Our long-term objective is to understand the cellular mechanisms underlying taxol toxicity in cancer patients and to identify treatments that may mediate the toxicity. Taxol has shown great promise in the treatment of certain solid tumors but major cardiotoxic side effects including cardiac arrhythmias and possible myocardial infarction are observed in more than one-third of all treated patients. Taxol's effects on microtubule organization and microtubule-dependent vesicle transport are potentially the cause of this toxicity. Microtubule-dependent vesicle transport is involved in the recycling and secretion of plasma membrane proteins. We have observed that microtubule-dependent vesicle transport in endothelial and fibroblast cells is significantly inhibited by taxol treatment. Of considerable interest is our recent preliminary data on the ability of okadaic acid, a protein phosphatase inhibitor to reverse taxol's inhibition of vesicle motility in fibroblasts. We hypothesize that by inhibition of vesicle transport, taxol could alter plasma membrane composition of cells. In endothelial and cardiac cells that are implicated in the observed cardiotoxicity, alteration of plasma membrane composition could lead to altered plasma membrane distribution of ion channels and other membrane receptors essential for normal function. Our investigations will focus on endothelial and fibroblast cells.
Our specific aims are: a. To define taxol's effect on microtubule-dependent vesicle transport. b. To examine the ability of protein phosphatase inhibitors to reverse taxol's inhibition of vesicle motility. c. To characterize taxol's effects on plasma membrane composition.
These aims will be investigated using video-enhanced DIC and fluorescence microscopy and biochemical methodologies. Video-enhanced DIC microscopy will facilitate analysis of vesicle transport in intact cells and in vitro using extracts and purified proteins for reconstitution of vesicle movement on microtubules. Biochemical techniques include those required for protein purification, fractionation of subcellular membrane compartments, and biochemical quantification of membrane receptors. This integrated approach to an understanding of the cellular and potentially molecular basis of the side effects of taxol can provide important clues for treatments which would reverse taxol's toxic side effects while not altering its ability to block cell division.
