Abstract B47: Drug-induced EMT is fundamentally distinct from pathologic EMT

Saeed Salehin Akhand; Michael Wendt

doi:10.1158/1538-7445.TUMMET15-B47

Abstract Background: Receptor tyrosine kinase ErbB2 (Her2) is overexpressed in an estimated 25% of the breast cancer patients. A therapeutic regime of Her2 targeted therapies, consists of Trastuzumab followed by the dual EGFR/Her2 tyrosine kinase inhibitor, Lapatinib. These therapies continue to show success in blocking the progression of this breast cancer subtype. Of clinical concern, however, cancer cells eventually develop pathways to evade Lapatinib-mediated tumor inhibition and acquire drug resistance. In this study, we sought to understand the molecular mechanism of resistance towards Lapatinib. Methods: We used Her2 overexpressing human mammary epithelial cells to generate Lapatinib resistant cell lines (LapR) in vitro. The LapR cells acquired growth persistence within the continued presence of Lapatinib for four weeks. We analyzed the resultant cell populations via cell viability assays, immunoblotting, flow cytometry, 3D cell culture and Nano-string gene expression analysis. Results: Generated LapR cell lines showed robust epithelial-mesenchymal transition (EMT) phenotypes that included upregulated FGFR1 and preservation of their Her2 expression. We tested the viability of LapR cells using dose response assays with Afatinib, a second-generation covalent ErbB kinase inhibitor, and Trastuzumab emtansine (T-DM1), an anti-Her2 antibody drug conjugate. We also utilized recently developed covalent inhibitors of FGFR. LapR cells were highly resistant to Afatinib but did show sensitivity to T-DM1 and FGFR kinase inhibition. Importantly, resistance to ErbB inhibition was maintained in the LapR cells for several weeks following removal of Lapatinib. This stable phenotype was consistent with a maintained yet distinct regulation of the cancer stem cell markers (CD44/CD24) compared to a transient EMT induced by transforming growth factor β (TGF-β). Strikingly, we observed that unlike their parental counterparts LapR cells were unable to grow in a compliant 3D matrix. Furthermore, unlike TGF-β-treated cells, LapR cells were unable to metastasize in vivo. By using Nano-string pan cancer progression panel analysis, we characterized key differences in the LapR EMT phenotype as compared to a TGF-β induced EMT. Future Directions: Currently, we are analyzing the expression data of LapR and TGF-β-induced EMT to identify key targets required for EMT-induced tumor progression. Overall, our studies seek to define what aspects of EMT are involved in cellular persistence in response to anti-cancer drugs versus those utilized by cells during metastatic progression. Citation Format: Saeed Salehin Akhand, Michael Wendt. Drug-induced EMT is fundamentally distinct from pathologic EMT. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Metastasis; 2015 Nov 30-Dec 3; Austin, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(7 Suppl):Abstract nr B47.

Abstract B47: Drug-induced EMT is fundamentally distinct from pathologic EMT

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