Rapamycin potentiates the effects of paclitaxel in endometrial cancer cells through inhibition of cell proliferation and induction of apoptosis.

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Introduction: Treatment of women with advanced endometrial cancer using cytotoxic chemotherapeutic agents has been met with limited success. Thus, the search has been for an additional agent which could be used in combination with more traditional therapies to dramatically increase efficacy while not increasing toxicities. Many novel agents are being investigated that target specific cellular signaling pathways thought to be essential in endometrial cancer progression and metastasis. One of the most promising of these for endometrial cancer are inhibitors of the mammalian target of rapamycin (mTOR). In this study, our objective was to examine the effect of combined therapy with rapamycin and paclitaxel in endometrial cancer cells.
 Methods: Cell proliferation was assessed in ECC-1 and Ishikawa endometrial cancer cell lines after exposure to rapamycin or paclitaxel alone and in combination, using the MTT assay. Median-effect plot analyses and calculation of the multiple drug effect/combination index (CI) was performed by the method of Chou and Talalay. Apoptosis was evaluated using a dsDNA assay. Western blotting was performed to assess the expression of phosphorylated S6, a downstream target of mTOR. Immunoflourescent staining was performed to evaluate α-tubulin expression.
 Results: Paclitaxel inhibited proliferation in a dose-dependent manner in both cell lines with IC₅₀ values of 0.1-0.5 nM and 1-5 nM for Ishikawa and ECC-1 cells, respectively. Simultaneous exposure of cells to various doses of paclitaxel in combination with rapamycin (1 nM) resulted in a significant synergistic anti-proliferative effect (CI<1, range .49-.01). Rapamycin alone did not induce apoptosis, but combined treatment with paclitaxel increased apoptosis over that of paclitaxel alone. S6 phosphorylation was decreased with rapamycin and combination treatment, but not decreased by paclitaxel alone. Paclitaxel dramatically decreased α-tubulin expression, and combination treatment with rapamycin appeared to enhance this effect. Rapamycin alone had little effect on α-tubulin expression.
 Conclusions: We demonstrate that rapamycin potentiates the effects of paclitaxel in endometrial cancer cells through inhibition of cell proliferation, induction of apoptosis and decreased α-tubulin expression. Although rapamycin did not induce apoptosis in these endometrial cancer cell lines, treatment with rapamycin and paclitaxel together dramatically increased induction of apoptosis to well above the effects of paclitaxel alone. Thus, mTOR inhibition by rapamycin may have the potential to inhibit tumor growth as well as enhance the effects of paclitaxel at lower doses, culminating in overall decreased toxicities for endometrial cancer patients. This suggests that the combination of rapamycin and paclitaxel may be a promising effective targeted therapy for this disease.