CYTOTOXIC ACTIVITY OF INVESTIGATIONAL DRUGS ON THE BREAST CANCER CELL LINE MDA-MB-435: PACLITAXEL INDUCES APOPTOSIS AND MODIFIES THE EXPRESSION OF BCL-2

Abstract

Intensive drug screening and research are underway to identify drugs with increased potency and efficacy toward eradicating breast cancer cells. Initial experiments described here were designed to compare the ability of several agents to induce cytostatic and cytocidal activity in the human breast cancer cell line MDA-MB-435. Two standardized in vitro assays, a microculture tetrazolium assay and a soft agar colony formation assay, were used to evaluate the activities of the following agents: vinblastine sulfate, etoposide, doxorubicin, actinomycin D, bleomycin, mitomycin C, paclitaxel, carmustine, chlorambucil, 5-fluorouracil and cisplatin. Results indicated that on a molar basis paclitaxel, doxorubicin and actinomycin D were the most potent at inducing cytostatic and/or cytocidal activity. Additional experiments investigated a possible mechanism by which paclitaxel induced the killing of MDA-MB-435 cells. Cell cycle analysis revealed that paclitaxel caused arrest of these cells in the G₂/M phase of the cell cycle and the accumulation of cells with an aneuploid content of DNA indicative of apoptosis. Indeed, paclitaxel induced dose-dependent apoptotic death as indicated by gel electrophoretic analysis of DNA fragmentation and measurement of cytoplasmic histone-associated DNA. Northern and western blot analyses were performed to determine whether paclitaxel regulated the expression of factors involved in the induction or suppression of apoptosis. Paclitaxel did not affect the expression of p53, Bax, Bc1-Xs/l or FAS proteins in MDA-MB-435. In contrast, paclitaxel modified the expression of Bc1-2 protein. Overall, the findings contained herein confirm and extend accumulating data on the potential usefulness of various chemotherapeutic agents in the treatment of breast cancer. Moreover, they implicate apoptosis as a mechanism of cytotoxic action of paclitaxel and suggest a biochemical pathway by which paclitaxel induces apoptotic death in breast cancer cells.