REVIEW ARTICLE
HER2 Testing in Gastric and Gastroesophageal Adenocarcinomas Efsevia Vakiani, MD, PhD
Abstract: The human epidermal growth factor receptor 2 (HER2) is overexpressed in 10% to 35% of gastric and gastroesophageal junction (GEJ) adenocarcinomas. In 2010, the phase III Trastuzumab for Gastric Cancer (ToGA) trial showed that addition of the anti-HER2 monoclonal antibody trastuzumab to chemotherapy significantly improved survival of patients with advanced or metastatic tumors that were positive for HER2 overexpression. As a result, HER2 testing is now recommended for all patients with advanced or metastatic disease, although there is still some debate as to the optimal methods of assessment. HER2 expression in gastric and GEJ tumors shows several differences compared with breast tumors and, for this reason, the proposed criteria for scoring HER2 expression in biopsies and resections of gastric and GEJ carcinomas differ from those used in breast carcinomas. This review discusses what is currently known about the patterns of HER2 expression in gastric and GEJ adenocarcinomas, summarizes the findings of the ToGA trial and its clinical implications, and provides an overview of the recommended guidelines for the most accurate evaluation of HER2 status in gastric and GEJ cancer. Key Words: HER2, gastric cancer, gastroesophageal cancer, immunohistochemistry, in situ hybridization, ToGA trial
(Adv Anat Pathol 2015;22:194–201)
HER2 EXPRESSION IN GASTRIC AND GASTROESOPHAGEAL ADENOCARCINOMAS The human epidermal growth factor receptor 2 (HER2, HER2/neu, ERBB2) is a protooncogene located on chromosome 17q21, which encodes for a 185 kD transmembrane tyrosine kinase receptor protein. It belongs to the human epidermal growth factor receptor superfamily, which is composed of 4 members sharing the same basic molecular structure: HER1 (more commonly known as EGFR), HER2, HER3, and HER4. The binding of a ligand to the extracellular domain triggers homodimerization or heterodimerization, phosphorylation of the receptor, and initiation of signal transduction cascades that affect a variety of cellular processes including apoptosis, proliferation, and differentiation.1,2 In contrast to the other family members, HER2 does not bind to a known ligand. HER2 is overexpressed in a number of different cancer types, where it promotes tumorigenesis, including breast, stomach, esophagus, colon, bladder, and ovary.3–8 Amplification of the HER2 gene and overexpression of the HER2 protein was first described in gastric cancer in 1986 and these findings have been confirmed in multiple studies since that From the Department of Pathology, Memorial Sloan-Kettering Cancer Center, New York, NY. The author has no NIH funding or conflicts of interest to disclose. Reprints: Efsevia Vakiani, MD, PhD, Department of Pathology, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY 10065 (e-mail: [email protected]). Copyright r 2015 Wolters Kluwer Health, Inc. All rights reserved.
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time.9–11 The reported frequency of protein overexpression by immunohistochemistry (IHC) varies significantly among studies ranging from 8.2% in one Japanese study to 53.4% in one German study.12,13 This high variation is likely secondary to different methodologies used, differences among ethnic groups, as well as different tumor histologies and locations among various cohorts. Results of studies using in situ hybridization (ISH) to look for HER2 gene amplification have been somewhat more consistent with amplification being reported in 10% to 27% of cases.11 Studies from the United States using both IHC and ISH have reported rates of HER2 overexpression that are on the lower end of the spectrum. For example, Kunz et al14 in their study of 169 primary gastric and gastroesophageal (GEJ) adenocarcinomas using tissue microarrays reported a HER2 positivity rate of 11.2%, whereas Tafe et al15 in their study of 135 primary gastric and GEJ adenocarcinomas using mostly biopsy material found a HER2 positivity rate of 15%. HER2 overexpression is significantly more frequent in intestinal-type adenocarcinomas compared with diffuse-type adenocarcinomas, a finding that seems to be consistent among studies. In one Finnish study of 231 gastric and GEJ tumors, HER2 overexpression was seen in 21.5% of intestinal-type adenocarcinomas compared with 2% of diffuse-type adenocarcinomas and 5% of mixed-type adenocarcinomas.6 An Australian study of 178 tumors reported HER2 positivity in 30% of intestinal-type tumors compared with 3.8% of diffusetype tumors and 9% of mixed-type tumors, whereas the study by Tafe et al15 found positivity in 22.1% of intestinal-type adenocarcinomas compared with 2.7% of diffuse-type adenocarcinomas.16 HER2 expression in unusual histologic types requires further study as there are only a few reports in the literature, sometimes with conflicting results.17–19 For example, Yano et al19 reported that all 6 tested papillary gastric adenocarcinomas showed strong HER2 overexpression, whereas in another study all 5 papillary adenocarcinomas were HER2 negative.17 HER2 overexpression appears to be more frequent among GEJ tumors compared with gastric tumors, although the difference has not been statistically significant in all studies and it is not known whether this may simply be a reflection of the fact that many gastric adenocarcinomas are diffuse type. HER2 overexpression in gastric/GEJ adenocarcinomas tends to show more heterogeneity compared with what is observed in breast adenocarcinomas. The frequency of heterogeneity ranges among studies from 5% to 50%. This high variability is likely due to differences in the definition of heterogeneity. Hofmann et al20 reported heterogeneity in 4.8% of cases referring to cases that showed strong positivity in 10% of tumor cells; or strong and circumferential membranous staining within r10% of tumor cells Circumferential membranous staining that is complete, intense, and within >10% of tumor cells
Interpretation Negative
Negative
Equivocal
Positive
GEJ indicates gastroesophageal junction.
carcinoma43 in 2 important ways. First, gastric/GEJ tumor cells may only show HER2 staining at the basolateral or lateral membrane regions. For this reason, complete membranous staining which is a requirement in breast cancer is not required in gastric/GEJ cancer. Second, due to the presence of heterogeneity, it was felt that the 10% cutoff for the number of reactive cells would be retained for surgical specimens but would not be appropriate for biopsies. Hofmann and colleagues initially suggested that in biopsies the pattern of reactivity should be scored as positive irrespective of the relative number of IHC-positive cells. Subsequently, Ruschoff and colleagues found that interobserver disagreement increased when few (< 5) cells were evaluated on biopsies as such small cell groups tend to show nonspecific staining. They, therefore, proposed that the cutoff of immunoreactive cells on biopsies should be 1 tumor cell cluster, the latter being defined as at least 5 cohesive cells. Distinction between 1 + , 2 + , and 3 + IHC scores is based on the HER2 staining intensity. Faint or barely perceptible staining corresponds to 1 + staining (Fig. 1A), weak to moderate staining should be scored as 2 + (Fig. 1B), and moderate to strong staining is 3 + (Fig. 1C). In an attempt to make intensity scoring more reproducible and increase interobserver agreement, Ruschoff and colleagues suggested the magnification rule. According to this rule, if one can visualize unequivocal membranous staining at low power ( 2.5, 5), then staining is considered “strong” and the case can be scored as 3 + if staining is present in 10% of cells (surgical specimen) or in a tumor cell cluster (biopsy specimen). If the unequivocal membranous staining can only be appreciated at medium magnification (10 to 20), the staining is considered weak to
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moderate and the case should be scored 2 + . Finally, if membranous staining can only be confirmed at high power ( 40), it is considered faint or barely perceptible and the case should be scored as 1 + . Interlaboratory agreement in the IHC scoring of gastric/GEJ tumors is generally good when following these criteria, although there seems to be room for improvement. In a study from Australia looking at the IHC assessment of 100 tissue samples from 9 laboratories, Fox et al37 found an overall agreement of 84% when IHC 0 and 1 + cases were grouped together as negative. In a study from China, agreement in the IHC assessment of 721 samples between 11 local laboratories and a central laboratory was 86.7%.60 In the latter study, discordant results were attributed to misinterpretation of staining results due to unfamiliarity with scoring criteria and potential pitfalls. In a recent study from North America, Sheffield and colleagues reported excellent agreement among 37 laboratories in IHC 3 + cases and very good agreement in IHC 0 or 1 + cases. Agreement was lower when scoring 2 + cases, nonetheless the overall sensitivity and specificity of HER2 IHC were 99% and 100%, respectively, when measured against expert review and consensus score.61
HER2 IMMUNOHISTOCHEMISTRY PITFALLS AND SPECIAL SITUATIONS Cytoplasmic background staining can be observed in normal or metaplastic mucosa and this is particularly common with the 4B5 antibody (Fig. 1E).48 This is usually not a problem as tumor cells can be readily distinguished from non-neoplastic epithelial cells under the microscope. A more common pitfall in IHC scoring is cytoplasmic and/
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or nuclear staining in tumor cells (Fig. 1F). This should not be misinterpreted as a positive result as only membranous staining should be scored. Tumor cells showing cytoplasmic and/or nuclear staining have not been found to show amplification by ISH,46,48 nonetheless there might be cases where there is concern that cytoplasmic staining might obscure some membranous staining. In these cases, it would be prudent to subject the tumor to analysis by ISH. Assessment of HER2 overexpression should be performed on invasive carcinoma. In some cases, however, particularly biopsies, there might be difficulty in distinguishing morphologically between high-grade dysplasia and invasive carcinoma. HER2 appears to be overexpressed in dysplastic lesions, but there are little data on the concordance between HER2 status in dysplastic lesions and their corresponding invasive adenocarcinomas.16,56,62–64 In a study by Lee et al,16 there was no clear association between the HER2 status of dysplastic and invasive components. The authors of that study encountered both cases where HER2 expression was seen in dysplasia but not in the invasive carcinoma as well as cases where the dysplastic component was HER2 negative, but the invasive component showed overexpression. In a more recent study, concordance between high-grade dysplasia and invasive carcinoma was seen only in 4/9 (44%) cases.56 These findings raise the issue of falsepositive results in biopsies due to HER2-positive dysplastic epithelium being misinterpreted as invasive carcinoma. Differentiating between weak 2 + and 1 + staining by IHC can also be challenging, especially in resection specimens where there might be a range of staining intensities. This can be an important distinction as a score of 1 + is not sent for ISH testing in many institutions. If there is uncertainty, it is recommended that the case be scored as 1 to 2 + and material sent for ISH analysis to prevent a false-negative result. As already mentioned, some cases show strong HER2 staining in 6 HER2 gene copies should be considered a positive result. They suggested that copy number is an important consideration especially in cases showing borderline amplification. In a single-probe assay if there are 4 to 6 HER2 gene copies then the use of a dual probe is recommended. Interestingly, a recent study correlating the level of HER2 amplification and response to trastuzumab identified a HER2/CEP17 ratio of 4.7 as the optimal cutoff value discriminating sensitive and refractory patients.67 Over the last few years, there is growing support for the use of bright-field ISH methods for the detection of HER2 amplification. Such methods have several advantages over FISH, including use of a regular microscope instead of a dark-field fluorescence microscope, better signal preservation over time, and better evaluation of morphology. Bright-field techniques may be particularly useful in the evaluation of gastric/GEJ tumors because of heterogeneity as they allow for alignment of IHC slides with the ISH slides. In this way it is easier to score by ISH specific areas showing focal HER2 overexpression. CISH uses an IHC-like peroxidase reaction to detect DNA probes instead of fluorescence. Several studies have reported very good concordance between FISH and CISH.27,50,55,68 For example, Kiyose et al68 used single-color CISH to study 198 gastric tumors and reported a concordance of 99% between FISH and CISH. Two discordant cases were observed one of which showed polysomy. In another study, Yan et al50 used a dual-color CISH assay in 199 samples of gastric cancer and found a 100% concordance with FISH results. A high level of agreement has also been reported between FISH and SISH, which is another brightfield ISH method that is based upon the deposition of silver at the target site after an enzymatic reaction.46,48,69 In a study of 166 gastric cancer samples, 96.4% of cases showed agreement between FISH and dual-color SISH. Six discordant cases were SISH positive and FISH negative and all showed polysomy.69 It is worth noting that the clinical significance of polysomy in gastric cancer is not known and in breast cancer polysomy has been reported in FISH-negative cases that responded to trastuzumab.70
CONCLUSIONS HER2 testing is now recommended for all patients with advanced gastric/GEJ adenocarcinoma to identify HER2positive tumors that can be treated with trastuzumab. Many guidelines recommend that assessment of HER2 status be performed using first IHC, with ISH analysis being reserved for IHC equivocal cases. HER2 expression in gastric/GEJ adenocarcinomas shows some differences compared with what is observed in breast adenocarcinomas, most notably basolateral or lateral membranous positivity and increased
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heterogeneity. Therefore, the IHC scoring criteria for gastric/ GEJ adenocarcinomas differ somewhat from those used in breast. It is recommended that pathologists familiarize themselves with these IHC scoring criteria and potential pitfalls, such as cytoplasmic or nuclear staining in tumor cells. Although there are no strict guidelines as to how many and which samples should be tested, testing >1 sample from the same patient may be justified, especially if available specimens are limited in tumor content or if there is suspicion of morphologic heterogeneity. A good understanding of the patterns of HER2 expression in gastric/GEJ tumors and scoring guidelines will ensure that HER2 status and eligibility for treatment with trastuzumab is determined as accurately as possible.
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HER2 Testing in Gastric and Gastroesophageal Adenocarcinomas Efsevia Vakiani, MD, PhD
Abstract: The human epidermal growth factor receptor 2 (HER2) is overexpressed in 10% to 35% of gastric and gastroesophageal junction (GEJ) adenocarcinomas. In 2010, the phase III Trastuzumab for Gastric Cancer (ToGA) trial showed that addition of the anti-HER2 monoclonal antibody trastuzumab to chemotherapy significantly improved survival of patients with advanced or metastatic tumors that were positive for HER2 overexpression. As a result, HER2 testing is now recommended for all patients with advanced or metastatic disease, although there is still some debate as to the optimal methods of assessment. HER2 expression in gastric and GEJ tumors shows several differences compared with breast tumors and, for this reason, the proposed criteria for scoring HER2 expression in biopsies and resections of gastric and GEJ carcinomas differ from those used in breast carcinomas. This review discusses what is currently known about the patterns of HER2 expression in gastric and GEJ adenocarcinomas, summarizes the findings of the ToGA trial and its clinical implications, and provides an overview of the recommended guidelines for the most accurate evaluation of HER2 status in gastric and GEJ cancer. Key Words: HER2, gastric cancer, gastroesophageal cancer, immunohistochemistry, in situ hybridization, ToGA trial
(Adv Anat Pathol 2015;22:194–201)
HER2 EXPRESSION IN GASTRIC AND GASTROESOPHAGEAL ADENOCARCINOMAS The human epidermal growth factor receptor 2 (HER2, HER2/neu, ERBB2) is a protooncogene located on chromosome 17q21, which encodes for a 185 kD transmembrane tyrosine kinase receptor protein. It belongs to the human epidermal growth factor receptor superfamily, which is composed of 4 members sharing the same basic molecular structure: HER1 (more commonly known as EGFR), HER2, HER3, and HER4. The binding of a ligand to the extracellular domain triggers homodimerization or heterodimerization, phosphorylation of the receptor, and initiation of signal transduction cascades that affect a variety of cellular processes including apoptosis, proliferation, and differentiation.1,2 In contrast to the other family members, HER2 does not bind to a known ligand. HER2 is overexpressed in a number of different cancer types, where it promotes tumorigenesis, including breast, stomach, esophagus, colon, bladder, and ovary.3–8 Amplification of the HER2 gene and overexpression of the HER2 protein was first described in gastric cancer in 1986 and these findings have been confirmed in multiple studies since that From the Department of Pathology, Memorial Sloan-Kettering Cancer Center, New York, NY. The author has no NIH funding or conflicts of interest to disclose. Reprints: Efsevia Vakiani, MD, PhD, Department of Pathology, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY 10065 (e-mail: [email protected]). Copyright r 2015 Wolters Kluwer Health, Inc. All rights reserved.
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time.9–11 The reported frequency of protein overexpression by immunohistochemistry (IHC) varies significantly among studies ranging from 8.2% in one Japanese study to 53.4% in one German study.12,13 This high variation is likely secondary to different methodologies used, differences among ethnic groups, as well as different tumor histologies and locations among various cohorts. Results of studies using in situ hybridization (ISH) to look for HER2 gene amplification have been somewhat more consistent with amplification being reported in 10% to 27% of cases.11 Studies from the United States using both IHC and ISH have reported rates of HER2 overexpression that are on the lower end of the spectrum. For example, Kunz et al14 in their study of 169 primary gastric and gastroesophageal (GEJ) adenocarcinomas using tissue microarrays reported a HER2 positivity rate of 11.2%, whereas Tafe et al15 in their study of 135 primary gastric and GEJ adenocarcinomas using mostly biopsy material found a HER2 positivity rate of 15%. HER2 overexpression is significantly more frequent in intestinal-type adenocarcinomas compared with diffuse-type adenocarcinomas, a finding that seems to be consistent among studies. In one Finnish study of 231 gastric and GEJ tumors, HER2 overexpression was seen in 21.5% of intestinal-type adenocarcinomas compared with 2% of diffuse-type adenocarcinomas and 5% of mixed-type adenocarcinomas.6 An Australian study of 178 tumors reported HER2 positivity in 30% of intestinal-type tumors compared with 3.8% of diffusetype tumors and 9% of mixed-type tumors, whereas the study by Tafe et al15 found positivity in 22.1% of intestinal-type adenocarcinomas compared with 2.7% of diffuse-type adenocarcinomas.16 HER2 expression in unusual histologic types requires further study as there are only a few reports in the literature, sometimes with conflicting results.17–19 For example, Yano et al19 reported that all 6 tested papillary gastric adenocarcinomas showed strong HER2 overexpression, whereas in another study all 5 papillary adenocarcinomas were HER2 negative.17 HER2 overexpression appears to be more frequent among GEJ tumors compared with gastric tumors, although the difference has not been statistically significant in all studies and it is not known whether this may simply be a reflection of the fact that many gastric adenocarcinomas are diffuse type. HER2 overexpression in gastric/GEJ adenocarcinomas tends to show more heterogeneity compared with what is observed in breast adenocarcinomas. The frequency of heterogeneity ranges among studies from 5% to 50%. This high variability is likely due to differences in the definition of heterogeneity. Hofmann et al20 reported heterogeneity in 4.8% of cases referring to cases that showed strong positivity in 10% of tumor cells; or strong and circumferential membranous staining within r10% of tumor cells Circumferential membranous staining that is complete, intense, and within >10% of tumor cells
Interpretation Negative
Negative
Equivocal
Positive
GEJ indicates gastroesophageal junction.
carcinoma43 in 2 important ways. First, gastric/GEJ tumor cells may only show HER2 staining at the basolateral or lateral membrane regions. For this reason, complete membranous staining which is a requirement in breast cancer is not required in gastric/GEJ cancer. Second, due to the presence of heterogeneity, it was felt that the 10% cutoff for the number of reactive cells would be retained for surgical specimens but would not be appropriate for biopsies. Hofmann and colleagues initially suggested that in biopsies the pattern of reactivity should be scored as positive irrespective of the relative number of IHC-positive cells. Subsequently, Ruschoff and colleagues found that interobserver disagreement increased when few (< 5) cells were evaluated on biopsies as such small cell groups tend to show nonspecific staining. They, therefore, proposed that the cutoff of immunoreactive cells on biopsies should be 1 tumor cell cluster, the latter being defined as at least 5 cohesive cells. Distinction between 1 + , 2 + , and 3 + IHC scores is based on the HER2 staining intensity. Faint or barely perceptible staining corresponds to 1 + staining (Fig. 1A), weak to moderate staining should be scored as 2 + (Fig. 1B), and moderate to strong staining is 3 + (Fig. 1C). In an attempt to make intensity scoring more reproducible and increase interobserver agreement, Ruschoff and colleagues suggested the magnification rule. According to this rule, if one can visualize unequivocal membranous staining at low power ( 2.5, 5), then staining is considered “strong” and the case can be scored as 3 + if staining is present in 10% of cells (surgical specimen) or in a tumor cell cluster (biopsy specimen). If the unequivocal membranous staining can only be appreciated at medium magnification (10 to 20), the staining is considered weak to
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moderate and the case should be scored 2 + . Finally, if membranous staining can only be confirmed at high power ( 40), it is considered faint or barely perceptible and the case should be scored as 1 + . Interlaboratory agreement in the IHC scoring of gastric/GEJ tumors is generally good when following these criteria, although there seems to be room for improvement. In a study from Australia looking at the IHC assessment of 100 tissue samples from 9 laboratories, Fox et al37 found an overall agreement of 84% when IHC 0 and 1 + cases were grouped together as negative. In a study from China, agreement in the IHC assessment of 721 samples between 11 local laboratories and a central laboratory was 86.7%.60 In the latter study, discordant results were attributed to misinterpretation of staining results due to unfamiliarity with scoring criteria and potential pitfalls. In a recent study from North America, Sheffield and colleagues reported excellent agreement among 37 laboratories in IHC 3 + cases and very good agreement in IHC 0 or 1 + cases. Agreement was lower when scoring 2 + cases, nonetheless the overall sensitivity and specificity of HER2 IHC were 99% and 100%, respectively, when measured against expert review and consensus score.61
HER2 IMMUNOHISTOCHEMISTRY PITFALLS AND SPECIAL SITUATIONS Cytoplasmic background staining can be observed in normal or metaplastic mucosa and this is particularly common with the 4B5 antibody (Fig. 1E).48 This is usually not a problem as tumor cells can be readily distinguished from non-neoplastic epithelial cells under the microscope. A more common pitfall in IHC scoring is cytoplasmic and/
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or nuclear staining in tumor cells (Fig. 1F). This should not be misinterpreted as a positive result as only membranous staining should be scored. Tumor cells showing cytoplasmic and/or nuclear staining have not been found to show amplification by ISH,46,48 nonetheless there might be cases where there is concern that cytoplasmic staining might obscure some membranous staining. In these cases, it would be prudent to subject the tumor to analysis by ISH. Assessment of HER2 overexpression should be performed on invasive carcinoma. In some cases, however, particularly biopsies, there might be difficulty in distinguishing morphologically between high-grade dysplasia and invasive carcinoma. HER2 appears to be overexpressed in dysplastic lesions, but there are little data on the concordance between HER2 status in dysplastic lesions and their corresponding invasive adenocarcinomas.16,56,62–64 In a study by Lee et al,16 there was no clear association between the HER2 status of dysplastic and invasive components. The authors of that study encountered both cases where HER2 expression was seen in dysplasia but not in the invasive carcinoma as well as cases where the dysplastic component was HER2 negative, but the invasive component showed overexpression. In a more recent study, concordance between high-grade dysplasia and invasive carcinoma was seen only in 4/9 (44%) cases.56 These findings raise the issue of falsepositive results in biopsies due to HER2-positive dysplastic epithelium being misinterpreted as invasive carcinoma. Differentiating between weak 2 + and 1 + staining by IHC can also be challenging, especially in resection specimens where there might be a range of staining intensities. This can be an important distinction as a score of 1 + is not sent for ISH testing in many institutions. If there is uncertainty, it is recommended that the case be scored as 1 to 2 + and material sent for ISH analysis to prevent a false-negative result. As already mentioned, some cases show strong HER2 staining in 6 HER2 gene copies should be considered a positive result. They suggested that copy number is an important consideration especially in cases showing borderline amplification. In a single-probe assay if there are 4 to 6 HER2 gene copies then the use of a dual probe is recommended. Interestingly, a recent study correlating the level of HER2 amplification and response to trastuzumab identified a HER2/CEP17 ratio of 4.7 as the optimal cutoff value discriminating sensitive and refractory patients.67 Over the last few years, there is growing support for the use of bright-field ISH methods for the detection of HER2 amplification. Such methods have several advantages over FISH, including use of a regular microscope instead of a dark-field fluorescence microscope, better signal preservation over time, and better evaluation of morphology. Bright-field techniques may be particularly useful in the evaluation of gastric/GEJ tumors because of heterogeneity as they allow for alignment of IHC slides with the ISH slides. In this way it is easier to score by ISH specific areas showing focal HER2 overexpression. CISH uses an IHC-like peroxidase reaction to detect DNA probes instead of fluorescence. Several studies have reported very good concordance between FISH and CISH.27,50,55,68 For example, Kiyose et al68 used single-color CISH to study 198 gastric tumors and reported a concordance of 99% between FISH and CISH. Two discordant cases were observed one of which showed polysomy. In another study, Yan et al50 used a dual-color CISH assay in 199 samples of gastric cancer and found a 100% concordance with FISH results. A high level of agreement has also been reported between FISH and SISH, which is another brightfield ISH method that is based upon the deposition of silver at the target site after an enzymatic reaction.46,48,69 In a study of 166 gastric cancer samples, 96.4% of cases showed agreement between FISH and dual-color SISH. Six discordant cases were SISH positive and FISH negative and all showed polysomy.69 It is worth noting that the clinical significance of polysomy in gastric cancer is not known and in breast cancer polysomy has been reported in FISH-negative cases that responded to trastuzumab.70
CONCLUSIONS HER2 testing is now recommended for all patients with advanced gastric/GEJ adenocarcinoma to identify HER2positive tumors that can be treated with trastuzumab. Many guidelines recommend that assessment of HER2 status be performed using first IHC, with ISH analysis being reserved for IHC equivocal cases. HER2 expression in gastric/GEJ adenocarcinomas shows some differences compared with what is observed in breast adenocarcinomas, most notably basolateral or lateral membranous positivity and increased
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heterogeneity. Therefore, the IHC scoring criteria for gastric/ GEJ adenocarcinomas differ somewhat from those used in breast. It is recommended that pathologists familiarize themselves with these IHC scoring criteria and potential pitfalls, such as cytoplasmic or nuclear staining in tumor cells. Although there are no strict guidelines as to how many and which samples should be tested, testing >1 sample from the same patient may be justified, especially if available specimens are limited in tumor content or if there is suspicion of morphologic heterogeneity. A good understanding of the patterns of HER2 expression in gastric/GEJ tumors and scoring guidelines will ensure that HER2 status and eligibility for treatment with trastuzumab is determined as accurately as possible.
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