E. Gleeson1, H. Glatthorn1, D. Zimmerman1, C. Carballo1, S. Mahmood4, P. Love1, M. F. Shaikh1, W. F. Morano1, S. D. Richard2, M. R. Pincus3, W. B. Bowne1 1Drexel University College Of Medicine,Surgery,Philadelphia, Pa, USA 2Drexel University College Of Medicine,Gynecologic Oncology,Philadelphia, Pa, USA 3New York Harbor Healthcare System VAMC,Pathology,New York, NY, USA 4University Of Pittsburg,Pittsburgh, PA, USA
Introduction: Of an estimated 22, 000 women diagnosed with ovarian cancer in the United States each year, 80% will develop lethal multi-drug resistance (MDR) to conventional chemotherapy. HDM-2 over-expression in the cancer cell membrane may be a potential anti-cancer target. We tested this target using a p53-derived, HDM2-binding peptide for anti-cancer activity in MDR ovarian cancer.
Methods: Western blot analysis was used to demonstrate expression of HDM-2 in MDR SKOV-3 human ovarian cancer cell membranes compared to the membranes of untransformed fibroblasts. 5×104 cells were treated with HDM2-binding construct PNC–27 and control peptide. Anti-cancer activity and mechanism of PNC-27 were studied for cell viability (MTT), necrosis (LDH), apoptosis (Caspase-3) and co-localization of HDM-2 with PNC-27 (immunofluorescence).
Results: HDM-2 was strongly expressed in the cell membranes of MDR SKOV-3 by western blot analysis. Accordingly, PNC–27 was selectively cytotoxic to these cells, inducing nearly 100% reduction in cell proliferation compared to control PNC–29 and untreated cells (p<0.001; See Figure). We observed a rapid (4h) cellular necrosis with a 2.5 fold increase in LDH (p<0.001) and no caspase-3 activity. This was shown to occur in a dose-dependent manner. In contrast, PNC-27 had no effect on untransformed fibroblasts. This observed effect was due to over-expression of HDM-2 in MDR SKOV-3 cells which demonstrated co-localization of PNC–27 and HDM–2 along their membranes.
Conclusion: These findings demonstrate that HDM-2 over-expression in MDR ovarian cancer cells may be used as a potential target for anti-cancer therapy. Therapies that target HDM-2 in ovarian cancer cell membranes may increase our armamentarium for the treatment of this lethal disease.
