Loss of Efficacy of EGFR Inhibitor in Caspase 3 deficient Triple Negative Breast Cancer
Ruya Zhao, BS1, Xinjiang Liu, PhD2, Scott T. Hollenbeck, MD2, Chuan-yuan Li, PhD2.
1Duke University School of Medicine, Durham, NC, USA, 2Duke Medical Center, Durham, NC, USA.
Introduction: Triple negative breast cancer (TNBC) is an aggressive subtype, with few current effective therapies. Since EGFR is often overexpressed in TNBC, it could be used as a therapeutic target. However, multiple clinical trials using EGFR inhibitors failed due to low response rate. The underlying mechanism of drug resistance to EGFR inhibitors is not well understood. In this study, we investigate the role of caspase 3, a major player in apoptosis, in modulating TNBC sensitivity to cancer treatment.
Methods: Caspase 3 gene was knockdown using CRIPSR/Cas9-mediated system in triple negative breast cancer cells (MDA-MB-468). IC50 of gefitinib in MDA-MB-468 is determined using MTT assay. Both vector control and Caspase 3-knockdown group were treated with gefinitib at 5μM and 10μM for 72hrs. Cell survival was determined using CTG luminescent cell viability assay.
Results: MTT assay showed that the IC50 of gefitinib of MDA-MB-468 cells is 15.22μmol/L (Figure 1A). Western blot demonstrated 41.8% knockdown of Caspase 3 gene expression using CRIPSR/Cas9-mediated system (Figure 1B). Cell survival was significantly higher in Caspase-3 deficient TNBC cells in response to EGFR inhibitor treatment at both 5μM and 10μM (Figure 1C; 3214.3±154 vs 1814.7±164 cells survival P=0.0016 at 5μM; 864.0±90 vs 459.2±45 cells survival, P=0.0071 at 10μM).
Conclusion: Caspase-mediated apoptosis is crucial in treatment responses of cancer therapy. Up-regulation or re-expression of caspases could serve as an adjuvant therapeutic approach to high-risk breast cancer resistant to chemotherapy.
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