Abstract C177: Src and ADAM17-mediated shedding of heregulin is a mechanism of acute resistance to chemotherapy in gastro-oesophageal cancer cells

Abstract

Abstract Background: We previously reported that, in response to chemotherapy, gastro-oesophageal cancer cells (GEC) activate a human epidermal receptor (HER)-mediated survival response abrogated by HER inhibitors. In order to elucidate potential biomarkers for response to chemotherapy treatment and potential novel targets, we investigated the pathways which mediate chemo-induced HER activity in GEC cells. Methods: Apoptosis was measured by Flow Cytometry or PARP, Caspase 8/3 cleavage. HER1/HER2/HER3, Akt expression/activity and Heregulin (HRG) expression levels were determined by Western blotting. ADAM17 activity was measured using a fluorometric (MCA) assay kit while sTGF-α levels were analysed by ELISA. Results: Following treatment with chemotherapy, we found that increased HER1/HER2/HER3 activity was associated with increased Src-family kinase (SFK) activity. Using the SFK inhibitor AZD0530, we found that chemotherapy-induced HER1 activation was abolished, indicating that HER1 activation was mediated via SFKs. Furthermore, we found that the dual HER1/HER2 inhibitor lapatinib had no effect on chemo-induced SFK activity, indicating that SFKs act upstream of HER. Furthermore, we found that chemotherapy treatment resulted in increased ADAM-17 activity, TGF-α and heregulin levels in culture medium of GEC cells. ADAM17 inhibition attenuated chemotherapy-induced TGF-α and heregulin levels and HER1/3 activity. When cells were co-treated with heregulin-neutralizing antibodies, we found strongest inhibition of chemotherapy-induced HER1/HER3 and Akt activity. In addition, following treatment with AZD0530, we found complete inhibition of chemotherapy-induced ADAM17 activity and ligand shedding. Finally, when HER1/HER2/HER3, ADAM17 or SFK inhibitors were combined with chemotherapy, we found a potent increase in apoptosis in GEC cells. Conclusions: We provide strong evidence that chemotherapy induces HER1/HER2/HER3 activation via a ligand dependent mechanism in GEC cells and have demonstrated that SFK's, ADAM17 and heregulin are critical mediators of HER1/HER2/HER3 activation following chemotherapy. We propose a model in which chemotherapy results in SFK activation that in turn activates ADAM17-mediated shedding of heregulin. Heregulin then activates HER1/HER2/HER3 pro-survival signalling in an autocrine and paracrine fashion. Thus, inhibiting HER1/HER2/HER3, SFKs or ADAM17 may have therapeutic potential for sensitizing GEC tumors to chemotherapy. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):C177.