Dietary isothiocyanates suppress microtubule dynamic instability in association with mitotic arrest and inhibition of tumor cell proliferation.

摘要

Microtubules are highly dynamic polymers that play fundamental roles in a number of cellular functions. Their essential role in mitosis makes them an excellent target for the development of effective anticancer chemotherapeutic drugs, toxic molecules that are mostly derived from natural products. Many anti-mitotic agents such as taxanes and the vinca alkaloids suppress microtubule dynamic instability at low concentrations that arrest mitosis and inhibit cell proliferation, and alter microtubule polymer mass at higher concentrations. They bind to either soluble tubulin or directly to the microtubules, interfering with microtubule function, leading to the disruption of mitotic spindles in dividing cells, cell cycle arrest, and ultimately cell death by apoptosis.;Isothiocyanates are naturally occurring phytochemicals found in cruciferous vegetables that inhibit cancer cell proliferation, cause cell cycle arrest and apoptosis; however the mechanisms of their anticancer effects remain poorly understood. Using quantitative fluorescence microscopy, I discovered that sulforaphane, a major isothiocyanate, stabilized microtubules in breast cancer cells by suppressing microtubule dynamic instability at concentrations that inhibited cell proliferation and induced mitotic arrest. In addition, I used biochemical techniques to show that sulforaphane suppressed the dynamicity of purified microtubules in vitro, and caused the conformational changes of tubulin, indicating that the suppression of dynamic instability by sulforaphane in cells is due to a direct effect on the microtubules. The results indicate that sulforaphane arrests proliferation and mitosis by stabilizing microtubules in a manner weaker than but similar to more powerful clinically used anti-mitotic anticancer drugs, strongly supporting the hypothesis that inhibition of mitosis by microtubule stabilization is important for the chemopreventive activity of isothiocyanates.;I also investigated the mechanism of action of eribulin, a novel synthetic analogue of halichondrin B in Phase III clinical trials for breast cancer that failed taxane chemotherapy. The studies summarized here indicate that eribulin suppresses centromere dynamics during mitotic metaphase in human osteocarcoma cells at concentrations that arrest mitosis. Using specific beta-tubulin preparations available to us, I found that eribulin does not preferentially modulate the in vitro dynamics of microtubules assembled from a particular beta-tubulin isotype, indicating that its effectiveness in paclitaxel-resistant cells is attributed to other mechanisms.