Resident Short Review. HER2/neu Gene Amplification and Protein Overexpression in Gastric and Gastroesophageal Junction Adenocarcinoma

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1 Resident Short Review HER2/neu Gene Amplification and Protein Overexpression in Gastric and Gastroesophageal Junction Adenocarcinoma A Review of Histopathology, Diagnostic Testing, and Clinical Implications N Amplification of the human epidermal growth factor receptor 2/neu (HER2/neu) gene and overexpression of the HER2 protein (HER2) have been shown to occur in gastric and gastroesophageal junction adenocarcinoma in a number of studies. With a dismal survival rate, patients with these cancers stand to benefit from the identification of possible molecular targets such as HER2 for both prognostic and therapeutic purposes. Although these and other carcinomas that overexpress HER2 may have a poorer prognosis and exhibit resistance to conventional chemotherapy, they have also recently been shown to respond to targeted therapy with the anti-her2 antibody trastuzumab. Here, we briefly review the molecular biology, histopathology, diagnostic techniques, and interpretation, as well as the clinical implications, of HER2 amplification/overexpression in gastric and gastroesophageal adenocarcinoma. (Arch Pathol Lab Med. 2012;136: ; doi: / arpa rs) Amplification of the human epidermal growth factor receptor 2 (HER2/neu) gene and/or overexpression of the HER2 protein has been observed in many human carcinomas, including breast, colon, endometrial, cervical, urothelial, lung adenocarcinoma, ovarian, and, more recently, gastric and gastroesophageal junction (GEJ) adenocarcinomas (GGEACs). 1 3 Many of these tumors with amplification of the HER2 gene tend to have a poorer prognosis and invite different, targeted therapy approaches. 4 Gastric and GEJ adenocarcinomas are major global health concerns, with combined yearly incidence of 1.4 million new diagnoses per year and 5-year survival rate less than 20%. Unfortunately, conventional chemotherapeutic agents such as 5-fluorouracil, cisplatin, epirubicin, and docetaxel have been of limited success, and efforts have been focused in identifying better molecular targets Accepted for publication July 7, From the Department of Pathology, Mount Sinai School of Medicine, New York, New York. The authors have no relevant financial interest in the products or companies described in this article. Reprints: Jaclyn F. Hechtman, MD, Department of Pathology, Mount Sinai School of Medicine, One Gustave L Levy Place, Box 1194, New York, NY ( jaclyn.hechtman@mountsinai.org). Jaclyn F. Hechtman, MD; Alexandros D. Polydorides, MD, PhD for treatment. 3 HER2/neu gene amplification and protein overexpression in GGEAC were first described in ,6 and were confirmed in subsequent studies. Gastroesophageal junction carcinomas possess a 1.8 to 2.6 times higher prevalence of HER2 gene amplification (24% 32%) compared to gastric carcinomas (9.5% 18%). 7 9 The reported rate of HER2 protein overexpression in GEJ carcinoma ranges from 2% to 45%, 10 and that of gastric carcinomas ranges from 8.2% to 53.4%. 3,11 In addition to tumor site, this variance in reported prevalence of HER2/neu gene amplification and protein overexpression may be due to sample size, population diversity, interobserver variability, specimen processing, tumor sampling, as well as differences in methodology. MOLECULAR BIOLOGY The HER2 proto-oncogene is located on chromosome 17q21 and encodes a 185-kDa transmembrane tyrosine kinase receptor, HER2 (also known as HER2/neu, ERBB2, p185). This protein is a member of the epidermal growth factor receptor superfamily, which, when activated by ligand binding, dimerizes and regulates intracellular signal transduction through the mitogen-activated protein kinase and phosphatidylinositol 3-kinase pathways. 1,12 The final target of these pathways is the regulation of gene expression for various proteins that play roles in a multitude of cellular processes such as differentiation, proliferation, and survival. HER2 is unique because, unlike other members of the epidermal growth factor receptor family, it has no known direct ligand. This receptor may thereby be capable of constitutive activity, producing its biologic response constantly and without the need for a bound, activating ligand. This ligandindependent activity of HER2 suggests that its overexpression may be capable of independently inducing malignant transformation and driving tumor growth through increased cell survival and proliferation, even in the absence of growth factor ligand(s). 13 HISTOPATHOLOGY Gastric carcinomas of the antrum, body, and fundus are divided into 2 histologic subtypes according to the Lauren classification: intestinal and diffuse. Carcinomas of the GEJ may be of either esophageal or gastric cardiac origin, Arch Pathol Lab Med Vol 136, June 2012 HER2 Overexpression in Gastric Carcinoma Hechtman & Polydorides 691

2 Interpretation of HER2 Immunohistochemical Staining in Gastric and Gastroesophageal Junction Carcinoma (GGEAC) in Comparison to Breast Carcinoma a Score Interpretation Criteria for GGEAC Biopsy Specimens 0 Negative No staining or membrane staining in clusters of,5 tumor cells 1+ Negative Cluster(s) of at least 5 cohesive tumor cells with weak (generally visible only at 3400 magnification) complete, basolateral, or lateral membrane staining, irrespective of tumor volume percentage 2+ Equivocal Cluster(s) of at least 5 cohesive tumor cells with moderate (generally visible at magnification) complete, basolateral, or lateral membrane staining, irrespective of tumor volume percentage 3+ Positive Cluster(s) of at least 5 cohesive tumor cells with strong (generally visible at magnification) complete, basolateral, or lateral membrane staining, irrespective of tumor volume percentage a Data derived from Ruschoff et al, 19 Wolff et al, 22 and Hofmann et al. 23 Criteria for GGEAC Resection Specimens No staining or membrane staining in,10% of tumor cells Weak (generally visible only at 3400 magnification) complete, basolateral, or lateral membrane staining in.10% of tumor cells Moderate (generally visible at magnification) complete, basolateral, or lateral membrane staining in.10% of tumor cells Strong (generally visible at magnification) complete, basolateral, or lateral membrane staining in.10% of tumor cells Criteria for Breast Carcinoma Specimens No membranous reactivity in tumor cells Noncircumferential, weak membranous reactivity in any percentage of tumor cells Heterogenous or weak (with circumferential distribution in $10% of cells) membranous reactivity; or strong and circumferential membranous reactivity in #30% of tumor cells Homogeneous, strong, and circumferential membranous reactivity in.30% of tumor cells and the most common clinical association is gastroesophageal reflux, which contrasts with the dietary and infectious (Helicobacter pylori) associations of distal gastric carcinomas. 12,14,15 Recent studies report that the intestinal type of gastric carcinoma has higher prevalence of HER2 overexpression (16% 34%) compared to the diffuse type (2% 7%). 1,4,9,16 Mixed intestinal-diffuse type cancers have HER2 overexpression rates similar to the intestinal type and, generally, overexpression in these tumors is usually limited to areas of intestinal morphology. 4 This trend of intestinal-type histomorphology carrying higher rates of HER2 overexpression is also true for GEJ carcinomas. 15 Rarer histologic types of gastric adenocarcinomas require further study, as reports place the prevalence of HER2 overexpression anywhere from 0 to 100%. 17,18 Yano et al studied a group of 6 papillary and 193 tubular carcinomas of the stomach and showed that all papillary carcinomas (100%) were positive for HER2 overexpression compared to only 20.6% of tubular carcinomas. 17 More recently, 11 cases of the newly described micropapillary variant of gastric carcinoma were evaluated and were all found to lack HER2 overexpression. 18 Even though these studies represent case series with a relatively small sample size, nevertheless the variability among histologic subtypes supports the hypothesis that separate genetic alterations belie different phenotypes in terms of HER2 overexpression, and there is therefore clinicopathologic utility in studying more rare morphologic types. DIAGNOSTIC TECHNIQUES AND INTERPRETATION Immunohistochemistry HER2 overexpression is most often evaluated by immunohistochemistry and various reagents are available, including HercepTest (Dako Denmark A/S, Glostrup, Denmark), SP3 (Ventana Medical Systems, Tucson, Arizona), and 4B5 (Labvision, Thermo Fisher Scientific, Fremont, California) antibodies. HercepTest is a US Food and Drug Administration approved polyclonal antibody that was introduced prior to the newer 4B5 and SP3 rabbit monoclonal antibodies. The latter 2 are purported to have lower background staining and higher avidity, as well as excellent correlation with in situ hybridization studies. 19,20 The 4B5 antibody is a US Food and Drug Administration approved antibody directed against the extracellular domain of the HER2 receptor. The SP3 antibody is directed against the intracellular domain, resulting in clearer staining yet possibly lower sensitivity. 21 The HercepTest scoring system was originally developed for the detection of HER2 protein overexpression in breast carcinoma, and it is the most commonly referenced test in the literature. This scoring system evaluates membranous immunostaining of tumor cells using criteria that include intensity of staining, extent of staining around the cell membrane (complete versus incomplete, including lateral and basolateral staining), and the percentage of immunoreactive tumor cells. Scores range from 0 (absent immunostaining) to 3+ (strong immunostaining). The American Society of Clinical Oncology/College of American Pathologists 2007 guidelines for HER2 immunohistochemical interpretation in breast carcinoma 22 form the basis for the interpretation of HER2 immunohistochemistry in GGEAC (Table). However, 2 major differences in HER2 staining patterns between breast carcinoma and GGEAC require modification of immunohistochemical interpretation criteria. First, incomplete membrane staining in a basolateral (or U-shaped) pattern is frequently observed in GGEAC. 23 It has been theorized that this basolateral pattern of membranous staining is due to the absence of growth factor receptors on the luminal surface of the cell. 24 Second, tumor heterogeneity with cases having areas of moderate or strong immunostaining was observed in 4.8% of GGEAC compared to 1.4% of breast carcinomas. 23 Although heterogeneity can sometimes be seen within a single gland, up to a third of gastric carcinomas are of the mixed intestinal-diffuse type, with 692 Arch Pathol Lab Med Vol 136, June 2012 HER2 Overexpression in Gastric Carcinoma Hechtman & Polydorides

3 Figure 1. HER2 immunoreactivity patterns in gastric and gastroesophageal junction adenocarcinoma. A, Gastroesophageal resection specimen with faint, barely perceptible (1+) basolateral or U-shaped (arrows) membranous immunoreactivity for HER2, visible only at high magnification (immunohistochemistry: 4B5 rabbit monoclonal antibody, Ventana Medical Systems, Inc, Tucson, Arizona). B, Gastric biopsy specimen with stronger membranous HER2 immunoreactivity (2+) in cohesive cell clusters of 5 or more tumor cells. C, Gastroesophageal junction adenocarcinoma with strong (3+) HER2 immunoreactivity, with inset showing strong basolateral membranous staining pattern. D, Gastric intratumoral heterogeneity for HER2 staining with absent (asterisk), 1+ (1 arrow), 2+ (2 arrows), and 3+ (3 arrows) areas. E, Nontumor areas showing artifactual cytoplasmic and nuclear HER2 staining. F, Positive fluorescence in situ hybridization with HER2 (red signal) to centromere 17 (green signal) amplification ratio calculated as 11.1 (original magnifications 3400 [A, C inset, and E], 3200 [B] and [D], 340 [C], and [F]). PathVysion probes by Vysis, Abbott Laboratories, Abbott Park, Illinois; photo F courtesy of Vesna Najfeld, PhD, Tumor Cytogenetics, Mount Sinai School of Medicine, New York, New York. Arch Pathol Lab Med Vol 136, June 2012 HER2 Overexpression in Gastric Carcinoma Hechtman & Polydorides 693

4 only the areas of intestinal morphology having positive staining. 24 In contrast, diffuse-type carcinomas are much less common in tumors of the GEJ. 14 Gastric and GEJ adenocarcinoma samples, if interpreted using the criteria for HER2 overexpression developed for breast carcinoma, would be interpreted as negative in both of these instances and would have led to the disqualification of patients who may have otherwise benefited from anti-her2 therapy. 19 Furthermore, Barros-Silva et al 25 and Bang et al 26 evaluated a series of gastric carcinomas using the HER2 scoring criteria for breast carcinoma and found a significantly lower percentage of cases meeting criteria for a positive result by immunohistochemistry (5.4% versus 11% in the Trastuzumab for Gastric Cancer trial). In 2008, Hofmann et al 23 modified the breast carcinoma criteria in a study that evaluated GGEAC cases for HER2 overexpression by immunohistochemistry compared to fluorescence in situ hybridization (FISH). The modifications were the following (Table): in resection specimens, staining limited to the basolateral membrane, or U-shaped (which would have been considered incomplete staining and thus negative in breast carcinoma), in $10% oftumorcellsisto be considered immunoreactive and further scored 1+ to 3+ according to its intensity (Figure 1, A through C) and interpreted in terms of HER2 overexpression as above; in biopsy specimens, cohesive clusters of at least 5 immunopositive tumor cells (Figure 1, D) replace the resection specimen criteria of $10% of tumor cells, because of the small tissue sample size in biopsies. 23 These modified criteria have been subsequently validated for gastric carcinoma by Rüschoff et al, 19 who studied HercepTest and 4B5 in a group of 547 gastric carcinoma cores using tissue microarrays. The antibody 4B5 tended to have higher sensitivity than HercepTest for the detection of tumors positive for HER2 overexpression without an increased rate of false positives. They also validated these guidelines in terms of interlaboratory and interobserver agreement, adding the following recommendations: the grading of membranous staining intensity should take into account the degree of magnification necessary for visible staining (ie, strong staining should only require 325 to 350 magnification, and any tumor staining requiring 3400 magnification should be graded as weak). These magnification rule recommendations resulted in less interobserver variability and have been incorporated into the Belgian guidelines for HER2 testing in gastric carcinoma. 19,24 The Belgian guidelines also included a statement that equivocally staining cases should require 3100 to 3200 magnification. 24 Attempting to achieve a balance between interobserver reproducibility and sensitivity, Ruschoff et al 19 recommended that a cluster of 5 or more positively staining tumor cells be the minimum required for the interpretation of an immunohistochemical result as positive. No specific recommendations regarding FISH interpretation on such limited tumor samples have been put forth. The current American Society of Clinical Oncology/College of American Pathologists guidelines recommend scoring at least 20 cells for FISH. It may, therefore, be prudent to document cases where HER2 status is based on the scoring of a limited number of tumor cells and retest these cases if additional material becomes available. The concept of intratumoral heterogeneity has been addressed in a number of studies. For example, Boers et al claimed that HER2 overexpression is a late event in GEJ and distal gastric carcinomas because of its observed intratumoral heterogeneity. 27 Marx et al 2 have reported homogeneous HER2 overexpression within tumors arising in the gastric cardia, corpus, and antrum, but there was no correlation between intratumoral homogeneity and tumor localization in the stomach. Intratumoral homogeneity of HER2 staining may suggest that overexpression occurs as an early event in tumor progression. In addition, Haas et al 28 studied a group of 52 GEJ carcinomas and found no correlation between HER2 overexpression/amplification and tumor stage. Significantly, the pattern of immunoreactivity may depend on the antibody used. Although the monoclonal antibodies SP3 and 4B5 have increased sensitivity (75% and 96%, respectively), cystoplasmic and nuclear background staining (Figure 1, E) has been observed with high frequency in benign gastric foveolar epithelium, reactive epithelial cells near ulcers, and areas of intestinal metaplasia, and within areas of carcinoma. 19,27 Only membranous staining in tumor cells should be considered positive. Nuclear and/or cytoplasmic staining in any cell as well as any staining in benign cells should be interpreted as negative. 19,27 Fluorescence and Chromogenic In Situ Hybridization Fluorescence in situ hybridization technology uses fluorescent-labeled DNA probes that are highly specific against the target region (ie, gene) of a particular chromosome. The same chromosome is tagged with a contrasting probe against the centromere or other location standard and then observed under fluorescence microscopy. Thus, the ratio of the number of copies of the target gene (eg, Her2/neu) to the total number of chromosomes is recorded per tumor cell, and an average amplification ratio is calculated from measurements in 20 nuclei. 20,22 The result is considered positive (Figure 1, F), negative, or equivocal according to established cutoffs in these ratios. In comparison to immunohistochemistry, FISH is more reproducible, but also more labor-intensive and timeconsuming. In addition to these technical issues, results of the Trastuzumab for Gastric Cancer trial revealed that patients with GGEAC positive for HER2 amplification yet negative for HER2 protein overexpression by immunohistochemistry did not benefit as much from trastuzumab therapy as patients with tumors equivocal or positive for HER2 protein overexpression. Therefore, using FISH as the only (or first) HER2 expression assay may lead to the treatment of a large number of would-be nonresponders and also exclude rare cases that are immunohistochemistry positive but FISH negative. 26,29 FISH is commonly used as a method for verifying results in cases designated as equivocal (moderate staining) by immunohistochemistry. As with immunohistochemical methods, FISH studies may have equivocal results. In the American Society of Clinical Oncology/College of American Pathologists 2007 guidelines for HER2 testing in breast carcinoma, HER2/ centromeric probe for chromosome 17 (CEP 17) ratios between 1.8 and 2.2 (or average gene copy numbers between 4.0 and 6.0 in systems without an internal control probe) were considered equivocal. 22 The Trastuzumab for Gastric Cancer trial, 26 as well as Yano et al, 17 has eschewed the equivocal category in FISH testing for gastric carcinoma and GGEAC respectively, because the significance of these ratios has not been independently studied. Instead, a HER2:CEP17 ratio of $2.0 is considered positive 694 Arch Pathol Lab Med Vol 136, June 2012 HER2 Overexpression in Gastric Carcinoma Hechtman & Polydorides

5 Figure 2. Diagnostic algorithm for the evaluation of HER2 protein overexpression and gene amplification in gastric and gastroesophageal adenocarcinomas. The first step involves interpretation of immunohistochemistry results (see Table). Scores of 0 or 1+ are considered negative for HER2 overexpression, whereas tumors with 3+ immunostaining are deemed positive for overexpression and are therefore eligible for anti-her2 therapy. Specimens with a HER2 immunostaining score of 2+ are equivocal and require further evaluation by fluorescence in situ hybridization (FISH). Gene amplification by FISH is expressed by the ratio of HER2 signal to the centromeric probe signal (HER2:- CEP17): a ratio of,2 is considered negative for HER2 gene amplification whereas a HER2:CEP17 ratio $2 is a positive FISH result. Cases that are immunohistochemistry 2+ and FISH positive are also eligible for anti- HER2 therapy. for Her2/neu gene amplification in GGEAC, whereas a ratio of,2.0 is considered negative. Similar techniques using alternative chromophore, such as chromogenic in situ hybridization (CISH) and silver in situ hybridization, are less commonly used and less extensively studied than FISH. Chromogenic in situ hybridization was originally introduced as an alternative to FISH for HER2 amplification detection in breast cancer in It uses an immunoperoxidase reaction to detect specific DNA probes. This makes visualization by conventional bright field microscopy possible. Large peroxidase-positive intranuclear clusters or.10 individual small signals in.50% of tumor cells (at least 20 cells counted) indicate positive CISH for HER2 amplification. CISH benefits include increased ease of use for pathologists not trained in fluorescence microscopy as well as decreased cost. 30 Perfect concordance between CISH and FISH has been reported in gastric carcinoma. 4 However, CISH or silver in situ hybridization may also facilitate the identification of tumor cells during scoring, owing to the partial preservation of tumor architecture compared to FISH. Validation and Concordance A small internal revalidation of both immunohistochemical and in situ hybridization assays for GGEAC is currently required according to the College of American Pathologists. 31 because although assay conditions need not vary between breast carcinoma and GGEAC, interpretation criteria do. The immunohistochemistry laboratory director should decide on the number of cases included in the revalidation, and 90% to 95% concordance for both positive and negative HER2 scores should be achieved. Ongoing validation should be performed biannually. 31 In addition, Ruschoff et al 19 have recommended the use of only US Food and Drug Administration approved immunohistochemical assays. The rate of agreement between immunohistochemical measurement of HER2 protein overexpression and in situ hybridization based analysis of Her2/neu gene amplification has gathered much interest. Studies in breast cancer reveal that approximately 95% of HER2 overexpression is due to increased gene copy number, or amplification. 32 However, such strong correlation has not been observed in gastric carcinoma. Difficulties of immunohistochemical methods, including varying fixation protocols, selected antibodies, and interobserver variability, mandate confirmatory FISH testing in equivocal cases. 3,19 With the criteria for HER2 overexpression in gastric carcinoma relatively recently published, studies have set out to assess concordance rates. Marx et al 2 studied HercepTest in gastric cardia and noncardia tumors using the modified Arch Pathol Lab Med Vol 136, June 2012 HER2 Overexpression in Gastric Carcinoma Hechtman & Polydorides 695

6 scoring criteria by Hofmann et al 23 and found that all (22 of 22) cases that showed strong (3+) staining for HER2 by immunohistochemistry and 5 of 6 cases with moderate staining were also positive for HER2 amplification by FISH, whereas none (0 of 138) of the cases with weak to absent staining were positive for gene amplification. Because GGEACs with immunohistochemical scores of 2+ tend to have varying gene amplification results, these equivocal cases should be further analyzed by FISH, in order to identify patients with the greatest likelihood of responding to anti-her2 therapy (see diagnostic algorithm in Figure 2). CLINICAL IMPLICATIONS Similar to other cancers, 4 GGEACs with HER2 overexpression/amplification tend to have a poorer prognosis. HER2 overexpression has been associated with shorter overall survival intervals in gastric carcinoma by univariate analysis: 6.6 months in HER2 positive gastric carcinomas compared to 12.7 months in those without HER2 overexpression amplification. 9 This was also observed in resected GEJ carcinomas by univariate analysis, but only when tumor stage was not taken into account. 15 HER2 staining intensity has also been correlated with tumor size, serosal invasion, and lymph node metastases. 33 Of note, in these studies, most HER2 overexpression was observed in late-stage tumors, but the prevalence and prognostic impact of HER2 overexpression/amplification as well as the role of anti-her2 therapy in less advanced (ie, intramucosal) GGEAC has yet to be thoroughly investigated. Although HER2 status may have prognostic implications in GGEAC, the more important clinical implication concerns the potential impact of HER2 status on the therapeutic management of patients with these tumors. Trastuzumab, an injectable monoclonal antibody that targets HER2 and induces antibody-dependent cellular toxicity, was recently investigated in a landmark clinical trial, the Trastuzumab for Gastric Cancer study. 26 The results showed a 2.7-month increase in median overall survival and a 1.2-month increase in disease-free interval for gastric carcinoma patients receiving trastuzumab, compared to those who did not. The relation between the extent of protein overexpression and the degree of clinical response to trastuzumab was also investigated in this trial by evaluating the intensity of HER2 protein immunostaining. Bang et al 26 found that when GGEAC cases positive for HER2 amplification were subdivided according to immunohistochemical staining intensity, those with strong staining (2+ and 3+) had a larger increase in median survival (4.2 versus 1.3 months) compared to those with absent to weak immunostaining (0 and 1+). It is apparent that the detection and quantitation of HER2 protein overexpression and gene amplification has become an essential component during the diagnosis of GGEAC in order to predict response to existing and forthcoming anti-her2 agents, which are now regarded as standard of care for HER2-positive GGEAC. The differences in HER2 overexpression observed between primary and metastatic carcinoma samples raise some concerns about the clinical utility of trastuzumab. This heterogeneity is a recognized cause of treatment failure after resection of the primary tumor. However, Reichelt et al 34 found 100% concordance in HER2 overexpression between 84 pairs of primary and metastatic esophageal adenocarcinoma. Marx et al 2 found similarly high concordance rates between HER2 overexpression at the primary site and metastases in cases of both cardiac and distal gastric carcinoma. The extent as well as the mechanisms conferring resistance to trastuzumab therapy in GGEAC remain to be investigated. 12 Nevertheless, combination therapy with newly developed drugs has shown some promise. Recently, lapatinib has been investigated as a therapeutic option for tumors with HER2 overexpression. It is a small molecule that targets and inhibits the tyrosine kinase domain of both epidermal growth factor receptor and HER2 and leads to apoptosis and decreased cell proliferation. Wainberg et al 35 investigated lapatinib in tumor cell lines and mouse tumor xenografts of HER2-amplified gastric carcinoma and found it to be of therapeutic value, both by itself and in combination with trastuzumab in synergistic fashion. Clinical trials with this drug are currently under way and the effect in HER2 overexpressing human GGEAC (both trastuzumab-sensitive and trastuzumab-resistant) remains to be seen. CONCLUSIONS Up to a fifth of gastric carcinomas and up to a third of GEJ carcinomas, primarily of intestinal morphology, are positive for HER2 overexpression amplification. 7 9 Differences exist in the interpretation of HER2 amplification and protein overexpression in GGEAC compared to breast carcinoma. Current recommendations for the evaluation of HER2 overexpression include the use of immunohistochemical methods initially on resection or biopsy specimens with cases having equivocal results further analyzed for Her2/neu gene amplification by in situ hybridization. 29,36 Targeted therapy against the HER2 receptor has been shown in clinical trials to increase the overall survival of patients with advanced gastric and GEJ carcinomas by 4.2 and 1.3 months in cases of high and low levels of HER2 overexpression, respectively. 26 Therefore, HER2 overexpression/amplification should be evaluated in advanced cases of GGEAC in order to assess patient eligibility for anti-her2 therapy. The authors would like to thank Ira Bleiweiss, MD, and Vesna Najfeld, PhD, from the Department of Pathology and the Division of Tumor Cytogenics, respectively, at the Mount Sinai School of Medicine. References 1. Gravalos C, Jimeno A. HER2 in gastric cancer: a new prognostic factor and a novel therapeutic target. Ann Oncol. 2008;19(9): Marx AH, Tharun L, Muth J, et al. HER-2 amplification is highly homogenous in gastric cancer. Hum Pathol. 2009;40(6): Jørgensen JT. Targeted HER2 treatment in advanced gastric cancer. Oncology. 2010;78(1): Yan B, Yau EX, Bte Omar SS, et al. A study of HER2 gene amplification and protein expression in gastric cancer. J Clin Pathol. 2010;63(9): Fukushige S, Matsubara K, Yoshida M, et al. Localization of a novel c-erbbrelated gene, c-erbb-2, on human chromosome 17 and its amplification in a gastric cancer cell line. Mol Cell Biol. 1986;6(3): Sakai K, Mori S, Kawamoto T, et al. 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Published Accessed May 9, Arch Pathol Lab Med Vol 136, June 2012 HER2 Overexpression in Gastric Carcinoma Hechtman & Polydorides 697