Benzyl isothiocyanate (BITC), a constituent of edible cruciferous vegetables, is a promising agent for chemoprevention of breast cancer as evidenced by efficacy against xenografted breast cancer cells (e.g., MDA-MB-231 cells) and in a transgenic mouse model (MMTV-neu). We have shown previously that mammary cancer chemoprevention with dietary BITC administration in the MMTV-neu mice is associated with apoptosis induction. In cellular models of human breast cancer (MCF-7 and MDA-MB-231 cells), the BITC-induced apoptosis is initiated by reactive oxygen species (ROS) resulting from inhibition of complex III of the mitochondrial respiratory chain. The present study builds upon these observations to provide insights into the molecular circuitry of BITC-induced apoptosis downstream of ROS production. Because BITC-induced apoptosis in different cell types is associated with activation of c-Jun N-terminal kinase (JNK), we initially focused on Bim as this protein is a downstream mediator of JNK-regulated apoptosis. Bim protein expression (extra-long form) was increased by about 2-3-fold upon 24-hour treatment of MDA-MB-231 cells with pharmacologically relevant concentrations of BITC (2.5 and 5 μM BITC). However, RNA interference of Bim had no effect on BITC-induced apoptosis as judged by analysis of cytoplasmic histone-associated DNA fragment release into the cytosol. Moreover, the BITC-treated MCF-7 cells exhibited a marked decrease in Bim (extra-long form) protein level. BITC treatment resulted in induction of PUMA in both MCF-7 and MDA-MB-231 cells, but as expected, this response was more pronounced in the MCF-7 cells which express wild-type p53. The BITC-induced apoptosis was partially but statistically significantly attenuated by RNA interference of PUMA in MCF-7 cells. Consistent with these observations, genetic depletion of PUMA in HCT-116 cells conferred partial but significant protection against BITC-induced apoptosis. Attenuation of BITC-induced apoptosis in PUMA knockout cells was accompanied by enhanced G2/M phase cell cycle arrest compared with isogenic wild-type HCT-116 cells. The BITC-mediated inhibition of MDA-MB-231 xenograft growth in vivo was associated with PUMA induction in the tumor as evidenced by immunohistochemistry and western blotting. In conclusion, these results indicate that Bim-independent apoptosis by BITC is partly mediated by PUMA. This investigation was supported by the US PHS grant CA129347 awarded by the National Cancer Institute.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 564. doi:1538-7445.AM2012-564

©2012 American Association for Cancer Research
2012