ESTRAMUSTINE-BINDING PROTEIN (EMBP) CONTENT IN 4 DIFFERENT CELL-LINESAND ITS CORRELATION TO ESTRAMUSTINE INDUCED METAPHASE ARREST

It is known that estramustine (EM) accumulates in cells at the G2/M-ph ase and causes metaphase arrest of various cell types. The inhibitory effect is mediated by interaction with microtubule-associated proteins (MAPs) and/or tubulin. Estramustine-binding protein (EMBP) is a secre tory protein which has been found in a number of different tumor cells and has been shown to facilitate the uptake of EM into cells. In this study the efficacy of EM in arresting cells at metaphase was studied, using four different human cell lines; the prostatic cancer cell line DU 145, the breast cancer cell line MDA 231, the colon cancer cell li ne Colon 320, and the urinary bladder cancer cell line RT4. The cells were incubated with EM at a concentration of 10 mu g/ml for 24 hours. The data reveal an increase in metaphase arrests in the DU 145 and in Colon 320 cell lines. Both of these cell lines were found to contain h igh amounts of EMBP using a dot-blot assay. The other two cell lines, MDA 231 and RT4 had undetectable intracellular amounts of the protein and exhibited a low increase in metaphase arrests. The cell lines were analysed regarding S-phase fraction with flow-cytometry (FCM) to excl ude the growth rate of the cells as a limiting factor. The results fro m the FCM confirmed the cytogenic analysis, that is a higher percentag e of cells were in the G2/M phase in both the DU 145 and Colon 320 cel l line compared to MDA 231 and RT4. EM causes miotic arrest in those c ell lines that contain detectable amounts of EMBP.