats, the metabolism of nutrients, disease, and stress pathways and also behavioral traits. The drugs which affect the physiology of the fishes can be tested easily in zebrafish especially their effect on a range of alleles to assess their genetic property [113]. Many researches have been done regarding the improvement of diet and their husbandry to improve the growth rate and reduce stress and disease in many fish species like gilthead seabream, seabass, rainbow trout, Atlantic salmon, tilapia, catfish, cod, etc. [10]. The zebrafish disease models are being used in various infections of aquaculture, for instance, tuberculosis and streptococcal and salmonella infections [114].

#### **7. Conclusion**

Zebrafish is a successful and versatile animal model system, offering a tool to model gene function, development of various organ systems, cancer studies,

**11**

provided the original work is properly cited.

\*Address all correspondence to: insectqh11@gmail.com

*Zebrafish (Danio rerio) as a Model Organism DOI: http://dx.doi.org/10.5772/intechopen.81517*

**Conflict of interest**

**Author details**

Farmanur Rahman Khan1

Al Qassim, Buraidah, Saudi Arabia

toxicology, drug discovery, human disease and disorders and also in aquaculture, etc. because low cost and easy maintenance, transparent embryo, easy manipulation, high fecundity, and rapid embryonic development favor the zebrafish as an attractive model for in vivo assays with simplicity and versatility of in vitro assays over mammalian models which lack all of these benefits. The future of zebrafish as model organism is very bright. In coming years, an increased number of reports are

expected on the application of zebrafish as an effective bioindicator.

The author declares that there is no conflict of interest.

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

\* and Saleh Sulaiman Alhewairini2

1 Department of Biology, Deanship of Educational Services, Qassim University,

2 Department of Plant Production and Protection, College of Agriculture and Veterinary Medicine, Qassim University, Al Qassim, Buraidah, Saudi Arabia

*Zebrafish (Danio rerio) as a Model Organism DOI: http://dx.doi.org/10.5772/intechopen.81517*

toxicology, drug discovery, human disease and disorders and also in aquaculture, etc. because low cost and easy maintenance, transparent embryo, easy manipulation, high fecundity, and rapid embryonic development favor the zebrafish as an attractive model for in vivo assays with simplicity and versatility of in vitro assays over mammalian models which lack all of these benefits. The future of zebrafish as model organism is very bright. In coming years, an increased number of reports are expected on the application of zebrafish as an effective bioindicator.

### **Conflict of interest**

*Current Trends in Cancer Management*

**4.4 Cardiotoxicity**

disorder [112].

and salmonella infections [114].

vascular system in transparent zebrafish embryo [84].

have potential therapeutic or toxic effects.

**5. Human disease and zebrafish**

or PTK787 can be assessed well and visualized through the development of the

In drug development, cardiotoxicity is one of the major concerns. Through the transparent zebrafish embryo, various cardiac functions like heart rate, rhythm, contraction, circulation, etc. can be assessed directly. It has been demonstrated well that toxic effects of ten cardiotoxic agents in zebrafish embryos have similar impact as in humans [100]. Treatment with terfenadine and clomipramine caused severe impairment of cardiac functions, edema, hemorrhage, arrested heartbeat, and even death. These results in zebrafish exhibit similarities with humans [101]. Another group of researchers proposed to use a transgenic model for high-throughput testing of small molecules that modulate the heart rate of the zebrafish embryo [102]. Thus, zebrafish is a suitable model for preliminary screening of molecules which

Most of the tissues and organs found in humans and zebrafish are the same except lungs and prostrate and mammary glands. The cloning of mutated genes screened for specific phenotypes in zebrafish has similarities in humans and thus serves as model for human disease and to study underlying mechanisms. The first human disease identified using zebrafish was a blood disorder involving specific

Many other mutants which show phenotypic similarities to human disease have been screened and identified. These include neurological disorders [104], hematological disorder [105, 106], cardiovascular diseases [107], muscle disease [108] and cancers [109, 110], Parkinson's disease [111], anxiety, and posttraumatic stress

Among different fish species of interest to aquaculture, zebrafish is genetically more tractable. The zebrafish model is used commercially in many areas of aquaculture such as in the identification of genes involved in the development of the muscles, bones, and fats, the metabolism of nutrients, disease, and stress pathways and also behavioral traits. The drugs which affect the physiology of the fishes can be tested easily in zebrafish especially their effect on a range of alleles to assess their genetic property [113]. Many researches have been done regarding the improvement of diet and their husbandry to improve the growth rate and reduce stress and disease in many fish species like gilthead seabream, seabass, rainbow trout, Atlantic salmon, tilapia, catfish, cod, etc. [10]. The zebrafish disease models are being used in various infections of aquaculture, for instance, tuberculosis and streptococcal

Zebrafish is a successful and versatile animal model system, offering a tool to model gene function, development of various organ systems, cancer studies,

defect in hemoglobin production through ALAS2 mutated gene [103].

**6. Zebrafish as a model organism for aquaculture species**

**10**

**7. Conclusion**

The author declares that there is no conflict of interest.

### **Author details**

Farmanur Rahman Khan1 \* and Saleh Sulaiman Alhewairini2

1 Department of Biology, Deanship of Educational Services, Qassim University, Al Qassim, Buraidah, Saudi Arabia

2 Department of Plant Production and Protection, College of Agriculture and Veterinary Medicine, Qassim University, Al Qassim, Buraidah, Saudi Arabia

\*Address all correspondence to: insectqh11@gmail.com

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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[16] Walker C, Streisinger G. Induction of mutations by gamma-rays in pregonial germ cells of zebrafish embryos. Genetics. 1983;**103**:125-136

[17] Grunwald DJ, Eisen JS. Headwaters of the zebrafish—Emergence of a new model vertebrate. Nature Reviews. Genetics. 2002;**3**:717-724

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**13**

*Zebrafish (Danio rerio) as a Model Organism DOI: http://dx.doi.org/10.5772/intechopen.81517*

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[31] Chávez MN, Aedo G, Fierro FA, Allende ML, Egaña JT. Zebrafish as an emerging model organism to study angiogenesis in development and regeneration. Frontiers in Physiology. 2016;**7**:56. DOI: 10.3389/

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Mawdsley DJ, White SJ, Shin J, Appel B, et al. Formation of the digestive system in zebrafish: III. Intestinal epithelium morphogenesis. Developmental Biology.

[22] Wallace AS, Burns AJ. Development of the enteric nervous system, smooth muscle and interstitial cells of Cajal in the human gastrointestinal tract. Cell and Tissue Research. 2005;**319**:367-382

[23] Muncan V, Faro A, Haramis AP, Hurlstone AF, Wienholds E, van Es J, et al. T-cell factor 4 (Tcf7l2) maintains proliferative compartments in zebrafish intestine. EMBO Reports.

[24] Kelsh RN, Eisen JS. The zebrafish colourless gene regulates development of non-ectomesenchymal neural crest derivatives. Development.

[25] Elworthy S, Pinto JP, Pettifer A, Cancela ML, Kelsh RN. Phox2b function in the enteric nervous system is conserved in zebrafish and is sox10-dependent. Mechanisms of Development. 2005;**122**:659-669

[26] Furness JB. The Enteric Nervous System. Oxford: John Wiley and Sons

[27] Olden T, Akhtar T, Beckman SA, Wallace KN. Differentiation of the zebrafish enteric nervous system and intestinal smooth muscle. Genesis.

[28] Shepherd IT, Pietsch J, Elworthy S, Kelsh RN, Raible DW. Roles for GFRalpha1 receptors in zebrafish

*Zebrafish (Danio rerio) as a Model Organism DOI: http://dx.doi.org/10.5772/intechopen.81517*

[20] Holmberg A, Olsson C, Hennig GW. TTX-sensitive and TTX-insensitive control of spontaneous gut motility in the developing zebrafish (*Danio rerio*) larvae. The Journal of Experimental Biology. 2007;**210**:1084-1091

[21] Ng AN, de Jong-Curtain TA, Mawdsley DJ, White SJ, Shin J, Appel B, et al. Formation of the digestive system in zebrafish: III. Intestinal epithelium morphogenesis. Developmental Biology. 2005;**286**:114-135

[22] Wallace AS, Burns AJ. Development of the enteric nervous system, smooth muscle and interstitial cells of Cajal in the human gastrointestinal tract. Cell and Tissue Research. 2005;**319**:367-382

[23] Muncan V, Faro A, Haramis AP, Hurlstone AF, Wienholds E, van Es J, et al. T-cell factor 4 (Tcf7l2) maintains proliferative compartments in zebrafish intestine. EMBO Reports. 2007;**8**:966-973

[24] Kelsh RN, Eisen JS. The zebrafish colourless gene regulates development of non-ectomesenchymal neural crest derivatives. Development. 2000;**127**:515-525

[25] Elworthy S, Pinto JP, Pettifer A, Cancela ML, Kelsh RN. Phox2b function in the enteric nervous system is conserved in zebrafish and is sox10-dependent. Mechanisms of Development. 2005;**122**:659-669

[26] Furness JB. The Enteric Nervous System. Oxford: John Wiley and Sons Ltd.; 2006

[27] Olden T, Akhtar T, Beckman SA, Wallace KN. Differentiation of the zebrafish enteric nervous system and intestinal smooth muscle. Genesis. 2008;**46**:484-498

[28] Shepherd IT, Pietsch J, Elworthy S, Kelsh RN, Raible DW. Roles for GFRalpha1 receptors in zebrafish

enteric nervous system development. Development. 2004;**131**:241-249

[29] Tiffany AH, Shepherd IT, Burns AJ. Enteric nervous system development in avian and zebrafish models. Developmental Biology. 2016;**417**(2):129-138. DOI: 10.1016/j. ydbio.2016.05.017

[30] Pandya N, Dhalla N, Santani D. Angiogenesis—A new target for future therapy. Vascular Pharmacology. 2006;**44**:265-274. DOI: 10.1016/j. vph.2006.01.005

[31] Chávez MN, Aedo G, Fierro FA, Allende ML, Egaña JT. Zebrafish as an emerging model organism to study angiogenesis in development and regeneration. Frontiers in Physiology. 2016;**7**:56. DOI: 10.3389/ fphys.2016.00056

[32] Isogai S, Horiguchi M, Weinstein BM. The vascular anatomy of the developing zebrafish: An atlas of embryonic and early larval development. Developmental Biology. 2001;**230**:278-301. DOI: 10.1006/ dbio.2000.9995

[33] Ellertsdóttir E, Lenard A, Blum Y, Krudewig A, Herwig L, Affolter M. Vascular morphogenesis in the zebrafish embryo. Developmental Biology. 2001;**341**:56-65. DOI: 10.1016/j. ydbio.2009.10.035

[34] Küchler AM, Gjini E, Peterson-Maduro J, Cancilla B, Wolburg H, Schulte-Merker S. Development of the zebrafish lymphatic system requires VEGFC signaling. Current Biology. 2006;**16**:1244-1248. DOI: 10.1016/j. cub.2006.05.026

[35] Coffindaffer-Wilson M, Craig MP, Hove JR. Determination of lymphatic vascular identity and developmental timecourse in zebrafish (*Danio rerio*). Lymphology. 2011;**44**:1-12

**12**

*Current Trends in Cancer Management*

[1] Gore AV, Monzo K, Cha YR, Pan W, Weinstein BM. Vascular development in the zebrafish. Cold Spring Harbor Perspectives in Medicine. [10] Carpio Y, Estrada MP. Zebrafish as a genetic model organism. Biotecnologia

[11] Pelegri F. Mutagenesis. In: Nusslein-Volhard C, Dahm R, editors. Zebrafish: A Practical Approach. Oxford: Oxford University Press; 2002. pp. 145-174

[12] Lele Z, Krone PH. The zebrafish as a model system in developmental,

[13] Kimmel CB. Genetics and early development of zebrafish. Trends in

[14] Chakrabarti S, Streisinger G, Singer F, Walker C. Frequency of gamma-ray induced specific locus and recessive lethal mutations in mature germ cells of the zebrafish, *Brachydanio rerio*.

[15] Streisinger G, Singer F, Walker C, Knauber D, Dower N. Segregation analyses and gene-centromere distances in zebrafish. Genetics. 1986;**112**:311-319

[16] Walker C, Streisinger G. Induction

[17] Grunwald DJ, Eisen JS. Headwaters of the zebrafish—Emergence of a new model vertebrate. Nature Reviews.

[18] Nüsslein-Volhard C, Wieschaus E. Mutations affecting segment number and polarity in Drosophila. Nature.

[19] Wallace KN, Akhter S, Smith EM, Lorent K, Pack M. Intestinal growth and differentiation in zebrafish. Mechanisms of Development.

of mutations by gamma-rays in pregonial germ cells of zebrafish embryos. Genetics. 1983;**103**:125-136

Genetics. 2002;**3**:717-724

1980;**287**:795-801

2005;**122**:157-173

toxicological and transgenic research. Biotechnology Advances.

Genetics. 1989;**5**:283-288

Genetics. 1983;**103**:109-123

1996;**14**(1):57-72

Aplicada. 2006;**23**:265-270

[2] Kanungo J, Cuevas E, Ali SF, Paule MG. Zebrafish model in drug safety assessment. Current Pharmaceutical Design. 2014;**20**(34):5416-5429

[3] Kalueff AV, Stewart AM, Gerlai R. Zebrafish as an emerging model for studying complex brain disorders. Trends in Pharmacological Sciences.

[4] Guyon JR, Steffen LS, Howell MH, Pusack TJ, Lawrence C, Kunkel LM. Modeling human muscle disease in zebrafish. Biochimica et Biophysica

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[6] Lieschke GJ, Oates AC, Crowhurst MO, Ward AC, Layton JE. Morphologic and functional characterization of granulocytes and macrophages in embryonic and adult zebrafish. Blood.

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[104] Gama Sosa MA, De Gasperi R, Elder GA. Modeling human neurodegenerative diseases in transgenic systems. Human Genetics. 2012;**131**:535-563

[105] Berman J, Payne E, Hall C. The zebrafish as a tool to study hematopoiesis, human blood diseases, and immune function. Advances in Hematology. 2012;**2012**:2. Article ID 425345. https://doi. org/10.1155/2012/425345

[106] Brownlie A, Donovan A, Pratt SJ, Paw BH, Oates AC, et al. Positional cloning of the zebrafish sauternes gene: A model for congenital sideroblastic anaemia. Nature Genetics. 1998;**20**:244-250

[107] Sehnert AJ, Huq A, Weinstein BM, Walker C, Fishman M, et al. Cardiac troponin T is essential in sarcomere assembly and cardiac contractility. Nature Genetics. 2002;**31**:106-110

[108] Lin YY. Muscle diseases in the zebrafish. Neuromuscular Disorders. 2012;**22**:673-684

[109] Liu S, Leach SD. Zebrafish models for cancer. Annual Review of Pathology. 2011;**6**:71-93

[110] Patton EE, Widlund HR, Kutok JL, Kopani KR, Amatruda JF, et al. BRAF mutations are sufficient to promote nevi formation and cooperate with p53 in the genesis of melanoma. Current Biology. 2005;**15**:249-254

[111] Sarath Babu N, Murthy CLN, Kakara S, Sharma R, Swamy B, Cherukuvada V, et al. 1-Methyl-4 phenyl-1, 2,3, 6-tetrahydropyridine induced Parkinson's disease in zebrafish. Proteomics. 2016;**16**:1407-1420

[112] Chakravarty S, Reddy BR, Sudhakar SR, Saxena S, Das T, Meghah V, et al. Chronic unpredictable stress (CUS)-induced anxiety and related

mood disorders in a zebrafish model: Altered brain proteome profile implicates mitochondrial dysfunction. PLoS One. 2013;**8**:e63302

[113] Dahm R, Geisler R. Learning from small fry: The zebrafish as a genetic model organism for aquaculture fish species. Marine Biotechnology. 2006;**8**(4):329-345

[114] Van der Sar AM, Appelmelk BJ, Vandenbroucke-Grauls CM, Bitter W. A star with stripes: Zebrafish as an infection model. Trends in Microbiology. 2004;**12**:451-457

**19**

**Chapter 2**

Cancer

*Aida Karachi*

**1. Introduction**

to stop immune evasion of cancer cells.

**Abstract**

Immunotherapy for Treatment of

Cancer is known to be second cause of death worldwide despite aggressive therapeutic measures such as surgical resection of tumors, radiation therapy, and chemotherapy. The failure of currently available therapeutics for cancers, has led to increasing interest in alternative approaches including immunotherapy. Immunotherapy for cancer treatment is enhancing immune responses to fight cancer cells. Monoclonal antibodies, immune checkpoint blockades, targeted therapy, adoptive cell therapy, CAR T cells, and cancer vaccines are the most current and efficient parts of immunotherapy armamentarium. Immunotherapy has tremendous success in the treatment of cancers and is considered as a standard care of treatment or recurrence preventive therapy for variety of cancers. In this chapter,

we discuss different types of immunotherapy for cancer treatment in detail.

**Keywords:** immunotherapy, immune checkpoint blockades, cancer vaccines,

Cancer is the second most common cause of death in the world that has threatened health for thousands of years. Several aggressive measures such as surgical resection of tumors, chemotherapy, and radiotherapy are used to cure cancers. Although these therapeutics can minimize and inhibit cancer cells proliferation and metastasis, they have not been able to effectively defeat cancers until now. The efficacy of conventional treatments for cancer management is limited by factors such as recurrence of tumors and severe toxicities induced by therapeutics. Immunotherapy has become a tempting approach a long time after William Coley described the first immune stimulation by live bacteria for the treatment of cancer in 1893 [1]. Immunotherapy harnesses patients' own immune system to kill cancer cells thereby reducing toxic effects of traditional chemotherapy and radiotherapy. Immune cells can identify cancer cells by recognizing tumor-associated antigens. The ability of cancer cells to escape from immune system has limited the efficacy of immunotherapy. Current novel approaches have been involved in immunotherapy

Immunotherapy includes several therapies such as monoclonal antibodies, tumor cell vaccines, immune cell vaccines, and adoptive cell therapy. Monoclonal antibodies, which block cytotoxic T lymphocyte-associated protein-4 (CTLA-4), programmed cell death-1 (PD-1), dendritic cell vaccines, and chimeric antigen receptor (CAR) T cells have shown a tremendous success in clinical trials for several

adoptive cell therapy, CAR T cell, personalized immunotherapy

#### **Chapter 2**

*Current Trends in Cancer Management*

[104] Gama Sosa MA, De Gasperi R,

mood disorders in a zebrafish model: Altered brain proteome profile

implicates mitochondrial dysfunction.

[113] Dahm R, Geisler R. Learning from small fry: The zebrafish as a genetic model organism for aquaculture fish species. Marine Biotechnology.

[114] Van der Sar AM, Appelmelk BJ, Vandenbroucke-Grauls CM, Bitter W. A star with stripes: Zebrafish as an infection model. Trends in Microbiology. 2004;**12**:451-457

PLoS One. 2013;**8**:e63302

2006;**8**(4):329-345

transgenic systems. Human Genetics.

[105] Berman J, Payne E, Hall C. The zebrafish as a tool to study hematopoiesis, human blood diseases, and immune function. Advances in Hematology. 2012;**2012**:2.

Article ID 425345. https://doi. org/10.1155/2012/425345

[106] Brownlie A, Donovan A, Pratt SJ, Paw BH, Oates AC, et al. Positional cloning of the zebrafish sauternes gene: A model for congenital sideroblastic anaemia. Nature Genetics.

[107] Sehnert AJ, Huq A, Weinstein BM, Walker C, Fishman M, et al. Cardiac troponin T is essential in sarcomere assembly and cardiac contractility. Nature Genetics. 2002;**31**:106-110

[108] Lin YY. Muscle diseases in the zebrafish. Neuromuscular Disorders.

[109] Liu S, Leach SD. Zebrafish models for cancer. Annual Review of Pathology.

[110] Patton EE, Widlund HR, Kutok JL, Kopani KR, Amatruda JF, et al. BRAF mutations are sufficient to promote nevi formation and cooperate with p53 in the genesis of melanoma. Current Biology.

[111] Sarath Babu N, Murthy CLN, Kakara S, Sharma R, Swamy B, Cherukuvada V, et al. 1-Methyl-4 phenyl-1, 2,3, 6-tetrahydropyridine induced Parkinson's disease in zebrafish.

Proteomics. 2016;**16**:1407-1420

[112] Chakravarty S, Reddy BR,

Sudhakar SR, Saxena S, Das T, Meghah V, et al. Chronic unpredictable stress (CUS)-induced anxiety and related

Elder GA. Modeling human neurodegenerative diseases in

2012;**131**:535-563

1998;**20**:244-250

2012;**22**:673-684

2011;**6**:71-93

2005;**15**:249-254

**18**

## Immunotherapy for Treatment of Cancer

*Aida Karachi*

#### **Abstract**

Cancer is known to be second cause of death worldwide despite aggressive therapeutic measures such as surgical resection of tumors, radiation therapy, and chemotherapy. The failure of currently available therapeutics for cancers, has led to increasing interest in alternative approaches including immunotherapy. Immunotherapy for cancer treatment is enhancing immune responses to fight cancer cells. Monoclonal antibodies, immune checkpoint blockades, targeted therapy, adoptive cell therapy, CAR T cells, and cancer vaccines are the most current and efficient parts of immunotherapy armamentarium. Immunotherapy has tremendous success in the treatment of cancers and is considered as a standard care of treatment or recurrence preventive therapy for variety of cancers. In this chapter, we discuss different types of immunotherapy for cancer treatment in detail.

**Keywords:** immunotherapy, immune checkpoint blockades, cancer vaccines, adoptive cell therapy, CAR T cell, personalized immunotherapy

#### **1. Introduction**

Cancer is the second most common cause of death in the world that has threatened health for thousands of years. Several aggressive measures such as surgical resection of tumors, chemotherapy, and radiotherapy are used to cure cancers. Although these therapeutics can minimize and inhibit cancer cells proliferation and metastasis, they have not been able to effectively defeat cancers until now. The efficacy of conventional treatments for cancer management is limited by factors such as recurrence of tumors and severe toxicities induced by therapeutics. Immunotherapy has become a tempting approach a long time after William Coley described the first immune stimulation by live bacteria for the treatment of cancer in 1893 [1]. Immunotherapy harnesses patients' own immune system to kill cancer cells thereby reducing toxic effects of traditional chemotherapy and radiotherapy. Immune cells can identify cancer cells by recognizing tumor-associated antigens. The ability of cancer cells to escape from immune system has limited the efficacy of immunotherapy. Current novel approaches have been involved in immunotherapy to stop immune evasion of cancer cells.

Immunotherapy includes several therapies such as monoclonal antibodies, tumor cell vaccines, immune cell vaccines, and adoptive cell therapy. Monoclonal antibodies, which block cytotoxic T lymphocyte-associated protein-4 (CTLA-4), programmed cell death-1 (PD-1), dendritic cell vaccines, and chimeric antigen receptor (CAR) T cells have shown a tremendous success in clinical trials for several cancers. It is shown that immunotherapy has the potential to move to the front-line of therapeutic options in most cancers. Despite the benefits of immunotherapy, some treatments have severe side effects such as nausea, fever, and diarrhea [2]. The aim of this chapter is to study the concept of immunotherapy for cancer treatment and to provide a thorough review on immunotherapy's developments for both oncologists and cancer immunologists.

#### **2. Monoclonal antibodies**

One of the mechanisms of immune system to defeat pathogens or cancers is to identify foreign substances or malignancies and generate antibodies against them. These antibodies can recognize pathogens and cancer cells by the antigens expressed on their surface. Antibodies have the ability to attach to the specific antigens and destroy foreign particles or malignancies. In the laboratory, scientists can generate many copies of antibodies that are specific to certain antigens on cancer cells. These are known as monoclonal antibodies. In 1997, the first monoclonal antibody, rituximab, was approved for treatment of non-Hodgkin's lymphoma. Beneficial outcomes of rituximab treatment resulted in emergence and development of monoclonal antibodies as a therapeutic approach for various hematological and solid cancers [3]. The most important step in generating monoclonal antibodies for cancer treatment is identifying right antigens on cancer cells. High mutation capacity of cancer cells and existence of various antigens make this task challenging. So far, monoclonal antibodies therapy has been more beneficial against some cancers than others.

Monoclonal antibodies can defeat cancer in different ways. Some monoclonal antibodies can recognize antigens expressed by cancer cells and mark them as a target that should be destroyed by immune system. This monoclonal antibody treatment is also known as targeted therapy [4]. Some of monoclonal antibodies cause apoptosis in cancer cells by directly attaching to the cancer cells. Preventing cell proliferation, destroying cell membrane, delivering radiation or chemotherapy to cancer cells, and inhibiting blood vessel growth are other functions of monoclonal antibodies to stop cancer cells. Monoclonal antibodies can robust, mimic or maintain the immune system's response on cancer cells in different ways, and some particular monoclonal antibodies act by more than one function [3]. Monoclonal antibodies can be categorized to three groups such as naked monoclonal antibodies (**Table 1**), conjugated monoclonal antibodies (**Table 2**), and bispecific monoclonal antibodies. Naked monoclonal antibodies act by just a single function. This single function can either be directly affecting cancer cells or by improving immune system against cancer cells. Trastuzumab is an example of monoclonal antibodies with direct effect on cancer cells. Trastuzumab can identify and block HER2 antigen, which is highly expressed on breast and stomach cancer cells. HER2 antigen is responsible for growth and proliferation of cancer cells. By blocking HER2 antigens, cancer cells are not able to expand and proliferate and spread in the body [5]. Immune check point inhibitors are monoclonal antibodies which improve immune system function. This group of antibodies will be discussed in detail later on this chapter. Some monoclonal antibodies can trigger immune system by attaching to immune cells and activating immune cells to destroy cancer cells. Alemtuzumab, which is a monoclonal antibody to treat chronic lymphocytic leukemia, binds to CD25 marker on the surface of lymphocytes and attracts immune cells to destroy cancer cells [6]. Conjugated monoclonal antibodies, also known as tagged antibodies or loaded antibodies, are antibodies that are being used to deliver either chemotherapy drugs or radioactive particles to cancer cells. These monoclonal

**21**

**Table 1.**

*cancer therapy.*

antibodies reduce the toxic effects of systemic chemotherapy and radiotherapy by directly homing the toxic drugs to tumor microenvironment [7, 8]. Ibritumomab tiuxetan is a radio-immunotherapeutic drug which directly delivers radio isotopes to cancerous B cells in non-Hodgkin lymphoma. Ibritumomab tiuxetan is a radiolabeled monoclonal antibody against CD20 antigen, which is expressed on B cell surface. By attaching Ibritumomab tiuxetan to CD20 on the B cells and killing cancer cells, the drug is able to eliminate lymphoma [7]. Chemolabeled antibodies are monoclonal antibodies that are attached to chemotherapy drugs. Brentuximab

*Unconjugated monoclonal antibodies currently approved by the Food and Drug Administration (FDA) for* 

monoclonal antibody

kappa monoclonal antibody

**Target Type Approval** 

Rituximab CD20 Chimeric IgG1 1997 B cell non-Hodgkin

Trastuzumab EGF Humanized IgG1 1998 Breast cancer

Alemtuzumab CD52 Humanized IgG1 2001 B cell chronic

Cetuximab VEGFR Chimeric IgG1 2004 Merkel cell carcinoma Bevacizumab VEGF Humanized IgG1 2004 Colon cancer Panitumumab EGFR Human IgG2 2006 Colorectal Ca

hybrid

Ofatumumab CD20 Human IgG1 2009 B cell chronic

Ipilimumab CTLA-4 Human IgG1 2011 Melanoma Brentuximab Vedotin CD30 2011 Hodgkin lymphoma Pertuzumab HER2 Humanized IgG1 2012 Breast cancer

Obinutuzumab CD20 2013 B cell chronic

Denosumab Human IgG2 2013 Osteoclastoma Ramucirumab VEGFR2 Human IgG1 2014 Gastric Ca Pembrolizumab PD-1 Humanized IgG1 2014 Melanoma Nivolumab PD-1 Human IgG1 2014 Melanoma Dinutuximab GD2 Chimeric IgG1 2015 Neuroblastoma Daratumumab CD38 Human IgG1 2015 Multiple myeloma Necitumumab EGFR Human IgG1 2015 Lung cancer Elotuzumab SLAMF7 Humanized IgG1 2015 Multiple myeloma Atezolizumab PD-L1 Humanized IgG1 2016 Urothelial cancer

Catumaxomab CD3 Chimeric mouse-rat

Avelumab (14) PD-L1 human IgG1

Durvalumab PD-L1 human IgG1

**year**

CD33 2000 Acute myeloid leukemia

CD20 2002 B cell non-Hodgkin

HER2 Humanized IgG1 2013 Breast cancer

**Cancer**

lymphoma

lymphocytic leukemia

lymphoma

lymphocytic leukemia

lymphocytic leukemia

2017 Metastatic merkel cell

2018 Urothelial carcinoma/

carcinoma

non-small cell lung cancer

2009 Malignant ascites

*Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

**Monoclonal antibody**

Gemtuzumab Ozogamicin

Ibritumomab Tiuxetan

Ado-Trastuzumab Emtansine

*Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

*Current Trends in Cancer Management*

oncologists and cancer immunologists.

**2. Monoclonal antibodies**

some cancers than others.

cancers. It is shown that immunotherapy has the potential to move to the front-line of therapeutic options in most cancers. Despite the benefits of immunotherapy, some treatments have severe side effects such as nausea, fever, and diarrhea [2]. The aim of this chapter is to study the concept of immunotherapy for cancer treatment and to provide a thorough review on immunotherapy's developments for both

One of the mechanisms of immune system to defeat pathogens or cancers is to identify foreign substances or malignancies and generate antibodies against them. These antibodies can recognize pathogens and cancer cells by the antigens expressed on their surface. Antibodies have the ability to attach to the specific antigens and destroy foreign particles or malignancies. In the laboratory, scientists can generate many copies of antibodies that are specific to certain antigens on cancer cells. These are known as monoclonal antibodies. In 1997, the first monoclonal antibody, rituximab, was approved for treatment of non-Hodgkin's lymphoma. Beneficial outcomes of rituximab treatment resulted in emergence and development of monoclonal antibodies as a therapeutic approach for various hematological and solid cancers [3]. The most important step in generating monoclonal antibodies for cancer treatment is identifying right antigens on cancer cells. High mutation capacity of cancer cells and existence of various antigens make this task challenging. So far, monoclonal antibodies therapy has been more beneficial against

Monoclonal antibodies can defeat cancer in different ways. Some monoclonal antibodies can recognize antigens expressed by cancer cells and mark them as a target that should be destroyed by immune system. This monoclonal antibody treatment is also known as targeted therapy [4]. Some of monoclonal antibodies cause apoptosis in cancer cells by directly attaching to the cancer cells. Preventing cell proliferation, destroying cell membrane, delivering radiation or chemotherapy to cancer cells, and inhibiting blood vessel growth are other functions of monoclonal antibodies to stop cancer cells. Monoclonal antibodies can robust, mimic or maintain the immune system's response on cancer cells in different ways, and some particular monoclonal antibodies act by more than one function [3]. Monoclonal antibodies can be categorized to three groups such as naked monoclonal antibodies (**Table 1**), conjugated monoclonal antibodies (**Table 2**), and bispecific monoclonal antibodies. Naked monoclonal antibodies act by just a single function. This single function can either be directly affecting cancer cells or by improving immune system against cancer cells. Trastuzumab is an example of monoclonal antibodies with direct effect on cancer cells. Trastuzumab can identify and block HER2 antigen, which is highly expressed on breast and stomach cancer cells. HER2 antigen is responsible for growth and proliferation of cancer cells. By blocking HER2 antigens, cancer cells are not able to expand and proliferate and spread in the body [5]. Immune check point inhibitors are monoclonal antibodies which improve immune system function. This group of antibodies will be discussed in detail later on this chapter. Some monoclonal antibodies can trigger immune system by attaching to immune cells and activating immune cells to destroy cancer cells. Alemtuzumab, which is a monoclonal antibody to treat chronic lymphocytic leukemia, binds to CD25 marker on the surface of lymphocytes and attracts immune cells to destroy cancer cells [6]. Conjugated monoclonal antibodies, also known as tagged antibodies or loaded antibodies, are antibodies that are being used to deliver either chemotherapy drugs or radioactive particles to cancer cells. These monoclonal

**20**


**Table 1.**

*Unconjugated monoclonal antibodies currently approved by the Food and Drug Administration (FDA) for cancer therapy.*

antibodies reduce the toxic effects of systemic chemotherapy and radiotherapy by directly homing the toxic drugs to tumor microenvironment [7, 8]. Ibritumomab tiuxetan is a radio-immunotherapeutic drug which directly delivers radio isotopes to cancerous B cells in non-Hodgkin lymphoma. Ibritumomab tiuxetan is a radiolabeled monoclonal antibody against CD20 antigen, which is expressed on B cell surface. By attaching Ibritumomab tiuxetan to CD20 on the B cells and killing cancer cells, the drug is able to eliminate lymphoma [7]. Chemolabeled antibodies are monoclonal antibodies that are attached to chemotherapy drugs. Brentuximab


#### **Table 2.**

*Conjugated monoclonal antibodies currently approved by the Food and Drug Administration (FDA) for cancer therapy.*

vedotin is a chemolabeled monoclonal antibody specific for CD30 antigen on lymphocytes that delivers monomethyl auristatin E chemotherapy to cancer cells for treatment of Hodgkin lymphoma and anaplastic large cell lymphoma [9]. Adotrastuzumab emtansine is another chemolabeled antibody attached to Mertansine (DM1) chemotherapy with ability to target HER2 molecules on breast cancer cells [10]. Immunotoxin monoclonal antibodies are a new class of monoclonal antibodies that are attached to highly toxic protein molecules of a plant or bacteria. Immunotoxins can specifically bind to their target and deliver potent toxins to cancer cells [11]. The most recent group of antibodies is bispecific monoclonal antibodies that consist of two separate antibodies targeting different specific antigens. Blinatumomab is a bispecific monoclonal antibody with the ability to bind to CD19 on lymphoma and leukemia cells and CD3 on T cells. This antibody is usually used for treatment of acute lymphocytic leukemia. By binding to two antigens on separate cells, Blinatumomab is able to bring immune cells and cancer cells together and ease the pathway for immune cells to find, attack, and kill cancer cells [12].

Based on the genetically engineering techniques, four groups of monoclonal antibodies have been developed. Murine monoclonal antibodies, which were derived from mice, were the first generation of antibodies. They were quickly eliminated from clinical studies as they were not able to interact with human immune system. Chimeric monoclonal antibodies are another category of monoclonal antibodies, consist of constant regions mostly derived from human source and variable regions entirely derived from murine source [13]. There is a subtype of chimeric non-humanized monoclonal antibodies also known as rat-mouse hybrid monoclonal antibodies with murine Fc portion that have specificities for binding to three different tumor cells, T cells and also accessory cells [14]. On the other hand, chimeric humanized monoclonal antibodies, that comprise human Fc portion, are

**23**

*Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

developed with more efficient interaction with human immune system and less immunogenicity [15]. Less immunogenic and more efficient monoclonal antibodies have been developed as humanized monoclonal antibodies, which predominantly originated from human source excluding Fab portion which is derived from murine source. Human monoclonal antibodies that are fully human and are derived from transgenic mice known to be the most efficient and the least immunogenic [16]. Although monoclonal antibodies are being used for treatment of cancer, they may increase the risk of immune reactions or adverse effects. The immune reactions including acute anaphylactic reaction, serum sickness, or cytokine release syndrome (CRS) generally occur after first infusion of monoclonal antibodies. Adverse effects of monoclonal antibodies are the result of immunodeficiency mediated by blockade of specific targets. Infections such as reactivation of tuberculosis or progressive multifocal leukoencephalopathy, autoimmune diseases such as lupus and thyroid disease, cancer, dermatitis, and organ-specific adverse effects are other risks of monoclonal antibodies administration [13]. The other problem of monoclonal antibodies are constant mutation of cancer cells which results in formation of different or neoantigens that already available antibodies cannot function against them. Generation of different or neoantigens lead to absence of responsiveness to monoclonal antibodies. Developed genome sequencing techniques is promising for identifying neoantigens and producing monoclonal antibodies against this targets [3]. Monoclonal antibodies have been proven to remarkably shrink solid tumors, suppress malignancies, diminish metastasis, and increase overall survival in patients [17, 18]. Monoclonal antibodies are promising for treatment of cancers in

both monotherapy and in combinatorial therapeutic approaches.

It was believed that cancer cells were completely resistant to immune system till 1800s when researchers reported regression or total elimination of some solid tumors in patients who had streptococcal skin infections or were infused with bacterial extracts [1, 19]. These studies were not continued until Sharma and Allison noticed that blocking of cytotoxic T lymphocyte-associated protein 4 (CTLA-4) enhances tumor killing capacity of T cells [20]. This hypothesis pops up that some bacterial or organisms' extracts have the ability to block molecules on immune cells, known as checkpoints, which promote immune cells' functionality against cancer cells. These observations led to more in-depth studies to identify immune checkpoints which their blockade can trigger robust anticancer immune responses.

One type of monoclonal antibodies that bind to immune check points is referred

as immune checkpoint blockades. Checkpoints or coinhibitory receptors are molecules on immune cells that bind to their ligands expressed on normal cells. Under normal circumstances, immune checkpoints recognize healthy cells as non-pathogenic by binding to the ligands on normal cells and prevent activity of the immune system against its own tissue. Some cancer cells express check points ligands which help them to escape from recognition and elimination by immune system. By blocking immune checkpoints, immune cells gain a robust response against cancer cells. Immune check point blockades have been proven to be effective in many cancers and are promising because they are targeting immune cells by

CTLA-4 is a coinhibitory receptor on T cells that prevent T cells activation. During T cells activation, antigen-presenting cells (APCs) present processed antigens on their major histocompatibility complex (MHC) molecules to T cell receptors. After the initial phase of activation, B7-1 or B7-2 molecules of APCs

**3. Immune checkpoint blockades**

removing inhibitory pathways [21].

#### *Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

*Current Trends in Cancer Management*

Ositumomab CD20

CD20 Radionucleotide (Yttrium90 or Indium111)

Radionucleotide (Iodine131)

Trastuzumab DM1

**Monoclonal antibody**

Ibritumomab tiuxetan

Brentuximab vedotin

Trastuzumab emtansine

Tositumomab; Iodine I 131 Tositumomab

Capromab pendetide

**Table 2.**

*therapy.*

vedotin is a chemolabeled monoclonal antibody specific for CD30 antigen on lymphocytes that delivers monomethyl auristatin E chemotherapy to cancer cells for treatment of Hodgkin lymphoma and anaplastic large cell lymphoma [9]. Adotrastuzumab emtansine is another chemolabeled antibody attached to Mertansine (DM1) chemotherapy with ability to target HER2 molecules on breast cancer cells [10]. Immunotoxin monoclonal antibodies are a new class of monoclonal antibodies that are attached to highly toxic protein molecules of a plant or bacteria. Immunotoxins can specifically bind to their target and deliver potent toxins to cancer cells [11]. The most recent group of antibodies is bispecific monoclonal antibodies that consist of two separate antibodies targeting different specific antigens. Blinatumomab is a bispecific monoclonal antibody with the ability to bind to CD19 on lymphoma and leukemia cells and CD3 on T cells. This antibody is usually used for treatment of acute lymphocytic leukemia. By binding to two antigens on separate cells, Blinatumomab is able to bring immune cells and cancer cells together and ease the pathway for immune cells to find, attack, and kill cancer

**Target Type Approval** 

Murine IgG2a

IgG1 Drug (auristatin E)

Humanized IgG1 Drug (mertansine)

IgG2a

Arcitumomab Diagnostic Murine IgG1 Colorectal cancer

*Conjugated monoclonal antibodies currently approved by the Food and Drug Administration (FDA) for cancer* 

Diagnostic Murine IgG1 Prostate cancer

CD30 Chimeric

CD19+ CD3 Murine

**year**

Murine IgG1 2002 B cell non-Hodgkin's lymphoma/

**Cancers**

lymphoproliferative disorder

systemic anaplastic large cell lymphoma

2003 Non-Hodgkin lymphoma

2011 Hodgkin lymphoma and

2014 Acute lymphoblastic leukemia

2013 Breast cancer

Based on the genetically engineering techniques, four groups of monoclonal antibodies have been developed. Murine monoclonal antibodies, which were derived from mice, were the first generation of antibodies. They were quickly eliminated from clinical studies as they were not able to interact with human immune system. Chimeric monoclonal antibodies are another category of monoclonal antibodies, consist of constant regions mostly derived from human source and variable regions entirely derived from murine source [13]. There is a subtype of chimeric non-humanized monoclonal antibodies also known as rat-mouse hybrid monoclonal antibodies with murine Fc portion that have specificities for binding to three different tumor cells, T cells and also accessory cells [14]. On the other hand, chimeric humanized monoclonal antibodies, that comprise human Fc portion, are

**22**

cells [12].

developed with more efficient interaction with human immune system and less immunogenicity [15]. Less immunogenic and more efficient monoclonal antibodies have been developed as humanized monoclonal antibodies, which predominantly originated from human source excluding Fab portion which is derived from murine source. Human monoclonal antibodies that are fully human and are derived from transgenic mice known to be the most efficient and the least immunogenic [16].

Although monoclonal antibodies are being used for treatment of cancer, they may increase the risk of immune reactions or adverse effects. The immune reactions including acute anaphylactic reaction, serum sickness, or cytokine release syndrome (CRS) generally occur after first infusion of monoclonal antibodies. Adverse effects of monoclonal antibodies are the result of immunodeficiency mediated by blockade of specific targets. Infections such as reactivation of tuberculosis or progressive multifocal leukoencephalopathy, autoimmune diseases such as lupus and thyroid disease, cancer, dermatitis, and organ-specific adverse effects are other risks of monoclonal antibodies administration [13]. The other problem of monoclonal antibodies are constant mutation of cancer cells which results in formation of different or neoantigens that already available antibodies cannot function against them. Generation of different or neoantigens lead to absence of responsiveness to monoclonal antibodies. Developed genome sequencing techniques is promising for identifying neoantigens and producing monoclonal antibodies against this targets [3]. Monoclonal antibodies have been proven to remarkably shrink solid tumors, suppress malignancies, diminish metastasis, and increase overall survival in patients [17, 18]. Monoclonal antibodies are promising for treatment of cancers in both monotherapy and in combinatorial therapeutic approaches.

#### **3. Immune checkpoint blockades**

It was believed that cancer cells were completely resistant to immune system till 1800s when researchers reported regression or total elimination of some solid tumors in patients who had streptococcal skin infections or were infused with bacterial extracts [1, 19]. These studies were not continued until Sharma and Allison noticed that blocking of cytotoxic T lymphocyte-associated protein 4 (CTLA-4) enhances tumor killing capacity of T cells [20]. This hypothesis pops up that some bacterial or organisms' extracts have the ability to block molecules on immune cells, known as checkpoints, which promote immune cells' functionality against cancer cells. These observations led to more in-depth studies to identify immune checkpoints which their blockade can trigger robust anticancer immune responses.

One type of monoclonal antibodies that bind to immune check points is referred as immune checkpoint blockades. Checkpoints or coinhibitory receptors are molecules on immune cells that bind to their ligands expressed on normal cells. Under normal circumstances, immune checkpoints recognize healthy cells as non-pathogenic by binding to the ligands on normal cells and prevent activity of the immune system against its own tissue. Some cancer cells express check points ligands which help them to escape from recognition and elimination by immune system. By blocking immune checkpoints, immune cells gain a robust response against cancer cells. Immune check point blockades have been proven to be effective in many cancers and are promising because they are targeting immune cells by removing inhibitory pathways [21].

CTLA-4 is a coinhibitory receptor on T cells that prevent T cells activation. During T cells activation, antigen-presenting cells (APCs) present processed antigens on their major histocompatibility complex (MHC) molecules to T cell receptors. After the initial phase of activation, B7-1 or B7-2 molecules of APCs attach to CD28 on T cells. TCR signal and costimulatory B7-CD28 induce complete T cell activation that result in cytokine release from activated T cells [22]. Besides, inhibitory signals induce by CTLA-4 act in an opposite way [23]. CTLA-4 molecule expressed on T cells has a higher affinity to bind to B7 compare to CD28. In a competition between CD28 and CTLA-4, CTLA-4 predominantly binds to B7 and generates an inhibitory signal during T cells activation. Inhibitory signals of CTLA-4 halt T cells activation and induce immune tolerance. Blocking of CTLA-4 by Ipilimumab (CTLA-4 blockade) was first approved by FDA due to success of CTLA-4 blockade in treatment of melanoma patients [24]. Ipilimumab boosts immune responses to cancer cells mediated by T cells activation. Most of patients experience Ipilimumab-related side effects like diarrhea, vomiting, skin rashes, nausea, and even life-threatening effects. All patients receiving this drug are always monitored closely and side effects are managed by corticosteroids [25].

In cancer, T cells are constantly exposed to antigen stimulation which result in gradual deterioration of their function by losing cytokine production ability and persistent increase in expression of inhibitory receptors. Defects in T cell activation, cytokine production, and proliferation is defined as exhaustion. Inhibitory receptors are highly expressed on exhausted T cells. Cancer cells have a high expression of inhibitory ligands that increase the chance of exhaustion in T cells. Programmed cell death-1 (PD-1) is an inhibitory molecule known as the receptor for cell death and have regulatory inhibitory role in activation of T cells. Physiologically, PD-1/PD-1 ligand (PD-L1) signaling pathway is a way to control excessive inflammation to protect normal tissues by induction of immune tolerance [26]. Interaction of PD-1 and PD-L1, which is highly expressed on tumor cells, causes exhaustion and dysfunctionality in T cells that avoid immune response against cancer cells. PD-1 or PD-L1 inhibitors pharmacologically prevent interaction of these molecules and efficiently maintain T cells function and facilitate them to kill tumor cells. Both PD-1 and PD-L1 immune checkpoint blockades have been proven to be effective for many malignancies but still it is not obvious that whether blocking of PD-1 on T cells or PD-L1 on tumors is more effective for cancer treatment. Patients' characteristics such as type of tumor, mutation burden of tumor, and metastases of tumor affect efficacy of PD-1/PD-L1 inhibitors [27]. PD-L1 is not constantly expressed on different tumors and even in different stages of tumor growth. Therefore, efficacy of PD-L1 blockade depends on the type of tumor, stage of tumor, location of the tumor, and many other factors [28, 29]. Atezolizumab, the first FDA-approved PD-L1 blockade, has been used as the first-line treatment of metastatic non-small lung carcinoma and cisplatin-resistant metastatic urothelial carcinoma. Avelumab, is another FDA-approved PD-L1 blockade for metastatic merkel cell carcinoma that lack efficient response to chemotherapy [30]. Nivolumab and Pembrolizumab are PD-1 blockers and are successfully used in Phase I clinical trial on patients with non-small-cell lung cancer and renal cell carcinoma. Nivolumab was approved by FDA for treatment of advanced melanoma patients after significant improved response in phase III trial. Also, Pembrolizumab is the first-line immune checkpoint blockade for the treatment of metastatic melanoma and metastatic non-small cell lung cancer [31]. These drugs have significantly increased survival of patients with minimal side effects in other solid tissue tumors. To improve benefits from immune checkpoint blockades, combinatorial strategies are under study. Combination regimens include administration of two immune checkpoint blockades together or a monoclonal antibody with chemotherapy or radiotherapy [32]. Combinatorial strategies enhance anticancer responses because each treatment works through targeting different pathways. Combination therapy of Ipilimumab/Nivolumab is approved by FDA for treatment of melanoma [33]. Pembrolizumab plus chemotherapy (pemetrexed/carboplatin) is approved for

**25**

*Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

are under investigation [32].

**4. Cancer vaccines**

vaccines, and genetic vaccines.

treatment of non-small cell lung carcinoma [34]. Several combination therapies including either two different checkpoint blockades or with chemo/radiotherapy

for patients receiving immune checkpoint blockades [35].

Immune checkpoint blockades have changed the treatment strategies for cancer with dramatic improves in many cancers. PD-1, PD-L1, and CTLA-4 inhibitors are able to change immune responses and it may cause adverse immune reactions. These immune reactions are usually better tolerated than chemotherapy drugs but still recognition and proactive treatments should be included in the treatment strategy

Cancer vaccines are a new generation of vaccines different to traditional prophylactic vaccines which were administered to healthy people. Cancer vaccines are administered to either prevent cancer in high-risk individuals or to treat cancer in patients with malignancies. Therapeutic cancer vaccines are able to enhance immune system to attack cancer cells. Two prophylactic vaccines were approved for cancers that are caused by virus infections. One of the prophylactic vaccines is for hepatitis B virus (HBV) infection that can cause liver cancers such as cirrhosis and hepatocellular carcinoma in those who suffer from chronic infections. Another prophylactic vaccine is against human papilloma virus (HPV) that mediates cervical, anal, vaginal, vulvar, and throat cancers as well as genital warts. Until now, preventive vaccines were only available for the cancers that are caused by infections. Therapeutic vaccines are meant to enhance immune system in order to interfere with cancer cells, stop their growth and proliferation, and kill cancer cells. Therapeutic cancers are divided to several categories of cell vaccines, peptide

Tumor cell vaccines are a type of cell vaccines including autologous tumor cell vaccines and allogenic tumor cell vaccines. Autologous tumor cell vaccines are isolated from patient-derived tumor cells and prepared in vitro for administration to the patient from whom the tumor cells were isolated. Preparation of tumor cells for vaccination includes irradiation of tumor cells or combining tumor cells with an immune stimulatory adjuvant such as recombinant granulocyte monocyte-colony stimulating factor (GM-CSF) [36]. Autologous cell vaccines are able to present a wide range of tumor-associated antigens to cytotoxic T cells, resulting in a robust antitumor activity. Modification of autologous tumor cells to induce higher levels of immune stimulation has been studied by many researchers. Autologous tumor cell vaccines in animal tumor models of lymphoma and melanoma were more potent when tumor cell vaccines were infected with Newcastle disease virus [37]. In another study, tumor cell vaccines were genetically modified to express higher levels of IL-2 which induced activation of T cells and natural killer (NK) cells [38]. Autologous tumor cell vaccines transduced with GM-CSF, named GVAX, are able to get involved with dendritic cells (DCs), and induce maturation of DCs. GVAXmediated matured DCs activate cytotoxic T cells and improve T cells response to cancer [39]. Autologous tumor cell vaccines have been extensively investigated in clinical and preclinical studies on several cancers and approximately 20% of patients survived for a long time [40]. The advantage of autologous tumor cell vaccines is that the vaccines can target the patient's own tumor-associated antigens and excludes the step to select specific antigens. One major problem in preparing autologous tumor cell vaccines is the time-consuming process of harvesting sufficient amount of tumor cells, which is a restriction for certain tumors. Appose to autologous tumor cell vaccines, allogeneic tumor cell vaccines are easy and less

#### *Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

treatment of non-small cell lung carcinoma [34]. Several combination therapies including either two different checkpoint blockades or with chemo/radiotherapy are under investigation [32].

Immune checkpoint blockades have changed the treatment strategies for cancer with dramatic improves in many cancers. PD-1, PD-L1, and CTLA-4 inhibitors are able to change immune responses and it may cause adverse immune reactions. These immune reactions are usually better tolerated than chemotherapy drugs but still recognition and proactive treatments should be included in the treatment strategy for patients receiving immune checkpoint blockades [35].

#### **4. Cancer vaccines**

*Current Trends in Cancer Management*

attach to CD28 on T cells. TCR signal and costimulatory B7-CD28 induce complete T cell activation that result in cytokine release from activated T cells [22]. Besides, inhibitory signals induce by CTLA-4 act in an opposite way [23]. CTLA-4 molecule expressed on T cells has a higher affinity to bind to B7 compare to CD28. In a competition between CD28 and CTLA-4, CTLA-4 predominantly binds to B7 and generates an inhibitory signal during T cells activation. Inhibitory signals of CTLA-4 halt T cells activation and induce immune tolerance. Blocking of CTLA-4 by Ipilimumab (CTLA-4 blockade) was first approved by FDA due to success of CTLA-4 blockade in treatment of melanoma patients [24]. Ipilimumab boosts immune responses to cancer cells mediated by T cells activation. Most of patients experience Ipilimumab-related side effects like diarrhea, vomiting, skin rashes, nausea, and even life-threatening effects. All patients receiving this drug are always

monitored closely and side effects are managed by corticosteroids [25].

In cancer, T cells are constantly exposed to antigen stimulation which result in gradual deterioration of their function by losing cytokine production ability and persistent increase in expression of inhibitory receptors. Defects in T cell activation, cytokine production, and proliferation is defined as exhaustion. Inhibitory receptors are highly expressed on exhausted T cells. Cancer cells have a high expression of inhibitory ligands that increase the chance of exhaustion in T cells. Programmed cell death-1 (PD-1) is an inhibitory molecule known as the receptor for cell death and have regulatory inhibitory role in activation of T cells. Physiologically, PD-1/PD-1 ligand (PD-L1) signaling pathway is a way to control excessive inflammation to protect normal tissues by induction of immune tolerance [26]. Interaction of PD-1 and PD-L1, which is highly expressed on tumor cells, causes exhaustion and dysfunctionality in T cells that avoid immune response against cancer cells. PD-1 or PD-L1 inhibitors pharmacologically prevent interaction of these molecules and efficiently maintain T cells function and facilitate them to kill tumor cells. Both PD-1 and PD-L1 immune checkpoint blockades have been proven to be effective for many malignancies but still it is not obvious that whether blocking of PD-1 on T cells or PD-L1 on tumors is more effective for cancer treatment. Patients' characteristics such as type of tumor, mutation burden of tumor, and metastases of tumor affect efficacy of PD-1/PD-L1 inhibitors [27]. PD-L1 is not constantly expressed on different tumors and even in different stages of tumor growth. Therefore, efficacy of PD-L1 blockade depends on the type of tumor, stage of tumor, location of the tumor, and many other factors [28, 29]. Atezolizumab, the first FDA-approved PD-L1 blockade, has been used as the first-line treatment of metastatic non-small lung carcinoma and cisplatin-resistant metastatic urothelial carcinoma. Avelumab, is another FDA-approved PD-L1 blockade for metastatic merkel cell carcinoma that lack efficient response to chemotherapy [30]. Nivolumab and Pembrolizumab are PD-1 blockers and are successfully used in Phase I clinical trial on patients with non-small-cell lung cancer and renal cell carcinoma. Nivolumab was approved by FDA for treatment of advanced melanoma patients after significant improved response in phase III trial. Also, Pembrolizumab is the first-line immune checkpoint blockade for the treatment of metastatic melanoma and metastatic non-small cell lung cancer [31]. These drugs have significantly increased survival of patients with minimal side effects in other solid tissue tumors. To improve benefits from immune checkpoint blockades, combinatorial strategies are under study. Combination regimens include administration of two immune checkpoint blockades together or a monoclonal antibody with chemotherapy or radiotherapy [32]. Combinatorial strategies enhance anticancer responses because each treatment works through targeting different pathways. Combination therapy of Ipilimumab/Nivolumab is approved by FDA for treatment of melanoma [33]. Pembrolizumab plus chemotherapy (pemetrexed/carboplatin) is approved for

**24**

Cancer vaccines are a new generation of vaccines different to traditional prophylactic vaccines which were administered to healthy people. Cancer vaccines are administered to either prevent cancer in high-risk individuals or to treat cancer in patients with malignancies. Therapeutic cancer vaccines are able to enhance immune system to attack cancer cells. Two prophylactic vaccines were approved for cancers that are caused by virus infections. One of the prophylactic vaccines is for hepatitis B virus (HBV) infection that can cause liver cancers such as cirrhosis and hepatocellular carcinoma in those who suffer from chronic infections. Another prophylactic vaccine is against human papilloma virus (HPV) that mediates cervical, anal, vaginal, vulvar, and throat cancers as well as genital warts. Until now, preventive vaccines were only available for the cancers that are caused by infections. Therapeutic vaccines are meant to enhance immune system in order to interfere with cancer cells, stop their growth and proliferation, and kill cancer cells. Therapeutic cancers are divided to several categories of cell vaccines, peptide vaccines, and genetic vaccines.

Tumor cell vaccines are a type of cell vaccines including autologous tumor cell vaccines and allogenic tumor cell vaccines. Autologous tumor cell vaccines are isolated from patient-derived tumor cells and prepared in vitro for administration to the patient from whom the tumor cells were isolated. Preparation of tumor cells for vaccination includes irradiation of tumor cells or combining tumor cells with an immune stimulatory adjuvant such as recombinant granulocyte monocyte-colony stimulating factor (GM-CSF) [36]. Autologous cell vaccines are able to present a wide range of tumor-associated antigens to cytotoxic T cells, resulting in a robust antitumor activity. Modification of autologous tumor cells to induce higher levels of immune stimulation has been studied by many researchers. Autologous tumor cell vaccines in animal tumor models of lymphoma and melanoma were more potent when tumor cell vaccines were infected with Newcastle disease virus [37]. In another study, tumor cell vaccines were genetically modified to express higher levels of IL-2 which induced activation of T cells and natural killer (NK) cells [38]. Autologous tumor cell vaccines transduced with GM-CSF, named GVAX, are able to get involved with dendritic cells (DCs), and induce maturation of DCs. GVAXmediated matured DCs activate cytotoxic T cells and improve T cells response to cancer [39]. Autologous tumor cell vaccines have been extensively investigated in clinical and preclinical studies on several cancers and approximately 20% of patients survived for a long time [40]. The advantage of autologous tumor cell vaccines is that the vaccines can target the patient's own tumor-associated antigens and excludes the step to select specific antigens. One major problem in preparing autologous tumor cell vaccines is the time-consuming process of harvesting sufficient amount of tumor cells, which is a restriction for certain tumors. Appose to autologous tumor cell vaccines, allogeneic tumor cell vaccines are easy and less

expensive to produce in large scales. Allogeneic whole tumor cell vaccines consist of at least two human tumor cell lines and have unlimited tumor-specific antigens. Canavaxin is an allogeneic tumor cell vaccine consisting of three irradiated allogeneic melanoma cell lines combined with adjuvant Bacillus Calmette-Guérin (BCG). Despite Canavaxin increased overall survival of melanoma patients in phase II of trials, clinical trials were terminated because of failure of the vaccine in stages III and IV [41]. Allogeneic GVAX vaccine has been studied for treatment of prostate cancer [42], breast cancer [43], and pancreatic cancer [44]. Combination of GVAX vaccine with CTLA-4 antibody (Ipilimumab) was approved by FDA for treatment of metastatic melanoma [45]. Belagenpumatucel-L is another allogeneic tumor cell vaccine formed from four non-small cell lung carcinoma (NSCLC) cell lines transfected with plasmid containing a transforming growth factor (TGF)-beta2 antisense transgene. This genetically modified vaccine secretes TGF-beta and is used for treatment of NSCLC [46].

#### **5. Dendritic cell vaccines**

Dendritic cell (DC) vaccines emerged as a potent cancer vaccine. DCs are professional antigen-presenting cells (APCs) that act as a bridge between innate and adoptive immune system [47]. DCs uptake pathogens, process them, and present pathogen antigens on their MHC molecules. Processed antigens on DCs are directly recognized by T cells which induce antigen-specific immune responses. Different subtypes of DCs exist in human body based on CD8, CD103, or CD11b expressions. DCs are in both non-lymphoid organs and lymphoid organs such as lymph nodes, spleens, and bone marrow. Classical DCs (cDCs) are divided to CD8+, CD103+, and CD11b+ DCs. Non-classic DCs include monocyte-derived DCs, plasmacytoid DCs, and Langerhans cells. These categories are based on expression of molecules and the location of DCs in body [48]. Studies showed that different subsets of DCs can prime and expand various T cells. For example, CD8+ CD205+ DCs present antigens on both MHC-I and MHC-II and are able to prime CD4+ T cell and CD8+ T cells but CD8-33D1+ DCs present antigens just on MHC-II and prime CD4+ T cells [49]. DCs act as a double-edged sword that can induce both immune tolerance and immune activation depending on which receptors on DCs are engaged [50]. Maturation and migration of DCs play a critical role in characteristics of DCs [51]. Matured DCs migrate to lymphoid organs and prime T cells to enhance antitumor responses. Loading of MHC molecules with cancer antigens, up regulation of costimulatory molecules such as CD40, CD80, and CD86 on DCs, and cytokine production of DCs are critically required for activation of T cells DCs [52, 53]. DC vaccines include ex vivo generation of DCs from CD34+ hematopoietic progenitor cells or peripheral blood-derived monocytes (PBMC) [53]. Ex vivo-generated DCs are loaded with appropriate source of tumor antigens and are subsequently activated with adjuvants and are administered back to patients to kill tumors. Tumor antigens derived from total tumor [54], DNA/RNA virus [55], tumor proteins, or peptides [56, 57] are utilized for DC vaccines. Moreover, some DC vaccines are composed of fusion of tumor cells and ex vivo-generated DCs [58]. Autologous DC vaccine pulsed with HLA-A0201 peptide (prostate-specific antigen) was among the first dendritic cell vaccines used in clinical trials with promising results [56]. DC vaccines have been studies in many clinical trials on various cancers. FDA-approved Sipuleucel-T DC vaccine for the first time for the treatment of metastatic castrate-resistant prostate cancer [59]. Sipuleucel-T composed of PBMC-derived DCs loaded with PA2024 (prostatic acid phosphate) fused to GM-CSF, which significantly increased patients survival. Although DC vaccines were successful in prostate cancer treatment, their

**27**

vaccines [91].

**6. Genetic vaccines**

*Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

to cancer cells.

efficacy in other cancers was modest. Researchers conduct studies to enhance DC vaccines potency by modulating stimulatory and inhibitory molecules on DCs. Modulation of costimulatory molecules such as CD40L, CD70, GITRL, CD137L, and OX40L [60–63] or inflammatory markers of IL-12p70, IL-18, IL-12, CXCL10, and CCR7 on DCs improve DCs maturation and T cell priming characteristics [64–68]. The other way to enhance anticancer T cell response by DCs is to suppress inhibitory molecules on DCs. Genetically silencing of ubiquitin-editing enzyme A20 [69], suppressor of cytokine signaling 1 (SOCS1) [70], and scavenger receptor SRA/CD204 [71] improve DCs function and subsequently enhance T cell response

Two of the most important limitations of cancer cell vaccines and DC vaccines are limited source of specimen and complicated procedure to generate these vaccines. New vaccines generated by tumor-associated antigen peptides combined with an adjuvant seemed to solve the restrictions of cancer cell and DC vaccines. The first encoded human tumor-associated antigen peptide was named MAGE-1 [72]. Different types of tumor-associated antigen peptides are studied. Cancer testis antigens are a group of genes available in both healthy and cancerous tissues. These genes such as MAGE, BAGE, NY-ESO-1, and SSX-2 are scant in normal tissues but are highly expressed in tumors [73–75]. Tissue differentiation antigens are available and active in both healthy tissues and tumors-like PSA and PAP in prostate cancer [76, 77], gp100, Melan-A/Mart-1, and tyrosinase in melanoma [78–80], and mammaglobin-A in breast carcinomas [81]. Tumor-specific antigens or -mutated oncogenes are a group of antigens expressed on both normal tissues and tumors with a unique up regulation in tumors such as CEA [82], MUC-1 [83], HER2/Neu [84], and certain antiapoptotic proteins (i.e. livin and survivin) [85, 86]. Clinical trials mostly focused on effects of peptide vaccines that target cancer testis antigens, and differentiation-associated antigens. To produce an effective peptide vaccine, addition of immune stimulatory adjuvant is required for an efficient immune response as tumor-associated antigens are not immunogenic. Some adjuvants used for peptide vaccine generation are aluminum salt, pathogen-associated molecular patterns (PAMPs), TLR agonists [87], BCG [88], and monophosphoryl lipid A (MPL) [89]. Cervarix is the first peptide vaccine for human papillomavirus composed of MPL and aluminum salt [90]. The advantage of peptide vaccines to DC vaccines and cancer cell vaccines is that peptide vaccines are more cost effective, but they may also appear to be less potent because they only target one or few epitopes of tumor-associated antigens. Formulation of peptide vaccines, route of delivery, and selection of immunogenic adjuvants can influence efficacy of peptide

Genetic vaccines are another approach for carrying tumor-associated antigens

to patients by utilizing plasmid DNA vectors. Genetic vaccines transfect DCs and directly present tumor-associated antigens to cytotoxic T cells or they can transfect somatic cells and indirectly cross prime T cells. Each genetic vaccine can deliver many tumor-associated antigens to patients and induce a robust anticancer immunity [92]. DNA vaccines are composed of bacterial plasmids that carry genes of interest under the control of mammalian promoter. DNA vaccines are able to initiate innate immunity and based on the site of delivery, they can trigger cellular and humoral immunity [93]. Usually the transgene is cytomegalovirus (CMV) immediate early promoter and its intron A sequence [94]. Optimizing codon usage can increase the transduction of antigens. In the intra muscular administration of DNA

#### *Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

*Current Trends in Cancer Management*

used for treatment of NSCLC [46].

**5. Dendritic cell vaccines**

expensive to produce in large scales. Allogeneic whole tumor cell vaccines consist of at least two human tumor cell lines and have unlimited tumor-specific antigens. Canavaxin is an allogeneic tumor cell vaccine consisting of three irradiated allogeneic melanoma cell lines combined with adjuvant Bacillus Calmette-Guérin (BCG). Despite Canavaxin increased overall survival of melanoma patients in phase II of trials, clinical trials were terminated because of failure of the vaccine in stages III and IV [41]. Allogeneic GVAX vaccine has been studied for treatment of prostate cancer [42], breast cancer [43], and pancreatic cancer [44]. Combination of GVAX vaccine with CTLA-4 antibody (Ipilimumab) was approved by FDA for treatment of metastatic melanoma [45]. Belagenpumatucel-L is another allogeneic tumor cell vaccine formed from four non-small cell lung carcinoma (NSCLC) cell lines transfected with plasmid containing a transforming growth factor (TGF)-beta2 antisense transgene. This genetically modified vaccine secretes TGF-beta and is

Dendritic cell (DC) vaccines emerged as a potent cancer vaccine. DCs are professional antigen-presenting cells (APCs) that act as a bridge between innate and adoptive immune system [47]. DCs uptake pathogens, process them, and present pathogen antigens on their MHC molecules. Processed antigens on DCs are directly recognized by T cells which induce antigen-specific immune responses. Different subtypes of DCs exist in human body based on CD8, CD103, or CD11b expressions. DCs are in both non-lymphoid organs and lymphoid organs such as lymph nodes, spleens, and bone marrow. Classical DCs (cDCs) are divided to CD8+, CD103+, and CD11b+ DCs. Non-classic DCs include monocyte-derived DCs, plasmacytoid DCs, and Langerhans cells. These categories are based on expression of molecules and the location of DCs in body [48]. Studies showed that different subsets of DCs can prime and expand various T cells. For example, CD8+ CD205+ DCs present antigens on both MHC-I and MHC-II and are able to prime CD4+ T cell and CD8+ T cells but CD8-33D1+ DCs present antigens just on MHC-II and prime CD4+ T cells [49]. DCs act as a double-edged sword that can induce both immune tolerance and immune activation depending on which receptors on DCs are engaged [50]. Maturation and migration of DCs play a critical role in characteristics of DCs [51]. Matured DCs migrate to lymphoid organs and prime T cells to enhance antitumor responses. Loading of MHC molecules with cancer antigens, up regulation of costimulatory molecules such as CD40, CD80, and CD86 on DCs, and cytokine production of DCs are critically required for activation of T cells DCs [52, 53]. DC vaccines include ex vivo generation of DCs from CD34+ hematopoietic progenitor cells or peripheral blood-derived monocytes (PBMC) [53]. Ex vivo-generated DCs are loaded with appropriate source of tumor antigens and are subsequently activated with adjuvants and are administered back to patients to kill tumors. Tumor antigens derived from total tumor [54], DNA/RNA virus [55], tumor proteins, or peptides [56, 57] are utilized for DC vaccines. Moreover, some DC vaccines are composed of fusion of tumor cells and ex vivo-generated DCs [58]. Autologous DC vaccine pulsed with HLA-A0201 peptide (prostate-specific antigen) was among the first dendritic cell vaccines used in clinical trials with promising results [56]. DC vaccines have been studies in many clinical trials on various cancers. FDA-approved Sipuleucel-T DC vaccine for the first time for the treatment of metastatic castrate-resistant prostate cancer [59]. Sipuleucel-T composed of PBMC-derived DCs loaded with PA2024 (prostatic acid phosphate) fused to GM-CSF, which significantly increased patients survival. Although DC vaccines were successful in prostate cancer treatment, their

**26**

efficacy in other cancers was modest. Researchers conduct studies to enhance DC vaccines potency by modulating stimulatory and inhibitory molecules on DCs. Modulation of costimulatory molecules such as CD40L, CD70, GITRL, CD137L, and OX40L [60–63] or inflammatory markers of IL-12p70, IL-18, IL-12, CXCL10, and CCR7 on DCs improve DCs maturation and T cell priming characteristics [64–68]. The other way to enhance anticancer T cell response by DCs is to suppress inhibitory molecules on DCs. Genetically silencing of ubiquitin-editing enzyme A20 [69], suppressor of cytokine signaling 1 (SOCS1) [70], and scavenger receptor SRA/CD204 [71] improve DCs function and subsequently enhance T cell response to cancer cells.

Two of the most important limitations of cancer cell vaccines and DC vaccines are limited source of specimen and complicated procedure to generate these vaccines. New vaccines generated by tumor-associated antigen peptides combined with an adjuvant seemed to solve the restrictions of cancer cell and DC vaccines. The first encoded human tumor-associated antigen peptide was named MAGE-1 [72]. Different types of tumor-associated antigen peptides are studied. Cancer testis antigens are a group of genes available in both healthy and cancerous tissues. These genes such as MAGE, BAGE, NY-ESO-1, and SSX-2 are scant in normal tissues but are highly expressed in tumors [73–75]. Tissue differentiation antigens are available and active in both healthy tissues and tumors-like PSA and PAP in prostate cancer [76, 77], gp100, Melan-A/Mart-1, and tyrosinase in melanoma [78–80], and mammaglobin-A in breast carcinomas [81]. Tumor-specific antigens or -mutated oncogenes are a group of antigens expressed on both normal tissues and tumors with a unique up regulation in tumors such as CEA [82], MUC-1 [83], HER2/Neu [84], and certain antiapoptotic proteins (i.e. livin and survivin) [85, 86]. Clinical trials mostly focused on effects of peptide vaccines that target cancer testis antigens, and differentiation-associated antigens. To produce an effective peptide vaccine, addition of immune stimulatory adjuvant is required for an efficient immune response as tumor-associated antigens are not immunogenic. Some adjuvants used for peptide vaccine generation are aluminum salt, pathogen-associated molecular patterns (PAMPs), TLR agonists [87], BCG [88], and monophosphoryl lipid A (MPL) [89]. Cervarix is the first peptide vaccine for human papillomavirus composed of MPL and aluminum salt [90]. The advantage of peptide vaccines to DC vaccines and cancer cell vaccines is that peptide vaccines are more cost effective, but they may also appear to be less potent because they only target one or few epitopes of tumor-associated antigens. Formulation of peptide vaccines, route of delivery, and selection of immunogenic adjuvants can influence efficacy of peptide vaccines [91].

#### **6. Genetic vaccines**

Genetic vaccines are another approach for carrying tumor-associated antigens to patients by utilizing plasmid DNA vectors. Genetic vaccines transfect DCs and directly present tumor-associated antigens to cytotoxic T cells or they can transfect somatic cells and indirectly cross prime T cells. Each genetic vaccine can deliver many tumor-associated antigens to patients and induce a robust anticancer immunity [92]. DNA vaccines are composed of bacterial plasmids that carry genes of interest under the control of mammalian promoter. DNA vaccines are able to initiate innate immunity and based on the site of delivery, they can trigger cellular and humoral immunity [93]. Usually the transgene is cytomegalovirus (CMV) immediate early promoter and its intron A sequence [94]. Optimizing codon usage can increase the transduction of antigens. In the intra muscular administration of DNA

vaccines, DNA plasmids transfect both myocytes and DCs. The plasmids act as an immunogenic and activate T cells via toll-like receptors [95]. DNA sensors in cytosol of cells such as DAI, H2B, IFI16, DDX41, LRRFIP1, and cGAS are able to detect presence of DNA vaccines. DNA sensors send signal to STING-TBK1 signaling cascade and activate interferon regulatory factor 3 which results in expression of type I interferons. TLR9 recognizes unmethylated CpG DNA and activates interferon regulatory factor 7 that induce expression of interferons. DCs phagocyte antigenexpressing cell (myocytes) and cross present antigens on MHC-I to CD8 T cells. Moreover, interferons promote this pathway. If DNA vaccines directly transfect DCs, DCs are able to uptake, process, and present antigens on MHC-I to CD8 T cells [96]. Transfection of the vector with multiple gene sequences increases the immunization and induces humoral [97] and CD8 T cell response [98]. Combination of DC vaccines with other immune stimulatory agents such as TLR agonists [99], or monoclonal antibodies [100] increase anticancer immunity. RNA vaccines are safe vaccines compared to DNA vaccines as they degrade and clear quickly in body. Total tumor RNAs are isolated from tumor tissues and they can induce a potent immune response. RNA vaccines are composed of various tumor antigens which reduce the possibility of immune escape by tumor cells. The first use of RNA vaccines was to immunize patients with mRNAs that encode tumor-associated antigens. Furthermore, RNA vaccines can be produced for personalized cancer treatment. Patients' neoantigens can be identified by tumor exome analyzing and personalized RNA vaccine can be specifically generated. In addition to direct use of mRNAs for vaccine generation, RNAs are utilized in cell therapies. Transfecting patient-derived cells with RNAs and giving manipulated cells back to patients are another form of utilizing mRNAs. For example, transfection of patient-derived DCs with mRNA of tumor-associated antigens can induce an antigen-specific T cell response in cancer patients. Transfection of patient-derived T cells with mRNA of chimeric antigen receptors, triggers T cells to identify specific antigens on cancer cells which quickly deteriorate cancer [101]. Liposomes and protaminase are adjuvants of RNA vaccines and help to stabilize RNAs [102].

#### **7. Adoptive cell therapy**

Adoptive T cell therapy (ACT) is a treatment that enhances T cells' ability to kill cancer cells by transferring immune system-derived cells to patients. The cells used for ACT can originate from the same patient or another individual. In 1988, the first ACT reduced metastatic melanoma tumors with transferring of autologous CD4+ and CD8+ tumor infiltrating lymphocytes (TILs) to the patients [103, 104]. Both peripheral blood T cells and TILs extracted from tumors are utilized to generate specific T cells for ACT. These T cells can be modified and then transferred to patients or directly administered in their natural state. TILs by their own nature have an antitumor activity as they are specific for tumor cells. TILs can recognize tumor antigens such as cancer germline antigens, neoantigens, and viral proteins and kill cancer cells [104]. After tumors are resected, the tumor tissues digest into fragments and each fragment is cultured in the presence of IL-2. The T cells are expanded and each clone is monitored for its reactivity against tumor cells. Proliferating lymphocytes kill tumor cells and produce a pure population of T cells. Cancer reactive T cells are infused back to patients. Moreover, T cells that express a TCR specific for tumor antigens can be selected in vitro from peripheral blood and expanded. Antigen-specific T cells are selected by coculturing of T cells with APCs loaded with tumor particles such as RNAs. By expansion of antigen-specific T cells, a specific antitumor T cell clone can be generated [105]. T cells with TCR targeting

**29**

*Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

tumorigenic mutations such as Ras mutations have shown promise in cancer treatment. Ras is commonly mutated at the onset of tumorigenesis in the dominant population of tumor cells. Targeting Ras mutations and killing tumor cells with Ras-specific ACT may have profound effects on cancers with Ras mutations [106]. TCRs targeting KRAS G12D, a common proto-oncogene encoding GTPase, have anti-tumorigenic effects on patients with colorectal cancer [107]. Also, genetically modified antitumor T cell clones can be produced by infecting T cells with viruses that carry genetically engineered TCRs [108]. TCR-transduced T cells are generated by cloning specific TCRs into a retrovirus. Patients derived PBMCs are activated with CD3 and IL-2 and are transduced with the retrovirus encoding the antigen-specific TCR. The T cells are expanded and injected back to the individuals. Peripheral blood T cells transfected with retrovirus encoding MART-1 TCR regress tumors in melanoma [103]. Genetically engineering techniques can modify TCRs to target-specific antigens. For example, T cells with modified TCRs that target NY-ESO-1, a cancer germline antigen, were successfully used as ACT for treatment of patients with synovial cell sarcoma and melanoma [109]. One major limitation of ACTs is that they induce short-lasting responses in immune system. Administration of T cells after chemotherapy increases cancer regression due to repopulation of host T cells with antigen-specific T cells. Lymphodepletion induced by chemotherapy helps T cells from ACT to proliferate during hemostatic proliferative phase and persist for months after infusion [109]. It was also shown that high doses of IL-2 therapy contribute to expansion of the transferred cells [110, 111]. The first signal in T cell activation begins with binding of TCR to MHC molecules on APCs. Furthermore, MHC expression downregulates on APCs in cancers so that they can escape immunity [112]. In 1989, first chimeric antigen receptors (CARs) were developed to avoid interaction of T cells with MHC molecules. CAR T cells are designed to identify cancer cells and attack them without mediation of APCs. As a result, CARs act independent of any stimulatory and TCR signaling. CAR composed of a ligand-binding domain and a signaling domain. Ligand-binding domain is the extracellular part of CAR that includes B cell receptor derived single chain variable fragment. The signaling domain is made of costimulatory molecules and CD3f and 1 [112]. CD19 CAR T cells were used in clinical trial for patients with refractory B cell lymphoma and hematological malignancies. No acute graft versus host disease (GVHD) has been reported in patients except for one mild chronic ocular GVHD that was observed 2 years after CAR T cells infusion [113]. In 2017, FDA-approved Tisagenlecleucel, CD19 CAR T cell, for the treatment of acute lymphoblastic

leukemia (ALL). Excellent results with these trials, increased interests in CAR T cell immunotherapy approach [114, 115]. Cytokine release syndrome (CRS) is one of the side effects of CAR T Cells. CRS is a storm of inflammatory cytokines including IL-6, IL-10, and IFN-γ that happens after the infusion of CAR T cells [2]. Patients may show symptoms such as hypotension, pulmonary edema, multi-organ failure, and even CRS-related death. Treatment of CRS includes administration of corticosteroids and IL-6 blockade. Using corticosteroids for treatment of CRS symptoms is controversial as corticosteroids dramatically decrease inflammatory cytokines and mitigate CAR T cells efficacy [116]. Another problem with CAR T cells is that they cannot penetrate into solid tumors. Studies are underway to alleviate limitations of

CAR T cells and improve their efficacy for treatment of solid tumors [117].

Many cancer patients do not benefit from immunotherapies they are receiving. Recently, many studies are focusing on identifying predictive and prognostic

**8. Developing personalized immunotherapy**

#### *Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

*Current Trends in Cancer Management*

and help to stabilize RNAs [102].

**7. Adoptive cell therapy**

vaccines, DNA plasmids transfect both myocytes and DCs. The plasmids act as an immunogenic and activate T cells via toll-like receptors [95]. DNA sensors in cytosol of cells such as DAI, H2B, IFI16, DDX41, LRRFIP1, and cGAS are able to detect presence of DNA vaccines. DNA sensors send signal to STING-TBK1 signaling cascade and activate interferon regulatory factor 3 which results in expression of type I interferons. TLR9 recognizes unmethylated CpG DNA and activates interferon regulatory factor 7 that induce expression of interferons. DCs phagocyte antigenexpressing cell (myocytes) and cross present antigens on MHC-I to CD8 T cells. Moreover, interferons promote this pathway. If DNA vaccines directly transfect DCs, DCs are able to uptake, process, and present antigens on MHC-I to CD8 T cells [96]. Transfection of the vector with multiple gene sequences increases the immunization and induces humoral [97] and CD8 T cell response [98]. Combination of DC vaccines with other immune stimulatory agents such as TLR agonists [99], or monoclonal antibodies [100] increase anticancer immunity. RNA vaccines are safe vaccines compared to DNA vaccines as they degrade and clear quickly in body. Total tumor RNAs are isolated from tumor tissues and they can induce a potent immune response. RNA vaccines are composed of various tumor antigens which reduce the possibility of immune escape by tumor cells. The first use of RNA vaccines was to immunize patients with mRNAs that encode tumor-associated antigens. Furthermore, RNA vaccines can be produced for personalized cancer treatment. Patients' neoantigens can be identified by tumor exome analyzing and personalized RNA vaccine can be specifically generated. In addition to direct use of mRNAs for vaccine generation, RNAs are utilized in cell therapies. Transfecting patient-derived cells with RNAs and giving manipulated cells back to patients are another form of utilizing mRNAs. For example, transfection of patient-derived DCs with mRNA of tumor-associated antigens can induce an antigen-specific T cell response in cancer patients. Transfection of patient-derived T cells with mRNA of chimeric antigen receptors, triggers T cells to identify specific antigens on cancer cells which quickly deteriorate cancer [101]. Liposomes and protaminase are adjuvants of RNA vaccines

Adoptive T cell therapy (ACT) is a treatment that enhances T cells' ability to kill cancer cells by transferring immune system-derived cells to patients. The cells used for ACT can originate from the same patient or another individual. In 1988, the first ACT reduced metastatic melanoma tumors with transferring of autologous CD4+ and CD8+ tumor infiltrating lymphocytes (TILs) to the patients [103, 104]. Both peripheral blood T cells and TILs extracted from tumors are utilized to generate specific T cells for ACT. These T cells can be modified and then transferred to patients or directly administered in their natural state. TILs by their own nature have an antitumor activity as they are specific for tumor cells. TILs can recognize tumor antigens such as cancer germline antigens, neoantigens, and viral proteins and kill cancer cells [104]. After tumors are resected, the tumor tissues digest into fragments and each fragment is cultured in the presence of IL-2. The T cells are expanded and each clone is monitored for its reactivity against tumor cells. Proliferating lymphocytes kill tumor cells and produce a pure population of T cells. Cancer reactive T cells are infused back to patients. Moreover, T cells that express a TCR specific for tumor antigens can be selected in vitro from peripheral blood and expanded. Antigen-specific T cells are selected by coculturing of T cells with APCs loaded with tumor particles such as RNAs. By expansion of antigen-specific T cells, a specific antitumor T cell clone can be generated [105]. T cells with TCR targeting

**28**

tumorigenic mutations such as Ras mutations have shown promise in cancer treatment. Ras is commonly mutated at the onset of tumorigenesis in the dominant population of tumor cells. Targeting Ras mutations and killing tumor cells with Ras-specific ACT may have profound effects on cancers with Ras mutations [106]. TCRs targeting KRAS G12D, a common proto-oncogene encoding GTPase, have anti-tumorigenic effects on patients with colorectal cancer [107]. Also, genetically modified antitumor T cell clones can be produced by infecting T cells with viruses that carry genetically engineered TCRs [108]. TCR-transduced T cells are generated by cloning specific TCRs into a retrovirus. Patients derived PBMCs are activated with CD3 and IL-2 and are transduced with the retrovirus encoding the antigen-specific TCR. The T cells are expanded and injected back to the individuals. Peripheral blood T cells transfected with retrovirus encoding MART-1 TCR regress tumors in melanoma [103]. Genetically engineering techniques can modify TCRs to target-specific antigens. For example, T cells with modified TCRs that target NY-ESO-1, a cancer germline antigen, were successfully used as ACT for treatment of patients with synovial cell sarcoma and melanoma [109]. One major limitation of ACTs is that they induce short-lasting responses in immune system. Administration of T cells after chemotherapy increases cancer regression due to repopulation of host T cells with antigen-specific T cells. Lymphodepletion induced by chemotherapy helps T cells from ACT to proliferate during hemostatic proliferative phase and persist for months after infusion [109]. It was also shown that high doses of IL-2 therapy contribute to expansion of the transferred cells [110, 111]. The first signal in T cell activation begins with binding of TCR to MHC molecules on APCs. Furthermore, MHC expression downregulates on APCs in cancers so that they can escape immunity [112]. In 1989, first chimeric antigen receptors (CARs) were developed to avoid interaction of T cells with MHC molecules. CAR T cells are designed to identify cancer cells and attack them without mediation of APCs. As a result, CARs act independent of any stimulatory and TCR signaling. CAR composed of a ligand-binding domain and a signaling domain. Ligand-binding domain is the extracellular part of CAR that includes B cell receptor derived single chain variable fragment. The signaling domain is made of costimulatory molecules and CD3f and 1 [112]. CD19 CAR T cells were used in clinical trial for patients with refractory B cell lymphoma and hematological malignancies. No acute graft versus host disease (GVHD) has been reported in patients except for one mild chronic ocular GVHD that was observed 2 years after CAR T cells infusion [113]. In 2017, FDA-approved Tisagenlecleucel, CD19 CAR T cell, for the treatment of acute lymphoblastic leukemia (ALL). Excellent results with these trials, increased interests in CAR T cell immunotherapy approach [114, 115]. Cytokine release syndrome (CRS) is one of the side effects of CAR T Cells. CRS is a storm of inflammatory cytokines including IL-6, IL-10, and IFN-γ that happens after the infusion of CAR T cells [2]. Patients may show symptoms such as hypotension, pulmonary edema, multi-organ failure, and even CRS-related death. Treatment of CRS includes administration of corticosteroids and IL-6 blockade. Using corticosteroids for treatment of CRS symptoms is controversial as corticosteroids dramatically decrease inflammatory cytokines and mitigate CAR T cells efficacy [116]. Another problem with CAR T cells is that they cannot penetrate into solid tumors. Studies are underway to alleviate limitations of CAR T cells and improve their efficacy for treatment of solid tumors [117].

#### **8. Developing personalized immunotherapy**

Many cancer patients do not benefit from immunotherapies they are receiving. Recently, many studies are focusing on identifying predictive and prognostic biomarkers in cancers as a beneficial guide for treatment decisions. This will stop administration of drugs for those patients who does not benefit from them and improve treatment in patients that are most likely respond to specific immunotherapies. Selecting the appropriate immunotherapy for each cancer patient is still a challenge. Scientists and oncologists are developing methods in genomic testing to discover cell signaling and biomarkers involved in responding to immunotherapy. It has been shown that cancers identified by specific quantity or pattern of mutations in the tumor microenvironment or surrounding area are more responsive to immune checkpoint blockades. Of note, scientists are trying to exploit other drugs to alter the tumor microenvironment of less immune responsive tumors, known as cold tumors, and turn them to check point blockades susceptible tumors that are defined as hot tumors [32]. Altering tumor microenvironment and surrounding tissues can increase the number of patients who can benefit from immune checkpoint blockades. Immunopharmacogenomics approach is providing a significant hope for personalized immunotherapy [118].

#### **9. Conclusion**

In summary, immunotherapy shows a tremendous potential in treatment of cancer. Different immunotherapies have been approved by FDA for prevention and treatment of cancers. Despite the breakthroughs achieved by immunotherapy, many cancers still do not respond to immunotherapy. Monotherapy of immune checkpoint blockades or other immunotherapies failed in treatment of some cancers. Finding the efficient treatment by combinatorial immunotherapies or combination of immunotherapy and traditional chemotherapy and radiotherapy are under investigation. Development of DCs and cancer vaccines, immune checkpoint blockades, CAR T cells, and ACT requires an in-depth understanding of tumor microenvironment and identifying tumor-specific antigens. More studies to develop immunotherapy can provide improved efficacy in cancer treatments.

#### **Author details**

Aida Karachi UF Brain Tumor Immunotherapy Program, Lillian S. Wells Department of Neurosurgery, Preston A. Wells Jr. Center for Brain Tumor Therapy, University of Florida, Gainesville, FL, USA

\*Address all correspondence to: aidakarachi@yahoo.com

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

**31**

*Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

[1] Coley WB. II. Contribution to the knowledge of sarcoma. Annals of Surgery. 1891;**14**(3):199-220

I study. Journal of Clinical Oncology.

[10] Amiri-Kordestani L, Blumenthal GM, Xu QC, et al. FDA approval: Ado-trastuzumab emtansine for the treatment of patients with HER2-positive metastatic breast cancer. Clinical Cancer Research.

[11] Ehrlich D, Wang B, Lu W, Dowling P, Yuan R. Intratumoral anti-HuD immunotoxin therapy for small cell lung cancer and neuroblastoma. Journal of Hematology & Oncology. 2014;**7**:91

[13] Hansel TT, Kropshofer H, Singer T, Mitchell JA, George AJ. The safety and side effects of monoclonal antibodies. Nature Reviews. Drug Discovery.

[14] Seimetz D. Novel monoclonal antibodies for cancer treatment: The trifunctional antibody catumaxomab (removab). Journal of Cancer.

[15] Reichert JM, Rosensweig CJ, Faden LB, Dewitz MC. Monoclonal antibody successes in the clinic. Nature Biotechnology. 2005;**23**(9):1073-1078

[16] Lonberg N. Human monoclonal antibodies from transgenic mice. Handbook of Experimental Pharmacology. 2008;**181**:69-97

[17] Topalian SL, Sznol M, McDermott DF, et al. Survival, durable tumor remission, and long-term safety in patients with advanced melanoma receiving nivolumab. Journal of Clinical Oncology. 2014;**32**(10):1020-1030

2014;**32**(28):3137-3143

2014;**20**(17):4436-4441

[12] Kaplan JB, Grischenko M, Giles FJ. Blinatumomab for the treatment of acute lymphoblastic leukemia. Investigational New Drugs.

2015;**33**(6):1271-1279

2010;**9**(4):325-338

2011;**2**:309-316

[2] Maude SL, Barrett D, Teachey DT, Grupp SA. Managing cytokine release syndrome associated with novel T cell-engaging therapies. Cancer Journal.

[3] Coulson A, Levy A, Gossell-Williams M. Monoclonal antibodies in cancer therapy: Mechanisms, successes and limitations. The West Indian Medical

[4] Vanneman M, Dranoff G. Combining immunotherapy and targeted therapies in cancer treatment. Nature Reviews.

**References**

2014;**20**(2):119-122

Journal. 2014;**63**(6):650-654

Cancer. 2012;**12**(4):237-251

2013;**22**(Suppl 2):S152-S155

[7] Steiner M, Neri D. Antibodyradionuclide conjugates for cancer therapy: Historical considerations and new trends. Clinical Cancer Research.

[8] Flygare JA, Pillow TH, Aristoff P. Antibody-drug conjugates for the treatment of cancer. Chemical Biology & Drug Design. 2013;**81**(1):113-121

Forero-Torres A, et al. Brentuximab vedotin in the front-line treatment of patients with CD30+ peripheral T-cell lymphomas: Results of a phase

[9] Fanale MA, Horwitz SM,

2011;**17**(20):6406-6416

[5] Pinto AC, Ades F, de Azambuja E, Piccart-Gebhart M. Trastuzumab for patients with HER2 positive breast cancer: Delivery, duration and combination therapies. Breast.

[6] Fiegl M, Stauder R, Steurer M, et al. Alemtuzumab in chronic lymphocytic leukemia: Final results of a large observational multicenter study in mostly pretreated patients. Annals of Hematology. 2014;**93**(2):267-277

*Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

#### **References**

*Current Trends in Cancer Management*

personalized immunotherapy [118].

**9. Conclusion**

biomarkers in cancers as a beneficial guide for treatment decisions. This will stop administration of drugs for those patients who does not benefit from them and improve treatment in patients that are most likely respond to specific immunotherapies. Selecting the appropriate immunotherapy for each cancer patient is still a challenge. Scientists and oncologists are developing methods in genomic testing to discover cell signaling and biomarkers involved in responding to immunotherapy. It has been shown that cancers identified by specific quantity or pattern of mutations in the tumor microenvironment or surrounding area are more responsive to immune checkpoint blockades. Of note, scientists are trying to exploit other drugs to alter the tumor microenvironment of less immune responsive tumors, known as cold tumors, and turn them to check point blockades susceptible tumors that are defined as hot tumors [32]. Altering tumor microenvironment and surrounding tissues can increase the number of patients who can benefit from immune checkpoint blockades. Immunopharmacogenomics approach is providing a significant hope for

In summary, immunotherapy shows a tremendous potential in treatment of cancer. Different immunotherapies have been approved by FDA for prevention and treatment of cancers. Despite the breakthroughs achieved by immunotherapy, many cancers still do not respond to immunotherapy. Monotherapy of immune checkpoint blockades or other immunotherapies failed in treatment of some cancers. Finding the efficient treatment by combinatorial immunotherapies or combination of immunotherapy and traditional chemotherapy and radiotherapy are under investigation. Development of DCs and cancer vaccines, immune checkpoint blockades, CAR T cells, and ACT requires an in-depth understanding of tumor microenvironment and identifying tumor-specific antigens. More studies to develop immunotherapy can provide improved efficacy in cancer treatments.

**30**

**Author details**

Florida, Gainesville, FL, USA

Aida Karachi

provided the original work is properly cited.

\*Address all correspondence to: aidakarachi@yahoo.com

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

UF Brain Tumor Immunotherapy Program, Lillian S. Wells Department of Neurosurgery, Preston A. Wells Jr. Center for Brain Tumor Therapy, University of [1] Coley WB. II. Contribution to the knowledge of sarcoma. Annals of Surgery. 1891;**14**(3):199-220

[2] Maude SL, Barrett D, Teachey DT, Grupp SA. Managing cytokine release syndrome associated with novel T cell-engaging therapies. Cancer Journal. 2014;**20**(2):119-122

[3] Coulson A, Levy A, Gossell-Williams M. Monoclonal antibodies in cancer therapy: Mechanisms, successes and limitations. The West Indian Medical Journal. 2014;**63**(6):650-654

[4] Vanneman M, Dranoff G. Combining immunotherapy and targeted therapies in cancer treatment. Nature Reviews. Cancer. 2012;**12**(4):237-251

[5] Pinto AC, Ades F, de Azambuja E, Piccart-Gebhart M. Trastuzumab for patients with HER2 positive breast cancer: Delivery, duration and combination therapies. Breast. 2013;**22**(Suppl 2):S152-S155

[6] Fiegl M, Stauder R, Steurer M, et al. Alemtuzumab in chronic lymphocytic leukemia: Final results of a large observational multicenter study in mostly pretreated patients. Annals of Hematology. 2014;**93**(2):267-277

[7] Steiner M, Neri D. Antibodyradionuclide conjugates for cancer therapy: Historical considerations and new trends. Clinical Cancer Research. 2011;**17**(20):6406-6416

[8] Flygare JA, Pillow TH, Aristoff P. Antibody-drug conjugates for the treatment of cancer. Chemical Biology & Drug Design. 2013;**81**(1):113-121

[9] Fanale MA, Horwitz SM, Forero-Torres A, et al. Brentuximab vedotin in the front-line treatment of patients with CD30+ peripheral T-cell lymphomas: Results of a phase I study. Journal of Clinical Oncology. 2014;**32**(28):3137-3143

[10] Amiri-Kordestani L, Blumenthal GM, Xu QC, et al. FDA approval: Ado-trastuzumab emtansine for the treatment of patients with HER2-positive metastatic breast cancer. Clinical Cancer Research. 2014;**20**(17):4436-4441

[11] Ehrlich D, Wang B, Lu W, Dowling P, Yuan R. Intratumoral anti-HuD immunotoxin therapy for small cell lung cancer and neuroblastoma. Journal of Hematology & Oncology. 2014;**7**:91

[12] Kaplan JB, Grischenko M, Giles FJ. Blinatumomab for the treatment of acute lymphoblastic leukemia. Investigational New Drugs. 2015;**33**(6):1271-1279

[13] Hansel TT, Kropshofer H, Singer T, Mitchell JA, George AJ. The safety and side effects of monoclonal antibodies. Nature Reviews. Drug Discovery. 2010;**9**(4):325-338

[14] Seimetz D. Novel monoclonal antibodies for cancer treatment: The trifunctional antibody catumaxomab (removab). Journal of Cancer. 2011;**2**:309-316

[15] Reichert JM, Rosensweig CJ, Faden LB, Dewitz MC. Monoclonal antibody successes in the clinic. Nature Biotechnology. 2005;**23**(9):1073-1078

[16] Lonberg N. Human monoclonal antibodies from transgenic mice. Handbook of Experimental Pharmacology. 2008;**181**:69-97

[17] Topalian SL, Sznol M, McDermott DF, et al. Survival, durable tumor remission, and long-term safety in patients with advanced melanoma receiving nivolumab. Journal of Clinical Oncology. 2014;**32**(10):1020-1030

[18] Naidoo J, Page DB, Li BT, et al. Toxicities of the anti-PD-1 and anti-PD-L1 immune checkpoint antibodies. Annals of Oncology. 2016;**27**(7):1362

[19] Littman DR. Releasing the brakes on cancer immunotherapy. Cell. 2015;**162**(6):1186-1190

[20] Sharma P, Allison JP. Immune checkpoint targeting in cancer therapy: Toward combination strategies with curative potential. Cell. 2015;**161**(2):205-214

[21] Tsai HF, Hsu PN. Cancer immunotherapy by targeting immune checkpoints: Mechanism of T cell dysfunction in cancer immunity and new therapeutic targets. Journal of Biomedical Science. 2017;**24**(1):35

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[23] Collins AV, Brodie DW, Gilbert RJ, et al. The interaction properties of costimulatory molecules revisited. Immunity. 2002;**17**(2):201-210

[24] Hodi FS, O'Day SJ, McDermott DF, et al. Improved survival with ipilimumab in patients with metastatic melanoma. The New England Journal of Medicine. 2010;**363**(8):711-723

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PD-1 and PD-L1 pathway. Trends in Molecular Medicine. 2015;**21**(1):24-33

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[32] Karachi A, Dastmalchi F, Mitchell D, Rahman M. Temozolomide for immunomodulation in the treatment of glioblastoma. Neuro-Oncology. 2018; noy072. https://doi.org/10.1093/neuonc/ noy072

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[43] Emens LA, Asquith JM, Leatherman JM, et al. Timed sequential treatment with cyclophosphamide, doxorubicin, and an allogeneic granulocytemacrophage colony-stimulating factor-secreting breast tumor vaccine: A chemotherapy dose-ranging factorial study of safety and immune activation. Journal of Clinical Oncology. 2009;**27**(35):5911-5918

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[46] Nemunaitis J, Nemunaitis M, Senzer N, et al. Phase II trial of Belagenpumatucel-L, a TGF-beta2 antisense gene modified allogeneic tumor vaccine in advanced non small cell lung cancer (NSCLC) patients. Cancer Gene Therapy. 2009;**16**(8):620-624

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**32**

661-672

*Current Trends in Cancer Management*

[18] Naidoo J, Page DB, Li BT, et al. Toxicities of the anti-PD-1 and anti-PD-L1 immune checkpoint antibodies. Annals of Oncology. 2016;**27**(7):1362

PD-1 and PD-L1 pathway. Trends in Molecular Medicine. 2015;**21**(1):24-33

oncometabolite, 2-hydroxyglutarate, may affect DNA methylation and expression of PD-L1 in gliomas. Frontiers in Molecular Neuroscience.

[29] Karachi A, Azari H, Flores C, Yang C, Dastmalchi F, Mitchell D, et al. TMZ results in priming of host immunity and changes in GBM tumor PDL-1 expression in a dose dependent fashion that can be leveraged for combination with immune checkpoint blockade: December 2016. Neuro-Oncology;**18**(Suppl 6):vi202

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[32] Karachi A, Dastmalchi F, Mitchell D, Rahman M. Temozolomide for immunomodulation in the treatment of glioblastoma. Neuro-Oncology. 2018; noy072. https://doi.org/10.1093/neuonc/

[33] Postow MA, Chesney J, Pavlick AC, et al. Nivolumab and ipilimumab versus ipilimumab in untreated melanoma. The New England Journal of Medicine.

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KEYNOTE-021 study. Lancet Oncology.

2015;**372**(21):2006-2017

2016;**17**(11):1497-1508

Expert Review of Clinical

noy072

Pharmacology. 2018;**11**(4):345-359

[28] Mu L, Long Y, Yang C, et al. The IDH1 mutation-induced

2018;**11**:82

[19] Littman DR. Releasing the brakes on cancer immunotherapy. Cell.

[20] Sharma P, Allison JP. Immune checkpoint targeting in cancer therapy: Toward combination

strategies with curative potential. Cell.

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[23] Collins AV, Brodie DW, Gilbert RJ, et al. The interaction properties of costimulatory molecules revisited. Immunity. 2002;**17**(2):201-210

[24] Hodi FS, O'Day SJ, McDermott DF, et al. Improved survival with ipilimumab in patients with metastatic melanoma. The New England Journal of

Medicine. 2010;**363**(8):711-723

Cancer Medicine. 2015;**4**(5):

[26] McDermott J, Jimeno

Today (Barc). 2015;**51**(1):7-20

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A. Pembrolizumab: PD-1 inhibition as a therapeutic strategy in cancer. Drugs

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2015;**162**(6):1186-1190

2015;**161**(2):205-214

2008;**224**:166-182

[21] Tsai HF, Hsu PN. Cancer

[49] Dudziak D, Kamphorst AO, Heidkamp GF, et al. Differential antigen processing by dendritic cell subsets in vivo. Science. 2007;**315**(5808):107-111

[50] Karachi A, Fazeli M, Karimi MH, et al. Evaluation of immunomodulatory effects of mesenchymal stem cells soluble factors on miR-155 and miR-23b expression in mice dendritic cells. Immunological Investigations. 2015;**44**(5):427-437

[51] Dastmalchi F, Karachi A, Azari H, Mitchell D, Rahman M. Strategy to enhance DC migration for increased efficacy of dendritic cells vaccine immunotherapy. Neuro-Oncology. 2017;**19**(suppl\_6):vi125

[52] Frankenberger B, Schendel DJ. Third generation dendritic cell vaccines for tumor immunotherapy. European Journal of Cell Biology. 2012;**91**(1):53-58

[53] Dastmalchi F, Karachi A, Mitchell D, Rahman M. Dendritic Cell Therapy. In: eLS. Chichester: John Wiley & Sons, Ltd; 2018. DOI: 10.1002/9780470015902.a0024243

[54] Batich KA, Reap EA, Archer GE, et al. Long-term survival in glioblastoma with cytomegalovirus pp65-targeted vaccination. Clinical Cancer Research. 2017;**23**(8):1898-1909

[55] Nair SK, Morse M, Boczkowski D, et al. Induction of tumor-specific cytotoxic T lymphocytes in cancer patients by autologous tumor RNAtransfected dendritic cells. Annals of Surgery. 2002;**235**(4):540-549

[56] Murphy G, Tjoa B, Ragde H, Kenny G, Boynton A. Phase I clinical trial: T-cell therapy for prostate cancer using autologous dendritic cells pulsed with HLA-A0201-specific peptides from prostate-specific membrane antigen. The Prostate. 1996;**29**(6):371-380

[57] Schuler-Thurner B, Schultz ES, Berger TG, et al. Rapid induction

of tumor-specific type 1 T helper cells in metastatic melanoma patients by vaccination with mature, cryopreserved, peptide-loaded monocyte-derived dendritic cells. The Journal of Experimental Medicine. 2002;**195**(10):1279-1288

[58] Rosenblatt J, Vasir B, Uhl L, et al. Vaccination with dendritic cell/tumor fusion cells results in cellular and humoral antitumor immune responses in patients with multiple myeloma. Blood. 2011;**117**(2):393-402

[59] Longo DL. New therapies for castration-resistant prostate cancer. The New England Journal of Medicine. 2010;**363**(5):479-481

[60] Bonehill A, Tuyaerts S, Van Nuffel AM, et al. Enhancing the T-cell stimulatory capacity of human dendritic cells by co-electroporation with CD40L, CD70 and constitutively active TLR4 encoding mRNA. Molecular Therapy. 2008;**16**(6):1170-1180

[61] Dannull J, Nair S, Su Z, et al. Enhancing the immunostimulatory function of dendritic cells by transfection with mRNA encoding OX40 ligand. Blood. 2005;**105**(8):3206-3213

[62] Grunebach F, Kayser K, Weck MM, Muller MR, Appel S, Brossart P. Cotransfection of dendritic cells with RNA coding for HER-2/neu and 4-1BBL increases the induction of tumor antigen specific cytotoxic T lymphocytes. Cancer Gene Therapy. 2005;**12**(9):749-756

[63] Tuyaerts S, Aerts JL, Corthals J, et al. Current approaches in dendritic cell generation and future implications for cancer immunotherapy. Cancer Immunology, Immunotherapy. 2007;**56**(10):1513-1537

**35**

*Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

protective and therapeutic effects of IL-12 and IL-18 gene-transduced dendritic neuroblastoma fusion cells on liver metastasis of murine neuroblastoma. Journal of Immunology. directly inhibiting ubiquitination of tumor necrosis factor (TNF) receptor-associated factor 6. The Journal of Biological Chemistry. 2011;**286**(21):18795-18806

[72] van der Bruggen P, Traversari C, Chomez P, et al. A gene encoding an antigen recognized by cytolytic T lymphocytes on a human melanoma. Science. 1991;**254**(5038):1643-1647

[73] De Smet C, Lurquin C, van der Bruggen P, De Plaen E, Brasseur F, Boon T. Sequence and expression pattern of the human MAGE2 gene. Immunogenetics. 1994;**39**(2):121-129

[74] Gnjatic S, Ritter E, Buchler MW, et al. Seromic profiling of ovarian and pancreatic cancer. Proceedings of the National Academy of Sciences of the United States of America.

[75] Karbach J, Neumann A, Atmaca A, et al. Efficient in vivo priming by vaccination with recombinant NY-ESO-1 protein and CpG in antigen naive prostate cancer patients. Clinical

[76] Correale P, Walmsley K, Nieroda C, et al. In vitro generation of human cytotoxic T lymphocytes specific for peptides derived from prostate-specific antigen. Journal of the National Cancer

[77] Kantoff PW, Higano CS, Shore ND, et al. Sipuleucel-T immunotherapy for castration-resistant prostate cancer. The New England Journal of Medicine.

2010;**107**(11):5088-5093

Cancer Research. 2011;**17**(4):

Institute. 1997;**89**(4):293-300

[78] Bakker AB, Schreurs MW, de Boer AJ, et al. Melanocyte lineage-specific antigen gp100 is recognized by melanoma-derived tumor-infiltrating lymphocytes. The Journal of Experimental Medicine.

2010;**363**(5):411-422

1994;**179**(3):1005-1009

861-870

[65] Kang TH, Bae HC, Kim SH, et al. Modification of dendritic cells with interferon-gamma-inducible protein-10 gene to enhance vaccine potency. The Journal of Gene Medicine.

[66] Minkis K, Kavanagh DG, Alter G, et al. Type 2 bias of T cells expanded from the blood of melanoma patients switched to type 1 by IL-12p70 mRNAtransfected dendritic cells. Cancer Research. 2008;**68**(22):9441-9450

[67] Ogawa F, Iinuma H, Okinaga K. Dendritic cell vaccine therapy by immunization with fusion cells of interleukin-2 gene-transduced, spleenderived dendritic cells and tumour cells. Scandinavian Journal of Immunology.

[68] Okada N, Mori N, Koretomo R, et al. Augmentation of the migratory ability of DC-based vaccine into

regional lymph nodes by efficient CCR7 gene transduction. Gene Therapy.

[69] Song XT, Evel-Kabler K, Shen L, Rollins L, Huang XF, Chen SY. A20 is an antigen presentation attenuator, and its inhibition overcomes regulatory T cellmediated suppression. Nature Medicine.

[70] Palmer DC, Restifo NP. Suppressors of cytokine signaling (SOCS) in T cell differentiation, maturation, and function. Trends in Immunology.

[71] Yu X, Yi H, Guo C, et al. Pattern recognition scavenger receptor CD204 attenuates Toll-like receptor 4-induced NF-kappaB activation by

2006;**176**(6):3461-3469

2009;**11**(10):889-898

2004;**59**(5):432-439

2005;**12**(2):129-139

2008;**14**(3):258-265

2009;**30**(12):592-602

[64] Iinuma H, Okinaga K, Fukushima R, et al. Superior *Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

*Current Trends in Cancer Management*

[49] Dudziak D, Kamphorst AO,

2015;**44**(5):427-437

2017;**19**(suppl\_6):vi125

2012;**91**(1):53-58

2017;**23**(8):1898-1909

[52] Frankenberger B, Schendel DJ. Third generation dendritic cell vaccines for tumor immunotherapy. European Journal of Cell Biology.

[53] Dastmalchi F, Karachi A, Mitchell D, Rahman M. Dendritic Cell Therapy. In: eLS. Chichester: John Wiley & Sons, Ltd; 2018. DOI: 10.1002/9780470015902.a0024243

[54] Batich KA, Reap EA, Archer GE, et al. Long-term survival in glioblastoma with cytomegalovirus pp65-targeted vaccination. Clinical Cancer Research.

[55] Nair SK, Morse M, Boczkowski D, et al. Induction of tumor-specific cytotoxic T lymphocytes in cancer patients by autologous tumor RNAtransfected dendritic cells. Annals of Surgery. 2002;**235**(4):540-549

[56] Murphy G, Tjoa B, Ragde H, Kenny G, Boynton A. Phase I clinical trial: T-cell therapy for prostate cancer using autologous dendritic cells pulsed with HLA-A0201-specific peptides from prostate-specific membrane antigen. The Prostate. 1996;**29**(6):371-380

[57] Schuler-Thurner B, Schultz ES, Berger TG, et al. Rapid induction

Heidkamp GF, et al. Differential antigen processing by dendritic cell subsets in vivo. Science. 2007;**315**(5808):107-111

of tumor-specific type 1 T helper cells in metastatic melanoma

2002;**195**(10):1279-1288

Blood. 2011;**117**(2):393-402

2010;**363**(5):479-481

2008;**16**(6):1170-1180

2005;**12**(9):749-756

2007;**56**(10):1513-1537

[64] Iinuma H, Okinaga K, Fukushima R, et al. Superior

[59] Longo DL. New therapies for castration-resistant prostate cancer. The New England Journal of Medicine.

[60] Bonehill A, Tuyaerts S, Van Nuffel AM, et al. Enhancing the T-cell stimulatory capacity of human dendritic cells by co-electroporation with CD40L, CD70 and constitutively active TLR4 encoding mRNA. Molecular Therapy.

[61] Dannull J, Nair S, Su Z, et al. Enhancing the immunostimulatory

[62] Grunebach F, Kayser K, Weck MM, Muller MR, Appel S, Brossart P. Cotransfection of dendritic cells with RNA coding for HER-2/neu and 4-1BBL increases the induction of tumor antigen specific cytotoxic T lymphocytes. Cancer Gene Therapy.

[63] Tuyaerts S, Aerts JL, Corthals J, et al. Current approaches in dendritic cell generation and future implications for cancer immunotherapy. Cancer Immunology, Immunotherapy.

function of dendritic cells by transfection with mRNA encoding OX40 ligand. Blood. 2005;**105**(8):3206-3213

patients by vaccination with mature, cryopreserved, peptide-loaded monocyte-derived dendritic cells. The Journal of Experimental Medicine.

[58] Rosenblatt J, Vasir B, Uhl L, et al. Vaccination with dendritic cell/tumor fusion cells results in cellular and humoral antitumor immune responses in patients with multiple myeloma.

[50] Karachi A, Fazeli M, Karimi MH, et al. Evaluation of immunomodulatory effects of mesenchymal stem cells soluble factors on miR-155 and miR-23b expression in mice dendritic cells. Immunological Investigations.

[51] Dastmalchi F, Karachi A, Azari H, Mitchell D, Rahman M. Strategy to enhance DC migration for increased efficacy of dendritic cells vaccine immunotherapy. Neuro-Oncology.

**34**

protective and therapeutic effects of IL-12 and IL-18 gene-transduced dendritic neuroblastoma fusion cells on liver metastasis of murine neuroblastoma. Journal of Immunology. 2006;**176**(6):3461-3469

[65] Kang TH, Bae HC, Kim SH, et al. Modification of dendritic cells with interferon-gamma-inducible protein-10 gene to enhance vaccine potency. The Journal of Gene Medicine. 2009;**11**(10):889-898

[66] Minkis K, Kavanagh DG, Alter G, et al. Type 2 bias of T cells expanded from the blood of melanoma patients switched to type 1 by IL-12p70 mRNAtransfected dendritic cells. Cancer Research. 2008;**68**(22):9441-9450

[67] Ogawa F, Iinuma H, Okinaga K. Dendritic cell vaccine therapy by immunization with fusion cells of interleukin-2 gene-transduced, spleenderived dendritic cells and tumour cells. Scandinavian Journal of Immunology. 2004;**59**(5):432-439

[68] Okada N, Mori N, Koretomo R, et al. Augmentation of the migratory ability of DC-based vaccine into regional lymph nodes by efficient CCR7 gene transduction. Gene Therapy. 2005;**12**(2):129-139

[69] Song XT, Evel-Kabler K, Shen L, Rollins L, Huang XF, Chen SY. A20 is an antigen presentation attenuator, and its inhibition overcomes regulatory T cellmediated suppression. Nature Medicine. 2008;**14**(3):258-265

[70] Palmer DC, Restifo NP. Suppressors of cytokine signaling (SOCS) in T cell differentiation, maturation, and function. Trends in Immunology. 2009;**30**(12):592-602

[71] Yu X, Yi H, Guo C, et al. Pattern recognition scavenger receptor CD204 attenuates Toll-like receptor 4-induced NF-kappaB activation by directly inhibiting ubiquitination of tumor necrosis factor (TNF) receptor-associated factor 6. The Journal of Biological Chemistry. 2011;**286**(21):18795-18806

[72] van der Bruggen P, Traversari C, Chomez P, et al. A gene encoding an antigen recognized by cytolytic T lymphocytes on a human melanoma. Science. 1991;**254**(5038):1643-1647

[73] De Smet C, Lurquin C, van der Bruggen P, De Plaen E, Brasseur F, Boon T. Sequence and expression pattern of the human MAGE2 gene. Immunogenetics. 1994;**39**(2):121-129

[74] Gnjatic S, Ritter E, Buchler MW, et al. Seromic profiling of ovarian and pancreatic cancer. Proceedings of the National Academy of Sciences of the United States of America. 2010;**107**(11):5088-5093

[75] Karbach J, Neumann A, Atmaca A, et al. Efficient in vivo priming by vaccination with recombinant NY-ESO-1 protein and CpG in antigen naive prostate cancer patients. Clinical Cancer Research. 2011;**17**(4): 861-870

[76] Correale P, Walmsley K, Nieroda C, et al. In vitro generation of human cytotoxic T lymphocytes specific for peptides derived from prostate-specific antigen. Journal of the National Cancer Institute. 1997;**89**(4):293-300

[77] Kantoff PW, Higano CS, Shore ND, et al. Sipuleucel-T immunotherapy for castration-resistant prostate cancer. The New England Journal of Medicine. 2010;**363**(5):411-422

[78] Bakker AB, Schreurs MW, de Boer AJ, et al. Melanocyte lineage-specific antigen gp100 is recognized by melanoma-derived tumor-infiltrating lymphocytes. The Journal of Experimental Medicine. 1994;**179**(3):1005-1009

[79] Kawakami Y, Eliyahu S, Sakaguchi K, et al. Identification of the immunodominant peptides of the MART-1 human melanoma antigen recognized by the majority of HLA-A2-restricted tumor infiltrating lymphocytes. The Journal of Experimental Medicine. 1994;**180**(1):347-352

[80] Parkhurst MR, Fitzgerald EB, Southwood S, Sette A, Rosenberg SA, Kawakami Y. Identification of a shared HLA-A\*0201-restricted T-cell epitope from the melanoma antigen tyrosinaserelated protein 2 (TRP2). Cancer Research. 1998;**58**(21):4895-4901

[81] Jaramillo A, Majumder K, Manna PP, et al. Identification of HLA-A3 restricted CD8+ T cell epitopes derived from mammaglobin-A, a tumorassociated antigen of human breast cancer. International Journal of Cancer. 2002;**102**(5):499-506

[82] Tsang KY, Zaremba S, Nieroda CA, Zhu MZ, Hamilton JM, Schlom J. Generation of human cytotoxic T cells specific for human carcinoembryonic antigen epitopes from patients immunized with recombinant vaccinia-CEA vaccine. Journal of the National Cancer Institute. 1995;**87**(13):982-990

[83] Finn OJ, Gantt KR, Lepisto AJ, Pejawar-Gaddy S, Xue J, Beatty PL. Importance of MUC1 and spontaneous mouse tumor models for understanding the immunobiology of human adenocarcinomas. Immunologic Research. 2011;**50**(2-3):261-268

[84] Disis ML, Wallace DR, Gooley TA, et al. Concurrent trastuzumab and HER2/neu-specific vaccination in patients with metastatic breast cancer. Journal of Clinical Oncology. 2009;**27**(28):4685-4692

[85] Schmollinger JC, Vonderheide RH, Hoar KM, et al. Melanoma inhibitor of apoptosis protein (ML-IAP)

is a target for immune-mediated tumor destruction. Proceedings of the National Academy of Sciences of the United States of America. 2003;**100**(6):3398-3403

[86] Schmidt SM, Schag K, Muller MR, et al. Survivin is a shared tumorassociated antigen expressed in a broad variety of malignancies and recognized by specific cytotoxic T cells. Blood. 2003;**102**(2):571-576

[87] Kawai T, Akira S. Toll-like receptors and their crosstalk with other innate receptors in infection and immunity. Immunity. 2011;**34**(5):637-650

[88] Heldwein KA, Liang MD, Andresen TK, et al. TLR2 and TLR4 serve distinct roles in the host immune response against Mycobacterium bovis BCG. Journal of Leukocyte Biology. 2003;**74**(2):277-286

[89] Mata-Haro V, Cekic C, Martin M, Chilton PM, Casella CR, Mitchell TC. The vaccine adjuvant monophosphoryl lipid A as a TRIFbiased agonist of TLR4. Science. 2007;**316**(5831):1628-1632

[90] Clarke MA, Wentzensen N, Mirabello L, et al. Human papillomavirus DNA methylation as a potential biomarker for cervical cancer. Cancer Epidemiology, Biomarkers & Prevention. 2012;**21**(12): 2125-2137

[91] Kumai T, Kobayashi H, Harabuchi Y, Celis E. Peptide vaccines in cancerold concept revisited. Current Opinion in Immunology. 2017;**45**:1-7

[92] Aurisicchio L, Ciliberto G. Genetic cancer vaccines: Current status and perspectives. Expert Opinion on Biological Therapy. 2012;**12**(8):1043-1058

[93] Liu MA. DNA vaccines: An historical perspective and view to

**37**

*Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

the future. Immunological Reviews.

[102] Fotin-Mleczek M, Zanzinger K, Heidenreich R, et al. Highly potent mRNA based cancer vaccines represent an attractive platform for combination therapies supporting an improved therapeutic effect. The Journal of Gene

personalized immunotherapy for human cancer. Science. 2015;**348**(6230):62-68

Medicine. 2012;**14**(6):428-439

[103] Rosenberg SA, Restifo NP. Adoptive cell transfer as

[104] Verdegaal EM. Adoptive cell therapy: A highly successful individualized therapy for melanoma

with great potential for other malignancies. Current Opinion in Immunology. 2016;**39**:90-95

[105] Yee C, Thompson JA, Byrd D, et al. Adoptive T cell therapy using antigen-specific CD8+ T cell clones for the treatment of patients with metastatic melanoma: in vivo persistence, migration, and antitumor effect of transferred T cells. Proceedings of the National Academy of Sciences of the United States of America. 2002;**99**(25):16168-16173

[106] Bryant KL, Mancias JD, Kimmelman AC, Der CJ. KRAS:

Trends in Biochemical Sciences.

[107] Tran E, Robbins PF, Lu YC, et al. T-cell transfer therapy targeting mutant KRAS in cancer. The New England Journal of Medicine. 2016;**375**(23):2255-2262

[108] Morgan RA, Dudley ME,

2006;**314**(5796):126-129

Wunderlich JR, et al. Cancer regression in patients after transfer of genetically engineered lymphocytes. Science.

[109] Robbins PF, Morgan RA, Feldman SA, et al. Tumor regression in patients with metastatic synovial cell sarcoma and melanoma using genetically engineered lymphocytes reactive with

2014;**39**(2):91-100

Feeding pancreatic cancer proliferation.

[94] Stratford R, Douce G, Zhang-Barber L, Fairweather N, Eskola J, Dougan G. Influence of codon usage on the immunogenicity of a DNA vaccine against tetanus. Vaccine.

2011;**239**(1):62-84

2000;**19**(7-8):810-815

2014;**10**(11):3153-3164

Biology. 1994;**10**:87-119

2008;**8**(2):108-120

[95] Barber GN. Cytoplasmic DNA innate immune pathways. Immunological Reviews. 2011;**243**(1):99-108

[96] Yang B, Jeang J, Yang A, Wu TC, Hung CF. DNA vaccine for cancer immunotherapy. Human Vaccines & Immunotherapeutics.

[97] Walter P, Johnson AE. Signal sequence recognition and protein targeting to the endoplasmic reticulum membrane. Annual Review of Cell

[98] Rice J, Ottensmeier CH, Stevenson FK. DNA vaccines: Precision tools for activating effective immunity against cancer. Nature Reviews. Cancer.

[99] Dharmapuri S, Aurisicchio L, Neuner P, Verdirame M, Ciliberto G, La Monica N. An oral TLR7 agonist is a potent adjuvant of DNA vaccination in transgenic mouse tumor models. Cancer Gene Therapy. 2009;**16**(5):462-472

[100] Orlandi F, Guevara-Patino JA, Merghoub T, Wolchok JD, Houghton AN, Gregor PD. Combination of epitope-optimized DNA vaccination and passive infusion of monoclonal antibody against HER2/neu leads to breast tumor regression in mice. Vaccine. 2011;**29**(20):3646-3654

[101] Fiedler K, Lazzaro S, Lutz J, Rauch S, Heidenreich R. mRNA cancer vaccines. Recent Results in Cancer

Research. 2016;**209**:61-85

*Immunotherapy for Treatment of Cancer DOI: http://dx.doi.org/10.5772/intechopen.81150*

the future. Immunological Reviews. 2011;**239**(1):62-84

*Current Trends in Cancer Management*

is a target for immune-mediated tumor destruction. Proceedings of the National Academy of Sciences of the United States of America.

[86] Schmidt SM, Schag K, Muller MR, et al. Survivin is a shared tumorassociated antigen expressed in a broad variety of malignancies and recognized by specific cytotoxic T cells. Blood.

[87] Kawai T, Akira S. Toll-like receptors and their crosstalk with other innate receptors in infection and immunity. Immunity. 2011;**34**(5):637-650

[88] Heldwein KA, Liang MD, Andresen

TK, et al. TLR2 and TLR4 serve distinct roles in the host immune response against Mycobacterium bovis BCG. Journal of Leukocyte Biology.

[89] Mata-Haro V, Cekic C, Martin M, Chilton PM, Casella CR, Mitchell TC. The vaccine adjuvant monophosphoryl lipid A as a TRIFbiased agonist of TLR4. Science. 2007;**316**(5831):1628-1632

[90] Clarke MA, Wentzensen N, Mirabello L, et al. Human

Prevention. 2012;**21**(12):

in Immunology. 2017;**45**:1-7

[92] Aurisicchio L, Ciliberto

[93] Liu MA. DNA vaccines: An historical perspective and view to

2012;**12**(8):1043-1058

2125-2137

papillomavirus DNA methylation as a potential biomarker for cervical cancer. Cancer Epidemiology, Biomarkers &

[91] Kumai T, Kobayashi H, Harabuchi Y, Celis E. Peptide vaccines in cancerold concept revisited. Current Opinion

G. Genetic cancer vaccines: Current status and perspectives. Expert Opinion on Biological Therapy.

2003;**100**(6):3398-3403

2003;**102**(2):571-576

2003;**74**(2):277-286

[80] Parkhurst MR, Fitzgerald EB, Southwood S, Sette A, Rosenberg SA, Kawakami Y. Identification of a shared HLA-A\*0201-restricted T-cell epitope from the melanoma antigen tyrosinaserelated protein 2 (TRP2). Cancer Research. 1998;**58**(21):4895-4901

[81] Jaramillo A, Majumder K, Manna PP, et al. Identification of HLA-A3 restricted CD8+ T cell epitopes derived from mammaglobin-A, a tumorassociated antigen of human breast cancer. International Journal of Cancer.

[82] Tsang KY, Zaremba S, Nieroda CA, Zhu MZ, Hamilton JM, Schlom J. Generation of human cytotoxic T cells specific for human carcinoembryonic antigen epitopes from patients

immunized with recombinant vaccinia-CEA vaccine. Journal of the National Cancer Institute. 1995;**87**(13):982-990

[83] Finn OJ, Gantt KR, Lepisto AJ, Pejawar-Gaddy S, Xue J, Beatty PL. Importance of MUC1 and

spontaneous mouse tumor models for understanding the immunobiology of human adenocarcinomas. Immunologic

Research. 2011;**50**(2-3):261-268

2009;**27**(28):4685-4692

[84] Disis ML, Wallace DR, Gooley TA, et al. Concurrent trastuzumab and HER2/neu-specific vaccination in patients with metastatic breast cancer. Journal of Clinical Oncology.

[85] Schmollinger JC, Vonderheide RH, Hoar KM, et al. Melanoma inhibitor of apoptosis protein (ML-IAP)

[79] Kawakami Y, Eliyahu S, Sakaguchi K, et al. Identification of the immunodominant peptides of the MART-1 human melanoma antigen recognized by the majority of HLA-A2-restricted tumor infiltrating lymphocytes. The Journal of Experimental Medicine.

1994;**180**(1):347-352

2002;**102**(5):499-506

**36**

[94] Stratford R, Douce G, Zhang-Barber L, Fairweather N, Eskola J, Dougan G. Influence of codon usage on the immunogenicity of a DNA vaccine against tetanus. Vaccine. 2000;**19**(7-8):810-815

[95] Barber GN. Cytoplasmic DNA innate immune pathways. Immunological Reviews. 2011;**243**(1):99-108

[96] Yang B, Jeang J, Yang A, Wu TC, Hung CF. DNA vaccine for cancer immunotherapy. Human Vaccines & Immunotherapeutics. 2014;**10**(11):3153-3164

[97] Walter P, Johnson AE. Signal sequence recognition and protein targeting to the endoplasmic reticulum membrane. Annual Review of Cell Biology. 1994;**10**:87-119

[98] Rice J, Ottensmeier CH, Stevenson FK. DNA vaccines: Precision tools for activating effective immunity against cancer. Nature Reviews. Cancer. 2008;**8**(2):108-120

[99] Dharmapuri S, Aurisicchio L, Neuner P, Verdirame M, Ciliberto G, La Monica N. An oral TLR7 agonist is a potent adjuvant of DNA vaccination in transgenic mouse tumor models. Cancer Gene Therapy. 2009;**16**(5):462-472

[100] Orlandi F, Guevara-Patino JA, Merghoub T, Wolchok JD, Houghton AN, Gregor PD. Combination of epitope-optimized DNA vaccination and passive infusion of monoclonal antibody against HER2/neu leads to breast tumor regression in mice. Vaccine. 2011;**29**(20):3646-3654

[101] Fiedler K, Lazzaro S, Lutz J, Rauch S, Heidenreich R. mRNA cancer vaccines. Recent Results in Cancer Research. 2016;**209**:61-85

[102] Fotin-Mleczek M, Zanzinger K, Heidenreich R, et al. Highly potent mRNA based cancer vaccines represent an attractive platform for combination therapies supporting an improved therapeutic effect. The Journal of Gene Medicine. 2012;**14**(6):428-439

[103] Rosenberg SA, Restifo NP. Adoptive cell transfer as personalized immunotherapy for human cancer. Science. 2015;**348**(6230):62-68

[104] Verdegaal EM. Adoptive cell therapy: A highly successful individualized therapy for melanoma with great potential for other malignancies. Current Opinion in Immunology. 2016;**39**:90-95

[105] Yee C, Thompson JA, Byrd D, et al. Adoptive T cell therapy using antigen-specific CD8+ T cell clones for the treatment of patients with metastatic melanoma: in vivo persistence, migration, and antitumor effect of transferred T cells. Proceedings of the National Academy of Sciences of the United States of America. 2002;**99**(25):16168-16173

[106] Bryant KL, Mancias JD, Kimmelman AC, Der CJ. KRAS: Feeding pancreatic cancer proliferation. Trends in Biochemical Sciences. 2014;**39**(2):91-100

[107] Tran E, Robbins PF, Lu YC, et al. T-cell transfer therapy targeting mutant KRAS in cancer. The New England Journal of Medicine. 2016;**375**(23):2255-2262

[108] Morgan RA, Dudley ME, Wunderlich JR, et al. Cancer regression in patients after transfer of genetically engineered lymphocytes. Science. 2006;**314**(5796):126-129

[109] Robbins PF, Morgan RA, Feldman SA, et al. Tumor regression in patients with metastatic synovial cell sarcoma and melanoma using genetically engineered lymphocytes reactive with

NY-ESO-1. Journal of Clinical Oncology. 2011;**29**(7):917-924

[110] Donohue JH, Rosenstein M, Chang AE, Lotze MT, Robb RJ, Rosenberg SA. The systemic administration of purified interleukin 2 enhances the ability of sensitized murine lymphocytes to cure a disseminated syngeneic lymphoma. Journal of Immunology. 1984;**132**(4):2123-2128

[111] Dastmalchi F, Karachi A, Allison JR, Basso K, Mitchell D, Rahman M. In vivo cellular tracking with 13C labeling of adopetive transferred T cells for the treatment of brain tumors. Neuro-Oncology. 2017;**19**(suppl\_6):vi119

[112] Leone P, Shin EC, Perosa F, Vacca A, Dammacco F, Racanelli V. MHC class I antigen processing and presenting machinery: Organization, function, and defects in tumor cells. Journal of the National Cancer Institute. 2013;**105**(16):1172-1187

[113] Brudno JN, Somerville RP, Shi V, et al. Allogeneic T cells that express an anti-CD19 chimeric antigen receptor induce remissions of B-cell malignancies that Progress after allogeneic hematopoietic stem-cell transplantation without causing graft-versus-host disease. Journal of Clinical Oncology. 2016;**34**(10):1112-1121

[114] Kochenderfer JN, Dudley ME, Kassim SH, et al. Chemotherapyrefractory diffuse large B-cell lymphoma and indolent B-cell malignancies can be effectively treated with autologous T cells expressing an anti-CD19 chimeric antigen receptor. Journal of Clinical Oncology. 2015;**33**(6):540-549

[115] Maude SL, Frey N, Shaw PA, et al. Chimeric antigen receptor T cells for sustained remissions in leukemia. The New England Journal of Medicine. 2014;**371**(16):1507-1517

[116] Porter DL, Levine BL, Kalos M, Bagg A, June CH. Chimeric antigen receptor-modified T cells in chronic lymphoid Leukemia; chimeric antigen receptor-modified T cells for acute lymphoid Leukemia; chimeric antigen receptor T cells for sustained remissions in Leukemia. The New England Journal of Medicine. 2016;**374**(10):998

[117] Ninomiya S, Narala N, Huye L, et al. Tumor indoleamine 2,3-dioxygenase (IDO) inhibits CD19- CAR T cells and is downregulated by lymphodepleting drugs. Blood. 2015;**125**(25):3905-3916

[118] Kakimi K, Karasaki T, Matsushita H, Sugie T. Advances in personalized cancer immunotherapy. Breast Cancer. 2017;**24**:16-24

Section 2

Digestive Cancers

39

### Section 2

## Digestive Cancers

*Current Trends in Cancer Management*

2011;**29**(7):917-924

NY-ESO-1. Journal of Clinical Oncology.

[116] Porter DL, Levine BL, Kalos M, Bagg A, June CH. Chimeric antigen receptor-modified T cells in chronic lymphoid Leukemia; chimeric antigen receptor-modified T cells for acute lymphoid Leukemia; chimeric antigen receptor T cells for sustained remissions in Leukemia. The New England Journal

of Medicine. 2016;**374**(10):998

2015;**125**(25):3905-3916

2017;**24**:16-24

[117] Ninomiya S, Narala N, Huye L, et al. Tumor indoleamine

2,3-dioxygenase (IDO) inhibits CD19- CAR T cells and is downregulated by lymphodepleting drugs. Blood.

[118] Kakimi K, Karasaki T, Matsushita H, Sugie T. Advances in personalized cancer immunotherapy. Breast Cancer.

[110] Donohue JH, Rosenstein M, Chang AE, Lotze MT, Robb RJ, Rosenberg SA. The systemic administration of purified interleukin 2 enhances the ability of sensitized murine lymphocytes to cure a disseminated syngeneic lymphoma. Journal of Immunology. 1984;**132**(4):2123-2128

[111] Dastmalchi F, Karachi A, Allison JR, Basso K, Mitchell D, Rahman M. In vivo cellular tracking with 13C labeling of adopetive transferred T cells for the treatment of brain tumors. Neuro-Oncology. 2017;**19**(suppl\_6):vi119

[112] Leone P, Shin EC, Perosa F, Vacca A, Dammacco F, Racanelli V. MHC class I antigen processing and presenting machinery: Organization, function, and defects in tumor cells. Journal of the National Cancer Institute.

[113] Brudno JN, Somerville RP, Shi V, et al. Allogeneic T cells that express an anti-CD19 chimeric antigen receptor induce remissions of B-cell malignancies

hematopoietic stem-cell transplantation without causing graft-versus-host disease. Journal of Clinical Oncology.

[114] Kochenderfer JN, Dudley ME, Kassim SH, et al. Chemotherapyrefractory diffuse large B-cell lymphoma and indolent B-cell

malignancies can be effectively treated with autologous T cells expressing an anti-CD19 chimeric antigen

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[115] Maude SL, Frey N, Shaw PA, et al. Chimeric antigen receptor T cells for sustained remissions in leukemia. The New England Journal of Medicine.

2013;**105**(16):1172-1187

that Progress after allogeneic

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2015;**33**(6):540-549

2014;**371**(16):1507-1517

**38**

Chapter 3

Abstract

1. Introduction

41

1.1 Incidence and mortality

Advances in the Treatment of

still unknown but may benefit in situations with positive margins.

Keywords: pancreatic cancer, neoadjuvant chemotherapy, FOLFIRINOX

in developed countries, 188,000 and 184,000 persons as compared to less

type of malignancy and the fourth leading cause of cancer deaths. There are estimated 55,440 new cases of pancreatic cancer that will be diagnosed in the USA in 2018 and result in estimated 44,330 deaths. Incidence rate of pancreatic cancer

developed regions, 150,000 and 146,000 persons [1].

has increased 1% per year from 2005 to 2014 [2].

on invasive pancreatic ductal adenocarcinoma [2].

Pancreatic cancer is the 13th most common malignancy worldwide, which was diagnosed in approximately 338,000 people in 2012. Pancreatic cancer is a very aggressive form of malignancy resulting in the seventh leading cause of cancer deaths worldwide, over 331,000 deaths in 2012 alone. The worldwide incidence of new pancreatic cancers was 4.9 in 100,000 persons with an associated mortality rate of approximately 4%. The incidence and deaths of pancreatic cancer is higher

Specifically looking at the USA, pancreatic cancer is the eighth most common

Most pancreatic cancers develop from the pancreatic exocrine tissue (94%), such as invasive ductal adenocarcinoma, while the remaining 6% of tumors stem from the hormone-producing islet cells, such as insulinomas, gastrinomas, and other pancreatic neuroendocrine tumors (pNETs). Those pNets will typically occur in younger patients with a better overall prognosis. The focus of this chapter will be

Pancreatic cancer is an aggressive solid organ malignancy with a high mortality rate. There has only been significant improvement in the overall survival until the last 5–10 years. The current trend toward the neoadjuvant approach of pancreatic cancer has shown success in tumor response, resection rate, and even overall survival. Using dedicated pancreatic protocol cross-sectional imaging, one can now follow the tumor and pancreatic parenchyma interface as well as tumor markers to predict treatment response. Aggressive combination chemotherapy regimens such as FOLFIRINOX (5-fluorouracil, leucovorin, oxaliplatin, and irinotecan), appropriate patient selection, and multidisciplinary treatment teams have made an impact in the current management of pancreatic cancer. Surgical intervention is still the mainstay treatment of pancreatic cancer. The role of routine radiation therapy is

Pancreatic Cancer

Michelle Marie Fillion

#### Chapter 3

## Advances in the Treatment of Pancreatic Cancer

Michelle Marie Fillion

#### Abstract

Pancreatic cancer is an aggressive solid organ malignancy with a high mortality rate. There has only been significant improvement in the overall survival until the last 5–10 years. The current trend toward the neoadjuvant approach of pancreatic cancer has shown success in tumor response, resection rate, and even overall survival. Using dedicated pancreatic protocol cross-sectional imaging, one can now follow the tumor and pancreatic parenchyma interface as well as tumor markers to predict treatment response. Aggressive combination chemotherapy regimens such as FOLFIRINOX (5-fluorouracil, leucovorin, oxaliplatin, and irinotecan), appropriate patient selection, and multidisciplinary treatment teams have made an impact in the current management of pancreatic cancer. Surgical intervention is still the mainstay treatment of pancreatic cancer. The role of routine radiation therapy is still unknown but may benefit in situations with positive margins.

Keywords: pancreatic cancer, neoadjuvant chemotherapy, FOLFIRINOX

#### 1. Introduction

#### 1.1 Incidence and mortality

Pancreatic cancer is the 13th most common malignancy worldwide, which was diagnosed in approximately 338,000 people in 2012. Pancreatic cancer is a very aggressive form of malignancy resulting in the seventh leading cause of cancer deaths worldwide, over 331,000 deaths in 2012 alone. The worldwide incidence of new pancreatic cancers was 4.9 in 100,000 persons with an associated mortality rate of approximately 4%. The incidence and deaths of pancreatic cancer is higher in developed countries, 188,000 and 184,000 persons as compared to less developed regions, 150,000 and 146,000 persons [1].

Specifically looking at the USA, pancreatic cancer is the eighth most common type of malignancy and the fourth leading cause of cancer deaths. There are estimated 55,440 new cases of pancreatic cancer that will be diagnosed in the USA in 2018 and result in estimated 44,330 deaths. Incidence rate of pancreatic cancer has increased 1% per year from 2005 to 2014 [2].

Most pancreatic cancers develop from the pancreatic exocrine tissue (94%), such as invasive ductal adenocarcinoma, while the remaining 6% of tumors stem from the hormone-producing islet cells, such as insulinomas, gastrinomas, and other pancreatic neuroendocrine tumors (pNETs). Those pNets will typically occur in younger patients with a better overall prognosis. The focus of this chapter will be on invasive pancreatic ductal adenocarcinoma [2].

The overall pancreatic cancer mortality rate has shown only slight improvement over the past 35 years. In 1975, pancreatic cancer mortality rate was observed at 3.1%, and in 2000, it increased to 5.2%. The largest incremental improvement in pancreatic cancer survival has occurred over the past 10 years (2008–2104), with the all stage 5 year survival between 8 and 8.5% [2–4].

For the small percentage of patients with early-stage localized pancreatic cancer (10%), the 5-year survival is between 32 and 34.3%. Once regional lymph node involvement has developed, the 5 year survival decreases to 11.5–12%. Unfortunately, most pancreatic cancer patients (52%) are diagnosed with distant metastatic disease, and that 5-year survival is only 3% [2, 3].

#### 1.2 Risk factors

At this point in the time, the cause of pancreatic cancer is still unknown. Increasing age is a significant risk factor for developing pancreatic cancer. The median age of diagnosis of pancreatic cancer in both sexes is at 70 years old. In addition, men have an increased incidence of developing pancreatic cancer as compared to women, 14.4 vs. 11.2 per 100,000 persons across all race and ethnicity [1–3].

There have been several risk factors to develop pancreatic cancer associated with race/ethnicity, environmental, dietary, medical, and genetic exposures identified (Table 1). Race is also another significant risk factor. African-Americans have the highest incidence (9.9 per 100,000 persons) and mortality (9.4 per 100,000 persons) of pancreatic cancer as compared to non-African-Americans [4]. In addition, Jews of Ashkenazi heritage also have an increased incidence of pancreatic cancer. The age standardized incidence rate of pancreatic cancer for Israeli Jews (7.2 per 100,000 males and 5.7 per 100,000 females) exceeds the incidence of Israeli non-Jews (4.0 per 100,000 males and 2.9 per 100,000 females) [5].

There are also several hereditary conditions associated with increased risk for pancreatic cancer (Table 2). While persons with these genetic syndromes are at increased risk for pancreatic cancer, they only account for 5% of all pancreatic diagnoses. Familial cases of pancreatic cancer are at increased incidence to develop secondary primary cancers as compared to non-familial-based cancers. Of those listed, Peutz-Jeghers and hereditary pancreatitis syndromes have the highest risk of developing pancreatic cancer [6–12].

> Tobacco use is the most well-established modifiable risk factor for developing pancreatic cancer and accounts for up to 30% of all pancreatic cancer cases. There is at least a twofold increase in risk for developing pancreatic cancer in cigarette smoker than a non-smoker. The risk also increases with an increase in the number of cigarettes and duration of smoking. It may take up to 20 years after cessation of cigarette smoking for one's risk of pancreatic cancer to be equal to take of a non-

> The current standard in pancreatic cancer staging is by use of a 64-slice multidetector computed tomography (CT). Specific CT pancreatic protocols can accurately stage the cancer and assess for resectability. These protocols include both the use of low-density oral contrast and nonionic iodinated contrast and scanned 30–45 seconds then again 60 seconds after injection to capture both arterial and

smoker [13].

43

Table 2.

2. Treatment of resectable disease

Genetic syndromes with increased risk of pancreatic cancer.

Advances in the Treatment of Pancreatic Cancer DOI: http://dx.doi.org/10.5772/intechopen.82074

2.1 Imaging considerations


Table 1. Risk factors for developing pancreatic cancer.


Table 2.

The overall pancreatic cancer mortality rate has shown only slight improvement over the past 35 years. In 1975, pancreatic cancer mortality rate was observed at 3.1%, and in 2000, it increased to 5.2%. The largest incremental improvement in pancreatic cancer survival has occurred over the past 10 years (2008–2104), with

For the small percentage of patients with early-stage localized pancreatic cancer (10%), the 5-year survival is between 32 and 34.3%. Once regional lymph node involvement has developed, the 5 year survival decreases to 11.5–12%. Unfortunately, most pancreatic cancer patients (52%) are diagnosed with distant metastatic

At this point in the time, the cause of pancreatic cancer is still unknown. Increasing age is a significant risk factor for developing pancreatic cancer. The median age of diagnosis of pancreatic cancer in both sexes is at 70 years old. In addition, men have an increased incidence of developing pancreatic cancer as compared to women,

There have been several risk factors to develop pancreatic cancer associated with race/ethnicity, environmental, dietary, medical, and genetic exposures identified (Table 1). Race is also another significant risk factor. African-Americans have the highest incidence (9.9 per 100,000 persons) and mortality (9.4 per 100,000 persons) of pancreatic cancer as compared to non-African-Americans [4]. In addition, Jews of Ashkenazi heritage also have an increased incidence of pancreatic cancer. The age standardized incidence rate of pancreatic cancer for Israeli Jews (7.2 per 100,000 males and 5.7 per 100,000 females) exceeds the incidence of Israeli non-Jews (4.0 per 100,000 males and 2.9 per 100,000 females) [5].

There are also several hereditary conditions associated with increased risk for pancreatic cancer (Table 2). While persons with these genetic syndromes are at increased risk for pancreatic cancer, they only account for 5% of all pancreatic diagnoses. Familial cases of pancreatic cancer are at increased incidence to develop secondary primary cancers as compared to non-familial-based cancers. Of those listed, Peutz-Jeghers and hereditary pancreatitis syndromes have the highest risk of

14.4 vs. 11.2 per 100,000 persons across all race and ethnicity [1–3].

the all stage 5 year survival between 8 and 8.5% [2–4].

Current Trends in Cancer Management

disease, and that 5-year survival is only 3% [2, 3].

developing pancreatic cancer [6–12].

Risk factors for developing pancreatic cancer.

Table 1.

42

1.2 Risk factors

Genetic syndromes with increased risk of pancreatic cancer.

Tobacco use is the most well-established modifiable risk factor for developing pancreatic cancer and accounts for up to 30% of all pancreatic cancer cases. There is at least a twofold increase in risk for developing pancreatic cancer in cigarette smoker than a non-smoker. The risk also increases with an increase in the number of cigarettes and duration of smoking. It may take up to 20 years after cessation of cigarette smoking for one's risk of pancreatic cancer to be equal to take of a nonsmoker [13].

#### 2. Treatment of resectable disease

#### 2.1 Imaging considerations

The current standard in pancreatic cancer staging is by use of a 64-slice multidetector computed tomography (CT). Specific CT pancreatic protocols can accurately stage the cancer and assess for resectability. These protocols include both the use of low-density oral contrast and nonionic iodinated contrast and scanned 30–45 seconds then again 60 seconds after injection to capture both arterial and

venous phases. The arterial phase will allow for good visualization of the celiac axis, common hepatic artery, superior mesenteric artery, and gastroduodenal artery. The venous phase will show enhanced visualization of the portal vein, superior mesenteric vein, splenic vein, pancreatic parenchyma, and the liver to assess for metastatic disease [14, 15].

#### 2.2 Surgical considerations

Only 20% of patients who present with pancreatic cancer can undergo surgical resection since most patients present with either unresectable or metastatic disease. The only chance for a curative treatment is with the inclusion of successful surgical removal of the cancer. To determine the patient's eligibility for pancreatic resection, an experienced pancreatic surgeon is required to review the dedicated pancreatic cross-sectional imaging. The relationship of the tumor to the major intra-abdominal vessels determines the resectability of the pancreatic cancer. Decisions regarding diagnostic and management and resectability should involve multidisciplinary consultation at high-volume center, at least 15–20 pancreatic resections per year [14].

In 2006, the National Comprehensive Cancer Network (NCCN) criteria initially defined pancreatic cancers resectability status into three classifications: resectable, borderline resectable, and unresectable. Since that time, there have been several varying definitions of tumor resectability that have evolved over the past decade. Several international surgical societies such as the American Hepato-Pancreatico-Biliary Association (AHPBA), Society of Surgical Oncology (SSO), Society for Surgery of the Alimentary Tract (SSAT), and International Association of Pancreatology (IAP) have issued consensus statements on the definition and criteria of resectability and borderline resectable pancreatic cancers as illustrated in Table 3 [14–16].

For a tumor to be considered resectable, it must not be in contact with the portal vein (PV) or superior mesenteric vein (SMV) per AHPBA/SSO/SSAT and IAP or less than 180° of contact with SMV/PV by NCCN criteria. By meeting these criteria, the surgeon believes there is a high likelihood of removing the cancer without leaving behind any residual tumor (R0 resection). When pancreatic cancers are classified as borderline resectable based on the vascular involvement, it means that there is a higher likelihood of having residual microscopic disease (R1 resection) if one was to proceed with upfront surgery. Borderline resectable means just that it is not quite resectable but not completely unresectable either. The criteria are less than 180° of arterial involvement of the superior mesenteric artery (SMA) or common hepatic artery (CHA) of celiac axis (CA). It also means there can be greater than 180° of involvement of SMV or even complete encasement of SMV or PV but still suitable for resection vascular reconstruction. Unresectable disease has greater than 180° of arterial involvement of SMA, CHA, or CA or nonreconstructable vein involvement including the first jejunal branch [14–16].

The best outcomes come from margin negative surgical resection with no residual microscopic disease (R0). There is current debate at the true definition of R1 resection as either no microscopic tumor cells at the resection margin or if tumor cells are less than 1 mm from the resection margin. It has been found that the 5-year survival rate has been improved in patients with greater than 1 mm of clearance as compared to those with less than 1 mm. Margins with 0 mm, less than 1 mm, or greater than 1 mm had 5-year survival rates at 16.3, 12.4, and 27.6%, respectively [17]. There is no benefit to performing a surgical resection if gross tumor (R2 resection) will be the result as the prognosis is similar to patients with non-operative management [18].

Table 3.

45

Definitions and criteria of resectable, borderline resectable and unresectable pancreatic cancer.

Advances in the Treatment of Pancreatic Cancer DOI: http://dx.doi.org/10.5772/intechopen.82074

Advances in the Treatment of Pancreatic Cancer DOI: http://dx.doi.org/10.5772/intechopen.82074

venous phases. The arterial phase will allow for good visualization of the celiac axis, common hepatic artery, superior mesenteric artery, and gastroduodenal artery. The venous phase will show enhanced visualization of the portal vein, superior mesenteric vein, splenic vein, pancreatic parenchyma, and the liver to assess for meta-

Only 20% of patients who present with pancreatic cancer can undergo surgical resection since most patients present with either unresectable or metastatic disease. The only chance for a curative treatment is with the inclusion of successful surgical removal of the cancer. To determine the patient's eligibility for pancreatic resection, an experienced pancreatic surgeon is required to review the dedicated pancreatic cross-sectional imaging. The relationship of the tumor to the major intra-abdominal vessels determines the resectability of the pancreatic cancer. Decisions regarding diagnostic and management and resectability should involve multidisciplinary consultation at high-volume center, at least 15–20 pancreatic resections per year [14]. In 2006, the National Comprehensive Cancer Network (NCCN) criteria initially defined pancreatic cancers resectability status into three classifications: resectable, borderline resectable, and unresectable. Since that time, there have been several varying definitions of tumor resectability that have evolved over the past decade. Several international surgical societies such as the American Hepato-Pancreatico-Biliary Association (AHPBA), Society of Surgical Oncology (SSO), Society for Surgery of the Alimentary Tract (SSAT), and International Association of Pancreatology (IAP) have issued consensus statements on the definition and criteria of resectability and borderline resectable pancreatic cancers as illustrated in

For a tumor to be considered resectable, it must not be in contact with the portal vein (PV) or superior mesenteric vein (SMV) per AHPBA/SSO/SSAT and IAP or less than 180° of contact with SMV/PV by NCCN criteria. By meeting these criteria, the surgeon believes there is a high likelihood of removing the cancer without leaving behind any residual tumor (R0 resection). When pancreatic cancers are classified as borderline resectable based on the vascular involvement, it means that there is a higher likelihood of having residual microscopic disease (R1 resection) if one was to proceed with upfront surgery. Borderline resectable means just that it is not quite resectable but not completely unresectable either. The criteria are less than 180° of arterial involvement of the superior mesenteric artery (SMA) or common hepatic artery (CHA) of celiac axis (CA). It also means there can be greater than 180° of involvement of SMV or even complete encasement of SMV or PV but still suitable for resection vascular reconstruction. Unresectable disease has

greater than 180° of arterial involvement of SMA, CHA, or CA or nonreconstructable vein involvement including the first jejunal branch [14–16]. The best outcomes come from margin negative surgical resection with no residual microscopic disease (R0). There is current debate at the true definition of R1 resection as either no microscopic tumor cells at the resection margin or if tumor cells are less than 1 mm from the resection margin. It has been found that the 5-year survival rate has been improved in patients with greater than 1 mm of clearance as compared to those with less than 1 mm. Margins with 0 mm, less than 1 mm, or greater than 1 mm had 5-year survival rates at 16.3, 12.4, and 27.6%, respectively

[17]. There is no benefit to performing a surgical resection if gross tumor (R2 resection) will be the result as the prognosis is similar to patients with

non-operative management [18].

44

static disease [14, 15].

Table 3 [14–16].

2.2 Surgical considerations

Current Trends in Cancer Management


Table 3.

Definitions and criteria of resectable, borderline resectable and unresectable pancreatic cancer.

Pancreatic surgery should involve high-volume surgeons with the expertise in pancreatic resection. Decisions regarding the management of pancreatic cancer patients require a multidisciplinary team. The location of the tumor and extent of disease will dictate the surgical approaches. Pancreatic head and uncinate tumors require pancreaticoduodenectomy (Whipple procedure) with reconstruction of the pancreas, bile duct, and stomach. If possible, the aim is to preserve the pylorus to limit bile acid reflux and gastric emptying. Tumors that exist in the body and tail of the pancreas will typically require a left-sided surgical resection, distal pancreatectomy, and splenectomy. Borderline resectable and locally advanced cancers may also require venous and/or arterial reconstruction at the time of surgical resection of the pancreatic cancer.

(1000 mg per square meter) weekly for 7 of 8 weeks and then on days 1, 8, and 15

The ESPAC-4 went on to compare adjuvant gemcitabine and capecitabine with gemcitabine monotherapy in patients with resected pancreatic cancer. Capecitabine is an orally active fluoropyrimidine carbamate which can provide prolonged fluorouracil exposure at lower peak concentrations. The 5-year overall survival with gemcitabine and capecitabine compared to gemcitabine alone was 29% vs. 16% [24]. The new standard of care quickly adopted doublet therapy as the new

The use of adjuvant FOLFIRINOX [fluorouracil (5-FU), leucovorin, irinotecan, oxaliplatin] has been extrapolated from the treatment of pancreatic cancer in the metastatic setting. In the ACCORD-11 trial, FOLFIRINOX was found to have a superior survival advantage over gemcitabine in metastatic pancreatic patients with median overall survival of 11.1 months vs. 6.8 months [25]. This study ultimately launched FOLFIRINOX into new treatment paradigms in the adjuvant and

The phase III PRODIGE 24/CCTG PA.6 trial compared a modified FOLFIRINOX regimen against single-agent gemcitabine therapy, as the results of the ESPAC-4 were not known at study design. This study used a modified FOLFIRINOX regimen:

, and irinotecan 180 mg/m<sup>2</sup> (dose

, leucovorin 400 mg/m<sup>2</sup>

reduced to 150 mg/m2 after patient 162) on day 1 and continuous fluorouracil infusion 2.4 gm/m<sup>2</sup> over 46 hours. This regimen was repeated every 2 weeks for 12 cycles. The gemcitabine regimen was 1000 mg/m<sup>2</sup> once per 3 of 4 weeks for 6 cycles [26]. The response rate was 31.18% in the mFOLFIRINOX group and 11.3% in the gemcitabine group. The disease-free survival (DFS) and overall survival (OS) in the mFOLFIRINOX arm were 21.6 and 54.4 months, while the gemcitabine arm were 17.7 and 35.0 months repetitively. Grade 3 or 4 adverse events (neutropenia, diarrhea, neuropathy) were significantly higher in the FOLFIRINOX treatment arm

mFOLFIRINOX has been the largest advancement in overall survival for resected pancreatic cancer patients, which more than doubled the previous median

There are mixed opinions regarding the routine use of radiation therapy in pancreatic cancer. The ESPAC-1 did not reveal any significant survival benefit with chemoradiation [19]. A meta-analysis of five randomized controlled trials using adjuvant chemoradiation in patients who underwent curation resection was performed to assess the survival benefit. It appeared that adjuvant chemoradiation had benefitted the subset of patients with a positive margin status; however, it was

The RTOG study looks to determine if the addition of gemcitabine to adjuvant fluorouracil chemoradiation improved survival as compared with fluorouracil. Patients were given either fluorouracil (continuous infusion 250 mg/m<sup>2</sup> per day) or 30 minutes infusion of gemcitabine (1000 mg/m<sup>2</sup> once a week) for 3 weeks prior to fluorouracil chemoradiation and for 12 weeks following chemoradiation. The median survival for the gemcitabine group was 20.5 months, while the median

every 4 weeks. The overall survival in the nab-paclitaxel-gemcitabine was 8.5 months as compared to gemcitabine alone with 6.7 months (p > 0.001). This doublet therapy did result in higher rates of myelosuppression and peripheral neuropathy than gemcitabine alone [23]. While this study was based on patients with metastatic disease, it advanced the adjuvant combined chemotherapy regimen in

resected pancreatic cancer.

Advances in the Treatment of Pancreatic Cancer DOI: http://dx.doi.org/10.5772/intechopen.82074

standard of care.

neoadjuvant settings.

oxaliplatin 85 mg/m<sup>2</sup>

overall survival.

47

2.4 Radiation therapy

than the gemcitabine arm [26].

not statistically significant [27].

#### 2.3 Chemotherapy

In patients with pancreatic cancer, the overall survival has improved due to systemic chemotherapy and combination therapies. It is still standard treatment to perform upfront surgical resection for resectable pancreatic cancer followed by adjuvant chemotherapy. However, there has been a shift toward upfront neoadjuvant chemotherapy in order to select out patients with latent metastatic disease or to downstage borderline and locally advanced cancers.

The ESPAC-1 (European Study Group for Pancreatic Cancer) showed that there was improvement in overall survival using surgery plus adjuvant 5-fluorouracil (5-FU) plus folinic acid (FA). This three-tracked trial compared patients treated with chemotherapy alone, surgical resection alone, or chemotherapy plus radiation therapy. The highest 5-year survival was seen in the chemotherapy arm 21% as compared to surgery alone 8% and chemoradiation 11%. This revealed the only significant survival benefit was with adjuvant chemotherapy [19].

The Charite Onkologie study (CONKO-001) from 2007 compared adjuvant gemcitabine therapy to observation in patients undergoing surgical resection of pancreatic cancer. In the treatment arm, patients received 6 cycles of adjuvant gemcitabine. Patients treated with adjuvant gemcitabine vs. surgery alone had statistically significant increased median overall survival of 22.8 months and 5-year survival of 21% compared to 20.2 months and 5-year survival of 9%. Gemcitabine also significantly delayed the development of recurrent disease as compared to observation alone [20].

The ESPAC-3 was a large randomized controlled trial which compared adjuvant 5-FU plus leucovorin or gemcitabine in patients who underwent RO or R1 resection of pancreatic cancer. This was initially a three-arm study comparing 5-FU plus leucovorin, gemcitabine, and observation; however, once the results of the ESPAC-1 were available, the observation arm was closed. Results of ESPAC-1, ESPAC-1 plus, and ESPAC-3 in subset analysis of 5-FU/FA vs. observation confirmed that adjuvant 5-FU/FA had superior overall survival as compared to observation after surgical resection. The 5 year survival for 5-FU/FA was 24% compared to observation which was 14% [21].

ESPAC-3 has enrolled 1088 patients and they were followed for over 6.5 years. Median survival for gemcitabine arm was 23.6 months while 5FU/leucovorin arm was 23 months [22]. This had shown that adjuvant gemcitabine had similar survival but less toxicity as compared to 5FU. At this point, there were now two different adjuvant treatment options for resected pancreatic cancer.

Van Hoof et al. (2013) performed a phase III trial in which metastatic pancreatic cancer patients were randomized to treatment with either nab-paclitaxel (125 mg per square meter of body surface area) plus gemcitabine (1000 mg per square meter) on days 1, 8, and 15 every 4 weeks or gemcitabine monotherapy

#### Advances in the Treatment of Pancreatic Cancer DOI: http://dx.doi.org/10.5772/intechopen.82074

Pancreatic surgery should involve high-volume surgeons with the expertise in pancreatic resection. Decisions regarding the management of pancreatic cancer patients require a multidisciplinary team. The location of the tumor and extent of disease will dictate the surgical approaches. Pancreatic head and uncinate tumors require pancreaticoduodenectomy (Whipple procedure) with reconstruction of the pancreas, bile duct, and stomach. If possible, the aim is to preserve the pylorus to limit bile acid reflux and gastric emptying. Tumors that exist in the body and tail of the pancreas will typically require a left-sided surgical resection, distal pancreatectomy, and splenectomy. Borderline resectable and locally advanced cancers may also require venous and/or arterial reconstruction at the time of surgical resection of

In patients with pancreatic cancer, the overall survival has improved due to systemic chemotherapy and combination therapies. It is still standard treatment to perform upfront surgical resection for resectable pancreatic cancer followed by adjuvant chemotherapy. However, there has been a shift toward upfront neoadjuvant chemotherapy in order to select out patients with latent metastatic

The ESPAC-1 (European Study Group for Pancreatic Cancer) showed that there

was improvement in overall survival using surgery plus adjuvant 5-fluorouracil (5-FU) plus folinic acid (FA). This three-tracked trial compared patients treated with chemotherapy alone, surgical resection alone, or chemotherapy plus radiation therapy. The highest 5-year survival was seen in the chemotherapy arm 21% as compared to surgery alone 8% and chemoradiation 11%. This revealed the only

The Charite Onkologie study (CONKO-001) from 2007 compared adjuvant gemcitabine therapy to observation in patients undergoing surgical resection of pancreatic cancer. In the treatment arm, patients received 6 cycles of adjuvant gemcitabine. Patients treated with adjuvant gemcitabine vs. surgery alone had statistically significant increased median overall survival of 22.8 months and 5-year survival of 21% compared to 20.2 months and 5-year survival of 9%. Gemcitabine also significantly delayed the development of recurrent disease as compared to

The ESPAC-3 was a large randomized controlled trial which compared adjuvant 5-FU plus leucovorin or gemcitabine in patients who underwent RO or R1 resection of pancreatic cancer. This was initially a three-arm study comparing 5-FU plus leucovorin, gemcitabine, and observation; however, once the results of the ESPAC-1 were available, the observation arm was closed. Results of ESPAC-1,

ESPAC-1 plus, and ESPAC-3 in subset analysis of 5-FU/FA vs. observation confirmed that adjuvant 5-FU/FA had superior overall survival as compared to observation after surgical resection. The 5 year survival for 5-FU/FA was 24%

ESPAC-3 has enrolled 1088 patients and they were followed for over 6.5 years. Median survival for gemcitabine arm was 23.6 months while 5FU/leucovorin arm was 23 months [22]. This had shown that adjuvant gemcitabine had similar survival but less toxicity as compared to 5FU. At this point, there were now two different

Van Hoof et al. (2013) performed a phase III trial in which metastatic pancreatic cancer patients were randomized to treatment with either nab-paclitaxel (125 mg per square meter of body surface area) plus gemcitabine (1000 mg per square meter) on days 1, 8, and 15 every 4 weeks or gemcitabine monotherapy

compared to observation which was 14% [21].

adjuvant treatment options for resected pancreatic cancer.

disease or to downstage borderline and locally advanced cancers.

significant survival benefit was with adjuvant chemotherapy [19].

the pancreatic cancer.

Current Trends in Cancer Management

observation alone [20].

46

2.3 Chemotherapy

(1000 mg per square meter) weekly for 7 of 8 weeks and then on days 1, 8, and 15 every 4 weeks. The overall survival in the nab-paclitaxel-gemcitabine was 8.5 months as compared to gemcitabine alone with 6.7 months (p > 0.001). This doublet therapy did result in higher rates of myelosuppression and peripheral neuropathy than gemcitabine alone [23]. While this study was based on patients with metastatic disease, it advanced the adjuvant combined chemotherapy regimen in resected pancreatic cancer.

The ESPAC-4 went on to compare adjuvant gemcitabine and capecitabine with gemcitabine monotherapy in patients with resected pancreatic cancer. Capecitabine is an orally active fluoropyrimidine carbamate which can provide prolonged fluorouracil exposure at lower peak concentrations. The 5-year overall survival with gemcitabine and capecitabine compared to gemcitabine alone was 29% vs. 16% [24]. The new standard of care quickly adopted doublet therapy as the new standard of care.

The use of adjuvant FOLFIRINOX [fluorouracil (5-FU), leucovorin, irinotecan, oxaliplatin] has been extrapolated from the treatment of pancreatic cancer in the metastatic setting. In the ACCORD-11 trial, FOLFIRINOX was found to have a superior survival advantage over gemcitabine in metastatic pancreatic patients with median overall survival of 11.1 months vs. 6.8 months [25]. This study ultimately launched FOLFIRINOX into new treatment paradigms in the adjuvant and neoadjuvant settings.

The phase III PRODIGE 24/CCTG PA.6 trial compared a modified FOLFIRINOX regimen against single-agent gemcitabine therapy, as the results of the ESPAC-4 were not known at study design. This study used a modified FOLFIRINOX regimen: oxaliplatin 85 mg/m<sup>2</sup> , leucovorin 400 mg/m<sup>2</sup> , and irinotecan 180 mg/m<sup>2</sup> (dose reduced to 150 mg/m2 after patient 162) on day 1 and continuous fluorouracil infusion 2.4 gm/m<sup>2</sup> over 46 hours. This regimen was repeated every 2 weeks for 12 cycles. The gemcitabine regimen was 1000 mg/m<sup>2</sup> once per 3 of 4 weeks for 6 cycles [26]. The response rate was 31.18% in the mFOLFIRINOX group and 11.3% in the gemcitabine group. The disease-free survival (DFS) and overall survival (OS) in the mFOLFIRINOX arm were 21.6 and 54.4 months, while the gemcitabine arm were 17.7 and 35.0 months repetitively. Grade 3 or 4 adverse events (neutropenia, diarrhea, neuropathy) were significantly higher in the FOLFIRINOX treatment arm than the gemcitabine arm [26].

mFOLFIRINOX has been the largest advancement in overall survival for resected pancreatic cancer patients, which more than doubled the previous median overall survival.

#### 2.4 Radiation therapy

There are mixed opinions regarding the routine use of radiation therapy in pancreatic cancer. The ESPAC-1 did not reveal any significant survival benefit with chemoradiation [19]. A meta-analysis of five randomized controlled trials using adjuvant chemoradiation in patients who underwent curation resection was performed to assess the survival benefit. It appeared that adjuvant chemoradiation had benefitted the subset of patients with a positive margin status; however, it was not statistically significant [27].

The RTOG study looks to determine if the addition of gemcitabine to adjuvant fluorouracil chemoradiation improved survival as compared with fluorouracil. Patients were given either fluorouracil (continuous infusion 250 mg/m<sup>2</sup> per day) or 30 minutes infusion of gemcitabine (1000 mg/m<sup>2</sup> once a week) for 3 weeks prior to fluorouracil chemoradiation and for 12 weeks following chemoradiation. The median survival for the gemcitabine group was 20.5 months, while the median

survival for the fluorouracil group was 16.9 months. There appeared to be a survival benefit, but it was not statistically significant [28].

margins, and negative lymph node metastasis are favorable prognostic indicators

The consensus for treatment of borderline resectable pancreatic cancer favors the neoadjuvant approach; however, it may vary per institution. After a multidisciplinary review at our hospital, the typical functional patient would undergo neoadjuvant chemotherapy (FOLFIRINOX) for 3–4 cycles followed by restaging with CT and CA 19–9. If stable or responding disease, then the patient would continue additional 3–4 cycles of FOLFIRINOX. The patient would again be restaged with CT and CA 19–9. Barring no metastatic disease developed and there was treatment response, the patient would then undergo surgical intervention. However, if the surgical margins were still threatened and there was concern for R1 resection, then the patient may undergo chemoradiation to "sterilize" the margins. Approximately 4–6 weeks after chemoradiation, the patient would ultimately

Unresectable pancreatic cancer means that the tumor cannot safely be removed due to vascular involvement or metastatic disease. Patients may undergo aggressive chemotherapy with FOLFIRINOX, and a few may be able to convert to a resectable cancer. It is of utmost importance for early palliative care interventions in these patients. For those with biliary obstruction, the use of endoscopic biliary stents and percutaneous biliary drains may provide relief from the jaundice. If the tumor is found to be unresectable in the operating room, then palliative hepaticojejunostomy may be performed. Gastric outlet obstruction may also be relieved with endoscopically placed luminal stents. Additionally, surgical bypass may be performed in

Pain can also become quite debilitating in patients with locally advanced unresectable pancreatic cancer. Celiac plexus neurolysis can be performed at the time of surgical exploration, or it may be performed by endoscopic or percutaneous

Irreversible electroporation (IRE) is a nonthermal ablative modality which relies on high voltage (maximum 3,000 volts) small microsecond pulse lengths. This is a novel option typically used in locally advanced pancreatic adenocarcinoma of the head or neck that is not amendable to resection. Some institutions are now using IRE to assist with the resection of locally advanced tumors, but this is not standard at this time. The procedure may be performed open or percutaneously. Patients will typical undergo several months of neoadjuvant chemotherapy to not miss occult metastatic disease prior to IRE. IRE can improve progression-free survival from 6 to

There are several active clinical trials investigating additional treatment options for pancreatic cancer. Several phase II trials are looking at the use of targeted agents

niraparib, PARP [poly (ADP-ribose) polymerase] inhibitor, in advanced pancreatic cancer patients [35]. Another clinical trial at the Massachusetts General Hospital is using the checkpoint inhibitor, nivolumab, as programed death-1 (PD-1) inhibition in combination with losartan, FOLFIRINOX, stereotactic body radiation therapy (SBRT), and surgery in advanced pancreatic cancer. This is a three-armed study:

in addition to systemic chemotherapy. One study is evaluating the safety of

for improved overall and disease-free survival.

Advances in the Treatment of Pancreatic Cancer DOI: http://dx.doi.org/10.5772/intechopen.82074

undergo surgical resection.

routes.

5. Clinical trials

49

4. Unresectable pancreatic cancer

laparoscopic or open fashion with a gastrojejunostomy.

14 months and overall survival from 23 to 20 months [34].

The LAP07 randomized clinical trial aimed to assess if chemoradiation would improve overall survival after 4 months of gemcitabine and to assess erlotinib's effect on survival as well as in patients with locally advanced pancreatic cancer. There was no difference in overall survival between the chemotherapy alone vs. the addition of chemoradiation, 16.5 months vs. 15.2 months [29].

While variations may occur at different institutions, a common approach for resectable pancreatic cancer would include the surgical resection of the cancer followed by adjuvant chemotherapy. The use of radiation may be used in the adjuvant setting for positive margins following chemotherapy after proving no metastatic disease developed.

#### 3. Treatment of borderline and locally advanced disease

#### 3.1 Neoadjuvant therapy

For patients that present with borderline resectable and locally advanced pancreatic cancer, neoadjuvant chemotherapy with or without chemoradiation allows for systemic control and may improve the likelihood of a R0 resection. The initial rationale for upfront therapies is to potentially downstage tumors to become resectable with a higher R0 resection rate and to allow potential latent metastatic disease to declare itself. In addition, the use of neoadjuvant chemoradiation may be used to "sterilize" the tumor margins near vessel involvement. This allows for selection of the most appropriate patients who have the highest likelihood of longterm survival.

Based on the ACCORD-11 trial showing superior response to FOLFIRINOX, this regimen has now been used effectively in the neoadjuvant setting for borderline and locally advanced pancreatic cancers. Several series have been published showing institutional success. The Massachusetts General Hospital reported that patients treated with mFOLFIRINOX have significantly smaller tumors and lower rates of lymphovascular invasion and perineural invasion. The R0 resection was 92% [30]. Similar reports from the Ohio State University were also noted. They were also able to convert locally advanced and unresectable pancreatic cancers to resectable in 51% of patients who underwent neoadjuvant mFOLFIRINOX with R0 resection of 86% [31].

While patients are undergoing neoadjuvant chemotherapy, serial imaging with pancreatic protocol CT is used to observe for treatment response. In those patients that develop metastatic disease or progression to unresectable disease while undergoing neoadjuvant, their poor biology of disease had declared itself, and they were spared the major morbidity of a surgical resection. For those demonstrating stable or treatment response, the radiologic imaging can be used to predict treatment response. The appearance of the tumor and pancreatic parenchyma interface that becomes more distinct indicates a cytotoxic response which ultimately translates to pathologic response. The ideal response is for the tumor to pull away from the vessels and no longer see haziness around the vessels, which may indicate an infiltrative process. Another prognostic marker of treatment response is normalization of CA 19–9 during neoadjuvant therapy [32].

A pathologic complete response (pCR) can be found in approximately 10% of patients treated in the neoadjuvant approach with FOLFIRINOX and chemoradiation. This is also an independent prognostic risk factor for improved overall and disease-free survival [33]. Additionally, small tumor size, negative

#### Advances in the Treatment of Pancreatic Cancer DOI: http://dx.doi.org/10.5772/intechopen.82074

survival for the fluorouracil group was 16.9 months. There appeared to be a survival

The LAP07 randomized clinical trial aimed to assess if chemoradiation would improve overall survival after 4 months of gemcitabine and to assess erlotinib's effect on survival as well as in patients with locally advanced pancreatic cancer. There was no difference in overall survival between the chemotherapy alone vs. the

While variations may occur at different institutions, a common approach for resectable pancreatic cancer would include the surgical resection of the cancer followed by adjuvant chemotherapy. The use of radiation may be used in the adjuvant setting for positive margins following chemotherapy after proving no

For patients that present with borderline resectable and locally advanced pancreatic cancer, neoadjuvant chemotherapy with or without chemoradiation allows for systemic control and may improve the likelihood of a R0 resection. The initial rationale for upfront therapies is to potentially downstage tumors to become resectable with a higher R0 resection rate and to allow potential latent metastatic disease to declare itself. In addition, the use of neoadjuvant chemoradiation may be used to "sterilize" the tumor margins near vessel involvement. This allows for selection of the most appropriate patients who have the highest likelihood of long-

Based on the ACCORD-11 trial showing superior response to FOLFIRINOX, this regimen has now been used effectively in the neoadjuvant setting for borderline and locally advanced pancreatic cancers. Several series have been published showing institutional success. The Massachusetts General Hospital reported that patients treated with mFOLFIRINOX have significantly smaller tumors and lower rates of lymphovascular invasion and perineural invasion. The R0 resection was 92% [30]. Similar reports from the Ohio State University were also noted. They were also able to convert locally advanced and unresectable pancreatic cancers to resectable in 51% of patients who underwent neoadjuvant mFOLFIRINOX with R0 resection

While patients are undergoing neoadjuvant chemotherapy, serial imaging with pancreatic protocol CT is used to observe for treatment response. In those patients that develop metastatic disease or progression to unresectable disease while undergoing neoadjuvant, their poor biology of disease had declared itself, and they were spared the major morbidity of a surgical resection. For those demonstrating stable or treatment response, the radiologic imaging can be used to predict treatment response. The appearance of the tumor and pancreatic parenchyma interface that becomes more distinct indicates a cytotoxic response which ultimately translates to pathologic response. The ideal response is for the tumor to pull away from the vessels and no longer see haziness around the vessels, which may indicate an infiltrative process. Another prognostic marker of treatment response is normaliza-

A pathologic complete response (pCR) can be found in approximately 10% of

chemoradiation. This is also an independent prognostic risk factor for improved overall and disease-free survival [33]. Additionally, small tumor size, negative

patients treated in the neoadjuvant approach with FOLFIRINOX and

tion of CA 19–9 during neoadjuvant therapy [32].

benefit, but it was not statistically significant [28].

metastatic disease developed.

Current Trends in Cancer Management

3.1 Neoadjuvant therapy

term survival.

of 86% [31].

48

addition of chemoradiation, 16.5 months vs. 15.2 months [29].

3. Treatment of borderline and locally advanced disease

margins, and negative lymph node metastasis are favorable prognostic indicators for improved overall and disease-free survival.

The consensus for treatment of borderline resectable pancreatic cancer favors the neoadjuvant approach; however, it may vary per institution. After a multidisciplinary review at our hospital, the typical functional patient would undergo neoadjuvant chemotherapy (FOLFIRINOX) for 3–4 cycles followed by restaging with CT and CA 19–9. If stable or responding disease, then the patient would continue additional 3–4 cycles of FOLFIRINOX. The patient would again be restaged with CT and CA 19–9. Barring no metastatic disease developed and there was treatment response, the patient would then undergo surgical intervention. However, if the surgical margins were still threatened and there was concern for R1 resection, then the patient may undergo chemoradiation to "sterilize" the margins. Approximately 4–6 weeks after chemoradiation, the patient would ultimately undergo surgical resection.

#### 4. Unresectable pancreatic cancer

Unresectable pancreatic cancer means that the tumor cannot safely be removed due to vascular involvement or metastatic disease. Patients may undergo aggressive chemotherapy with FOLFIRINOX, and a few may be able to convert to a resectable cancer. It is of utmost importance for early palliative care interventions in these patients. For those with biliary obstruction, the use of endoscopic biliary stents and percutaneous biliary drains may provide relief from the jaundice. If the tumor is found to be unresectable in the operating room, then palliative hepaticojejunostomy may be performed. Gastric outlet obstruction may also be relieved with endoscopically placed luminal stents. Additionally, surgical bypass may be performed in laparoscopic or open fashion with a gastrojejunostomy.

Pain can also become quite debilitating in patients with locally advanced unresectable pancreatic cancer. Celiac plexus neurolysis can be performed at the time of surgical exploration, or it may be performed by endoscopic or percutaneous routes.

Irreversible electroporation (IRE) is a nonthermal ablative modality which relies on high voltage (maximum 3,000 volts) small microsecond pulse lengths. This is a novel option typically used in locally advanced pancreatic adenocarcinoma of the head or neck that is not amendable to resection. Some institutions are now using IRE to assist with the resection of locally advanced tumors, but this is not standard at this time. The procedure may be performed open or percutaneously. Patients will typical undergo several months of neoadjuvant chemotherapy to not miss occult metastatic disease prior to IRE. IRE can improve progression-free survival from 6 to 14 months and overall survival from 23 to 20 months [34].

#### 5. Clinical trials

There are several active clinical trials investigating additional treatment options for pancreatic cancer. Several phase II trials are looking at the use of targeted agents in addition to systemic chemotherapy. One study is evaluating the safety of niraparib, PARP [poly (ADP-ribose) polymerase] inhibitor, in advanced pancreatic cancer patients [35]. Another clinical trial at the Massachusetts General Hospital is using the checkpoint inhibitor, nivolumab, as programed death-1 (PD-1) inhibition in combination with losartan, FOLFIRINOX, stereotactic body radiation therapy (SBRT), and surgery in advanced pancreatic cancer. This is a three-armed study:

Arm 1 with FOLFIRINOX, SBRT, and then surgery; Arm 2 with FOLFIRINOX plus losartan, SBRT plus losartan, and then surgery; and Arm 3 with FOLFIRINOX plus losartan, SBRT plus nivolumab and losartan, and then surgery [36].

Reviewing the past studies on chemoradiation, one must keep in mind these studies were using monotherapy chemotherapy and conventional fractionated radiation therapy. There are now several clinical trials assessing the role of radiation therapy, specifically SBRT in the setting of FOLFIRINOX. SBRT utilizes high doses of ablative radiotherapy in typically 1–5 fractions.

The ALLIANCE A021501 is a randomized controlled trial using modified FOLFIRINOX regimen (oxaliplatin 85 mg/m<sup>2</sup> , irinotecan 180 mg/m2 , leucovorin 400 mg/m<sup>2</sup> , and infusional 5-fluorouracil 2400 mg/m<sup>2</sup> over 2 days for 4 cycles) in borderline resectable pancreatic head adenocarcinomas. Arm 1 is delivering this regimen for 8 cycles, while Arm 2 is receiving 7 cycles followed by SBRT (33–40 Gy in 5 fractions). The patient then undergoes pancreaticoduodenectomy followed 4 cycles of adjuvant-modified FOLFOX6 (oxaliplatin 85 mg/m2 , leucovorin 400 mg/m<sup>2</sup> , bolus 5-fluorouracil 400 mg/m<sup>2</sup> , and infusional 5-fluorouracil 2400 mg/m<sup>2</sup> over 2 days for 4 cycles). The main aim of this study is to assess 18-month overall survival, R0 resection, and event-free survival [37].

Another randomized controlled trial by the Pancreatic Cancer Radiotherapy Study Group (PanCRS) is assessing the progression-free survival between mFOLFIRINOX alone vs. mFOLFIRINOX and SBRT in locally advanced unresectable pancreatic cancer [38].

Also, a novel class of drug, cancer stemness inhibitors, is being investigated as a potential new treatment for pancreatic cancer. Napabucasin is an oral small molecule that blocks stem cell activity by targeting the signal transducer and activator of transcription 3 pathway. This pathway is believed to be an important pathway in the propagation of stem-cell-mediated cancer cells [39].

#### 6. Conclusion

While pancreatic cancer is still an aggressive malignancy which is often lethal, there have been significant improvements in the systemic chemotherapy which has improved patients' overall survival. In addition, the radiographic quality has improved thus we are better able to appropriately stage patients for resectability from the onset. Future research in the use of targeted and immunotherapy and the promise of SBRT may control to improve the outcomes of pancreatic cancer patients. With the use of multidisciplinary treatment teams, aggressive combination chemotherapies and surgical resections, there is hope for the patients with pancreatic cancer.

Author details

51

Michelle Marie Fillion

New Hanover Regional Medical Center, Wilmington, North Carolina, USA

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

\*Address all correspondence to: michelle.fillion@nhrmc.org

provided the original work is properly cited.

Advances in the Treatment of Pancreatic Cancer DOI: http://dx.doi.org/10.5772/intechopen.82074

#### Conflict of interest

The authors declare that there are no conflicts of interest.

Advances in the Treatment of Pancreatic Cancer DOI: http://dx.doi.org/10.5772/intechopen.82074

Arm 1 with FOLFIRINOX, SBRT, and then surgery; Arm 2 with FOLFIRINOX plus losartan, SBRT plus losartan, and then surgery; and Arm 3 with FOLFIRINOX plus

Reviewing the past studies on chemoradiation, one must keep in mind these studies were using monotherapy chemotherapy and conventional fractionated radiation therapy. There are now several clinical trials assessing the role of radiation therapy, specifically SBRT in the setting of FOLFIRINOX. SBRT utilizes high doses

The ALLIANCE A021501 is a randomized controlled trial using modified

borderline resectable pancreatic head adenocarcinomas. Arm 1 is delivering this regimen for 8 cycles, while Arm 2 is receiving 7 cycles followed by SBRT

(33–40 Gy in 5 fractions). The patient then undergoes pancreaticoduodenectomy

uracil 2400 mg/m<sup>2</sup> over 2 days for 4 cycles). The main aim of this study is to assess

Another randomized controlled trial by the Pancreatic Cancer Radiotherapy

Also, a novel class of drug, cancer stemness inhibitors, is being investigated as a potential new treatment for pancreatic cancer. Napabucasin is an oral small molecule that blocks stem cell activity by targeting the signal transducer and activator of transcription 3 pathway. This pathway is believed to be an important pathway in

While pancreatic cancer is still an aggressive malignancy which is often lethal, there have been significant improvements in the systemic chemotherapy which has improved patients' overall survival. In addition, the radiographic quality has improved thus we are better able to appropriately stage patients for resectability from the onset. Future research in the use of targeted and immunotherapy and the promise of SBRT may control to improve the outcomes of pancreatic cancer patients. With the use of multidisciplinary treatment teams, aggressive combination chemotherapies and surgical resections, there is hope for the patients with pancre-

, bolus 5-fluorouracil 400 mg/m<sup>2</sup>

followed 4 cycles of adjuvant-modified FOLFOX6 (oxaliplatin 85 mg/m2

18-month overall survival, R0 resection, and event-free survival [37].

the propagation of stem-cell-mediated cancer cells [39].

The authors declare that there are no conflicts of interest.

Study Group (PanCRS) is assessing the progression-free survival between mFOLFIRINOX alone vs. mFOLFIRINOX and SBRT in locally advanced

, and infusional 5-fluorouracil 2400 mg/m<sup>2</sup> over 2 days for 4 cycles) in

, irinotecan 180 mg/m2

, leucovorin

,

, and infusional 5-fluoro-

losartan, SBRT plus nivolumab and losartan, and then surgery [36].

of ablative radiotherapy in typically 1–5 fractions.

FOLFIRINOX regimen (oxaliplatin 85 mg/m<sup>2</sup>

Current Trends in Cancer Management

400 mg/m<sup>2</sup>

leucovorin 400 mg/m<sup>2</sup>

6. Conclusion

atic cancer.

50

Conflict of interest

unresectable pancreatic cancer [38].

### Author details

Michelle Marie Fillion New Hanover Regional Medical Center, Wilmington, North Carolina, USA

\*Address all correspondence to: michelle.fillion@nhrmc.org

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

### References

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[5] Lynch HT, Deters CA, Lynch JF, et al. Familial pancreatic carcinoma in Jews. Familial Cancer. 2004;3:233-240

[6] Lowenfels AB, Maisonneuve P, Whitcomb DC, et al. Risk factors for cancer in hereditary pancreatitis. The Medical Clinics of North America. 2000;

[7] Lynch HT, Voorhees GJ, Lanspa SJ, et al. Pancreatic carcinoma and hereditary nonpolyposis colorectal cancer: A family study. British Journal of

[8] Goggins M, Schutte M, Lu J, et al. Germline BRCA2 gene mutations in patients with apparently sporadic pancreatic carcinomas. Cancer Research. 1996;56:5360-5364

[9] Lynch HT, Smyrk T, Kern SE, et al. Familial pancreatic cancer: A review. Seminars in Oncology. 1996;23:251-275

et al. Trends in pancreatic adenocarcinoma incidence and

Cancer. 2018;18:688-698

Cancer. 1985;52:271-273

84:565-573

52

[19] Neoptolemos JP, Dunn JA, Moffitt DD, et al. ESPAC-1: A European, randomized controlled trial of chemoradiotherapy and chemotherapy after resection of pancreatic cancer. The New England Journal of Medicine. 2004;350:1200-1210

[20] Oettle H, Post S, Neuhaus P, et al. Adjuvant chemotherapy with gemcitabine vs observation in patient undergoing curative intent resection of pancreatic cancer: A randomized controlled trial. JAMA. 2007;297(3): 267-277

[21] Neoptolemos JP, Stocken DD, Smith c T, et al. Adjuvant 5-fluorouracil and folinic acid vs observation for pancreatic cancer: Composite data from the ESPAC-1 and -3(v1) trials. British Journal of Cancer. 2009;100(2): 246-250

[22] Neopltolemos JP, Stocken DD, Bassi C, et al. Adjuvant chemotherapy with fluorouracil plus folinic acid vs gemcitabine following pancreatic cancer resection: A randomized controlled trial. JAMA. 2010;304(10):1073-1081

[23] Von Hoff DD, Ervin T, Arena FP, et al. Increased survival in pancreatic cancer with nab-paclitaxel plus gemcitabine. The New England Journal of Medicine. 2013;369: 1691-1703

[24] Neoptolemos JP, Palmer DH, Ghaneh EF, et al. Comparison of adjuvant gemcitabine and capecitabine with gemcitabine monotherapy in patients with resected pancreatic cancer (ESPAC-4): A multicenter, open label randomized, phase 3 trial. Lancet. 2017; 389:1011-1024

[25] Conroy T, Desseigne F, Ychou M, et al. FOLFIRINOX vs gemcitabine for metastatic pancreatic cancer. The New England Journal of Medicine. 2011;364: 1817-1825

[26] Conroy T. PRODIGE 24: Comparing adjuvant chemotherapy with gemcitabine versus mfolfirinox to treat resected pancreatic adenocarcinoma. In: ASCO Annual Meeting; Chicago, IL, USA. 2018

[27] Stocken DD, Buechler MW, Dervenis C, et al. Meta-analysis of randomised adjuvant therapy trials for pancreatic cancer. British Journal of Cancer. 2005;92(8):1372-1381

[28] Regine WF, Winter KA, Abrams RA, et al. Fluorouracil vs gemcitabine chemotherapy before and after fluorouracil-based chemoradiation following resection of pancreatic adenocarcinoma: A randomized controlled trial. JAMA. 2008;299(9): 1019-1026

[29] Hammel P, Huuguet F, Laethem JL, et al. Effect of chemoradiotherapy vs chemotherapy on survival in patients with locally advanced pancreatic cancer controlled after 4 months of gemcitabine with or without erlotinib: The LAP07 randomized clinical trial. JAMA. 2016;315(17):1844-1853

[30] Ferrone CR, Marchegiani G, Hong TS, et al. Radiological and surgical implications of neoadjuvant treatment with FOLFIRINOX for locally advanced and borderline resectable pancreatic cancer. Annals of Surgery. 2015;261(1): 12-17

[31] Blazer M, Wu C, Goldberg RM, et al. Neoadjuvant mFOLFIRINOX for locally advanced unresectable (LAPC) and borderline resectable (BRPC) adenocarcinoma of the pancreas. Annals of Surgical Oncology. 2015;22(40): 1153-1159

[32] Amer AM, Zais M, Chaudhury B, et al. Imaging-based biomarkers: Changes in the tumor interface of pancreatic ductal adenocarcinoma on computed tomography scans indicate response to cytotoxic therapy. Cancer. 2018;124(8):1701-1709

[33] He J, Blair AB, Groot VP, et al. Is a pathological complete response following neoadjuvant chemoradiation associated with prolonged survival in patients with pancreatic cancer? Annals of Surgery. 2018;268(1):1-8

[34] Martin RC. Use of irreversible electroporation in unresectable pancreatic cancer. Hepatobiliary Surgery and Nutrition. 2015;4(3): 211-215

[35] Cleary J. Niraparib in patients with pancreatic cancer. ClinicalTrials.gov Identifier: NCT03601923. 2018. Retrieved from: https://clinicaltrials. gov/ct2/show/NCT03601923

[36] Hong TS. Losartan and nivolumab in combination with FOLFIRINOX and SBRT in localized pancreatic cancer. ClinicalTrials.gov Identifier: NCT03563248. 2018. Retrieved from: https://clinicaltrials.gov/ct2/show/ NCT03563248

[37] Katz MHG, Herman JM, Ahmad SA, et al. Alliance for clinical trials in oncology (ALLIANCE) trial A021501: preoperative extended chemotherapy vs. chemotherapy plus hypofractionated radiation therapy for borderline resectable adenocarcinoma of the head of the pancreas. BMC Cancer. 2017; 17(1):505

[38] Chang DT. Phase III FOLFIRINOX (mFFX) +/– SBRT in locally advanced pancreatic cancer. ClinicalTrials.gov Identifier: NCT01926197. 2013. Retrieved from: https://clinicaltrials. gov/ct2/show/NCT01926197

[39] Hubbard JH, Grothey A. Napabucasin: An update on the first-inclass cancer stemness inhibitor. Drugs. 2017;77(10):1091-1103

**55**

**Chapter 4**

**Abstract**

vival rates.

prevention, treatment

**1. Introduction**

Colon Cancer

*Mehmet Ali Koc, Suleyman Utku Celik and Cihangir Akyol*

Colorectal cancers (CRCs) are commonly diagnosed malignancy in both men and women. Although it is a common disease, mortality rates decrease with widespread use of screening methods and novel developments in surgery. Physical examination, abdomen and pelvic computerized tomography, and chest imaging are necessary for preoperative staging and surgical planning of a newly diagnosed colon cancer. CRCs usually develop from adenomatous polyps. Although curative treatment of localized colon cancer is surgery, endoscopic polypectomy is sufficient when severe dysplasia or carcinoma in situ is detected on a polyp surface. Total mesorectal excision and neoadjuvant chemoradiotherapy in rectum cancers resulted in significant reductions in morbidity, mortality, and recurrence rates. Recently, complete mesocolic excision and central vascular ligation method has been described in the surgical treatment of colon cancer to achieve similar results. Unfortunately, metastatic colon cancer rate at presentation is approximately 20%. Surgery is a potentially curative option in selected patients with liver and lung metastasis. Pathologic stage of the tumor at presentation is the most important prognostic factor after resection. Therefore, early diagnosis of colon cancer by screening methods and new surgical techniques will lead to better results in sur-

**Keywords:** colon cancer, central vascular ligation, complete mesocolic excision,

Colorectal cancer (CRC) is the second most common cancer in women and third in men with an estimation of approximately 1.4 million new cases globally [1]. Men are more affected than women in most of the world with a higher incidence in North America and Europe and, lower incidence in South-Central Asia and Africa [1]. Although it is a common disease, mortality rates decrease with novel developments in surgery and widespread use of screening methods such as colonoscopy, computed tomography colonoscopy, fecal occult blood test. In the United States, decrease in CRC mortality rates has been shown in the Survey of Epidemiology and End Results (SEER) program [2]. Due to comprehensive researches about the biological and molecular characteristics of CRC, cancer pathogenesis has been well elucidated. Since CRC develops after a long process under the influence of both genetic and environmental factors, early diagnosis is possible and as a result there

Colorectal cancer incidence rises steadily after the age of 50 years and most of the cases are diagnosed in 6th and 7th decades. The incidence under age 40 years is only 5% [3]. Although it is recommended to initiate screening studies at 50 years

are better treatment outcomes and prognosis [1–3].

#### **Chapter 4**

[32] Amer AM, Zais M, Chaudhury B, et al. Imaging-based biomarkers: Changes in the tumor interface of pancreatic ductal adenocarcinoma on computed tomography scans indicate response to cytotoxic therapy. Cancer.

Current Trends in Cancer Management

[39] Hubbard JH, Grothey A.

2017;77(10):1091-1103

Napabucasin: An update on the first-inclass cancer stemness inhibitor. Drugs.

[33] He J, Blair AB, Groot VP, et al. Is a

pathological complete response following neoadjuvant chemoradiation associated with prolonged survival in patients with pancreatic cancer? Annals

of Surgery. 2018;268(1):1-8

gov/ct2/show/NCT03601923

ClinicalTrials.gov Identifier:

NCT03563248

17(1):505

54

211-215

[34] Martin RC. Use of irreversible electroporation in unresectable pancreatic cancer. Hepatobiliary Surgery and Nutrition. 2015;4(3):

[35] Cleary J. Niraparib in patients with pancreatic cancer. ClinicalTrials.gov Identifier: NCT03601923. 2018. Retrieved from: https://clinicaltrials.

[36] Hong TS. Losartan and nivolumab in combination with FOLFIRINOX and SBRT in localized pancreatic cancer.

NCT03563248. 2018. Retrieved from: https://clinicaltrials.gov/ct2/show/

[37] Katz MHG, Herman JM, Ahmad SA, et al. Alliance for clinical trials in oncology (ALLIANCE) trial A021501: preoperative extended chemotherapy vs. chemotherapy plus hypofractionated

radiation therapy for borderline resectable adenocarcinoma of the head of the pancreas. BMC Cancer. 2017;

gov/ct2/show/NCT01926197

[38] Chang DT. Phase III FOLFIRINOX (mFFX) +/– SBRT in locally advanced pancreatic cancer. ClinicalTrials.gov Identifier: NCT01926197. 2013. Retrieved from: https://clinicaltrials.

2018;124(8):1701-1709

## Colon Cancer

*Mehmet Ali Koc, Suleyman Utku Celik and Cihangir Akyol*

#### **Abstract**

Colorectal cancers (CRCs) are commonly diagnosed malignancy in both men and women. Although it is a common disease, mortality rates decrease with widespread use of screening methods and novel developments in surgery. Physical examination, abdomen and pelvic computerized tomography, and chest imaging are necessary for preoperative staging and surgical planning of a newly diagnosed colon cancer. CRCs usually develop from adenomatous polyps. Although curative treatment of localized colon cancer is surgery, endoscopic polypectomy is sufficient when severe dysplasia or carcinoma in situ is detected on a polyp surface. Total mesorectal excision and neoadjuvant chemoradiotherapy in rectum cancers resulted in significant reductions in morbidity, mortality, and recurrence rates. Recently, complete mesocolic excision and central vascular ligation method has been described in the surgical treatment of colon cancer to achieve similar results. Unfortunately, metastatic colon cancer rate at presentation is approximately 20%. Surgery is a potentially curative option in selected patients with liver and lung metastasis. Pathologic stage of the tumor at presentation is the most important prognostic factor after resection. Therefore, early diagnosis of colon cancer by screening methods and new surgical techniques will lead to better results in survival rates.

**Keywords:** colon cancer, central vascular ligation, complete mesocolic excision, prevention, treatment

#### **1. Introduction**

Colorectal cancer (CRC) is the second most common cancer in women and third in men with an estimation of approximately 1.4 million new cases globally [1]. Men are more affected than women in most of the world with a higher incidence in North America and Europe and, lower incidence in South-Central Asia and Africa [1]. Although it is a common disease, mortality rates decrease with novel developments in surgery and widespread use of screening methods such as colonoscopy, computed tomography colonoscopy, fecal occult blood test. In the United States, decrease in CRC mortality rates has been shown in the Survey of Epidemiology and End Results (SEER) program [2]. Due to comprehensive researches about the biological and molecular characteristics of CRC, cancer pathogenesis has been well elucidated. Since CRC develops after a long process under the influence of both genetic and environmental factors, early diagnosis is possible and as a result there are better treatment outcomes and prognosis [1–3].

Colorectal cancer incidence rises steadily after the age of 50 years and most of the cases are diagnosed in 6th and 7th decades. The incidence under age 40 years is only 5% [3]. Although it is recommended to initiate screening studies at 50 years

age, suspicious symptoms like rectal bleeding, unexplained anemia, change in bowel habits, and weight loss should be investigated regardless of the individual's age. Approximately 80% of CRCs are sporadic, 15% are non-syndromic familial, and 5% are syndromic familial cancers [3–5].

CRC is more common in developed societies that consume high-calorie diets rich in animal fat, red meat, processed meat, sweets, refined grains, and alcohol. However a diet rich in fiber, vegetables, fruits, fish, dairy products, and olive oil is beneficial to prevent CRC [4, 5]. The consumption of vegetable fibers shortens the period of contact of the carcinogenic substances with the colon mucosa and at the same time increases the fecal volume and leads to the dilution of the harmful substances so that the adverse effect on the mucosa is reduced. The fat-rich diet stimulates bile acid and cholesterol synthesis in the liver, the amount of these sterols in the colon increases. Due to colon bacteria, production of secondary bile acids and other toxic metabolites are increased and causes negative effects on the colon mucosa [3, 6]. The intake of A, C, E vitamins, calcium, selenium, and carotenoids is thought to reduce the risk of developing CRC [5, 6]. The risk of developing CRC in obese and sedentary individuals also increases like other cancers [7]. Furthermore, chronic alcohol consumption and smoking have been reported to increase the risk of colon adenomas.

The risk of developing CRC in individuals with long-standing inflammatory bowel disease is significantly increased [8]. It is thought that chronic inflammation of the mucosa is a predisposing factor for CRC. Extent and the duration of the colitis is closely related with the development of CRC. While the cumulative risk of CRC in ulcerative colitis patients with pancolitis or left-sided disease is 1.6% at 10 years, it increases approximately 5 times at 20 years (8.3%), and 11 times at 30 years (18.4%) [9]. A similar risk for CRC is also associated with Crohn's disease. Screening colonoscopy for CRC has been recommended annually for patients with inflammatory bowel disease, 8–10 years after the first symptoms of the disease [10].

#### **2. Pathogenesis**

Most CRCs usually develop from adenomatous polyps that become dysplastic (adenoma-carcinoma sequence) (**Figures 1** and **2**). The epithelium of small bowel is constantly renewed. During this renewal process, progressive deterioration leads primarily to adenomatous polyps and later to dysplasia and invasive cancer. Hypothesis that CRC are a result of adenoma-carcinoma sequence are supported by findings such as frequent early carcinoma detection in large adenomatous polyps, detection of adenomas in patients 10 years before cancer in both sporadic and familial cases, and reduction of CRC incidence by removal of polyps in controlled trials [11].

CRCs occur by accumulation of epigenetic and genetic changes over time [12]. These changes transform normal glandular epithelium into adenocarcinoma. In hereditary forms of CRC, individuals are born with mutant genes. That means the mutant gene is present in one allele in the zygote from the beginning (germ-line mutation) but a second hit needed. Hereditary non-polyposis colorectal cancer (HNPCC) and familial adenomatous polyposis (FAP) are the best known types of hereditary CRC. Sometimes mutations develop after birth due to environmental factors (somatic mutation) and sporadic cancers occur [10–13].

Tumor suppressor gene mutations may remove an inhibitory signal while oncogenic mutations may cause overexpression of a gene or pathway [11]. These changes

**57**

**Figure 2.**

that cause CRC, which is a different heterogenous disease in each person, affect the phenotype of the disease, prognosis and response to treatment. Chromosomal instability, microsatellite instability, and the methylator phenotype are the three major molecular pathways that involved in CRC development.

*Adenoma—carcinoma sequence (figure taken from with permission of Prof. Kuzu, Turkish Society of Colon and Rectal Surgery; Colon and Rectum Cancers.Eds, Baykan A, Zorluoglu A, Gecim E, Terzi C. 2010).*

*Colon Cancer*

**Figure 1.**

*Adenoma—carcinoma sequence.*

*DOI: http://dx.doi.org/10.5772/intechopen.81597*

*Current Trends in Cancer Management*

of colon adenomas.

of the disease [10].

**2. Pathogenesis**

controlled trials [11].

and 5% are syndromic familial cancers [3–5].

age, suspicious symptoms like rectal bleeding, unexplained anemia, change in bowel habits, and weight loss should be investigated regardless of the individual's age. Approximately 80% of CRCs are sporadic, 15% are non-syndromic familial,

CRC is more common in developed societies that consume high-calorie diets rich in animal fat, red meat, processed meat, sweets, refined grains, and alcohol. However a diet rich in fiber, vegetables, fruits, fish, dairy products, and olive oil is beneficial to prevent CRC [4, 5]. The consumption of vegetable fibers shortens the period of contact of the carcinogenic substances with the colon mucosa and at the same time increases the fecal volume and leads to the dilution of the harmful substances so that the adverse effect on the mucosa is reduced. The fat-rich diet stimulates bile acid and cholesterol synthesis in the liver, the amount of these sterols in the colon increases. Due to colon bacteria, production of secondary bile acids and other toxic metabolites are increased and causes negative effects on the colon mucosa [3, 6]. The intake of A, C, E vitamins, calcium, selenium, and carotenoids is thought to reduce the risk of developing CRC [5, 6]. The risk of developing CRC in obese and sedentary individuals also increases like other cancers [7]. Furthermore, chronic alcohol consumption and smoking have been reported to increase the risk

The risk of developing CRC in individuals with long-standing inflammatory bowel disease is significantly increased [8]. It is thought that chronic inflammation of the mucosa is a predisposing factor for CRC. Extent and the duration of the colitis is closely related with the development of CRC. While the cumulative risk of CRC in ulcerative colitis patients with pancolitis or left-sided disease is 1.6% at 10 years, it increases approximately 5 times at 20 years (8.3%), and 11 times at 30 years (18.4%) [9]. A similar risk for CRC is also associated with Crohn's disease. Screening colonoscopy for CRC has been recommended annually for patients with inflammatory bowel disease, 8–10 years after the first symptoms

Most CRCs usually develop from adenomatous polyps that become dysplastic (adenoma-carcinoma sequence) (**Figures 1** and **2**). The epithelium of small bowel is constantly renewed. During this renewal process, progressive deterioration leads primarily to adenomatous polyps and later to dysplasia and invasive cancer. Hypothesis that CRC are a result of adenoma-carcinoma sequence are supported by findings such as frequent early carcinoma detection in large adenomatous polyps, detection of adenomas in patients 10 years before cancer in both sporadic and familial cases, and reduction of CRC incidence by removal of polyps in

CRCs occur by accumulation of epigenetic and genetic changes over time [12]. These changes transform normal glandular epithelium into adenocarcinoma. In hereditary forms of CRC, individuals are born with mutant genes. That means the mutant gene is present in one allele in the zygote from the beginning (germ-line mutation) but a second hit needed. Hereditary non-polyposis colorectal cancer (HNPCC) and familial adenomatous polyposis (FAP) are the best known types of hereditary CRC. Sometimes mutations develop after birth due to environmental

Tumor suppressor gene mutations may remove an inhibitory signal while oncogenic mutations may cause overexpression of a gene or pathway [11]. These changes

factors (somatic mutation) and sporadic cancers occur [10–13].

**56**

**Figure 1.** *Adenoma—carcinoma sequence.*

#### **Figure 2.**

*Adenoma—carcinoma sequence (figure taken from with permission of Prof. Kuzu, Turkish Society of Colon and Rectal Surgery; Colon and Rectum Cancers.Eds, Baykan A, Zorluoglu A, Gecim E, Terzi C. 2010).*

that cause CRC, which is a different heterogenous disease in each person, affect the phenotype of the disease, prognosis and response to treatment. Chromosomal instability, microsatellite instability, and the methylator phenotype are the three major molecular pathways that involved in CRC development. Each pathway has unique characteristics, and multiple pathways may play a role in the development of CRC [12, 13].

#### **2.1 Chromosomal instability**

Chromosomes are unstable in chromosomal instability (CIN), because of a change in the chromosome structure or copy number (Loss of heterozygosity-LOH). CIN is the most common occurrence in CRC [13]. Approximately 80% of CRC patients have CIN. Vogelstein and Fearon described the classical adenoma-carcinoma sequence and their study supported that LOH was responsible for the sequence [14]. The main genes which play role in the carcinogenesis are the adenomatous polyposis coli (APC), K-ras, deleted in colon cancer (DCC), and P53 (**Figure 1**).

The APC is a tumor suppressor gene, therefore, mutation in both alleles are necessary for the initiation of the sequence. Mutated APC causes decreased production or lack of APC protein. Thus, translocation into the nucleus due to intracellular accumulation of β-caterin, which is controlled by the APC protein to regulate the WNT signaling pathway, causes alterations in cell signaling, proliferation, and adhesion [15]. The APC gene is first described in patients with FAP. However, it was then reported that majority of the sporadic CRC has the APC gene mutation and APC mutation is present in adenomas smaller than 0.5 cm [16].

K-ras is a cellular variant of RAS oncogenes and the most frequently mutated RAS proto-oncogene in CRC. Since K-ras is a proto-oncogene, mutation of only one allele is enough. K-ras gene encodes a G-protein (Guanine nucleotide binding protein) that is active when GTP bond state and inactive (GDP-bond state) after hydrolyzed by GTPase. This protein is involved in mitogen-activated protein kinase (MAPK) pathway which promotes cell growth and proliferation. RAS mutation results in an active GTP-bond protein, which is unable to switch off by GTPase, and leads to uncontrolled cell division. About 43% of non-hypermutated (Microsatellite stabile-MSS) CRC, which are nearly 80% of CRC, has RAS mutations [17].

DCC and SMAD4 mutations have been found in CRCs [18, 19]. Both are tumor suppressor genes. DCC gene product is thought to be involved in cell differentiation and adhesion in CRC [20]. *DCC* and SMAD4 (formerly PC4-deleted in pancreatic cancer) were both identified at 18q. SMAD4 mutations is thought to perturb TGF-beta signaling pathway which has an inhibitory influence on normal cell growth [19, 20].

TP53 gene on chromosome 17p encode P53 protein which arrests the cell cycle and facilitates DNA repair [21]. In all human cancers most of the mutations occur in TP53 gene. TP53 mutation occurs in about 75% of CRCs [14]. However it is not frequent in adenomas, therefore, it is considered to be a late event in CRC tumorigenesis and related with invasiveness [22, 23].

#### **2.2 Microsatellite instability (MSI)**

Microsatellites are non-coding DNA segments containing 1 to 4 repetitive nucleotide sequences. In normal individuals microsatellites are completely identical in all cells. But the failure of the DNA mismatch repair genes to function properly causes a change in the length of the microsatellite sites that are already prone to error during copying. This is called microsatellite instability. There are also short repetitive segments in various tumor suppressor genes (TSG), and accumulation of the mutations in TSGs due to the inactivity of MMR genes (most commonly MLH1 or MSH2) lead to the development of adenoma and subsequent carcinoma [21–23].

It is possible to detect microsatellite instability with current diagnostic procedures, as long as many cells carry the same abnormality, which means that the cells

**59**

*Colon Cancer*

neoplastic process [24].

*DOI: http://dx.doi.org/10.5772/intechopen.81597*

**2.3 CpG island methylator phenotype (CIMP)**

**3. Hereditary colorectal cancers**

MSI-H tumors [30].

belong to the clonal process. Clonal proliferation is a characteristic feature of the neoplastic process. And It should be understood that MSI is an indicator of a clonal

Cancers arising through MSI pathway are approximately 15% of all CRC and tend to be hypermutated, therefore, are also termed the mutator phenotype [17].

However, prognosis is better than cancers arising through CIN pathway.

However, Lynch-related CRCs only have K-ras mutations [28, 29].

**3.1 Hereditary non-polyposis colorectal cancer (HNPCC)**

and within 30 years this rate reaches to 62% [32].

10-year survival from CRC is 91% [33].

HNPCC is the most common hereditary colorectal cancer. HNPCC is also termed Lynch syndrome. In Lynch syndrome, CRC and endometrial cancer risk are significantly increased as well as several other malignancies. It accounts for 3% of all colorectal cancers [30]. Lynch is an autosomal dominant disease. Mutations in DNA repair genes (mismatch repair-MMR) are detected in affected individuals. In Lynch syndrome there is a germline mutation. Since this mutation is only in one allele, a second hit is necessary (mutation, loss of heterozygosity, or epigenetic silencing). Colorectal cancer develops in 80% of patients around 40 years of age. The most frequently mutated MMR genes in HNPCC were MLH 1 (37%), MSH 2 (41%), MSH 6, and PMS 2. CRCs developed in Lynch syndrome are

In patients with Lynch syndrome, the risk of synchronous and metachronous tumors is increased, and approximately 7% of patients have a second tumor at the time of diagnosis [31]. Metachronous tumors develop within 10 years in 16% of individuals who had previously undergone colon resection due to Lynch syndrome

Lynch-associated CRCs also evolve from adenomas like most CRCs. However the adenomas are more often proximally located, and more likely to be larger and flatter. And as compared with sporadic adenomas, high-grade dysplasia and/or villous histology are more often detected. It is also known that the adenoma-carcinoma sequence progresses more rapidly in Lynch syndrome. Fortunately, the overall

Tumors in HNPCC are more often found in the proximal colon than sporadic cancers. Unlike sporadic cancers, tumor in Lynch is poor differentiated and there is

Another common pathway in CRC is epigenetic instability. Epigenetic alterations such as hypermethylation of DNA promoter regions can silence gene transcription and contributes to diseases like cancer. Methylation of cytosine is normally an essential process and controls multiple processes [25]. There are cytosine-guanine (CpG) dinucleotide enriched areas in promoter regions. These CpG enriched regions of genes are called CpG islands and normally maintained in an unmethylated state. Several tumor suppressor genes contain CpG repetitive sequences in the promoter region. Aberrant methylation of these CpG islands silences gene transcription and contributes to cancer process [25, 26]. This phenomenon is called CpG island methylator phenotype (CIMP). Especially methylation of the MMR gene, hMLH1 causes approximately 80% of MSI CRCs [27]. Almost all MSI-high (MSI–H), CIMP+ cancers without K-ras mutations have BRAF mutations.

#### *Colon Cancer DOI: http://dx.doi.org/10.5772/intechopen.81597*

*Current Trends in Cancer Management*

the development of CRC [12, 13].

**2.1 Chromosomal instability**

Each pathway has unique characteristics, and multiple pathways may play a role in

Chromosomes are unstable in chromosomal instability (CIN), because of a change in the chromosome structure or copy number (Loss of heterozygosity-LOH). CIN is the most common occurrence in CRC [13]. Approximately 80% of CRC patients have CIN. Vogelstein and Fearon described the classical adenoma-carcinoma sequence and their study supported that LOH was responsible for the sequence [14]. The main genes which play role in the carcinogenesis are the adenomatous polyposis

The APC is a tumor suppressor gene, therefore, mutation in both alleles are necessary for the initiation of the sequence. Mutated APC causes decreased production or lack of APC protein. Thus, translocation into the nucleus due to intracellular accumulation of β-caterin, which is controlled by the APC protein to regulate the WNT signaling pathway, causes alterations in cell signaling, proliferation, and adhesion [15]. The APC gene is first described in patients with FAP. However, it was then reported that majority of the sporadic CRC has the APC gene mutation and

K-ras is a cellular variant of RAS oncogenes and the most frequently mutated RAS proto-oncogene in CRC. Since K-ras is a proto-oncogene, mutation of only one allele is enough. K-ras gene encodes a G-protein (Guanine nucleotide binding protein) that is active when GTP bond state and inactive (GDP-bond state) after hydrolyzed by GTPase. This protein is involved in mitogen-activated protein kinase (MAPK) pathway which promotes cell growth and proliferation. RAS mutation results in an active GTP-bond protein, which is unable to switch off by GTPase, and leads to uncontrolled cell division. About 43% of non-hypermutated (Microsatellite

DCC and SMAD4 mutations have been found in CRCs [18, 19]. Both are tumor suppressor genes. DCC gene product is thought to be involved in cell differentiation and adhesion in CRC [20]. *DCC* and SMAD4 (formerly PC4-deleted in pancreatic cancer) were both identified at 18q. SMAD4 mutations is thought to perturb TGF-beta signaling pathway which has an inhibitory influence on normal cell growth [19, 20]. TP53 gene on chromosome 17p encode P53 protein which arrests the cell cycle and facilitates DNA repair [21]. In all human cancers most of the mutations occur in TP53 gene. TP53 mutation occurs in about 75% of CRCs [14]. However it is not frequent in adenomas, therefore, it is considered to be a late event in CRC tumori-

Microsatellites are non-coding DNA segments containing 1 to 4 repetitive nucleotide sequences. In normal individuals microsatellites are completely identical in all cells. But the failure of the DNA mismatch repair genes to function properly causes a change in the length of the microsatellite sites that are already prone to error during copying. This is called microsatellite instability. There are also short repetitive segments in various tumor suppressor genes (TSG), and accumulation of the mutations in TSGs due to the inactivity of MMR genes (most commonly MLH1 or MSH2) lead to the development of adenoma and subsequent carcinoma [21–23]. It is possible to detect microsatellite instability with current diagnostic procedures, as long as many cells carry the same abnormality, which means that the cells

stabile-MSS) CRC, which are nearly 80% of CRC, has RAS mutations [17].

coli (APC), K-ras, deleted in colon cancer (DCC), and P53 (**Figure 1**).

APC mutation is present in adenomas smaller than 0.5 cm [16].

genesis and related with invasiveness [22, 23].

**2.2 Microsatellite instability (MSI)**

**58**

belong to the clonal process. Clonal proliferation is a characteristic feature of the neoplastic process. And It should be understood that MSI is an indicator of a clonal neoplastic process [24].

Cancers arising through MSI pathway are approximately 15% of all CRC and tend to be hypermutated, therefore, are also termed the mutator phenotype [17]. However, prognosis is better than cancers arising through CIN pathway.

#### **2.3 CpG island methylator phenotype (CIMP)**

Another common pathway in CRC is epigenetic instability. Epigenetic alterations such as hypermethylation of DNA promoter regions can silence gene transcription and contributes to diseases like cancer. Methylation of cytosine is normally an essential process and controls multiple processes [25]. There are cytosine-guanine (CpG) dinucleotide enriched areas in promoter regions. These CpG enriched regions of genes are called CpG islands and normally maintained in an unmethylated state. Several tumor suppressor genes contain CpG repetitive sequences in the promoter region. Aberrant methylation of these CpG islands silences gene transcription and contributes to cancer process [25, 26]. This phenomenon is called CpG island methylator phenotype (CIMP). Especially methylation of the MMR gene, hMLH1 causes approximately 80% of MSI CRCs [27]. Almost all MSI-high (MSI–H), CIMP+ cancers without K-ras mutations have BRAF mutations. However, Lynch-related CRCs only have K-ras mutations [28, 29].

#### **3. Hereditary colorectal cancers**

#### **3.1 Hereditary non-polyposis colorectal cancer (HNPCC)**

HNPCC is the most common hereditary colorectal cancer. HNPCC is also termed Lynch syndrome. In Lynch syndrome, CRC and endometrial cancer risk are significantly increased as well as several other malignancies. It accounts for 3% of all colorectal cancers [30]. Lynch is an autosomal dominant disease. Mutations in DNA repair genes (mismatch repair-MMR) are detected in affected individuals. In Lynch syndrome there is a germline mutation. Since this mutation is only in one allele, a second hit is necessary (mutation, loss of heterozygosity, or epigenetic silencing). Colorectal cancer develops in 80% of patients around 40 years of age. The most frequently mutated MMR genes in HNPCC were MLH 1 (37%), MSH 2 (41%), MSH 6, and PMS 2. CRCs developed in Lynch syndrome are MSI-H tumors [30].

In patients with Lynch syndrome, the risk of synchronous and metachronous tumors is increased, and approximately 7% of patients have a second tumor at the time of diagnosis [31]. Metachronous tumors develop within 10 years in 16% of individuals who had previously undergone colon resection due to Lynch syndrome and within 30 years this rate reaches to 62% [32].

Lynch-associated CRCs also evolve from adenomas like most CRCs. However the adenomas are more often proximally located, and more likely to be larger and flatter. And as compared with sporadic adenomas, high-grade dysplasia and/or villous histology are more often detected. It is also known that the adenoma-carcinoma sequence progresses more rapidly in Lynch syndrome. Fortunately, the overall 10-year survival from CRC is 91% [33].

Tumors in HNPCC are more often found in the proximal colon than sporadic cancers. Unlike sporadic cancers, tumor in Lynch is poor differentiated and there is peritumoral lymphocytic infiltration, and Crohn's-like reaction [34]. However, the prognosis is still better than sporadic colorectal cancer. The following three theories stand out for this reason: earlier diagnosis in HNPCC tumors, the genomic instability in HNPCC tumors leads to the continual increase of mutations and the loss of critical functions and metastatic ability of the tumor cell due to this mutation burden, and Crohn's-like lymphocytic infiltration around the tumor enhances host immunity by expressing IL-4, TNF-a [33, 34].

Two different forms of HNPCC have been described. Lynch Syndrome I is characterized with proximal colon tumor, young age, no extracolonic involvement. Generally same colonic segment is involved in other relatives. In Lynch Syndrome II, in addition to Lynch I, stomach, small intestine, pancreas, ovary, endometrium and urinary tract cancers may develop. The most important tool in diagnosing Lynch syndrome is family history of CRC or other cancers related with Lynch. Several family history-based criteria (Revised Bethesda Guidelines and Amsterdam II Criteria) have been used to determine the people at risk for HNPCC (**Table 1**) [34].

MSI is a characteristic of tumors in HNPCC and caused by a loss of DNA MMR. An MSI screening test is required for patients with a positive Bethesda criteria. Polymerase chain reaction is used to test for MSI by copying a panel of DNA sequences that contains nucleotide repeats [33–35]. Family members who meet the Amsterdam II criteria or revised Bethesda guidelines, or those with a diagnosis of endometrium cancer prior to the age of 50 years, or individuals with a MMR gene mutation in the first degree relatives are at risk for Lynch syndrome.

In Lynch syndrome, synchronous or metachronous cancer and polyp development are common. It is important to be cautious in this regard. Colonoscopy should be done every one or two years starting from the age of 20–25 [36, 37]. Gynecological examination and endometrial aspiration biopsy for endometrium cancer, and transvaginal ultrasonography for ovarian cancer should be done once a

#### **Amsterdam II Criteria and Revised Bethesda Guidelines**

*Amsterdam II Criteria* [35] All criteria must be met:


*Revised Bethesda Guidelines* [34]

One or more of the following criteria must be met:


*HNPCC: hereditary nonpolyposis colorectal cancer; MSI: microsatellite instability. \*HNPCC-related tumors include colorectal, endometrial, stomach, ovarian, pancreas, ureter and renal pelvis, biliary tract, and brain (usually glioblastoma as seen in Turcot syndrome) tumors, sebaceous gland adenomas and keratoacanthomas in Muir-Torre syndrome, and carcinoma of the small bowel. \*\*Presence of tumor-infiltrating lymphocytes, Crohn's-like lymphocytic reaction, mucinous/signet-ring cell differentiation, or medullary growth pattern.*

**61**

**Figure 3.**

*Colonoscopic familial adenomatous polyposis (FAP).*

*Colon Cancer*

*3.1.1 Treatment*

*DOI: http://dx.doi.org/10.5772/intechopen.81597*

year starting from the age of 30–35, or 3–5 years prior to the age of a relative diagnosed with HNPCC [37]. Upper gastrointestinal endoscopy should be done once every 2 years starting from the age of 30–35, with gastric biopsy and treatment for *Helicobacter pylori* infection when found on biopsy. Renal ultrasound, urine analysis and cytology should be done every year starting from the age of 30–35 [36, 37].

Prophylactic colectomy is not considered in patients without CRC. Tumor is usually located in the proximal colon. Total abdominal colectomy—ileorectal anastomosis is recommended in patients with Lynch syndrome. Considering the quality of life in elderly patients, or in patients who are not eligible for total colectomy a segmental colectomy may be recommended according to the location of the tumor [36]. However patients who undergo segmental colectomy are at increased risk for subsequent CRC as compared to patients with total abdominal colectomy—ileorectal anastomosis [38, 39]. Prophylactic hysterectomy and oophorectomy should be recommended at the time of colorectal surgery. However, it may be recommended

It accounts for 1% of all colorectal cancers and is an autosomal dominant disease. FAP is characterized by the presence of hundreds of adenomas in the colon (**Figure 3**). It is a broad spectrum disease with extraintestinal manifestations. It usually occurs after puberty and is diagnosed at a mean age of 29 years. Colorectal cancer develops at an average age of 39 years. Colorectal cancer is unavoidable in patients who do not undergo

Mutation in the APC (adenomatous polyposis coli) gene, which is located at the q21 locus of chromosome 5, is responsible for FAP (5q deletion) [40, 41]. Mutations in codons close to the 3'and 5' ends of the APC gene lead to attenuated FAP (AFAP), while mutations in the middle, between codons 169 to 1393, result in FAP. Generally, there is less than 100 polyps in AFAP. Unlike FAP, life-long risk for CRC development in AFAP is approximately 70%, and polyps and CRCs develop later in life than FAP. In AFAP, tumors mostly do not develop in the rectum and are characterized by a more proximal distribution in the colon. If germline mutation is absent in these patients, MMR gene mutation should be considered for HNPCC elimination [41]. In

for women who aged 35 years or older after family planning [39].

**3.2 Familial adenomatous polyposis (FAP)**

surgery. FAP is 80% familial, and 20% sporadic [40].

#### **Table 1.**

*Amsterdam II Criteria and Revised Bethesda Guidelines.*

#### *Colon Cancer DOI: http://dx.doi.org/10.5772/intechopen.81597*

year starting from the age of 30–35, or 3–5 years prior to the age of a relative diagnosed with HNPCC [37]. Upper gastrointestinal endoscopy should be done once every 2 years starting from the age of 30–35, with gastric biopsy and treatment for *Helicobacter pylori* infection when found on biopsy. Renal ultrasound, urine analysis and cytology should be done every year starting from the age of 30–35 [36, 37].

#### *3.1.1 Treatment*

*Current Trends in Cancer Management*

(**Table 1**) [34].

immunity by expressing IL-4, TNF-a [33, 34].

peritumoral lymphocytic infiltration, and Crohn's-like reaction [34]. However, the prognosis is still better than sporadic colorectal cancer. The following three theories stand out for this reason: earlier diagnosis in HNPCC tumors, the genomic instability in HNPCC tumors leads to the continual increase of mutations and the loss of critical functions and metastatic ability of the tumor cell due to this mutation burden, and Crohn's-like lymphocytic infiltration around the tumor enhances host

Two different forms of HNPCC have been described. Lynch Syndrome I is characterized with proximal colon tumor, young age, no extracolonic involvement. Generally same colonic segment is involved in other relatives. In Lynch Syndrome II, in addition to Lynch I, stomach, small intestine, pancreas, ovary, endometrium and urinary tract cancers may develop. The most important tool in diagnosing Lynch syndrome is family history of CRC or other cancers related with Lynch. Several family history-based criteria (Revised Bethesda Guidelines and Amsterdam II Criteria) have been used to determine the people at risk for HNPCC

MSI is a characteristic of tumors in HNPCC and caused by a loss of DNA MMR. An MSI screening test is required for patients with a positive Bethesda criteria. Polymerase chain reaction is used to test for MSI by copying a panel of DNA sequences that contains nucleotide repeats [33–35]. Family members who meet the Amsterdam II criteria or revised Bethesda guidelines, or those with a diagnosis of endometrium cancer prior to the age of 50 years, or individuals with a MMR gene

In Lynch syndrome, synchronous or metachronous cancer and polyp development are common. It is important to be cautious in this regard. Colonoscopy should be done every one or two years starting from the age of 20–25 [36, 37]. Gynecological examination and endometrial aspiration biopsy for endometrium cancer, and transvaginal ultrasonography for ovarian cancer should be done once a

• Three or more individuals with colorectal cancers or HNPCC-related cancers, and one of them being

• At least one relative has colorectal or HNPCC-related cancer diagnosed before the age of 50 years.

• Synchronous or metachronous colorectal cancer or other HNPCC-related tumors\*, regardless of age,

• Colorectal cancer diagnosed in one or more first degree relatives with HNPCC-related tumor, and one

• Colorectal cancer in 2 or more first- or second-degree relatives with HNPCC-associated tumors,

*\*HNPCC-related tumors include colorectal, endometrial, stomach, ovarian, pancreas, ureter and renal pelvis, biliary tract, and brain (usually glioblastoma as seen in Turcot syndrome) tumors, sebaceous gland adenomas and* 

*\*\*Presence of tumor-infiltrating lymphocytes, Crohn's-like lymphocytic reaction, mucinous/signet-ring cell* 

• Colorectal cancer with MSI-high histology\*\* diagnosed before the age of 60 years

mutation in the first degree relatives are at risk for Lynch syndrome.

**Amsterdam II Criteria and Revised Bethesda Guidelines**

a first-degree relative of the other two, • Two or more successive generations are affected,

One or more of the following criteria must be met:

• Colorectal cancer diagnosed before the age of 50 years,

of them being diagnosed before the age of 50 years,

*HNPCC: hereditary nonpolyposis colorectal cancer; MSI: microsatellite instability.*

*keratoacanthomas in Muir-Torre syndrome, and carcinoma of the small bowel.*

*Amsterdam II Criteria* [35] All criteria must be met:

*Revised Bethesda Guidelines* [34]

regardless of age.

*differentiation, or medullary growth pattern.*

*Amsterdam II Criteria and Revised Bethesda Guidelines.*

**60**

**Table 1.**

Prophylactic colectomy is not considered in patients without CRC. Tumor is usually located in the proximal colon. Total abdominal colectomy—ileorectal anastomosis is recommended in patients with Lynch syndrome. Considering the quality of life in elderly patients, or in patients who are not eligible for total colectomy a segmental colectomy may be recommended according to the location of the tumor [36]. However patients who undergo segmental colectomy are at increased risk for subsequent CRC as compared to patients with total abdominal colectomy—ileorectal anastomosis [38, 39]. Prophylactic hysterectomy and oophorectomy should be recommended at the time of colorectal surgery. However, it may be recommended for women who aged 35 years or older after family planning [39].

#### **3.2 Familial adenomatous polyposis (FAP)**

It accounts for 1% of all colorectal cancers and is an autosomal dominant disease. FAP is characterized by the presence of hundreds of adenomas in the colon (**Figure 3**). It is a broad spectrum disease with extraintestinal manifestations. It usually occurs after puberty and is diagnosed at a mean age of 29 years. Colorectal cancer develops at an average age of 39 years. Colorectal cancer is unavoidable in patients who do not undergo surgery. FAP is 80% familial, and 20% sporadic [40].

Mutation in the APC (adenomatous polyposis coli) gene, which is located at the q21 locus of chromosome 5, is responsible for FAP (5q deletion) [40, 41]. Mutations in codons close to the 3'and 5' ends of the APC gene lead to attenuated FAP (AFAP), while mutations in the middle, between codons 169 to 1393, result in FAP. Generally, there is less than 100 polyps in AFAP. Unlike FAP, life-long risk for CRC development in AFAP is approximately 70%, and polyps and CRCs develop later in life than FAP. In AFAP, tumors mostly do not develop in the rectum and are characterized by a more proximal distribution in the colon. If germline mutation is absent in these patients, MMR gene mutation should be considered for HNPCC elimination [41]. In

**Figure 3.** *Colonoscopic familial adenomatous polyposis (FAP).*

FAP, one inherited mutant allele is not enough to cause carcinoma. Carcinoma develops when the second allele of APC and other necessary gene mutations occur [42].

Extraintestinal manifestations of FAP include gastric, duodenal, and periampullary polyps, and less common manifestations such as epidermoid cysts, desmoid tumors, osteomas, and brain tumors. Gastric and duodenal polyps occur in about half of affected individuals. Most of the gastric polyps are hyperplasia of the fundus glands, rather than adenomatous polyps and their malignancy potential is limited [43]. However duodenal polyps are adenomatous. They are present in approximately 90% of FAP patients and should be considered premalignant [44, 45]. Periampullary tumor risk is higher in FAP patients. In patients who undergo total colectomy, the most important cause of cancer-related death is duodenal adenocancer. Adenomatous polyps and cancer are rarely found in the jejunum and ileum of FAP patients. Other rare extraintestinal malignancies in FAP patients are extrahepatic bile duct, gallbladder, pancreas, adrenal, thyroid, and liver cancers [45].

The likelihood of a desmoid tumor is increased especially in mutations in the 3' end of the APC gene [46]. Most of the desmoid tumors occur within the first 5 years in patients who have undergone abdominal surgery, presumably as an inflammatory response [47, 48]. In addition to abdominal surgery and APC mutation, pregnancy, female sex, and family history are other risk factors for desmoid tumors [49, 50]. Although desmoid tumors are slow growing, non-metastatic mesenchymal tumors, they may cause complications such as pain, bowel, and ureter obstruction by compressing and encasing adjacent structures [50].

An interesting marker for FAP is congenital hypertrophy of the retinal pigment epithelium (CHRPE), which can be determined by ophthalmoscopy in about 75% of patients [51]. Fundus examination with ophthalmoscopy reveals oval, pigmented lesions with regular borders in the retina. Lesions may be bilateral and multiple. CHRPE can be used as a clinical diagnostic tool in the screening of FAP and Gardner syndrome [51].

FAP has two subtypes with their own extracolonic manifestations. Gardner's syndrome is a variant of FAP and characterized by desmoid tumors, colonic polyps, osteoma, soft tissue sarcomas, and CHRPE [52]. Also, although very rare, an adenomatous polyposis coli may be associated with malignant tumors of the central nervous system (especially medulloblastoma and/or glioma), known as Turcot syndrome [53]. Turcot is the true variant form of FAP and has a familial character. Colonoscopy and brain scanning tests should also be performed on family members. APC mutations are also responsible for both syndromes.

Screening for FAP should be performed in individuals with an APC mutation and in individuals who are first-degree relatives of those with FAP, or who have >10 cumulative colorectal adenomas, or colorectal adenomas in combination with extracolonic features such as duodenal adenomas, desmoid tumors, osteomas, etc. [53, 54].

Screening for CRC should begin during puberty and flexible sigmoidoscopy, or genetic testing for APC mutations should be performed every 6 months or year. When positive genotype is detected by genetic screening, or adenomatous polyps are detected by sigmoidoscopy, full colonoscopy should be performed to evaluate the spread of the disease. Several polyps should also be sampled to confirm histology. First-degree relatives of FAP patients, who do not have a genetic diagnosis, may be removed from aggressive follow-up and included in standard general population screening programs if there is no polyp detected until 40 years of age on screening [54]. Screening of the upper gastrointestinal tract should be performed at the time of diagnosis, or before 25 years of age. In later periods, it should be done while the colon is being evaluated. Thyroid cancer is rarely seen in patients with FAP. Studies have shown that thyroid

**63**

*Colon Cancer*

*3.2.1 Genetic testing*

*3.2.2 Treatment*

prevention are unclear.

*3.2.2.2 Surgical treatment*

*DOI: http://dx.doi.org/10.5772/intechopen.81597*

history but without a FAP trait in the family [36].

*3.2.2.1 Medical treatment and chemoprevention*

mesenteric desmoids that may develop [54].

cancer reaches up to 2.6% and thyroid nodules up to 51.7%. Therefore, it is recom-

Identification of the APC mutation and its type in a patient diagnosed with FAP facilitates screening of other family members as well as recognition of possible phenotypic lesions that may result from different APC mutations. Commonly accepted indications for genetic testing include FAP cases, FAP in first degree relatives, APC mutation in first degree relatives, at least 10 cumulative colorectal adenomas, extracolonic involvement of FAP and multiple adenomas with colorectal cancer family

In cases with FAP non-steroid anti-inflammatory drugs (NSAIDs) are thought to achieve regression in number and size of polyps [57, 58]. The most commonly used agents for this purpose are sulindac and celecoxib. The role of chemopreventive agents in FAP patients is controversial, because the effects of these agents on cancer

Prophylactic colectomy should be performed in all cases with FAP. The timing of surgery is planned according to the number of polyps, number of adenomas, presence of dysplasia, size of polyps, symptoms and characteristics of the patient. In cases with mild to moderate polyposis and no other risk factors (low risk of cancer), surgery can be done at mid-puberty. However patients should continue to undergo annual CRC surveillance with colonoscopy while awaiting colectomy. Surgery should be performed in patients with severe polyposis, dysplasia, and polyp greater than 5 mm and in symptomatic cases without any time loss after diagnosis [58].

Surgical treatment options include subtotal colectomy with ileorectal anastomosis (IRA), total proctocolectomy with ileal pouch-anal anastomosis (IPAA), or total proctocolectomy and permanent ileostomy. Total proctocolectomy and permanent ileostomy is preferred in cases of rectal tumor that involved the sphincter complex, and in cases which IPAA is not technically feasible. Functional outcomes (quality of life) and the risk of developing rectal cancer, which is the result of leaving the rectum in place, are important in the selection of anorectal anastomosis or IPAA. Patients with a few rectal polyps which can be controlled endoscopically are ideal for IRA. Chemoprevention is recommended for these patients in the postoperative period. Since colon cancer mostly develops in proximal colon in AFAP patients, total abdominal colectomy and IRA is ideal for this group [58–60].

Risks of developing rectal cancer in the 10th and 25th years in patients undergoing IRA are 4–8% and 26–30% respectively [59, 60]. It is known that adenoma or cancer may develop in patients with IPAA, even in those with end ileostomy [61]. Therefore, the remaining rectum or pouch should be examined endoscopically at 6 months or 1 year intervals after whichever method is preferred (IPAA, IRA). IRA should be avoided in cases with family history of desmoid tumor, and IPAA should be preferred. Because in the case of a cancer or polyposis that may develop later in the rectum, revision of IRA to IPAA would be technically very difficult due to

mended to screen thyroid gland by ultrasonography once a year [55, 56].

cancer reaches up to 2.6% and thyroid nodules up to 51.7%. Therefore, it is recommended to screen thyroid gland by ultrasonography once a year [55, 56].

#### *3.2.1 Genetic testing*

*Current Trends in Cancer Management*

liver cancers [45].

syndrome [51].

compressing and encasing adjacent structures [50].

bers. APC mutations are also responsible for both syndromes.

FAP, one inherited mutant allele is not enough to cause carcinoma. Carcinoma develops when the second allele of APC and other necessary gene mutations occur [42]. Extraintestinal manifestations of FAP include gastric, duodenal, and periampullary polyps, and less common manifestations such as epidermoid cysts, desmoid tumors, osteomas, and brain tumors. Gastric and duodenal polyps occur in about half of affected individuals. Most of the gastric polyps are hyperplasia of the fundus glands, rather than adenomatous polyps and their malignancy potential is limited [43]. However duodenal polyps are adenomatous. They are present in approximately 90% of FAP patients and should be considered premalignant [44, 45]. Periampullary tumor risk is higher in FAP patients. In patients who undergo total colectomy, the most important cause of cancer-related death is duodenal adenocancer. Adenomatous polyps and cancer are rarely found in the jejunum and ileum of FAP patients. Other rare extraintestinal malignancies in FAP patients are extrahepatic bile duct, gallbladder, pancreas, adrenal, thyroid, and

The likelihood of a desmoid tumor is increased especially in mutations in the 3' end of the APC gene [46]. Most of the desmoid tumors occur within the first 5 years in patients who have undergone abdominal surgery, presumably as an inflammatory response [47, 48]. In addition to abdominal surgery and APC mutation, pregnancy, female sex, and family history are other risk factors for desmoid tumors [49, 50]. Although desmoid tumors are slow growing, non-metastatic mesenchymal tumors, they may cause complications such as pain, bowel, and ureter obstruction by

An interesting marker for FAP is congenital hypertrophy of the retinal pigment epithelium (CHRPE), which can be determined by ophthalmoscopy in about 75% of patients [51]. Fundus examination with ophthalmoscopy reveals oval, pigmented lesions with regular borders in the retina. Lesions may be bilateral and multiple. CHRPE can be used as a clinical diagnostic tool in the screening of FAP and Gardner

FAP has two subtypes with their own extracolonic manifestations. Gardner's syndrome is a variant of FAP and characterized by desmoid tumors, colonic polyps, osteoma, soft tissue sarcomas, and CHRPE [52]. Also, although very rare, an adenomatous polyposis coli may be associated with malignant tumors of the central nervous system (especially medulloblastoma and/or glioma), known as Turcot syndrome [53]. Turcot is the true variant form of FAP and has a familial character. Colonoscopy and brain scanning tests should also be performed on family mem-

Screening for FAP should be performed in individuals with an APC mutation and in individuals who are first-degree relatives of those with FAP, or who have >10 cumulative colorectal adenomas, or colorectal adenomas in combination with extracolonic features such as duodenal adenomas, desmoid tumors, osteomas, etc.

Screening for CRC should begin during puberty and flexible sigmoidoscopy, or genetic testing for APC mutations should be performed every 6 months or year. When positive genotype is detected by genetic screening, or adenomatous polyps are detected by sigmoidoscopy, full colonoscopy should be performed to evaluate the spread of the disease. Several polyps should also be sampled to confirm histology. First-degree relatives of FAP patients, who do not have a genetic diagnosis, may be removed from aggressive follow-up and included in standard general population screening programs if there is no polyp detected until 40 years of age on screening [54]. Screening of the upper gastrointestinal tract should be performed at the time of diagnosis, or before 25 years of age. In later periods, it should be done while the colon is being evaluated. Thyroid cancer is rarely seen in patients with FAP. Studies have shown that thyroid

**62**

[53, 54].

Identification of the APC mutation and its type in a patient diagnosed with FAP facilitates screening of other family members as well as recognition of possible phenotypic lesions that may result from different APC mutations. Commonly accepted indications for genetic testing include FAP cases, FAP in first degree relatives, APC mutation in first degree relatives, at least 10 cumulative colorectal adenomas, extracolonic involvement of FAP and multiple adenomas with colorectal cancer family history but without a FAP trait in the family [36].

#### *3.2.2 Treatment*

#### *3.2.2.1 Medical treatment and chemoprevention*

In cases with FAP non-steroid anti-inflammatory drugs (NSAIDs) are thought to achieve regression in number and size of polyps [57, 58]. The most commonly used agents for this purpose are sulindac and celecoxib. The role of chemopreventive agents in FAP patients is controversial, because the effects of these agents on cancer prevention are unclear.

#### *3.2.2.2 Surgical treatment*

Prophylactic colectomy should be performed in all cases with FAP. The timing of surgery is planned according to the number of polyps, number of adenomas, presence of dysplasia, size of polyps, symptoms and characteristics of the patient. In cases with mild to moderate polyposis and no other risk factors (low risk of cancer), surgery can be done at mid-puberty. However patients should continue to undergo annual CRC surveillance with colonoscopy while awaiting colectomy. Surgery should be performed in patients with severe polyposis, dysplasia, and polyp greater than 5 mm and in symptomatic cases without any time loss after diagnosis [58].

Surgical treatment options include subtotal colectomy with ileorectal anastomosis (IRA), total proctocolectomy with ileal pouch-anal anastomosis (IPAA), or total proctocolectomy and permanent ileostomy. Total proctocolectomy and permanent ileostomy is preferred in cases of rectal tumor that involved the sphincter complex, and in cases which IPAA is not technically feasible. Functional outcomes (quality of life) and the risk of developing rectal cancer, which is the result of leaving the rectum in place, are important in the selection of anorectal anastomosis or IPAA. Patients with a few rectal polyps which can be controlled endoscopically are ideal for IRA. Chemoprevention is recommended for these patients in the postoperative period. Since colon cancer mostly develops in proximal colon in AFAP patients, total abdominal colectomy and IRA is ideal for this group [58–60].

Risks of developing rectal cancer in the 10th and 25th years in patients undergoing IRA are 4–8% and 26–30% respectively [59, 60]. It is known that adenoma or cancer may develop in patients with IPAA, even in those with end ileostomy [61]. Therefore, the remaining rectum or pouch should be examined endoscopically at 6 months or 1 year intervals after whichever method is preferred (IPAA, IRA). IRA should be avoided in cases with family history of desmoid tumor, and IPAA should be preferred. Because in the case of a cancer or polyposis that may develop later in the rectum, revision of IRA to IPAA would be technically very difficult due to mesenteric desmoids that may develop [54].

#### **3.3 MutYH-associated polyposis (MAP)**

The number of polyps may range from 0 to 1000, but it is known that MAP usually contains less adenomatous polyps than FAP. MAP is an autosomal recessive disease. There is a biallelic mutation of the MutYH (MYH) gene on chromosome 1 [62]. MAP usually occurs in fifth or sixth decade with a polyp number of 10–100 [62, 63]. There are insufficient data on extraintestinal manifestations. However, gastric and duodenal polyps may be found in individuals with MAP. Unlike FAP, there is no association with desmoids, osteomas, and CHRPE in MAP [63].

MAP should be suspected in individuals with 10 or more cumulative adenomas as in other adenomatous polyposis syndromes. Germline MYH testing is recommended to those who have a family history of colorectal cancer or polyposis in recessive pattern, or who have a clinical FAP or AFAP phenotype but a negative APC mutation test result. In patients with biallelic MUTYH mutations, the cumulative lifetime risk of developing colorectal cancer is 75% in men and 72% in women by age 70 [64]. Most of the patients with MAP are diagnosed when they have cancer, but it is recommended to perform a colonoscopy every one to two years to individuals with known biallelic mutations, starting at 25–30 years of age [36].

CRC, adenomatous polyp with high-grade dysplasia that cannot be removed endoscopically, and a great number of polyps that cannot be controlled endoscopically are indications for surgery. It is recommended to remove the newly developed polyps by performing at least annual colonoscopy in patients not eligible for surgery. Surgical options include subtotal colectomy with IRA, total abdominal colectomy, or proctocolectomy with IPAA [63, 64].

#### **4. Clinical findings of colon cancer**

Patients frequently present with changes in bowel habits, rectal bleeding, anemia, and abdominal pain accompanying these findings. Patients may also suffer from weight loss, fatigue, nausea, vomiting, obstruction and perforation [65–69]. Clinical findings vary according to the tumor location. Abdominal pain, which can be seen in all localizations, is the most common clinical manifestation. The most common symptoms in right colon tumors are blunt, permanent lower quadrant pain and anemia of iron deficiency due to occult hemorrhage, fatigue, and anorexia and weight loss. Sometimes, a mass can be palpated in the lower right quadrant [24, 68, 69].

In the left colon, the diameter is smaller (especially sigmoid colon) and the content is solid. In addition, left colon cancers are scirrhous and annular. Therefore, obstructive symptoms are common. Obstruction may lead to perforation and peritonitis. According to Laplace's law, the most likely location of perforation as a result of obstruction of the sigmoid colon is the cecum of which the diameter is largest. A change in bowel habits and a progressive decrease in stool diameter may be the first symptoms. While rectal bleeding may be a finding as occult blood in feces on the right side, it may occur as hematochezia on the left side. In the presence of iron deficiency anemia in an adult male or postmenopausal woman, the diagnosis of colon cancer should be absolutely ruled out [67–70].

Patients may also present with metastatic disease. At the time of presentation, metastatic disease is detected in approximately 20 percent of patients in the United States [70]. Advanced, or often metastatic disease should be suspected in case of the presence of abdominal distention, ascites, early satiety, right upper quadrant pain, periumbilical nodules, or supraclavicular lymphadenopathy.

Colon cancer can spread in 4 different ways; directly through the neighborhood, lymphatic route, hematogen route, and through the peritoneal cavity by

**65**

*Colon Cancer*

**Figure 4.**

*Colon cancer dissemination.*

tumors) [70, 71].

performed, if necessary.

*DOI: http://dx.doi.org/10.5772/intechopen.81597*

gravity (seeding) (**Figure 4**). It should be remembered that tumor may also spread by implantation due to manipulation at the time of surgery. The most common metastasis is in the regional lymph nodes. The most important factor determining lymph node involvement is the T category. While lymph node metastasis is 5–20% in T1–T2 tumors, in T3–T4 tumors lymph node involvement increases to more than 50%. Tumor differentiation, presence of lymphovascular invasion and tumor size are other factors. Hematogenous spread occurs with portal system and the most common site for metastasis is the liver. The other common sites for metastasis after liver is the lung and bones. Seeding is caused by the placement of free tumor cells in the omentum, periton (peritoneal carcinomatosis), rectovesical pouch (Blumer's shelf tumors), and ovary (Krukenberg

**5. Diagnosis and preoperative evaluation in colon cancer**

Colon cancer may be suspected from vague but suspicious symptoms and signs,

The most accurate and preferred diagnostic test for colon cancer is colonoscopy since it can be used for detecting and sampling of lesions along the large bowel, examination of lesions by direct observation, treatment in appropriate patients, and detection of synchronous tumors. Synchronous CRCs occur in 4–5% of patients [71, 72]. In some individuals, a minority of neoplastic lesions are nonpolypoid and flat, and may be more challenging to detect by colonoscopy. However colonoscopy

Flexible rectosigmoidoscopy can be used for diagnostic purposes as well. It is mostly recommended for screening of CRC every 5 years, starting at the age of 50 with annual fecal occult blood test. In recent years there is an increase in right-sided or proximal colon cancers. Because of this and the likelihood of synchronous CRCs, it should be considered that flexible sigmoidoscopy may be an inadequate test for

or sometimes, especially asymptomatic CRC, may only be revealed by routine screening. Anamnesis, physical and rectal examination are valuable in diagnosis. If there is a suspicion of CRC in the patient after anamnesis and physical examination, the first diagnostic test should be colonoscopy or flexible sigmoidoscopy. In addition, barium enema and computed tomography colonography (CTC) may be

is still more sensitive in this situation than barium enema or CTC [72].

diagnosis of a patient suspected of having a CRC [72, 73].

*Current Trends in Cancer Management*

**3.3 MutYH-associated polyposis (MAP)**

The number of polyps may range from 0 to 1000, but it is known that MAP usually contains less adenomatous polyps than FAP. MAP is an autosomal recessive disease. There is a biallelic mutation of the MutYH (MYH) gene on chromosome 1 [62]. MAP usually occurs in fifth or sixth decade with a polyp number of 10–100 [62, 63]. There are insufficient data on extraintestinal manifestations. However, gastric and duodenal polyps may be found in individuals with MAP. Unlike FAP, there is no association with desmoids, osteomas, and CHRPE in MAP [63].

MAP should be suspected in individuals with 10 or more cumulative adenomas as in other adenomatous polyposis syndromes. Germline MYH testing is recommended to those who have a family history of colorectal cancer or polyposis in recessive pattern, or who have a clinical FAP or AFAP phenotype but a negative APC mutation test result. In patients with biallelic MUTYH mutations, the cumulative lifetime risk of developing colorectal cancer is 75% in men and 72% in women by age 70 [64]. Most of the patients with MAP are diagnosed when they have cancer, but it is recommended to perform a colonoscopy every one to two years to individu-

CRC, adenomatous polyp with high-grade dysplasia that cannot be removed endoscopically, and a great number of polyps that cannot be controlled endoscopically are indications for surgery. It is recommended to remove the newly developed polyps by performing at least annual colonoscopy in patients not eligible for surgery. Surgical options include subtotal colectomy with IRA, total abdominal

Patients frequently present with changes in bowel habits, rectal bleeding, anemia,

Patients may also present with metastatic disease. At the time of presentation, metastatic disease is detected in approximately 20 percent of patients in the United States [70]. Advanced, or often metastatic disease should be suspected in case of the presence of abdominal distention, ascites, early satiety, right upper quadrant pain,

Colon cancer can spread in 4 different ways; directly through the neighborhood, lymphatic route, hematogen route, and through the peritoneal cavity by

and abdominal pain accompanying these findings. Patients may also suffer from weight loss, fatigue, nausea, vomiting, obstruction and perforation [65–69]. Clinical findings vary according to the tumor location. Abdominal pain, which can be seen in all localizations, is the most common clinical manifestation. The most common symptoms in right colon tumors are blunt, permanent lower quadrant pain and anemia of iron deficiency due to occult hemorrhage, fatigue, and anorexia and weight loss. Sometimes, a mass can be palpated in the lower right quadrant [24, 68, 69]. In the left colon, the diameter is smaller (especially sigmoid colon) and the content is solid. In addition, left colon cancers are scirrhous and annular. Therefore, obstructive symptoms are common. Obstruction may lead to perforation and peritonitis. According to Laplace's law, the most likely location of perforation as a result of obstruction of the sigmoid colon is the cecum of which the diameter is largest. A change in bowel habits and a progressive decrease in stool diameter may be the first symptoms. While rectal bleeding may be a finding as occult blood in feces on the right side, it may occur as hematochezia on the left side. In the presence of iron deficiency anemia in an adult male or postmenopausal woman, the diagnosis

als with known biallelic mutations, starting at 25–30 years of age [36].

colectomy, or proctocolectomy with IPAA [63, 64].

of colon cancer should be absolutely ruled out [67–70].

periumbilical nodules, or supraclavicular lymphadenopathy.

**4. Clinical findings of colon cancer**

**64**

**Figure 4.** *Colon cancer dissemination.*

gravity (seeding) (**Figure 4**). It should be remembered that tumor may also spread by implantation due to manipulation at the time of surgery. The most common metastasis is in the regional lymph nodes. The most important factor determining lymph node involvement is the T category. While lymph node metastasis is 5–20% in T1–T2 tumors, in T3–T4 tumors lymph node involvement increases to more than 50%. Tumor differentiation, presence of lymphovascular invasion and tumor size are other factors. Hematogenous spread occurs with portal system and the most common site for metastasis is the liver. The other common sites for metastasis after liver is the lung and bones. Seeding is caused by the placement of free tumor cells in the omentum, periton (peritoneal carcinomatosis), rectovesical pouch (Blumer's shelf tumors), and ovary (Krukenberg tumors) [70, 71].

### **5. Diagnosis and preoperative evaluation in colon cancer**

Colon cancer may be suspected from vague but suspicious symptoms and signs, or sometimes, especially asymptomatic CRC, may only be revealed by routine screening. Anamnesis, physical and rectal examination are valuable in diagnosis. If there is a suspicion of CRC in the patient after anamnesis and physical examination, the first diagnostic test should be colonoscopy or flexible sigmoidoscopy. In addition, barium enema and computed tomography colonography (CTC) may be performed, if necessary.

The most accurate and preferred diagnostic test for colon cancer is colonoscopy since it can be used for detecting and sampling of lesions along the large bowel, examination of lesions by direct observation, treatment in appropriate patients, and detection of synchronous tumors. Synchronous CRCs occur in 4–5% of patients [71, 72]. In some individuals, a minority of neoplastic lesions are nonpolypoid and flat, and may be more challenging to detect by colonoscopy. However colonoscopy is still more sensitive in this situation than barium enema or CTC [72].

Flexible rectosigmoidoscopy can be used for diagnostic purposes as well. It is mostly recommended for screening of CRC every 5 years, starting at the age of 50 with annual fecal occult blood test. In recent years there is an increase in right-sided or proximal colon cancers. Because of this and the likelihood of synchronous CRCs, it should be considered that flexible sigmoidoscopy may be an inadequate test for diagnosis of a patient suspected of having a CRC [72, 73].

Computerized tomography is the most frequently used test for staging purposes. Positron emission tomography has no place in routine staging and screening. However, in suspicious cases, tumor and fibrous tissue are well separated. Abdominal ultrasonography has no place in the diagnosis of colon cancer [71].

Routine laboratory tests including complete blood count, liver function tests, etc. have no role in diagnosis. However In the presence of iron deficiency anemia in an adult male or postmenopausal woman, colon cancer should be ruled out. Although liver function tests has no role in diagnosis of liver metastasis, the increase in liver enzymes in patients with colon cancer should be taken into consideration to scan for metastasis [71, 72].

It is known that some tumor markers, especially CEA (carcinoembryonic antigen), are associated with CRC. Nevertheless, tumor markers such as CEA and CA 19-9 appears to have a low diagnostic yield to diagnose primary CRC, since these markers have low sensitivity for early-stage disease and may also increase in some benign diseases. However, both markers have prognostic significance. High CEA suggests the presence of metastasis. In addition, after appropriate treatment increase in CEA level in follow-up should be assessed in favor of recurrence or metastasis [73].

#### **6. Staging**

The local features (size, invasion, lymph node involvement) of the tumor are important in determining the resection margin. Therefore, preoperative clinical staging should be done properly. Physical examination and radiological tests are used for accurate clinical staging. It should not be forgotten that the accuracy of radiological detection of the stage is 85–90%, even in the best hands. Definitive staging can only be performed by pathological examination [74].

Pathologic staging in colorectal cancers is based on tumor depth, lymph node involvement, and the metastatic status. Dukes and Astler-Coller classifications are no longer used, instead the TNM staging system is preferred [74]. The most recent (8th edition) revision of the TNM staging classification contains few changes compared with the earlier edition (7th edition). T categories have been revised. Tis in the AJCC 8th edition refers only to intramucosal carcinoma, a lesion with involvement of lamina propria with no extension through muscularis mucosae. T4 is defined as tumor exceeds the visceral peritoneum either by continuous invasion or perforation of the tumor. N categories have not changed. Lastly, the M category has been expanded, with the addition of M1c for peritoneal metastases. Therefore, a new stage, IVc, have been added in stage grouping.

Stage of the disease is the most important prognostic parameter that determines the type of surgery and postoperative treatment options in colorectal cancers. Because of the different lymphatic drainage on the intestinal wall, colorectal cancer gains potential to make metastasis only when there is submucosal invasion. For this reason, colorectal carcinoma is diagnosed only in the presence of submucosal invasion [74].

#### **Dukes staging:**

**Dukes A:** Tumor is limited in the bowel wall.

**Dukes B:** Invasion through the bowel wall but no lymph node involvement.

**67**

**Figure 5.**

*specimen.*

*Colon Cancer*

**7. Treatment**

*DOI: http://dx.doi.org/10.5772/intechopen.81597*

**B3:** Lesion involves adjacent organs. **C1:** B1 + lymph node involvement. **C2:** B2 + lymph node involvement. **C3:** B3 + lymph node involvement.

Colon cancers mostly develop from polyps. Although curative treatment of localized colon cancer is surgery, endoscopic polypectomy is enough when carcinoma in situ or severe dysplasia presents on the polyp surface. However, surgery should be considered for the treatment of colon cancer especially in patients a polyp that cannot be removed endoscopically, and if there is continuity in resection

In the last decade, there have been major changes in colorectal cancer management. Total mesorectal excision and neoadjuvant chemoradiotherapy in rectum cancers resulted in significant reductions in morbidity, mortality, and recurrence rates. Recently, complete mesocolic excision (CME) and central vascular ligation (CVL) (open or laparoscopic) has been described in colon cancer treatment to achieve similar oncological results. In 2007, Hohenberger published the first article on CME with CVL for colon cancer and which was later published in English [75, 76]. The aim of CME with CVL method is to create an intact protective mesocolic fascia and avoid tumor spread within peritoneal cavity by dissection of the visceral fascia from the parietal (retroperitoneal) plane (**Figure 5**). The origin of colonic vessels is well exposed and ligated centrally at their origin using this technique. The specimens are characterized by a greater distance from the tumor to the high vascular tie, higher distance from the closest bowel wall to the high vascular tie, longer length of the colon and larger area of mesentery. Thus, maximum lymphatic tissue harvest is achieved [76, 77]. Increase in the patients who have a high number of lymph nodes, decrease in perioperative morbidity, reduction in local recurrence, and advancement in colon cancer-specific survival rate have been shown in recent studies regarding CME [76, 78–80]. This technique is a matter of controversy in colon surgery. Because longer operating times, autonomic nerve injury, and major vascular damage are disadvantages of routine implementation of CME. Although the technique has improved oncologic data, routine implementation of CME may decrease health-

*Complete mesocolic excision and central vascular ligation for the treatment of extended right hemicolectomy* 

**D:** Distant metastasis.

margin after polypectomy [75].

related quality of life (QoL) [76, 80].

**Dukes C:** Lymph node involvement.

**Dukes D:** Distant metastasis.

#### **Modified Astler-Coller staging:**

**A:** Tumor is limited to mucosa.

**B1:** Muscularis propria is invaded but not exceeded.

**B2:** Invades through muscularis propria (subserosal dissemination).

**B3:** Lesion involves adjacent organs. **C1:** B1 + lymph node involvement. **C2:** B2 + lymph node involvement. **C3:** B3 + lymph node involvement. **D:** Distant metastasis.

#### **7. Treatment**

*Current Trends in Cancer Management*

eration to scan for metastasis [71, 72].

**6. Staging**

Computerized tomography is the most frequently used test for staging purposes. Positron emission tomography has no place in routine staging and screening. However, in suspicious cases, tumor and fibrous tissue are well separated. Abdominal ultrasonography has no place in the diagnosis of colon cancer [71]. Routine laboratory tests including complete blood count, liver function tests, etc. have no role in diagnosis. However In the presence of iron deficiency anemia in an adult male or postmenopausal woman, colon cancer should be ruled out. Although liver function tests has no role in diagnosis of liver metastasis, the increase in liver enzymes in patients with colon cancer should be taken into consid-

It is known that some tumor markers, especially CEA (carcinoembryonic antigen), are associated with CRC. Nevertheless, tumor markers such as CEA and CA 19-9 appears to have a low diagnostic yield to diagnose primary CRC, since these markers have low sensitivity for early-stage disease and may also increase in some benign diseases. However, both markers have prognostic significance. High CEA suggests the presence of metastasis. In addition, after appropriate treatment increase in CEA level in follow-up should be assessed in favor of recurrence or metastasis [73].

The local features (size, invasion, lymph node involvement) of the tumor are important in determining the resection margin. Therefore, preoperative clinical staging should be done properly. Physical examination and radiological tests are used for accurate clinical staging. It should not be forgotten that the accuracy of radiological detection of the stage is 85–90%, even in the best hands. Definitive

Pathologic staging in colorectal cancers is based on tumor depth, lymph node involvement, and the metastatic status. Dukes and Astler-Coller classifications are no longer used, instead the TNM staging system is preferred [74]. The most recent (8th edition) revision of the TNM staging classification contains few changes compared with the earlier edition (7th edition). T categories have been revised. Tis in the AJCC 8th edition refers only to intramucosal carcinoma, a lesion with involvement of lamina propria with no extension through muscularis mucosae. T4 is defined as tumor exceeds the visceral peritoneum either by continuous invasion or perforation of the tumor. N categories have not changed. Lastly, the M category has been expanded, with the addition of M1c for peritoneal metastases. Therefore, a

Stage of the disease is the most important prognostic parameter that determines

the type of surgery and postoperative treatment options in colorectal cancers. Because of the different lymphatic drainage on the intestinal wall, colorectal cancer gains potential to make metastasis only when there is submucosal invasion. For this reason, colorectal carcinoma is diagnosed only in the presence of submucosal

**Dukes B:** Invasion through the bowel wall but no lymph node involvement.

**B2:** Invades through muscularis propria (subserosal dissemination).

staging can only be performed by pathological examination [74].

new stage, IVc, have been added in stage grouping.

**Dukes A:** Tumor is limited in the bowel wall.

**B1:** Muscularis propria is invaded but not exceeded.

**Dukes C:** Lymph node involvement. **Dukes D:** Distant metastasis. **Modified Astler-Coller staging: A:** Tumor is limited to mucosa.

**66**

invasion [74].

**Dukes staging:**

Colon cancers mostly develop from polyps. Although curative treatment of localized colon cancer is surgery, endoscopic polypectomy is enough when carcinoma in situ or severe dysplasia presents on the polyp surface. However, surgery should be considered for the treatment of colon cancer especially in patients a polyp that cannot be removed endoscopically, and if there is continuity in resection margin after polypectomy [75].

In the last decade, there have been major changes in colorectal cancer management. Total mesorectal excision and neoadjuvant chemoradiotherapy in rectum cancers resulted in significant reductions in morbidity, mortality, and recurrence rates. Recently, complete mesocolic excision (CME) and central vascular ligation (CVL) (open or laparoscopic) has been described in colon cancer treatment to achieve similar oncological results. In 2007, Hohenberger published the first article on CME with CVL for colon cancer and which was later published in English [75, 76]. The aim of CME with CVL method is to create an intact protective mesocolic fascia and avoid tumor spread within peritoneal cavity by dissection of the visceral fascia from the parietal (retroperitoneal) plane (**Figure 5**). The origin of colonic vessels is well exposed and ligated centrally at their origin using this technique. The specimens are characterized by a greater distance from the tumor to the high vascular tie, higher distance from the closest bowel wall to the high vascular tie, longer length of the colon and larger area of mesentery. Thus, maximum lymphatic tissue harvest is achieved [76, 77]. Increase in the patients who have a high number of lymph nodes, decrease in perioperative morbidity, reduction in local recurrence, and advancement in colon cancer-specific survival rate have been shown in recent studies regarding CME [76, 78–80]. This technique is a matter of controversy in colon surgery. Because longer operating times, autonomic nerve injury, and major vascular damage are disadvantages of routine implementation of CME. Although the technique has improved oncologic data, routine implementation of CME may decrease healthrelated quality of life (QoL) [76, 80].

#### **Figure 5.**

*Complete mesocolic excision and central vascular ligation for the treatment of extended right hemicolectomy specimen.*

Surgical resection of the tumor is the main curative treatment option. The colon segment where the tumor is located, the mesentery that contains the lymphatic drainage, and, if there is invasion, adjacent organs should be removed in one piece without deteriorating tumor integrity. If the tumor cannot be removed surgically, palliative surgical procedures such as limited resections, proximal diversion ostomies (colostomy, ileostomy), or bypass surgeries may be applied to relieve symptoms or prevent possible complications [79]. Right hemicolectomy (extended or not), transverse colectomy, left hemicolectomy (extended or not) sigmoid colectomy, and subtotal or total colectomy are preferred for surgical treatments of colon tumors according to involved bowel segment. Surgical intervention may be performed conventional (open) or laparoscopic, provided that it conforms to oncologic principles [80].

#### **7.1 Cecum and ascending colon tumors**

Right hemicolectomy is performed as a standard surgical treatment option in the right-sided colon tumors. In this operation, right branch of the middle colic, ileocolic, and right colic vessels are ligated as high as possible. The ascending colon, the hepatic flexure, the first third of the transverse colon, and distal part of the terminal ileum is resected (**Figure 6**). Then, ileocolonic anastomosis is performed between ileum and transverse segment of the colon.

#### **7.2 Hepatic flexure tumors**

To remove the entire lymphatic network, CVL of the middle colic, right colic, and ileocolic vessels is performed. This operation is called extended right hemicolectomy (**Figure 7**). When compared to the left hemicolectomy, the amount of transverse colon that is resected increases and only distal 1/3 of the transverse colon is left. An anastomosis should be avoided in areas of unreliable blood supply such as splenic flexure. In this case resection margins should be expanded and splenic

**69**

*Colon Cancer*

*DOI: http://dx.doi.org/10.5772/intechopen.81597*

flexure should be removed as well. Finally, anastomosis is created between the

The choice of surgery type in transverse colon tumors may be a matter of debate. The arterial supply of the transverse colon is provided by right colic and left colic and middle colic arteries. Ischemia usually does not occur in the anastomosis at the hepatic flexure due to branches from ileocolic and right colic arteries, even if middle colic artery is centrally ligated. However when middle colic artery is ligated, arterial supply of splenic flexure is only provided by left colic artery and there is an ischemia risk in the anastomosis at the splenic flexure. Therefore, transverse colectomy could be performed by CVL of the middle colic and left colic vessels for mid-transverse colon tumors. In this procedure, distal ascending, hepatic flexure, transverse, splenic flexure, and proximal descending colon are resected (**Figure 8**). Moreover, it is recommended surgical resection of the ascending colon and cecum to perform ileosigmoidal anastomosis because it is technically difficult to create an anastomosis between ascending and sigmoid colon. Consequently, it is necessary to consider both the arterial circulation and the lymphatic drainage in the selection of

Extended left hemicolectomy is recommended for splenic flexure tumors, as lymphatics may drain to the lymph nodes along the inferior mesenteric artery (IMA) and middle colic artery. The central ligation of the middle colic artery and IMA requires to remove all entire bowel from the proximal transverse colon to the proximal rectum. At the end of the surgical procedure, an anastomosis is performed between proximal transverse colon and proximal segment of the rectum (**Figure 9**).

ileum and the proximal end of the remaining colon.

the operation type in transverse colon tumors.

**7.4 Splenic flexure tumors**

**7.3 Transverse colon tumors**

*Resection margins in hepatic flexure tumors.*

**Figure 7.**

**Figure 6.** *Resection margins in cecum and ascending colon tumors.*

*Current Trends in Cancer Management*

**7.1 Cecum and ascending colon tumors**

**7.2 Hepatic flexure tumors**

between ileum and transverse segment of the colon.

Surgical resection of the tumor is the main curative treatment option. The colon segment where the tumor is located, the mesentery that contains the lymphatic drainage, and, if there is invasion, adjacent organs should be removed in one piece without deteriorating tumor integrity. If the tumor cannot be removed surgically, palliative surgical procedures such as limited resections, proximal diversion ostomies (colostomy, ileostomy), or bypass surgeries may be applied to relieve symptoms or prevent possible complications [79]. Right hemicolectomy (extended or not), transverse colectomy, left hemicolectomy (extended or not) sigmoid colectomy, and subtotal or total colectomy are preferred for surgical treatments of colon tumors according to involved bowel segment. Surgical intervention may be performed conventional (open) or laparoscopic, provided that it conforms to oncologic principles [80].

Right hemicolectomy is performed as a standard surgical treatment option in the right-sided colon tumors. In this operation, right branch of the middle colic, ileocolic, and right colic vessels are ligated as high as possible. The ascending colon, the hepatic flexure, the first third of the transverse colon, and distal part of the terminal ileum is resected (**Figure 6**). Then, ileocolonic anastomosis is performed

To remove the entire lymphatic network, CVL of the middle colic, right colic, and ileocolic vessels is performed. This operation is called extended right hemicolectomy (**Figure 7**). When compared to the left hemicolectomy, the amount of transverse colon that is resected increases and only distal 1/3 of the transverse colon is left. An anastomosis should be avoided in areas of unreliable blood supply such as splenic flexure. In this case resection margins should be expanded and splenic

**68**

**Figure 6.**

*Resection margins in cecum and ascending colon tumors.*

**Figure 7.** *Resection margins in hepatic flexure tumors.*

flexure should be removed as well. Finally, anastomosis is created between the ileum and the proximal end of the remaining colon.

#### **7.3 Transverse colon tumors**

The choice of surgery type in transverse colon tumors may be a matter of debate. The arterial supply of the transverse colon is provided by right colic and left colic and middle colic arteries. Ischemia usually does not occur in the anastomosis at the hepatic flexure due to branches from ileocolic and right colic arteries, even if middle colic artery is centrally ligated. However when middle colic artery is ligated, arterial supply of splenic flexure is only provided by left colic artery and there is an ischemia risk in the anastomosis at the splenic flexure. Therefore, transverse colectomy could be performed by CVL of the middle colic and left colic vessels for mid-transverse colon tumors. In this procedure, distal ascending, hepatic flexure, transverse, splenic flexure, and proximal descending colon are resected (**Figure 8**). Moreover, it is recommended surgical resection of the ascending colon and cecum to perform ileosigmoidal anastomosis because it is technically difficult to create an anastomosis between ascending and sigmoid colon. Consequently, it is necessary to consider both the arterial circulation and the lymphatic drainage in the selection of the operation type in transverse colon tumors.

#### **7.4 Splenic flexure tumors**

Extended left hemicolectomy is recommended for splenic flexure tumors, as lymphatics may drain to the lymph nodes along the inferior mesenteric artery (IMA) and middle colic artery. The central ligation of the middle colic artery and IMA requires to remove all entire bowel from the proximal transverse colon to the proximal rectum. At the end of the surgical procedure, an anastomosis is performed between proximal transverse colon and proximal segment of the rectum (**Figure 9**).

**Figure 8***. Resection margins in transverse colon tumors.*

**Figure 9.** *Resection margins in splenic flexure tumors.*

#### **7.5 Descending colon tumors**

Left hemicolectomy is recommended in patients with ascending colon tumor. IMA is centrally ligated without preserving the left colic artery. Splenic flexure, descending and sigmoid colons are removed. Then, anastomosis is established between transverse colon and proximal rectum (**Figure 10**).

**71**

**Figure 11.**

**7.6 Sigmoid colon tumors**

*Resection margins in sigmoid colon tumors.*

and colorectal anastomosis is created (**Figure 11**).

The appropriate operation for these tumors is sigmoid colon resection. IMA is centrally ligated while left colic artery is preserved. Sigmoid colon is then removed

*Colon Cancer*

**Figure 10.**

*Resection margins in descending colon tumors.*

*DOI: http://dx.doi.org/10.5772/intechopen.81597*

*Current Trends in Cancer Management*

*Resection margins in transverse colon tumors.*

**70**

**Figure 9.**

**Figure 8***.*

**7.5 Descending colon tumors**

*Resection margins in splenic flexure tumors.*

Left hemicolectomy is recommended in patients with ascending colon tumor. IMA is centrally ligated without preserving the left colic artery. Splenic flexure, descending and sigmoid colons are removed. Then, anastomosis is established

between transverse colon and proximal rectum (**Figure 10**).

**Figure 10.** *Resection margins in descending colon tumors.*

#### **Figure 11.**

*Resection margins in sigmoid colon tumors.*

#### **7.6 Sigmoid colon tumors**

The appropriate operation for these tumors is sigmoid colon resection. IMA is centrally ligated while left colic artery is preserved. Sigmoid colon is then removed and colorectal anastomosis is created (**Figure 11**).

#### **8. Prognosis**

Pathologic stage at presentation is the strongest prognostic factor. In patients with stage 1 colon tumor, the 5-year survival rate is approximately 90% while it drops to 15% in stage 4 patients [2]. Despite a curative surgery and modern adjuvant treatments, recurrence develops in approximately 40% of stage 2 and 3 patients [81]. Almost all recurrences develop within the first 5 years, and most of them are seen within the first 3 years [82].

Besides pathologic staging, the most important prognostic factors for CRC are histologic grade of differentiation, extramural tumor deposits, lymphovascular and perineural invasion, the preoperative carcinoembryonic antigen (CEA) level, MSI, and *RAS* and *BRAF* mutations. The local extent of disease independently influences survival [83, 84]. However tumor size has no significant impact on prognosis [84, 85]. One of the adverse prognostic factors is residual tumor after resection [86, 87]. There are three types of R designation for residual tumors in non-metastatic patients: R0 resection, complete resection of the tumor with histologically negative margins, R1 resection, incomplete tumor resection with positive microscopic margin involvement, R2 resection, and incomplete resection with macroscopic margin involvement [74].

Regional lymph node metastasis is the other important determinant of prognosis after distant metastasis. Lymph node involvement is alone an indication for postoperative adjuvant therapy to reduce the metastasis risk. Although the number of positive lymph nodes involved is a crucial predictor of outcome [74, 88], relationship between total number of the lymph nodes and the prognosis is not well understood. However, increased number of total lymph nodes in the surgical specimen may be an indicator for the quality of the surgical procedure [88].

Tumor deposits are separate nodules of tumor within the pericolic fat or mesentery. In the TNM staging they are staged as N1c which means there are no regional lymph nodes involved but the subserosa, mesentery, or nonperitonealized pericolic tissues contains tumor deposits(74). These deposits are strong adverse prognostic determinants, and there is a relation between extramural extranodal tumor deposits and extramural venous invasion [89, 90]. Lymphovascular involvement which is tumor invasion into veins, especially extramural veins, or lymphatics is thought to be an adverse prognostic factor [91–93]. Perineural invasion is also associated with an elevated risk of recurrence and poor prognosis [94, 95].

Several studies have provided evidence that preoperative high CEA levels have adverse impact on prognosis for colon cancer. It has been determined that higher CEA levels increase overall mortality and even prognosis is similar or worse in patients with higher CEA levels but lower stages when compared to patients with higher stages but lower CEA levels according to AJCC TNM staging [96, 97].

Metastatic disease is another significant clinical problem in patients with CRCs. The liver, lungs, lymph nodes, and peritoneum are the most frequently involved organs. Major developments in chemotherapy have increased survival rates in a serious manner, but 5-year survival rates are below 20% without resection or ablation of metastasis. Five-year survival rates are 36–58% in patients undergoing partial hepatectomy for hepatic metastases [98–102]. Lung involvement is less common than liver metastasis, but in carefully selected patients metastasectomy is a favorable option for treatment [102].

**73**

**Author details**

Mehmet Ali Koc1

Ankara, Turkey

Ankara, Turkey

*Colon Cancer*

*DOI: http://dx.doi.org/10.5772/intechopen.81597*

provided the original work is properly cited.

, Suleyman Utku Celik<sup>2</sup>

\*Address all correspondence to: cihangirakyol@gmail.com

1 Department of General Surgery, Ankara University School of Medicine,

2 Clinic of General Surgery, Gulhane Training and Research Hospital,

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

and Cihangir Akyol1

\*

#### **Conflict of interest**

The authors declare that there are no conflicts of interest.

*Current Trends in Cancer Management*

seen within the first 3 years [82].

Pathologic stage at presentation is the strongest prognostic factor. In patients with stage 1 colon tumor, the 5-year survival rate is approximately 90% while it drops to 15% in stage 4 patients [2]. Despite a curative surgery and modern adjuvant treatments, recurrence develops in approximately 40% of stage 2 and 3 patients [81]. Almost all recurrences develop within the first 5 years, and most of them are

Besides pathologic staging, the most important prognostic factors for CRC are histologic grade of differentiation, extramural tumor deposits, lymphovascular and perineural invasion, the preoperative carcinoembryonic antigen (CEA) level, MSI, and *RAS* and *BRAF* mutations. The local extent of disease independently influences survival [83, 84]. However tumor size has no significant impact on prognosis [84, 85]. One of the adverse prognostic factors is residual tumor after resection [86, 87]. There are three types of R designation for residual tumors in non-metastatic patients: R0 resection, complete resection of the tumor with histologically negative margins, R1 resection, incomplete tumor resection with positive microscopic margin involvement, R2 resection, and incomplete resection with macroscopic margin

Regional lymph node metastasis is the other important determinant of prognosis after distant metastasis. Lymph node involvement is alone an indication for postoperative adjuvant therapy to reduce the metastasis risk. Although the number of positive lymph nodes involved is a crucial predictor of outcome [74, 88], relationship between total number of the lymph nodes and the prognosis is not well understood. However, increased number of total lymph nodes in the surgical specimen

Tumor deposits are separate nodules of tumor within the pericolic fat or mesentery. In the TNM staging they are staged as N1c which means there are no regional lymph nodes involved but the subserosa, mesentery, or nonperitonealized pericolic tissues contains tumor deposits(74). These deposits are strong adverse prognostic determinants, and there is a relation between extramural extranodal tumor deposits and extramural venous invasion [89, 90]. Lymphovascular involvement which is tumor invasion into veins, especially extramural veins, or lymphatics is thought to be an adverse prognostic factor [91–93]. Perineural invasion is also associated with

Several studies have provided evidence that preoperative high CEA levels have adverse impact on prognosis for colon cancer. It has been determined that higher CEA levels increase overall mortality and even prognosis is similar or worse in patients with higher CEA levels but lower stages when compared to patients with higher stages but lower CEA levels according to AJCC TNM staging [96, 97].

Metastatic disease is another significant clinical problem in patients with CRCs. The liver, lungs, lymph nodes, and peritoneum are the most frequently involved organs. Major developments in chemotherapy have increased survival rates in a serious manner, but 5-year survival rates are below 20% without resection or ablation of metastasis. Five-year survival rates are 36–58% in patients undergoing partial hepatectomy for hepatic metastases [98–102]. Lung involvement is less common than liver metastasis, but in carefully selected patients metastasectomy is a

may be an indicator for the quality of the surgical procedure [88].

an elevated risk of recurrence and poor prognosis [94, 95].

The authors declare that there are no conflicts of interest.

favorable option for treatment [102].

**Conflict of interest**

**8. Prognosis**

involvement [74].

**72**

### **Author details**

Mehmet Ali Koc1 , Suleyman Utku Celik<sup>2</sup> and Cihangir Akyol1 \*

1 Department of General Surgery, Ankara University School of Medicine, Ankara, Turkey

2 Clinic of General Surgery, Gulhane Training and Research Hospital, Ankara, Turkey

\*Address all correspondence to: cihangirakyol@gmail.com

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

### **References**

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[2] Howlader N, Noone AM, Krapcho M, et al. SEER Cancer Statistics Review (CSR) 1975-2014. https://seer.cancer. gov/archive/csr/1975\_2014/

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[10] Shergill AK, Farraye FA. Toward a consensus on endoscopic surveillance of patients with colonic inflammatory bowel disease. Gastrointestinal Endoscopy Clinics of North America.

[11] Zauber AG, Winawer SJ, O'Brien MJ, et al. Colonoscopic polypectomy and long-term prevention of colorectalcancer deaths. The New England Journal of Medicine. 2012;**366**(8):687-696

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[13] Grady WM, Carethers JM. Genomic and epigenetic instability in colorectal cancer pathogenesis. Gastroenterology.

[14] Vogelstein B, Fearon ER, Hamilton SR, et al. Genetic alterations during colorectal-tumor development. The New England Journal of Medicine.

[15] Fodde R. The APC gene in colorectal cancer. European Journal of Cancer.

[16] Powell SM, Zilz N, Beazer-Barclay Y, et al. APC mutations occur early during colorectal tumorigenesis. Nature.

[17] Cancer Genome Atlas Network.

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Comprehensive molecular characterization of human colon and rectal cancer. Nature.

2012;**487**(7407):330-337

2014;**24**(3):469-481

2015;**60**(3):762-772

2008;**135**(4):1079-1099

1988;**319**(9):525-532

2002;**38**(7):867-871

**359**(6392):235-237

[2] Howlader N, Noone AM, Krapcho M, et al. SEER Cancer Statistics Review (CSR) 1975-2014. https://seer.cancer.

[3] Gordon PH. Malignant neoplasms of the colon. In: Gordon PH, Nivatvongs S, editors. Principles and Practice of Surgery for the Colon; Rectum; and Anus. 3rd ed. New York: Informa Healthcare USA Inc.; 2007. pp. 489-643

[4] Ryan-Harshman M, Aldoori W. Diet and colorectal cancer: Review of the evidence. Canadian Family Physician.

[5] Akin H, Tözün N. Diet, microbiota, and colorectal cancer. Journal of Clinical Gastroenterology. 2014;**48**(Suppl 1):

[6] Itzkowitz SH, Rochester J. Colonic polyps and polyposis syndromes. In: Feldman M, Fiedman LS, Brandt LJ, editors. Gastrointestinal and Liver Disease. 8th ed. Philadelphia: Saunders

Elsevier; 2006. pp. 2713-2757

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[8] Kim ER, Chang DK. Colorectal cancer in inflammatory bowel disease: The risk, pathogenesis, prevention and diagnosis. World Journal of Gastroenterology. 2014;**20**(29):9872-9881

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2007;**53**(11):1913-1920

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**81**

Section 3

Urogenital Cancers
