P53 Gene Therapy Sensitizes Resistant Breast Cancer Cells to Doxorubicin Chemotherapy

Ajay      Kumar; Srinath      Palakurthi

doi:10.2174/2210303111303030002

Abstract

Tumor-suppressor gene, p53, is the most commonly mutated gene in human cancer and it is considered as a major reason for resistance to chemotherapy. Replacement of mutated p53 gene with the wild-type gene (wt-p53) has been reported to restore cell-cycle control and inhibit tumorigenesis. Aim of this study was to investigate the effect of exogenously supplied wt-p53 on the doxorubicin cytotoxicity to doxorubicin-resistant cancer cells and the normal cells. Cancer cell specific p53 delivery was achieved using dendriplexes prepared with ornithine-conjugated PAMAMG4 dendrimers and a CMV promoter driven p53-GFP expressing plasmid DNA (pDNA). Sustenance of exogenously supplied GFP-p53 in different cell lines was assessed by flow cytometry and PCR. Effect of wt-p53 treatment on doxorubicin cytotoxicity in normal: MCF 12A (wt-p53), and cancer: MCF 7 (wt-p53), MDA-MB-231 (mt-p53) and NCI/ADR-RES (mt-p53 and doxorubicin resistant) cells was assessed by MTT assay. Flow cytometry and PCR results revealed that exogenous p53 expression lasted for 4 days both at protein and at mRNA levels. Annexin V binding and cytotoxicity assays revealed that p53 (pGFP-p53 dendriplexes) caused significantly more apoptosis and cytotoxicity (P < 0.0005) than the control (pGFP dendriplexes) in cancer cells. Combination treatment with wt-p53 and doxorubicin (500 nM), significantly increased doxorubicin cytotoxicity to MCF 7 and MDA-MB-231 cells, but no cytotoxicity was observed in MCF 12A cells. NCI/ADR-RES cells did not respond to lower doxorubicin concentration but at higher concentrations (5 and 50 µM), combination therapy showed a significant increase in cytotoxicity (P < 0.05 and 0.0005, respectively). It may be concluded from the results that p53 gene therapy followed by doxorubicin treatment may be a suitable combination therapy for doxorubicin-resistant cancer.

Keywords: Breast cancer, cytotoxicity, doxorubicin resistance, gene delivery, p53, PAMAMG4-ORN60, poly(amido)amine (PAMAM) dendrimers.

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