HER2-overexpressing Gastric Cancer Research Articles

HER2 in gastric cancer: a biomarker with clinical impact, but not without translational challenges

Twenty-fi ve years ago the amplifi cation of the HER2 gene and its protein overexpression was fi rst described in gastric cancer [1]. Twenty years later a group of Spanish investigators fi rst experienced the activity of trastuzumab in patients with HER2 overexpressing gastric cancer [2]. Meanwhile, a randomised multi-national trial (ToGA) made things even clearer and brought trastuzumab to regulatory approval and marketing authorisation in Europe and other parts of the world. ToGA could show that the addition of trastuzumab to standard chemotherapy can prolong the survival of patients with incurable HER2 overexpressing gastric cancer [3]. The benefi t is somewhere between 2.7 months (primary endpoint: all included patients) and 5.6 months (subgroup of patients with HER2 gene amplifi cation and score 3+ protein expression). Things look simple at figlance: Trastuzumab, a monoclonal antibody directed against the HER2 receptor protein, is effective in HER2-positive advanced gastric cancer and should be used in this disease in combination with standard platinum-5FU chemotherapy. But oncology would not be oncology if things remained so simple at a more in-depth analysis. Our current challenge with HER2-positive gastric cancer is to make the true nature of every individual disease visible and to translate our fi ndings to the best available treatment recommendation. Our translator in this critical venture is our highly respected partner, the molecular pathologist, who‐ more than ever‐is an integral part of the multidisciplinary treatment team. Pathologists and clinicians know that‐for the best of

Pertuzumab in Combination with Trastuzumab Shows Significantly Enhanced Antitumor Activity in HER2-Positive Human Gastric Cancer Xenograft Models

We investigated the antitumor activity of the combination of two different humanized monoclonal human epidermal growth factor receptor (HER) 2 antibodies, pertuzumab and trastuzumab, for gastric cancer. Tumor mouse xenograft models were used to examine antitumor activity. Cell proliferation was examined using crystal violet staining. HER family proteins' expression was analyzed by ELISA and immunohistochemistry. Phosphorylated proteins and heterodimers were detected by Western blotting and in situ proximity ligation assay (PLA), respectively. Apoptosis activity was examined by caspase 3/7 activity. Antibody-dependent cellular cytotoxicity (ADCC) activity was detected by xCELLigence. Microvessel density was examined by CD31 staining. Pertuzumab in combination with trastuzumab showed significant antitumor activity compared with each monotherapy in NCI-N87, an HER2-positive human gastric cancer xenograft model. The efficacy was stronger than that of the maximum effective dose with each monotherapy. Similar antitumor activity was shown in 4-1ST, another HER2-positive gastric cancer model, but not in MKN-28, an HER2-negative model. Combining pertuzumab with trastuzumab enhanced cell growth inhibition and apoptosis activity by inhibiting EGFR-HER2 heterodimerization and the phosphorylation of these receptors and their downstream factors. This effect was also seen in HER2-HER3 signaling. Furthermore, pertuzumab in combination with trastuzumab potentiated the ADCC activity of those antibodies and reduced tumor microvessel density. We showed the significantly enhanced efficacy of pertuzumab combining with trastuzumab for HER2 overexpressing gastric cancer through the potentiation of cell growth inhibition, apoptosis activity, cell killing activity by ADCC, and antiangiogenic activity. This study suggests the clinical benefit of combination therapy with pertuzumab and trastuzumab for patients with HER2-positive gastric cancers.

HER2 status as an independent prognostic marker in patients with advanced gastric cancer receiving adjuvant chemotherapy after curative gastrectomy.

4084 Background: It has been recently confirmed that trastuzumab, a monoclonal antibody against HER2, improves the overall survival in HER2 overexpressing gastric cancers. We investigated the prognostic significance of HER2 status in gastric cancers after curative gastrectomy and adjuvant chemotherapy. Methods: HER2 status was evaluated in 679 gastric cancer patients who recieved curative gastrectomy and adjuvant chemotherapy. All were enrolled in a phase III clinical trial from January 2002 to July 2006 in Asan Medical Center. Tissue microarrays were constructed using two independent cores form each case. Immunohistochemical (IHC) staining was scored by gastric cancer scoring system. Double silver-enhanced in situ hybridization (SISH) was done using HER2 probe and chromosome 17 centromere oligoprobe (CEP17). IHC score 2+ or 3+, and HER2/CEP17 ratio ≥2.0 were considered positive for IHC and SISH, respectively. IHC 3+ or IHC 2+ and SISH positive cases were defined as “HER2 status-positive”. Results: HER2 IHC-positivity (9.7%), SISH-positivity (12.2%) and HER2 status-positivity (8.3%) were all associated with differentiated or intestinal type histology and distal gastrectomy. IHC-positivity (P = 0.315), SISH-positivity (P = 0.382) and HER2 status-positivity (P = 0.277) showed negative correlation with the recurrence-free survival (RFS) but did not show statistical significance. However, in subgroup analyses according to the stage grouping (II vs. III+IV), HER2 status-positivity was a poor prognostic factor in advanced stage group (P = 0.001) but not in earlier stage (P = 0.827). The HER2 status-positivity remained as an independent prognostic factor after multivariate analysis (P = 0.023, HR = 1.709). Conclusions: HER2 status, taking both protein expression and gene amplification into consideration, was proved to be an independent poor prognostic marker for RFS in advanced stage gastric cancers after curative gastrectomy. Since it has been shown that HER2 status-positive patients benefited from trastuzumab therapy in unresectable gastric cancer patients, trastuzumab may play a role in locally advanced gastric cancer patients as adjuvant therapy.

Combination of exposure-response and case-control analyses in regulatory decision making.

4087 Background: In 2010, the FDA approved Herceptin (trastuzumab) as treatment for HER2-overexpressing metastatic gastric cancer. The purpose of this analysis was to further evaluate whether the a...

The search for optimal response prediction in cancer leads to a personalized approach

There is a clear need for better response prediction in the management of patients with cancer. Important progress has indeed been made in understanding the underlying mechanisms of tumor growth and targeting important new targets with novel drugs blocking receptors or interfering with one of the important pathways. A long list of new, targeted agents has been developed or is under development. With this comes also the need of new methods and new molecular markers for response prediction. The area of response prediction is booming: functional imaging is becoming a very important tool and aid and the knowledge on molecular markers is growing very rapidly. Functional imaging with positron emission tomography (PET) scan, dynamic contrast-enhanced ultrasonography (DCE-US) and magnetic resonance imaging (MRI) is coming into the research arena and is likely to be implemented in the future in clinical practice. Better validation of the clinical utility of functional imaging techniques is, however, still needed in many clinical situations. A major clinical problem for the use of novel molecular targeted agents is to determine their activity by the use of classic imaging evaluation methods such as computed tomography (CT) and by the use of standard tumor size assessment with Response Evaluation Criteria in Solid Tumors (RECIST). Therefore, functional imaging techniques that could be performed in the evaluation of the early activity and also in the determination of the optimal dose of the new targeted agents are necessary. Dynamic contrast-enhanced (DCE)-MRI and DCE-US, diffusion and perfusion MRI and PET-CT, using new tracers for proliferation, will probably be integrated gradually in the new generation of clinical trials and eventually also in clinical practice. Molecular markers clearly have the potential to help to select cancer patients that are suitable for treatment with a molecular targeted, agent if they are able to predict the activity or the lack of activity of these agents. Among examples of predictive biomarkers for response and clinical activity, there are cKIT mutations for imatinib therapy in gastrointestinal stromal tumors (GIST), epidermal growth factor receptor (EGFR) mutations for erlotinib and gefitinib treatment in nonsmall cell lung cancer (NSCLC), HER-2 gene amplification for the use of trastuzumab in breast cancer, as well as recently HER-2 overexpression in gastric cancer, KRAS mutations as predictors of lack of activity of cetuximab and panitumumab in colorectal cancer and, finally, BRAF mutations for treatment with selective BRAF inhibitors in melanoma. This list is not completed and will certainly be expanded in the near future. Further, some of these biomarkers are not only predictive for response to a given treatment, but they could have also a prognostic role. As an example, BRAF mutations correlate with poor prognosis, lack of cetuximab and panitumumab efficacy and poor response to chemotherapy in metastatic colorectal cancer. The discovery of whole gene signatures, if they would be confirmed predictive and/or prognostic, will for E. Van Cutsem (*) University Hospital Gasthuisberg, Leuven, Belgium e-mail: eric.vancutsem@uz.kuleuven.ac.be

The dual EGFR/HER‐2 tyrosine kinase inhibitor lapatinib sensitizes colon and gastric cancer cells to the irinotecan active metabolite SN‐38

Members of the human epidermal receptor (HER) family are frequently associated with aggressive disease and poor prognosis in multiple malignancies. Lapatinib is a dual tyrosine kinase inhibitor targeting the epidermal growth factor receptor (EGFR) and HER-2. This study evaluated the therapeutic potential of lapatinib, alone and in combination with SN-38, the active metabolite of irinotecan (CPT-11), in colon and gastric cancer cell lines. Concentration-dependent antiproliferative effects of both lapatinib and SN-38 were observed in all colon and gastric cancer cell lines tested but varied significantly between individual cell lines (lapatinib range 0.08-11.7 muM; SN-38 range 3.6-256 nM). Lapatinib potently inhibited the growth of a HER-2 overexpressing gastric cancer cell line and demonstrated moderate activity in gastric and colon cancer cells with detectable HER-2 expression. The combination of lapatinib and SN-38 interacted synergistically to inhibit cell proliferation in all colon and gastric cancer cell lines tested. Cotreatment with lapatinib and SN-38 also resulted in enhanced cell cycle arrest and the induction of apoptosis with subsequent cellular pharmacokinetic analysis demonstrating that lapatinib promoted the increased intracellular accumulation and retention of SN-38 when compared to SN-38 treatment alone. Finally, the combination of lapatinib and CPT-11 demonstrated synergistic antitumor efficacy in the LoVo colon cancer mouse xenograft model with no apparent increase in toxicity compared to CPT-11 monotherapy. These results provide compelling preclinical rationale indicating lapatinib to be a potentially efficacious chemotherapeutic combination partner for irinotecan in the treatment of gastrointestinal carcinomas.

Her2-PI3K-Akt signaling transducfion and gastric cancer

PI3K-Akt, one important pathway of：the Her2 signaling transduction, affects the survival and apoptosis of cells, and influences the aggression, transfer, prognosis and chemoreslstance of tumors by regulating some apoptosls- related factors such as NF-kappa B. The biological and clinical characters of gastric cancer are closely related to Her2-PI3K-Akt signaling transduction. Trastuzumab is a humanized monoclonal anti-body directed against the Her2 protein, and pre-clinical studies have proved its inhibitive effect on Her2-overexpressing gastric cancer cell lines and xenograft models. Key words: genes, erbB-2 ; NF- kappa B; Stomach neoplasms ; Antibodies, monoclonal

Molecular basis for sensitivity and acquired resistance to gefitinib in HER2-overexpressing human gastric cancer cell lines derived from liver metastasis

Gastric cancer metastasised to the liver was found to overexpress HER2 at a significantly higher incidence than primary gastric cancers. The purpose of the present study was to investigate the possibility of molecular therapy targeting HER2 overexpression in gastric cancer liver metastasis. We developed three new HER2-overexpressing gastric cancer cell lines (GLM-1, GLM-2, GLM-4) without epidermal growth factor receptor (EGFR) mutations derived from such liver metastasis, two of which had HER2 gene amplifications. All these GLM series of cell lines were highly sensitive to gefitinib in vitro, a specific inhibitor of EGFR tyrosine kinase (Iressa) rather than anti-HER2 antibody trastuzumab (Herceptin), whereas most of the HER2 low-expressing counterparts were not. In these HER2-overexpressing GLM series, protein kinase B (Akt), but not extracellular signal-regulated kinase 1/2 (ERK1/2), was constitutively phosphorylated, and gefitinib efficiently inhibited this Akt phosphorylation, induced strong apoptosis in vitro and exhibited antitumour activity in tumour xenografts in nude mice. This gefitinib-mediated antitumour effect in xenograft was significantly potentiated by trastuzumab treatment. On the other hand, gefitinib-resistant cells (GLM-1R) exhibited increased EGFR expression, followed by constitutive activation of mitogen-activated protein kinase (MAPK) pathway. These results suggest that the antitumour effect of gefitinib is due to the effective inhibition of HER2-driven constitutive activation of phosphatidylinositol-3-kinase (PI3K)/Akt pathway, and that the acquired resistance to gefitinib is due to the constitutive activation of Ras/MAPK pathway in compensation for PI3K/Akt pathway. Gastric cancer liver metastasis with HER2 overexpression would be a potential molecular target for gefitinib and trastuzumab.

Antitumor activity of trastuzumab in combination with chemotherapy in human gastric cancer xenograft models

To clarify the antitumor activity of trastuzumab and its potential as an effective treatment for gastric cancer patients. Levels of HER2 expression in tumor tissues of gastric cancer cell lines were examined using immunohistochemistry (IHC), fluorescence in situ hybridization (FISH), and mRNA quantification. Efficacy of trastuzumab was examined as a single agent or in combination with chemotherapeutic agents widely used clinically for gastric cancers in HER2-overexpressing human gastric cancer xenograft models. Two of nine human gastric cancer xenograft models, NCI-N87 and 4-1ST, showed overexpression of HER2 mRNA and protein by IHC (HercepTest) and HER2 gene amplification by FISH (Pathvysion). HER2 protein showed potent staining in peripheral membranes, similar to the staining pattern of breast cancer. FISH scores were also comparable to those of breast cancer models. Trastuzumab as a single agent inhibited the tumor growth in both of the HER2-overexpressing models but not in the HER2-negative models, GXF97 and MKN-45. In any combination with capecitabine, cisplatin, irinotecan, docetaxel, or paclitaxel, trastuzumab showed more potent antitumor activity than the anticancer agents alone. A three-drug combination of capecitabine, cisplatin, and trastuzumab showed remarkable tumor growth inhibition. In NCI-N87 in vitro, trastuzumab showed direct antiproliferative activity according to cell count or crystal violet dying, and showed indirect antitumor activity such as antibody-dependent cellular cytotoxicity. The antitumor activity of trastuzumab observed in human gastric cancer models warrants consideration of its use in clinical treatment regimens for human gastric cancer as a single agent or a combination drug with various chemotherapeutic agents.

Suppression of tumor growth in human gastric cancer with HER2 overexpression by an anti-HER2 antibody in a murine model.

New modalities are necessary for the treatment of patients with unresectable gastric cancer. The aim of this study was to investigate whether or not anti-HER2 antibody could suppress the growth of human gastric cancer cells with HER2 overexpression in vitro and in vivo. Four human gastric cancer cell lines, NCI-N87, MKN-45P, Kato-III, and MKN-1, were used in this study. The suppression of cell proliferation in vitro and of subcutaneous tumor growth in a nude mouse model after treatment with trastuzumab was examined. The expression of HER2 protein was investigated by Western blot analysis. The effect of trastuzumab on the survival rate of nude mice with peritoneal dissemination was examined. Trastuzumab significantly reduced proliferative activity in NCI-N87, a HER2-overexpressing human gastric cancer cell line, in vitro. In the nude mouse model with transplanted subcutaneous tumor, trastuzumab significantly suppressed the tumor growth of NCI-N87 cells, and then HER2 expression was reduced. Trastuzumab improved the survival rate of mice with peritoneal dissemination of MKN-45P cells. Trastuzumab therapy is a potential candidate for a novel treatment of HER2-overexpressing gastric cancer.

Monoclonal antibodies as effective therapeutic agents for solid tumors.

Monoclonal antibodies (mAbs) against growth factors or their receptors have been revealed to be effective therapeutic agents for solid tumors. Trastuzumab (humanized anti-HER2 mAb) is the first mAb approved for the treatment of a solid tumor, metastatic breast cancer. Large-scale phase III clinical trials are now ongoing to further evaluate the additive effects on chemotherapy and the efficacy as a maintenance monotherapy. Another anti-HER2 mAb CH401 that we developed also seems to have good potential. This chimeric mAb completely suppressed the growth of established human tumor xenografts in SCID mice after a single injection. Furthermore, CH401 characteristically showed much stronger induction of apoptosis in HER2-overexpressing gastric cancer cells compared to trastuzumab. Additional targets now being intensively evaluated are epidermal growth factor receptor (EGFR) and vascular endothelial growth factor (VEGF). Both cetuximab (chimeric anti-EGFR mAb) and bevacizumab (humanized anti-VEGF mAb) have recently been shown to be of clinical value for metastatic colorectal cancer. Anti-idiotype mAbs are unique as active immunotherapeutic agents, and survival benefits have been observed in clinical trials for solid tumors.