**Meet the editor**

Dr Tejinder Kataria is the Chairperson, Division of Radiation Oncology at Medanta-the Medicity, Gurgaon, India. After her graduation she pursued MD in Radiotherapy followed by a fellowship for intraluminal brachytherapy. She then went on to hone her skills in Brachytherapy for flexible Iridium 192 interstitial implants and permanent Iodine125 seed implants for

prostate, while on a Commonwealth scholarship. She returned to India and started the department of Radiotherapy in one of the premier cancer centers in Delhi. She trained at DKFZ and implemented Intensity Modulated Radiotherapy in the country in 2001-02.Subsequently the quest for learning took her to Mannheim Klinikum, in 2007 for image guided radiotherapy and in 2010 for volumetric arc therapy . She started frameless Linac based stereotaxy in 2010 after training at Wurzburg .She was instrumental in bringing the first the Cyberknife- VSI system to her country. Her efforts have been towards implementing conformal radiation therapy techniques and advancements for the benefit of her patients and training the younger generation in the art and science of cancer care.

Contents

**Preface IX** 

Chapter 2 **A Framework for Modeling** 

**Section 2 Dosimetry and Medical Physics 33** 

Chapter 4 **3D Dosimetric Tools in Radiotherapy for Photon Beams 53** 

Chapter 5 **A Respiratory Motion Prediction Based** 

Sugita and Makoto Yoshizawa

Chapter 6 **Neutron Dose Equivalent in Tissue Due to Linacs of Clinical Use 91**  S. Agustín Martínez Ovalle

**Section 1 Molecular Biology of Radiation Therapy 1** 

Chapter 1 **Histone Acetyltransferases (HATs) Involved in Non-Homologous End Joining as a Target for Radiosensitization 3** 

> Takahiro Oike, Hideaki Ogiwara, Takashi Nakano and Takashi Kohno

**the Cellular Defending Mechanisms** 

Chapter 3 **Applications to Radiotherapy Using Three Different** 

**Code and Treatment Planning System 35** 

**Against Genome Stress Under Radiotherapy 13**  Jin-Peng Qi, Yong-Sheng Ding and Xian-Hui Zeng

**Dosimetric Tools: MAGIC-***f* **Gel, PENELOPE Simulation** 

Thatiane Alves Pianoschi and Mirko Salomón Alva-Sánchez

Mirko Salomón Alva-Sánchez and Thatiane Alves Pianoschi

Kei Ichiji, Noriyasu Homma, Masao Sakai, Makoto Abe, Norihiro

**on Time-Variant Seasonal Autoregressive Model for Real-Time Image-Guided Radiotherapy 73** 

## Contents

#### **Preface XI**


S. Agustín Martínez Ovalle

#### X Contents

#### **Section 3 Clinical Radiotherapy 113**

	- **Section 4 Quality of Life Issues 195**

## Preface

Radiation therapy was applied for treatment of cancers empirically after the discovery of radium . The acute and long term side effects were recorded clinically as the patients started outliving the tumours. The combination of radiation before or after surgery and subsequent addition of chemotherapy has revolutionized the way we treat cancer today. Every tumour is approached through a multimodality discussion such that the toxicity of either treatment can be reduced with maximum curative potential. **Frontiers in Radiation Oncology** has been brought forth to understand the basics of radiation sensitization, cellular and genomic stress responses to radiation,inhibiting repair of subletahl damage along with an understanding of the dosimetric aspects of radiation physics. The chapters on clinical aspects have been designed to bring out the changing concepts of cure in metastatic disease with the advent of stereotactic body radiotherapy. The effects of radiation, concurrent chemotherapy and surgery are also the subjects of exploration in the clinical context. Any textbook on providing care to cancer patients cannot be complete without addressing the quality of life in cancer treatment and the nutritional needs of the high catabolic state in the cancer patients. The last two chapters address these needs.

The book is meant for physicians, physicists, dosimetrists and counselors who are trying to provide a holistic care in cancer while trying to understand the complex basics of radiation interaction within the human body.

> **Tejinder Kataria** Radiation Oncology, Medanta-The Medicity, India

**Section 1** 

**Molecular Biology of Radiation Therapy** 

**Molecular Biology of Radiation Therapy** 

**Chapter 1** 

© 2013 Kohno et al., licensee InTech. This is an open access chapter 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.

© 2013 Kohno et al., licensee InTech. This is a paper 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.

**Histone Acetyltransferases (HATs) Involved** 

Takahiro Oike, Hideaki Ogiwara, Takashi Nakano and Takashi Kohno

Radiation therapy is one of the most important treatment modalities for cancer therapy alongside surgery and chemotherapy. However, a major problem associated with radiation therapy in the clinical setting is that, in many cases, local control of the tumor cannot be achieved using this modality alone. This has driven researchers into radiosensitizers, i.e., compounds that enhance the intrinsic sensitivity of cancer cells to ionizing radiation (IR). Several compounds, including halogenated pyrimidines and nitroimidazole derivatives, show radiosensitizing effects in cancer cells [1]; however, clinical application of these compounds is limited because they are highly toxic to normal cell and tissues. Therefore,

The principal target for IR-induced killing of cancer cells is DNA [2]. Of the different types of DNA damage generated by IR, DNA double-strand breaks (DSBs) are thought to be the most cytotoxic. DSBs induced by IR are preferentially repaired by non-homologous end joining (NHEJ) [3, 4], which joins the two broken DNA ends without the need for sequence homology. To enable NHEJ, the chromatin needs to be remodeled into an 'open' state so that the DNA repair proteins can access the DSB sites [5]. We previously reported that acetylation of histone proteins at DSB sites by the histone acetyltransferases (HATs), TIP60, CBP and p300, facilitates NHEJ through chromatin remodeling [6]. This suggests that the inhibition of HAT activity will radiosensitize cancer cells by suppressing NHEJ. In line with this, we and others demonstrated that several natural compounds with HAT-inhibitory activity are able to radiosensitize cancer cells [6-12]. Since some of these compounds are safe when administered to humans [13-15], they could potentially be used as radiosensitizers in a clinical setting. Here, we discuss the role of HATs in NHEJ, the radiosensitizing effects of compounds with HAT-inhibitory activity, and

radiosensitizers with low toxicity to normal tissues are urgently needed.

the prospects for the clinical application of these compounds.

**in Non-Homologous End Joining** 

**as a Target for Radiosensitization** 

Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/56555

**1. Introduction** 

## **Histone Acetyltransferases (HATs) Involved in Non-Homologous End Joining as a Target for Radiosensitization**

Takahiro Oike, Hideaki Ogiwara, Takashi Nakano and Takashi Kohno

Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/56555

## **1. Introduction**

Radiation therapy is one of the most important treatment modalities for cancer therapy alongside surgery and chemotherapy. However, a major problem associated with radiation therapy in the clinical setting is that, in many cases, local control of the tumor cannot be achieved using this modality alone. This has driven researchers into radiosensitizers, i.e., compounds that enhance the intrinsic sensitivity of cancer cells to ionizing radiation (IR). Several compounds, including halogenated pyrimidines and nitroimidazole derivatives, show radiosensitizing effects in cancer cells [1]; however, clinical application of these compounds is limited because they are highly toxic to normal cell and tissues. Therefore, radiosensitizers with low toxicity to normal tissues are urgently needed.

The principal target for IR-induced killing of cancer cells is DNA [2]. Of the different types of DNA damage generated by IR, DNA double-strand breaks (DSBs) are thought to be the most cytotoxic. DSBs induced by IR are preferentially repaired by non-homologous end joining (NHEJ) [3, 4], which joins the two broken DNA ends without the need for sequence homology. To enable NHEJ, the chromatin needs to be remodeled into an 'open' state so that the DNA repair proteins can access the DSB sites [5]. We previously reported that acetylation of histone proteins at DSB sites by the histone acetyltransferases (HATs), TIP60, CBP and p300, facilitates NHEJ through chromatin remodeling [6]. This suggests that the inhibition of HAT activity will radiosensitize cancer cells by suppressing NHEJ. In line with this, we and others demonstrated that several natural compounds with HAT-inhibitory activity are able to radiosensitize cancer cells [6-12]. Since some of these compounds are safe when administered to humans [13-15], they could potentially be used as radiosensitizers in a clinical setting. Here, we discuss the role of HATs in NHEJ, the radiosensitizing effects of compounds with HAT-inhibitory activity, and the prospects for the clinical application of these compounds.

© 2013 Kohno et al., licensee InTech. This is an open access chapter 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. © 2013 Kohno et al., licensee InTech. This is a paper 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.

## **2. Radiosensitization by HAT inhibition**

## **2.1. HATs are involved in chromatin remodeling required for DNA repair**

Chromosomal DNA and histones form a highly condensed structure known as chromatin. During the repair of DNA DSBs, the accessibility of DNA repair proteins to the DSB sites on chromosomal DNA is regulated by the relaxation of the chromosome structure via chromatin remodeling. Remodeling is mediated by both covalent (histone modifications, e.g., acetylation) and non-covalent (ATPase-dependent chromatin remodeling) interactions **(Figure 1)**. Several studies show that the acetylation of histones located at the DSB sites is a critical step for DNA repair [16-18]; however, the HATs involved in NHEJ have not been fully identified.

Histone Acetyltransferases (HATs) Involved in Non-Homologous End Joining as a Target for Radiosensitization 5

reverse direction, was integrated into the chromosomal DNA of H1299 human lung cancer cells as a substrate for DSBs and subsequent NHEJ repair. Human genomic DNA does not contain *I-Sce*I sites; therefore, the *I-Sce*I protein transiently expressed after transfection of the *I-Sce*I expression plasmid specifically cleaves the two *I-Sce*I sites in the substrate DNA to yield DSBs with incompatible ends. This results in DSBs in the chromosomal DNA. NHEJ of the two broken DNA strands results in deletion of the herpes simplex virus-thymidine kinase (TK) open reading frame and leads to the production of a transcript that enables the

translation of enhanced green fluorescent protein (EGFP) instead of the TK protein.

**Figure 2.** Cell-based NHEJ activity assay [6]. Upper panel: Assay design. Two *I-Sce*I sites (in the reverse

promoter/enhancer; IRES, internal ribosome entry site; pA, polyA signal. Lower panel: Factors essential

direction) are indicated by the yellow arrow heads. The locations of the PCR primers used for quantitative PCR to monitor DSB introduction by *I-Sce*I (uncut DNA) and subsequent joining (joined

DNA) are indicated by the purple and red arrows, respectively. CMV, cytomegalovirus

for NHEJ of DSBs on chromosomal DNA.

**Figure 1.** Chromatin remodeling is required for DSB repair. Both histone modifications and ATPasedependent chromatin remodeling are needed for efficient repair.

#### **2.2. Cell-based NHEJ activity assay**

We developed a new assay system for evaluating NHEJ repair of DSBs in the chromosomal DNA in living human cells [6, 18] because the existing NHEJ assays used only nonchromosomal (i.e., plasmid) DNA. The assay design is outlined in **Figure 2**. The IRES-TK-EGFP plasmid, which contains two recognition sites for *I-Sce*I endonuclease [20] in the

**2. Radiosensitization by HAT inhibition** 

**2.1. HATs are involved in chromatin remodeling required for DNA repair** 

repair [16-18]; however, the HATs involved in NHEJ have not been fully identified.

**Figure 1.** Chromatin remodeling is required for DSB repair. Both histone modifications and ATPase-

We developed a new assay system for evaluating NHEJ repair of DSBs in the chromosomal DNA in living human cells [6, 18] because the existing NHEJ assays used only nonchromosomal (i.e., plasmid) DNA. The assay design is outlined in **Figure 2**. The IRES-TK-EGFP plasmid, which contains two recognition sites for *I-Sce*I endonuclease [20] in the

dependent chromatin remodeling are needed for efficient repair.

**2.2. Cell-based NHEJ activity assay** 

Chromosomal DNA and histones form a highly condensed structure known as chromatin. During the repair of DNA DSBs, the accessibility of DNA repair proteins to the DSB sites on chromosomal DNA is regulated by the relaxation of the chromosome structure via chromatin remodeling. Remodeling is mediated by both covalent (histone modifications, e.g., acetylation) and non-covalent (ATPase-dependent chromatin remodeling) interactions **(Figure 1)**. Several studies show that the acetylation of histones located at the DSB sites is a critical step for DNA reverse direction, was integrated into the chromosomal DNA of H1299 human lung cancer cells as a substrate for DSBs and subsequent NHEJ repair. Human genomic DNA does not contain *I-Sce*I sites; therefore, the *I-Sce*I protein transiently expressed after transfection of the *I-Sce*I expression plasmid specifically cleaves the two *I-Sce*I sites in the substrate DNA to yield DSBs with incompatible ends. This results in DSBs in the chromosomal DNA. NHEJ of the two broken DNA strands results in deletion of the herpes simplex virus-thymidine kinase (TK) open reading frame and leads to the production of a transcript that enables the translation of enhanced green fluorescent protein (EGFP) instead of the TK protein.

**Figure 2.** Cell-based NHEJ activity assay [6]. Upper panel: Assay design. Two *I-Sce*I sites (in the reverse direction) are indicated by the yellow arrow heads. The locations of the PCR primers used for quantitative PCR to monitor DSB introduction by *I-Sce*I (uncut DNA) and subsequent joining (joined DNA) are indicated by the purple and red arrows, respectively. CMV, cytomegalovirus promoter/enhancer; IRES, internal ribosome entry site; pA, polyA signal. Lower panel: Factors essential for NHEJ of DSBs on chromosomal DNA.

Therefore, the efficiency of NHEJ can be assessed by monitoring EGFP production. In addition, the DSBs produced by *I-Sce*I and the subsequent NHEJ of the two broken DNA strands can be monitored by quantitative PCR.

Histone Acetyltransferases (HATs) Involved in Non-Homologous End Joining as a Target for Radiosensitization 7

**Figure 3.** Suppression of NHEJ by HAT inhibitors [6, 8]. H1299-dA3-1 #1 cells pretreated with curcumin

(20 M), anacardic acid (50 M), or garcinol (12 M) were transfected with the *I-Sce*I expression plasmid. (a) The proportion of EGFP-positive cells assessed by fluorescence-activated cell sorting analysis. (b,c) Proportion of joined (b) and uncut (c) DNA in cells assessed by quantitative PCR. The proportion of EGFP-positive cells and the proportion of joined DNA after garcinol treatment versus those in cells treated with DMSO (expressed as a ratio). The proportion of uncut DNA remaining after drug treatment expressed as a ratio of the amount of uncut DNA present after *I-Sce*I transduction versus the amount present before transduction. Results 48 h after transfection of the *I-Sce*I expression plasmid are shown. The results represent the mean ± s.d from three independent experiments. siCTR, non-

targeting siRNA.

Nucleotide sequencing of the joined DNA revealed that ligation required no (or very little) sequence homology between the DNA ends, indicating that the DNA ends were joined via NHEJ. In addition, the contribution of other factors essential for the NHEJ of incompatible DNA ends (whose involvement was indicated by *in vitro* and *in vivo* plasmid-based assays) was also confirmed in the present chromosome-based *in vivo* assay. These factors are KU80 and DNA-PKcs (synapsis), Artemis and PALF (DNA end resection), POL and POL (gap filling), and LIG4 (ligation) [21, 22].

## **2.3. HATs involved in NHEJ**

To date, several distinct families of HAT proteins have been identified, including CBP, P300, PCAF, GCN5 and MYST (which includes TIP60) [23]. We investigated the effects of ablating *CBP*, *P300*, *PCAF* and *TIP60* on NHEJ using the cell-based NHEJ activity assay. A decrease in the number of EGFP-positive cells was observed upon transfection with siRNA specific for *CBP*, *P300* or *TIP60*, but not in cells transfected with siRNA specific for *PCAF* [6]. These results indicate that CBP, P300 and TIP60 are involved in NHEJ in human cells.

### **2.4. Natural compounds with HAT activity suppress NHEJ activity**

Several compounds derived from natural ingredients show HAT-inhibitory activity (**Table 1**). Curcumin, a major curcumanoid found in the spice turmeric, is a specific inhibitor of the homologous HATs, CBP and P300 [24]. Anacardic acid, derived from the shell of *Anacardium occidentale* ('cashew nut'), inhibits P300, PCAF and TIP60 [7, 25, 26]*,* and Garcinol, found in the rind of *Garcinia indica* (mangosteen), inhibits P300 and PCAF [27].


1Molecular weight; 2Dose enhancement ratio (as assessed in cell viability assays [8]).

**Table 1.** HAT inhibitors that suppress NHEJ in human cells.

The cell-based NHEJ activity assay was used to investigate the effects of curcumin, anacardic acid, and garcinol on NHEJ; the results showed that treatment with each compound decreased the proportion of EGFP-positive cells (**Figure 3**) [6, 8]. This indicates that these HAT inhibitors suppress NHEJ activity *in vivo*.

strands can be monitored by quantitative PCR.

filling), and LIG4 (ligation) [21, 22].

**2.3. HATs involved in NHEJ** 

Compound MW1 Target

Anacardic acid

Therefore, the efficiency of NHEJ can be assessed by monitoring EGFP production. In addition, the DSBs produced by *I-Sce*I and the subsequent NHEJ of the two broken DNA

Nucleotide sequencing of the joined DNA revealed that ligation required no (or very little) sequence homology between the DNA ends, indicating that the DNA ends were joined via NHEJ. In addition, the contribution of other factors essential for the NHEJ of incompatible DNA ends (whose involvement was indicated by *in vitro* and *in vivo* plasmid-based assays) was also confirmed in the present chromosome-based *in vivo* assay. These factors are KU80 and DNA-PKcs (synapsis), Artemis and PALF (DNA end resection), POL and POL (gap

To date, several distinct families of HAT proteins have been identified, including CBP, P300, PCAF, GCN5 and MYST (which includes TIP60) [23]. We investigated the effects of ablating *CBP*, *P300*, *PCAF* and *TIP60* on NHEJ using the cell-based NHEJ activity assay. A decrease in the number of EGFP-positive cells was observed upon transfection with siRNA specific for *CBP*, *P300* or *TIP60*, but not in cells transfected with siRNA specific for *PCAF* [6]. These

Several compounds derived from natural ingredients show HAT-inhibitory activity (**Table 1**). Curcumin, a major curcumanoid found in the spice turmeric, is a specific inhibitor of the homologous HATs, CBP and P300 [24]. Anacardic acid, derived from the shell of *Anacardium occidentale* ('cashew nut'), inhibits P300, PCAF and TIP60 [7, 25, 26]*,* and Garcinol, found in

pathway, AP-1 STAT, LOX-1

pathways, topoisomerase I/II

Other target proteins/pathways DER2

NF-B pathway, LOX-1, Xanthine oxidase, 1.51

1.23

1.61

results indicate that CBP, P300 and TIP60 are involved in NHEJ in human cells.

**2.4. Natural compounds with HAT activity suppress NHEJ activity** 

the rind of *Garcinia indica* (mangosteen), inhibits P300 and PCAF [27].

Curcumin 368.38 CBP, P300 NF-B pathway, PI3K/mTOR/ETS2

Garcinol 602.80 P300, PCAF NF-B pathway, Src, MAPK/ERK, PI3K/Akt

The cell-based NHEJ activity assay was used to investigate the effects of curcumin, anacardic acid, and garcinol on NHEJ; the results showed that treatment with each compound decreased the proportion of EGFP-positive cells (**Figure 3**) [6, 8]. This indicates

HATs

1Molecular weight; 2Dose enhancement ratio (as assessed in cell viability assays [8]).

342.47 P300, PCAF, TIP60

**Table 1.** HAT inhibitors that suppress NHEJ in human cells.

that these HAT inhibitors suppress NHEJ activity *in vivo*.

**Figure 3.** Suppression of NHEJ by HAT inhibitors [6, 8]. H1299-dA3-1 #1 cells pretreated with curcumin (20 M), anacardic acid (50 M), or garcinol (12 M) were transfected with the *I-Sce*I expression plasmid. (a) The proportion of EGFP-positive cells assessed by fluorescence-activated cell sorting analysis. (b,c) Proportion of joined (b) and uncut (c) DNA in cells assessed by quantitative PCR. The proportion of EGFP-positive cells and the proportion of joined DNA after garcinol treatment versus those in cells treated with DMSO (expressed as a ratio). The proportion of uncut DNA remaining after drug treatment expressed as a ratio of the amount of uncut DNA present after *I-Sce*I transduction versus the amount present before transduction. Results 48 h after transfection of the *I-Sce*I expression plasmid are shown. The results represent the mean ± s.d from three independent experiments. siCTR, nontargeting siRNA.

### **2.5. HAT inhibitors radiosensitize cancer cells**

Because DNA DSBs induced by IR are preferentially repaired by NHEJ [3, 4], and HAT inhibitors suppress the activity of NHEJ, it is thought that HAT inhibitors may enhance the intrinsic sensitivity of cancer cells to IR. In line with this, the radiosensitizing effects of curcumin, anacardic acid and garcinol have been studied by ourselves and others both *in vitro* and *in vivo* (see **Table 2**); however, it is not certain that the observed radiosensitizing effects of these compounds is entirely due to their HAT-inhibitory activity, since they may also affect many other proteins or pathways considered to be important for the cancer cell survival (see **Table 1)**. In our own study, garcinol showed the strongest radiosensitization effect of the compounds tested. A nontoxic concentration of garcinol (4 uM) inhibited NHEJ without significantly affecting the DNA damage checkpoint **(Table 1)** [8]. Further investigations into mechanisms underlying the radiosensitizing effects of HAT inhibitors are ongoing.

Histone Acetyltransferases (HATs) Involved in Non-Homologous End Joining as a Target for Radiosensitization 9

capecitabine *vs.* RT1,

II Curcumin, gemcitabine *vs*. gemcitabine

II Curcumin alone M.D. Anderson

I Curcumin alone Lao CD, et al.[14]

M.D. Anderson Cancer Center

Healthcare Campus

Majeed M, et al.[15]

Cancer Center

Rambam

al.[13]

Sabinsa Corporation

capecitabine

Compound Disease Phase Modality Sponsor/Author

II Curcumin, RT1,

Curcumin Colorectal cancer I Curcumin alone Sharma RA, et

 II Garcinol, HCA2 acid *vs.* HCA2

piperine

The growing incidence of cancer worldwide indicates that radiation therapy will become increasingly significant as a cancer treatment [28]. Enhancing the efficacy of IR against cancer cells is urgent needs local control of tumors. As discussed in this article, radiosensitization of cancer cells by compounds with HAT-inhibitory activity has been reported at the level of basic research. Clinical studies indicate that some of these compounds can be administered to human patients with low systemic toxicity. Taken together, the available data suggest that compounds with HAT-inhibitory activity are promising candidates for radiosensitizers that may be applicable in clinical settings. However, the detailed mechanisms by which these compounds radiosensitize cancer cells are still largely unknown. Moreover, it is unclear whether these compounds can achieve adequate levels of radiosensitization in humans at a dose that shows no (or at least low) toxicity. Further investigations will establish whether HAT inhibitors can be used clinically

Garcinol Obesity II Garcinol, HCA2, forskolin,

Curcumin Rectal cancer

Curcumin Pancreatic

Curcumin Pancreatic

Curcumin Healthy

Garcinol Obesity

cancer

cancer

volunteer

Details on the clinical trials are available for inspection at:

1Radiation therapy, 2Hydroxycitric acid.

**3. Conclusions/perspectives** 

to radiosensitize cancer cells.

http://www.clinicaltrials.gov/ and http://www.garcitrin.com/clinical/.

**Table 3.** Clinical studies using compounds with HAT-inhibitory activity.


**Table 2.** Radiosensitization by HAT inhibitors.

#### **2.6. Clinical studies using compounds with HAT-inhibitory activity**

There are several clinical studies reporting the administration of compounds with HATinhibitory to humans (**Table 3**). Curcumin has been used, either alone or combined with radiation therapy and/or chemotherapeutic agents, to treat cancer patients, and garcinol has been used for weight-loss therapy. Although not all of the studies were designed to specifically evaluate the radiosensitizing effects of these compounds, the data will be of help to estimate their toxicity. The available data indicate that the side effects of these compounds are tolerable, at least when used alone.


Details on the clinical trials are available for inspection at:

http://www.clinicaltrials.gov/ and http://www.garcitrin.com/clinical/.

1Radiation therapy, 2Hydroxycitric acid.

8 Frontiers in Radiation Oncology

ongoing.

Curcumin /anacardic acid

Curcumin Cells &

mice

Anacardic acid Cells SQ20B,

**Table 2.** Radiosensitization by HAT inhibitors.

compounds are tolerable, at least when used alone.

**2.5. HAT inhibitors radiosensitize cancer cells** 

Because DNA DSBs induced by IR are preferentially repaired by NHEJ [3, 4], and HAT inhibitors suppress the activity of NHEJ, it is thought that HAT inhibitors may enhance the intrinsic sensitivity of cancer cells to IR. In line with this, the radiosensitizing effects of curcumin, anacardic acid and garcinol have been studied by ourselves and others both *in vitro* and *in vivo* (see **Table 2**); however, it is not certain that the observed radiosensitizing effects of these compounds is entirely due to their HAT-inhibitory activity, since they may also affect many other proteins or pathways considered to be important for the cancer cell survival (see **Table 1)**. In our own study, garcinol showed the strongest radiosensitization effect of the compounds tested. A nontoxic concentration of garcinol (4 uM) inhibited NHEJ without significantly affecting the DNA damage checkpoint **(Table 1)** [8]. Further investigations into mechanisms underlying the radiosensitizing effects of HAT inhibitors are

Compound Cells/mice Cell lines Authors Year Journal

SCC35, HeLa

**2.6. Clinical studies using compounds with HAT-inhibitory activity** 

SCC1 Khafif A, *et al*.[9] 2009 The Laryngoscope

Sun Y, *et al*.[7] 2006 FEBS Lett

Biol Phys

Biol Phys

Cells HCT116 Sandur SK, *et al*.[10] 2009 Int J Radiat Oncol

Cells PC-3 Li M, *et al*.[11] 2007 Cancer Res Cells PC-3 Chendil D, *et al*.[12] 2004 Oncogene

Cells H1299 Ogiwara H, *et al*.[6] 2011 Oncogene

Garcinol Cells A549, HeLa Oike, *et al*.[8] 2012 Int J Radiat Oncol

There are several clinical studies reporting the administration of compounds with HATinhibitory to humans (**Table 3**). Curcumin has been used, either alone or combined with radiation therapy and/or chemotherapeutic agents, to treat cancer patients, and garcinol has been used for weight-loss therapy. Although not all of the studies were designed to specifically evaluate the radiosensitizing effects of these compounds, the data will be of help to estimate their toxicity. The available data indicate that the side effects of these **Table 3.** Clinical studies using compounds with HAT-inhibitory activity.

## **3. Conclusions/perspectives**

The growing incidence of cancer worldwide indicates that radiation therapy will become increasingly significant as a cancer treatment [28]. Enhancing the efficacy of IR against cancer cells is urgent needs local control of tumors. As discussed in this article, radiosensitization of cancer cells by compounds with HAT-inhibitory activity has been reported at the level of basic research. Clinical studies indicate that some of these compounds can be administered to human patients with low systemic toxicity. Taken together, the available data suggest that compounds with HAT-inhibitory activity are promising candidates for radiosensitizers that may be applicable in clinical settings. However, the detailed mechanisms by which these compounds radiosensitize cancer cells are still largely unknown. Moreover, it is unclear whether these compounds can achieve adequate levels of radiosensitization in humans at a dose that shows no (or at least low) toxicity. Further investigations will establish whether HAT inhibitors can be used clinically to radiosensitize cancer cells.

## **Author details**

#### Takahiro Oike

*Division of Genome Biology, National Cancer Center Research Institute, Tokyo, Japan Department of Radiation Oncology, Gunma University Graduate School of Medicine, Gunma, Japan*  Histone Acetyltransferases (HATs) Involved in Non-Homologous End Joining as a Target for Radiosensitization 11

[9] Khafif A, Lev-Ari S, Vexler A, *et al*. (2009) Curcumin: a potential radio-enhancer in head

[10] Sandur SK, Deorukhkar A, Pandey MK, *et al.* (2009) Curcumin modulates the radiosensitivity of colorectal cancer cells by suppressing constitutive and inducible NF-

[11] Mao Li, Zhuo Zhang, Donald L. Hill, *et al*. (2007) Curcumin, a dietary component, has anticancer, chemosensitization, and radiosensitization effects by down-regulating the MDM2 oncogene through the PI3K/mTOR/ETS2 pathway. Cancer Res. 67:1988-96. [12] Chendil D, Ranga RS, Meigooni D, *et al*. (2004) Curcumin confers radiosensitizing effect

[13] Sharma RA, Euden SA, Platton AL, *et al*. (2004) Phase I clinical trial of oral curcumin biomarkers of systemic activity and compliance. Clin. Cancer Res. 10:6847-1854. [14] Lao CD, Ruffin MT, Normolle D, *et al*. (2006) Dose escalation of a curcuminoid

[15] Majeed M, Badmaev V, Khan N, *et al.* (2009) A new class of phytonutrients for body

[16] Bird AW, Yu DY, Pray-Grant MG, *et al*. (2002) Acetylation of histone H4 by Esa1 is

[17] Tamburini BA, Tyler JK. (2005) Localized histone acetylation and deacetylation triggered by the homologous recombination pathway of double-strand DNA repair.

[18] Murr R, Loizou JI, Yang YG, *et al*. (2006) Histone acetylation by Trrap-Tip60 modulates loading of repair proteins and repair of DNA double-strand breaks. Nat. Cell. Biol.

[19] Lan L, Ui A, Nakajima S, Hatakeyama K, Hoshi M, Watanabe R, Janicki S, Ogiwara H, Kohno T, Kanno S, Yasui A. The ACF1 complex is required for DNA double-strand

[20] Jasin M. (1996) Genetic manipulation of genomes with rare-cutting endonucleases.

[21] Ogiwara H, Kohno T. Essential factors for incompatible DNA end joining at chromosomal DNA double strand breaks in vivo. PLoS One, 2011, 6(12) e28756. [22] Li S, Kanno S, Watanabe R, Ogiwara H, Kohno T, Watanabe G, Yasui A, Lieber MR. PALF acts as both a single-stranded DNA endonuclease and a single-stranded DNA 3' -exonuclease and can participate in DNA end joining in a biochemical system. J Biol

[23] Carrozza MJ, Utley RT, Workman JL, *et al*. (2003) The diverse functions of histone

[24] Balasubramanyam K, Varier RA, Altaf M, *et al*. (2004) Curcumin, a novel p300/CREBbinding protein-specific inhibitor of acetyltransferase, represses the acetylation of histone/nonhistone proteins and histone acetyltransferase-dependent chromatin

required for DNA double-strand break repair. Nature. 419:411–415.

and neck cancer. Laryngoscope. 119:2019-2026.

B activity. Int. J. Radiat. Oncol. Biol. Phys. 75:534-542.

in prostate cancer cell line PC-3. Oncogene. 23:1599-1607.

formulation. BMC Complment Altern. Med. 6:10.

break repair in human cells. Mol Cell 2010, 40: 976-987.

acetyltransferase complexes. Trends Genet. 19:321–329.

transcription. J. Biol. Chem. 279:51163-51171.

weight management. NUTRAfoods 8:17-26.

Mol. Cell. Biol. 25:4903–4913.

Trends Genet. 12:224–228.

Chem. 2011,286:36368-77.

8:91–99.

Hideaki Ogiwara and Takashi Kohno\* *Division of Genome Biology, National Cancer Center Research Institute, Tokyo, Japan* 

Takashi Nakano *Department of Radiation Oncology, Gunma University Graduate School of Medicine, Gunma, Japan* 

## **Acknowledgement**

This work was supported by Grants-in-Aid from the Ministry of Education, Culture, Sports, Science and Technology of Japan for Scientific Research on Innovative Areas (22131006) and from the Japan Society for the Promotion of Science for Young Scientists (B) KAKENHI (23701110); and the National Cancer Center Research and Development Fund.

## **4. References**


<sup>\*</sup> Corresponding Author

Histone Acetyltransferases (HATs) Involved in Non-Homologous End Joining as a Target for Radiosensitization 11

[9] Khafif A, Lev-Ari S, Vexler A, *et al*. (2009) Curcumin: a potential radio-enhancer in head and neck cancer. Laryngoscope. 119:2019-2026.

10 Frontiers in Radiation Oncology

Hideaki Ogiwara and Takashi Kohno\*

Williams Wilkins. pp. 419-431.

Radiat. Oncol. Biol. Phys. in press.

Lippincott Williams Wilkins. pp. 16-29.

in maintaining genomic integrity. DNA Repair. 8:1042-1048.

*Division of Genome Biology, National Cancer Center Research Institute, Tokyo, Japan* 

*Division of Genome Biology, National Cancer Center Research Institute, Tokyo, Japan* 

(23701110); and the National Cancer Center Research and Development Fund.

*Department of Radiation Oncology, Gunma University Graduate School of Medicine, Gunma, Japan* 

*Department of Radiation Oncology, Gunma University Graduate School of Medicine, Gunma, Japan* 

This work was supported by Grants-in-Aid from the Ministry of Education, Culture, Sports, Science and Technology of Japan for Scientific Research on Innovative Areas (22131006) and from the Japan Society for the Promotion of Science for Young Scientists (B) KAKENHI

[1] Hall EJ, Giaccia AJ. (2006) Radiosensitizers and bioreductive drugs. In: McAllister L, Bierig L, Barret K, editors. Radiobiology for the radiologist. Philadelphia: Lippincott

[2] Hall EJ, Giaccia AJ. (2006) DNA strand breaks and chromosomal aberrations. In: McAllister L, Bierig L, Barret K, editors. Radiobiology for the radiologist. Philadelphia:

[3] Burma S, Chen BPC, Chen DJ, *et al.* (2006) Role of non-homologous end joining (NHEJ)

[4] Lieber MR. (2008) The mechanism of human nonhomologous DNA end joining. J. Biol.

[5] Rossetto D, Truman AW, Kron SJ, *et al.* (2010) Epigenetic modifications in double-strand

[6] Ogiwara H, Ui A, Otsuka A, *et al.* (2011) Histone acetylation by CBP and p300 at double-strand break sites facilitates SWI/SNF chromatin remodeling and the

[7] Sun Y, Jiang X, Chen S, *et al.* (2006) Inhibition of histone acetyltransferase activity by anacardic acid sensitizes tumor cells to ionizing radiation. FEBS Lett. 580:4353–4356. [8] Oike T, Ogiwara H, Torikai K, *et al*. (2012) Garcinol, a histone acetyltransferase inhibitor, radiosensitizes cancer cells by inhibiting non-homologous end joining. Int. J.

break DNA damage signaling and repair. Clin. Cancer. Res. 15:4543-4552.

recruitment of non-homologous end joining factors. Oncogene. 5:2135-2146.

**Author details** 

Takahiro Oike

Takashi Nakano

**4. References** 

Chem. 283:1-5.

 \*

Corresponding Author

**Acknowledgement** 

	- [25] Hemshekhar M, Sebastin SM, Kemparaju K, *et al*. (2011) Emerging roles of anacardic acid and its derivatives: a pharmacological overview. Basic Clin. Pharmacol. Toxicol. doi: 10.1111/j.1742-7843.2011.00833.x. [Epub ahead of print]

**Chapter 2** 

© 2013 Qi et al., licensee InTech. This is an open access chapter 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.

© 2013 Qi et al., licensee InTech. This is a paper 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.

**A Framework for Modeling** 

Jin-Peng Qi, Yong-Sheng Ding and Xian-Hui Zeng

Additional information is available at the end of the chapter

biochemical control during and after radiotherapy [6-8].

http://dx.doi.org/10.5772/56623

**1. Introduction** 

**the Cellular Defending Mechanisms** 

**Against Genome Stress Under Radiotherapy** 

Like immunotherapy, chemotherapy, and surgery, radiotherapy is one of the major tools in fighting against cancer. As acute IR is applied, cell can trigger its self-defensive mechanisms in response to genome stresses [1]. As one of the pivotal anticancer genes within the cell, P53 can control the transcription and translation of series genes, and trigger cell cycle arrest and apoptosis through interaction with downstream genes and their complicated signal pathways [2]. Under radiotherapy, the outcomes of cellular response depend on the presence of functional P53 proteins to induce tumor regression through apoptotic pathways [3]. Conversely, the P53 tumor suppressor is the most commonly known specific target of mutation in tumorigenesis [4]. Abnormalities in the P53 have been identified in over 60% of human cancers and the status of P53 within tumor cells has been proposed to be one of the determinant response to anticancer therapies [3,4]. Controlled radiotherapy studies show the existence of a strong biologic basis for considering P53 status as a radiation predictor [3,5]. Therefore, the status of P53 in tumor cell can be considered as a predictor for long-term

Recently, several models have been proposed to explain the damped oscillations of P53 in cell populations [9-12]. However, the dynamic mechanism of the single-cell responses is not completely clear yet, and the complicated regulations among genes and their signal

Many studies have indicated that introducing novel mathematical and computational approaches can stimulate in-depth investigation into various complicated biological systems (see, e.g., [13-23]). These methods have provided useful tools for both basic research and

pathways need to be further addressed, particularly under the condition of acute IR.

