The effects of estramustine on metaphase and anaphase in DU 145 prostatic carcinoma cells.

The chemotherapeutic drug, estramustine, has been shown to cause the disassembly of microtubules via binding to microtubule-associated proteins. In this report, estramustine is shown to be a potent inhibitor of mitotic progression in the human prostatic carcinoma cell line, DU 145. Examination of individual living cells via video-enhanced differential interference contrast (DIC) optics shows that the drug delays the onset of anaphase, reduces anaphase spindle-pole elongation (anaphase B), and delays cytokinesis. In addition, immunofluorescent studies demonstrate that estramustine causes a rapid disorganization of the mitotic apparatus at significantly lower concentrations than those reported previously. Electron microscopic studies show that microtubule bundles are present in drug-treated mitotic cells in association with kinetochores and centrioles.

Mechanism based chemotherapy for prostate cancer.

Estramustine offers a mechanistically novel therapy for the treatment of prostatic cancer. The drug is composed of oestradiol coupled to nor-nitrogen mustard. The cytotoxic properties of estramustine act independently of its constituent molecules. In vivo and in vitro studies indicate that the drug binds microtubule associated proteins and inhibits microtubule regulated processes. We have lately shown that micromolar levels of estramustine inhibit selected processes in mitosis, suggesting that certain classes of microtubules and/or MAPs are differentially sensitive to the drug. New clinical trials using estramustine and vinblastine on patients with advanced hormone refractory disease suggest that the combination of two antimicrotubule agents acting by means of different target molecules may be important clinically.

Resistance to the antimitotic drug estramustine is distinct from the multidrug resistant phenotype.

Following EMS mutagenesis, three estramustine (EM) resistant DU 145 human prostatic carcinoma cell lines were clonally selected by exposure to incrementally increasing concentrations of the drug. Although only low levels of resistance (approximately 3-fold) were attainable, this resistance was stable in the absence of continuous drug exposure. These EM-resistant clones (EMR 4,9,12) did not exhibit cross resistance to vinblastine, taxol, or adriamycin, and had collateral sensitivity to cytochalasin B. None of the lines had elevated expression of P-glycoprotein mRNA or glutathione S-transferase activity, suggesting a phenotype distinct from the classic multi-drug resistance phenotype. This conclusion was supported further by the observation that two MDR cell lines (FLC mouse erythroleukaemic and SKOV3 human ovarian carcinoma cells) showed sensitivity to EM. Fluorescent activated cell sorting analysis of the effects of EM on cell cycle traverse revealed that at EM concentrations up to 20 microM an increasing percentage of wild type cells were blocked in G2/M; no such effect occurred in EMR lines. Differential interference contrast microscopy was employed to study EM's effect on mitosis. EMR lines were able to form functional, albeit smaller, spindles at EM concentrations that resulted in chromosomal disorganisation and inhibition of mitotic progression in wild type cells. EMR lines were able to progress through mitosis and cytokinesis at the same rate as untreated cells. Tritiated EM was used to evaluate potential drug uptake/efflux mutations in ERM clones. EMR 4 and 9 incorporate less EM than wild type cells; however, they have significantly decreased cellular volumes. The initial efflux rate constants for EMR clones were greater than for wild type cells. Within 5 min greater than 70% of the drug was lost from resistant cells compared to a 50% loss by the wild type. Although the specific mechanisms of resistance have yet to be defined, the lack of collateral resistance to other MDR/anti-microtubule agents could serve as the basis for the clinical use of EM in combination chemotherapy.