**3.4 Human equilibrative nucleoside transporter 1 (hENT1)**

Gemcitabine, a pyrimidine nucleoside analogue, has clinically important activity in advanced and metastatic pancreatic adenocarcinoma and for which it is now the standard of care (Maréchal et al., 2009)(Ansari et al., 2011). Gemcitabine is a prodrug that is phosphorylated by deoxycitidine kinase to its mononucleotide in the rate-limiting step of its cellular anabolism. Subsequent nucleotide kinases convert gemcitabine monophosphate to its active metabolites, gemcitabine diphosphate and triphosphate. Permeation of gemcitabine through the plasma membrane requires specialized integral membrane nucleoside transporter proteins. Among these transporters, the major mediators of gemcitabine uptake into human cells appear to be the human equilibrative nucleoside transporter 1 (hENT1) and to a lesser extent the human conservative nucleoside transporter 3 (hCNT3) (Maréchal et al., 2009). Recently, it was reported that tissue mRNA levels of the hENT1, which mediates the cellular entry of gemcitabine, correlated with survival (Ansari et al., 2011). Several subsequent immunohistochemically based studies demonstrated that hENT-1 holds promise as an independent predictive marker to identify those likely to benefit from gemcitabine based monotherapy (Morinaga et al., 2011)(Spratlin et al., 2004)(Farrell et al., 2009) and gemcitabine based chemoradiotherapy (Maréchal et al., 2009)(Murata et al., 2011). In addition, the expression of hENT1 provides independent prognostic information in untreated pancreatic carcinoma patients as well as those treated with adjuvant gemcitabine-based therapy (Kim et al., 2011b)(Maréchal et al., 2009). Whether these assays provide sufficient predictive information to guide treatment decision requires prospective evaluation in randomized clinical trials. However, the consistency and strength of the accumulating preclinical and translational data suggest that nucleoside transporters play an important role in clinical outcomes after gemcitabine adjuvant chemotherapy for pancreatic cancer (Maréchal et al., 2009).

### **3.5 Conclusion**

Although the tumor node metastasis classification provides important prognostic information, it permits only crude stratification of clinical outcome for patients with pancreatic cancer. Although some potential markers were identified, a high degree of inconsistency still exists between reports. Validation through large multicenter prospective studies using standardized protocols is still needed. Considering the complexity of the disease, it seems reasonable to hypothesize that panels of markers, rather than single proteins, might become useful (Ansari et al., 2011).

#### **4. Gastric cancer**

#### **4.1 Introduction**

Gastric cancer is one of the most common tumors and remains the second leading cause of cancer death worldwide (Gravalos & Jimeno, 2008)(Wagner & Moehler, 2009). Gastric cancer is a heterogeneous disease divided in at least two different tumor entities, the intestinal and the diffuse form, with difference in epidemiology, cause, pathogenesis and disorder. The development of the intestinal form, usually in older patients, is related to *Helicobacter pylori* and usually located in the corpus and the antrum and related to preexisting corpus predominant atrophic gastritis, followed by intestinal metaplasia. In contrast, the diffuse form is usually poorly differentiated, located most frequently in the proximal stomach and its incidence is rising at an alarming rate in overweight young men suffering from gastroesophageal reflux. Diffuse type cancers have usually a worse prognosis (Wagner & Moehler, 2009).

Surgical resection remains the mainstay of treatment and cure in localized, non-metastatic gastric cancer while no globally accepted consensus exists on the best treatment regimen to be used in advanced gastric cancer (De Vita et al., 2010)(Lorenzen & Lordick, 2011). At present, the combination of a fluoropyrimidine and a platinum analogue either alone or in combination with a third drug such as an antracycline or taxanes are the most effective combinations resulting in a median survival of 8-10 months (Lorenzen & Lordick, 2011). These observations suggest the need for new therapeutic approaches, based on the implementation of predictive biomarkers, to further improve the outcome of patients with advanced gastric cancer (De Vita et al., 2010)(Wagner & Moehler, 2009)(Lorenzen & Lordick, 2011). A better understanding of the molecular basis of cancer has contributed to the development of rationally designed molecular targeted therapies, which interfere with the signaling cascades involved in cell differentiation, proliferation and survival (Gravalos & Jimeno, 2008). Recently, the evidence that upregulation of signaling pathways of EGFRfamily plays a central role in cell differentiation, proliferation, and survival has supported the development of antitumor strategies against these targets (De Vita et al., 2010). One of the most considerable innovative targets in human cancer is the HER family.

#### **4.2 HER2**

62 Biomarker

Gemcitabine, a pyrimidine nucleoside analogue, has clinically important activity in advanced and metastatic pancreatic adenocarcinoma and for which it is now the standard of care (Maréchal et al., 2009)(Ansari et al., 2011). Gemcitabine is a prodrug that is phosphorylated by deoxycitidine kinase to its mononucleotide in the rate-limiting step of its cellular anabolism. Subsequent nucleotide kinases convert gemcitabine monophosphate to its active metabolites, gemcitabine diphosphate and triphosphate. Permeation of gemcitabine through the plasma membrane requires specialized integral membrane nucleoside transporter proteins. Among these transporters, the major mediators of gemcitabine uptake into human cells appear to be the human equilibrative nucleoside transporter 1 (hENT1) and to a lesser extent the human conservative nucleoside transporter 3 (hCNT3) (Maréchal et al., 2009). Recently, it was reported that tissue mRNA levels of the hENT1, which mediates the cellular entry of gemcitabine, correlated with survival (Ansari et al., 2011). Several subsequent immunohistochemically based studies demonstrated that hENT-1 holds promise as an independent predictive marker to identify those likely to benefit from gemcitabine based monotherapy (Morinaga et al., 2011)(Spratlin et al., 2004)(Farrell et al., 2009) and gemcitabine based chemoradiotherapy (Maréchal et al., 2009)(Murata et al., 2011). In addition, the expression of hENT1 provides independent prognostic information in untreated pancreatic carcinoma patients as well as those treated with adjuvant gemcitabine-based therapy (Kim et al., 2011b)(Maréchal et al., 2009). Whether these assays provide sufficient predictive information to guide treatment decision requires prospective evaluation in randomized clinical trials. However, the consistency and strength of the accumulating preclinical and translational data suggest that nucleoside transporters play an important role in clinical outcomes after gemcitabine adjuvant chemotherapy for

Although the tumor node metastasis classification provides important prognostic information, it permits only crude stratification of clinical outcome for patients with pancreatic cancer. Although some potential markers were identified, a high degree of inconsistency still exists between reports. Validation through large multicenter prospective studies using standardized protocols is still needed. Considering the complexity of the disease, it seems reasonable to hypothesize that panels of markers, rather than single

Gastric cancer is one of the most common tumors and remains the second leading cause of cancer death worldwide (Gravalos & Jimeno, 2008)(Wagner & Moehler, 2009). Gastric cancer is a heterogeneous disease divided in at least two different tumor entities, the intestinal and the diffuse form, with difference in epidemiology, cause, pathogenesis and disorder. The development of the intestinal form, usually in older patients, is related to *Helicobacter pylori* and usually located in the corpus and the antrum and related to preexisting corpus predominant atrophic gastritis, followed by intestinal metaplasia. In contrast, the diffuse

**3.4 Human equilibrative nucleoside transporter 1 (hENT1)** 

pancreatic cancer (Maréchal et al., 2009).

proteins, might become useful (Ansari et al., 2011).

**3.5 Conclusion** 

**4. Gastric cancer 4.1 Introduction** 

The epidermal growth factor receptor (EGFR) family is composed of four members: HER1 also known as EGFR1, HER2, HER3 and HER4, amongst which the EGFR1 and HER2 represents targets for drugs currently under development for gastric cancer (Wagner & Moehler, 2009). The HER2 protein is a 185 kDa transmembrane tyrosine kinase (TK) receptor encoded by a gene located on chromosome 17q21, with an extracellular ligand-binding domain, a short transmembrane domain and an intracellular domain with TK activity. Up to now, no ligands have been identified for its extracellular domain, but it seems to be the preferred heterodimerization partner for other members of the HER family (De Vita et al., 2010). HER2 functions as an oncogene and its amplification or overexpression plays a central role in the initiation, progression and metastasis of some common cancers. Aberrant HER2 expression or function has been implicated in about 10-34% of invasive breast cancers. In addition, HER 2 also appears to be overexpressed in colon, bladder, ovarian, endometrium, lung, uterine cervix, head and neck, and esophageal carcinomas. The first description of HER2 overexpression in gastric cancer, using IHC, was reported in 1986. Since then, a number of studies have confirmed these findings, reporting a HER2 positivity rate in a wide range (6-35%) of gastric carcinomas. Moreover, HER2 expression varies depending on histology and on primary tumor location (Lorenzen & Lordick, 2011)(De Vita et al., 2010). The randomized open-label, multinational phase III ToGA (Trastuzumb for Gastric Cancer) trial, in which by now the largest population of 3807 gastric cancers were centrally screened for *HER2* gene amplification (Fluorescent in situ hybridization (FISH)) and HER2 protein overexpression (IHC 3+), reported a HER2 positivity of 22.1%, with a high degree of concordance between IHC and FISH (87,2%). Furthermore, HER2 positivity rates were found to be higher in esophagogastric junction cancer than in gastric cancer and

Biomarkers in Gastrointestinal Cancer: Focus on Colon, Pancreatic and Gastric Cancer 65

Trastuzumab, a recombinant humanized IgG1 monoclonal antibody directed against the extracellular domain of HER2, induces antibody-dependent cellular cytotoxicity, inhibits HER2-mediated signaling and prevents cleavage of the extracellular domain of HER2. In HER2 positive breast cancer, trastuzumab has demonstrated survival benefits for patients with early and metastatic disease and is now the standard of care (Bang et al., 2010)(Croxtall & McKeage, 2010). Several studies indicate antitumor activity of trastuzumab in overexpressing HER2 human gastric cancer cell lines or xenograft models (Matsui et al., 2005)(Tanner et al., 2005)(Fujimoto-Ouchi et al., 2007). Most of these studies used the NCI-N87 and or 4-1ST gastric cell lines, which show HER2 expression in IHC and gene amplification on FISH. These studies showed that trastuzumab suppressed the growth of human gastric cancer with HER2 overexpression *in vitro* and *in vivo* and improved the survival of mice with peritoneal dissemination and ascites of gastric cancer. In addition, trastuzumab administered in combination with chemotherapy agents for gastric cancer showed potent antitumor activity, which was significantly greater than did trastuzumab or the chemotherapy agents as single treatments. A three-drug combination of capacetabine, cisplatin, and trastuzumab achieved remarkable tumor growth inhibition in the N87 model (Fujimoto-Ouchi et al., 2007)(Kim et al., 2008)(Gravalos & Jimeno, 2008). In addition, there are currently no data regarding resistance to trastuzumab in gastric cancer cells and there are no *in vitro* tests available to enable the prediction of resistance (Croxtall & McKeage,

Based on these results there was a strong rationale to investigate the clinical potential of trastuzumab in gastric cancer patients. Several preliminary single-arm phase II trials paved the way for the registration of the large, randomized controlled, open label, multicenter, international phase III trial which was undertaken in 24 centers in Aisa, Central and South America and Europe. The objective of this "Trastuzumab for Gastric Cancer" (ToGA) study was to assess the clinical efficacy and safety of trastuzumab added to chemotherapy for firstline treatment of advanced gastric or gastro-esophageal junction cancer with overexpression of HER2. Tumors were centrally tested for HER2 status with IHC (Hercep test) and FISH. Because of the inherent difference between breast and gastric tumors, notably tumor heterogeneity and the occurrence of baso(lateral) membrane staining, a new set of IHC scoring criteria were developed that are specific for gastric cancer. Patients were eligible if their tumor samples were scored as 3+ on IHC or if they were FISH positive (Bang et al., 2010) (Bang et al., 2010). As mentioned above, 22,1% of all gastric cancer screened in the ToGA trial were HER2-positive, which is broadly comparable with the incidence in breast cancer. Moreover, there was a high degree of concordance between IHC and FISH. Therefore, IHC is suitable for primary testing of HER2 positivity in gastric cancer with a score of IHC3+ indicating eligibility for treatment with trastuzumab, IHC2+ should be retested by FISH to confirm HER2 positivity and a score of IHC0 or 1+ should be considered as HER2-negative. The incidence of HER2 positivity differed according to tumor location and histological subtype. There was a significantly greater incidence of HER2-positive cancers of the oesophagogastric junction than the stomach and in intestinal than diffuse or mixed cancers. In addition, the incidence of HER2 positive gastric cancers was similar between Europe and Asia, but varied between countries (Croxtall & McKeage, 2010). Patients who satisfied all eligibility criteria (594 patients) were randomized in a 1:1 ratio and

**4.4 Trastuzumab and the predictive role of HER2 in gastric cancer** 

2010).

in intestinal cancer than diffuse or mixed type (Lorenzen & Lordick, 2011)(Croxtall & McKeage, 2010).

Trastuzumab, a recombinant humanized IgG1 monoclonal antibody directed against the extracellular domain of HER2, in combination with chemotherapy agents, has recently received approval in the EU and USA for treatment of metastatic HER2-positive gastric cancer without prior anti-cancer treatment for metastatic disease (Croxtall & McKeage, 2010).

#### **4.3 HER2 as prognostic factor in gastric cancer**

The TNM stage is the most important prognostic factor for gastric cancer. Prognosis, however, varies among patients in the same stage. Therefore, additional classification parameters, like HER2 need to be defined in addition to the TNM and the classical pathological characteristics of the tumor in order to better identify the biological subset of this disease (Gravalos & Jimeno, 2008). The role of HER2 as prognostic marker in gastric cancer has been controversial because some of the initial studies failed to find an association with outcome (Zhang et al., 2009a)(De Vita et al., 2010)(Gravalos & Jimeno, 2008)(Lorenzen & Lordick, 2011). Other authors, however, reported a direct correlation between HER2 overexpression and poor outcome (Ananiev et al., 2011)(Kim et al., 2011a)(Im et al., 2005). However, the largest study to date investigating the prognostic significance of HER2 expression in 924 gastric cancer patients showed that HER2 expression is not related to gastric cancer patients outcome (Grabsch et al., 2010). Chua et al. (Chua & Merrett, 2011) performed a systematic examination of the literature to identify translational studies that correlated HER2 with clinicopathologic markers and/or survival. This review included 49 studies totaling 11,337 patients. IHC was most commonly used to assess HER2 expression, identifying a median rate of 18% of gastric cancer demonstrating HER2 overexpression. In patients with and without HER2 overexpression, the median 3-year disease-free survival rate was 58% and 86%, respectively. Of the 35 studies reporting the impact of HER2 overexpression on survival, 20 studies (57%) reported no difference in overall survival, two studies (6%) reported significantly longer overall survival in patients with HER2 overexpression and 13 studies (37%) reported significantly poorer overall survival in patients with HER2 overexpression. HER2 overexpression appears to be associated with poorer survival and with intestinal-type gastric cancer in this group of patients for whom majority undergone curative gastrectomy. Results of the ToGA trial, which are discussed below, seem to refute this suggestion as demonstrated by the longer than expected survival of patients in the control arm, who received chemotherapy alone. However possible confounding factors, such as the wide use of second line treatment and the better prognosis associated with the intestinal histology, should be kept in mind when interpreting these results (Fornaro et al., 2011). In addition, these conflicting results could be due to the lack of a standardized definition of HER2 positivity in gastric cancer (De Vita et al., 2010)(Lorenzen & Lordick, 2011).

Hence, definitive answers about the prognostic role of HER2 in gastric cancer and gastricesophageal cancer cannot be derived from the available data, which thus emphasizes the need for further research in the field (Fornaro et al., 2011).

#### **4.4 Trastuzumab and the predictive role of HER2 in gastric cancer**

64 Biomarker

in intestinal cancer than diffuse or mixed type (Lorenzen & Lordick, 2011)(Croxtall &

Trastuzumab, a recombinant humanized IgG1 monoclonal antibody directed against the extracellular domain of HER2, in combination with chemotherapy agents, has recently received approval in the EU and USA for treatment of metastatic HER2-positive gastric cancer without prior anti-cancer treatment for metastatic disease (Croxtall & McKeage,

The TNM stage is the most important prognostic factor for gastric cancer. Prognosis, however, varies among patients in the same stage. Therefore, additional classification parameters, like HER2 need to be defined in addition to the TNM and the classical pathological characteristics of the tumor in order to better identify the biological subset of this disease (Gravalos & Jimeno, 2008). The role of HER2 as prognostic marker in gastric cancer has been controversial because some of the initial studies failed to find an association with outcome (Zhang et al., 2009a)(De Vita et al., 2010)(Gravalos & Jimeno, 2008)(Lorenzen & Lordick, 2011). Other authors, however, reported a direct correlation between HER2 overexpression and poor outcome (Ananiev et al., 2011)(Kim et al., 2011a)(Im et al., 2005). However, the largest study to date investigating the prognostic significance of HER2 expression in 924 gastric cancer patients showed that HER2 expression is not related to gastric cancer patients outcome (Grabsch et al., 2010). Chua et al. (Chua & Merrett, 2011) performed a systematic examination of the literature to identify translational studies that correlated HER2 with clinicopathologic markers and/or survival. This review included 49 studies totaling 11,337 patients. IHC was most commonly used to assess HER2 expression, identifying a median rate of 18% of gastric cancer demonstrating HER2 overexpression. In patients with and without HER2 overexpression, the median 3-year disease-free survival rate was 58% and 86%, respectively. Of the 35 studies reporting the impact of HER2 overexpression on survival, 20 studies (57%) reported no difference in overall survival, two studies (6%) reported significantly longer overall survival in patients with HER2 overexpression and 13 studies (37%) reported significantly poorer overall survival in patients with HER2 overexpression. HER2 overexpression appears to be associated with poorer survival and with intestinal-type gastric cancer in this group of patients for whom majority undergone curative gastrectomy. Results of the ToGA trial, which are discussed below, seem to refute this suggestion as demonstrated by the longer than expected survival of patients in the control arm, who received chemotherapy alone. However possible confounding factors, such as the wide use of second line treatment and the better prognosis associated with the intestinal histology, should be kept in mind when interpreting these results (Fornaro et al., 2011). In addition, these conflicting results could be due to the lack of a standardized definition of HER2 positivity in gastric cancer (De Vita et al., 2010)(Lorenzen

Hence, definitive answers about the prognostic role of HER2 in gastric cancer and gastricesophageal cancer cannot be derived from the available data, which thus emphasizes the

need for further research in the field (Fornaro et al., 2011).

McKeage, 2010).

& Lordick, 2011).

**4.3 HER2 as prognostic factor in gastric cancer** 

2010).

Trastuzumab, a recombinant humanized IgG1 monoclonal antibody directed against the extracellular domain of HER2, induces antibody-dependent cellular cytotoxicity, inhibits HER2-mediated signaling and prevents cleavage of the extracellular domain of HER2. In HER2 positive breast cancer, trastuzumab has demonstrated survival benefits for patients with early and metastatic disease and is now the standard of care (Bang et al., 2010)(Croxtall & McKeage, 2010). Several studies indicate antitumor activity of trastuzumab in overexpressing HER2 human gastric cancer cell lines or xenograft models (Matsui et al., 2005)(Tanner et al., 2005)(Fujimoto-Ouchi et al., 2007). Most of these studies used the NCI-N87 and or 4-1ST gastric cell lines, which show HER2 expression in IHC and gene amplification on FISH. These studies showed that trastuzumab suppressed the growth of human gastric cancer with HER2 overexpression *in vitro* and *in vivo* and improved the survival of mice with peritoneal dissemination and ascites of gastric cancer. In addition, trastuzumab administered in combination with chemotherapy agents for gastric cancer showed potent antitumor activity, which was significantly greater than did trastuzumab or the chemotherapy agents as single treatments. A three-drug combination of capacetabine, cisplatin, and trastuzumab achieved remarkable tumor growth inhibition in the N87 model (Fujimoto-Ouchi et al., 2007)(Kim et al., 2008)(Gravalos & Jimeno, 2008). In addition, there are currently no data regarding resistance to trastuzumab in gastric cancer cells and there are no *in vitro* tests available to enable the prediction of resistance (Croxtall & McKeage, 2010).

Based on these results there was a strong rationale to investigate the clinical potential of trastuzumab in gastric cancer patients. Several preliminary single-arm phase II trials paved the way for the registration of the large, randomized controlled, open label, multicenter, international phase III trial which was undertaken in 24 centers in Aisa, Central and South America and Europe. The objective of this "Trastuzumab for Gastric Cancer" (ToGA) study was to assess the clinical efficacy and safety of trastuzumab added to chemotherapy for firstline treatment of advanced gastric or gastro-esophageal junction cancer with overexpression of HER2. Tumors were centrally tested for HER2 status with IHC (Hercep test) and FISH. Because of the inherent difference between breast and gastric tumors, notably tumor heterogeneity and the occurrence of baso(lateral) membrane staining, a new set of IHC scoring criteria were developed that are specific for gastric cancer. Patients were eligible if their tumor samples were scored as 3+ on IHC or if they were FISH positive (Bang et al., 2010) (Bang et al., 2010). As mentioned above, 22,1% of all gastric cancer screened in the ToGA trial were HER2-positive, which is broadly comparable with the incidence in breast cancer. Moreover, there was a high degree of concordance between IHC and FISH. Therefore, IHC is suitable for primary testing of HER2 positivity in gastric cancer with a score of IHC3+ indicating eligibility for treatment with trastuzumab, IHC2+ should be retested by FISH to confirm HER2 positivity and a score of IHC0 or 1+ should be considered as HER2-negative. The incidence of HER2 positivity differed according to tumor location and histological subtype. There was a significantly greater incidence of HER2-positive cancers of the oesophagogastric junction than the stomach and in intestinal than diffuse or mixed cancers. In addition, the incidence of HER2 positive gastric cancers was similar between Europe and Asia, but varied between countries (Croxtall & McKeage, 2010). Patients who satisfied all eligibility criteria (594 patients) were randomized in a 1:1 ratio and

Biomarkers in Gastrointestinal Cancer: Focus on Colon, Pancreatic and Gastric Cancer 67

Ample data shows that only a limited portion of patients may benefit from anti-cancer treatments currently used in the clinic. Personalized cancer medicine, based on genetic profiling of individual tumors and biomarkers, is regarded as the treatment strategy of the future. In this review the most promising biomarkers in colorectal, pancreatic and gastric cancer were discussed (table 2). Currently, the only biomarker that has made it into clinical practice for colorectal cancer is *KRAS* mutation for the selection of patients eligible for cetuximab therapy. Furthermore, evidence shows that other molecular alterations, such as *BRAF*, *PIK3CA* (exon-20) mutations or loss of PTEN expression, could preclude response to

**Location Predictive Prognostic Stage** 

BRAF

hENT1 miRs GLUT

HER2

Clinical stage (since 2008) Preclinical stage Preclinical stage

Clinical stage Preclinical stage Preclinical stage Preclinical stage

Clinical stage (since 2010) Preclinical stage

PIK3CA exon 20 mut.

hENT1 miRs

Table 2. The most promising biomarkers in colorectal, pancreatic and gastric cancer

Improved screening for early diagnosis is essential in order to increase the rate of curatively resectable pancreatic carcinomas, thereby ameliorating patient's prognosis. A relatively noninvasive, cost efficient possibility could be provided by the measurement of disease-specific markers in peripheral blood. However, only a few markers have shown promising results in recent studies with CA19-9 being the most widely investigated and evaluated single marker

In patients with gastric cancer there is mounting evidence of the role of HER2 overexpression since it has been correlated to poor outcome and more aggressive disease. Furthermore, HER 2 overexpression was found to be predictive for treatment of gastric cancer patients with trastuzumab. Routine HER2 testing is now suggested for all patients

Given the importance of biomarkers in this era of targeted therapies more and especially

Allegra, C. J., Jessup, J. M., Somerfield, M. R., Hamilton, S. R., Hammond, E. H., Hayes, D.

F., McAllister, P. K., Morton, R. F. & Schilsky, R. L. (2009). American Society of Clinical Oncology provisional clinical opinion: testing for KRAS gene mutations in

**5. General conclusion** 

Colorectal cancer KRAS

Pancreatic cancer CA19-9

Gastric cancer HER2

EGFR moAb.

summarized.

(Bünger et al., 2011).

with advanced disease.

**6. References** 

prospective randomized trials are necessary.

584 patients received treatment with trastuzumab plus chemotherapy (5FU or capecitabine and cisplatin) for six cycles or chemotherapy alone (Lorenzen & Lordick, 2011). The primary objective was to compare overall survival in both treatment arms, and the secondary objectives were to compare progression-free survival, time to progression, overall response rate, control disease, duration of response, and quality of life (Gravalos & Jimeno, 2008). The primary endpoint of the study was met: trastuzumab significantly improved overall survival by nearly 3 months (median 11.1 vs 13.8 months) (Lorenzen & Lordick, 2011). In addition, an exploratory post-hoc analysis showed that trastuzumab plus chemotherapy substantially improved overall survival in patients with high expression of HER2 protein (IHC2+/FISH+ or IHC3+, 16 months) compared with patients with low expression of HER2 protein (IHC0 or 1+ and FISH+) (Bang et al., 2010). The secondary endpoints also showed significant improvements when trastuzumab was added to chemotherapy. The addition of trastuzumab to chemotherapy did not increase toxic effects associated with standard fluoropyrimidine-based and platinum-based chemotherapy and therefore trastuzumab can be combined with standard chemotherapy without affecting the overall safety profile. On the basis of these findings, trastuzumab can be considered as a new standard option for patients with HER2-positive advanced gastric or gastro-oesophageal junction cancer when combined with a chemotherapy regimen consisting of capecitabine plus cisplatin or fluorouracil plus cisplatin (Bang et al., 2010).

On the basis of this evidence, in January 2010 the EMEA and on 20 October 2010, the US FDA granted approval for trastuzumab in combination with cisplatin and fluoropyrimidine (either capecitabine or 5FU), for the treatment of metastatic HER-2 positive gastric or gastrooesophageal junction adenocarcinoma who have not received prior treatment for metastatic disease (Lorenzen & Lordick, 2011).

Further studies are necessary to investigate the role of trastuzumab in curative gastric cancer treatment, as well its role as monotherapy, maintenance therapy and second line treatment in the palliative setting. Furthermore, additional predictive markers are needed besides a HER2-positive status. In addition, there is an urgent need to improve the knowledge of the mechanisms involved in anti-HER2 sensitivity or resistance, in order to develop other rationally targeted agents in the near future (Fornaro et al., 2011).

#### **4.5 Conclusion**

Since there is no internationally accepted standard of care for gastric or gastro-esophageal cancer patients and survival remains poor, new therapeutic strategies are needed. There is mounting evidence of the role of HER2 overexpression in patients with gastric cancer. HER2 overexpression has been correlated to poor outcome and more aggressive disease. Furthermore, the positive results of the randomized phase III ToGA trial have opened up new frontiers. Trastuzumab not only represents a new and effective therapeutic option, but has also stimulated the search for predictive marker in order to refine patient selection (Fornaro et al., 2011). Trastuzumab represents a new reference treatment for patients with HER2-positive metastatic gastric or gastro-esophageal cancer. Routine HER2 testing is suggested for all patients with advanced disease. Other agents directed against members of the HER family (like lapatinib) are currently under investigation (Lorenzen & Lordick, 2011).
