Rapamycin Inhibits In Vitro Growth and Release of Angiogenetic Factors in Human Bladder Cancer

Abstract

To evaluate the in vitro effects of rapamycin in human transitional cell carcinoma and to clarify the possible rapamycin-hypoxia interactions. Monolayer cultures of RT112 (G1), RT4 (G1-2), T24 (G3), and SUP (G4) cells were incubated in medium with or without rapamycin (10 microM, 100 microM) in different gaseous conditions (1% oxygen plus 5% carbon dioxide plus 94% nitrogen or 95% oxygen plus 5% carbon dioxide or 21% oxygen plus 78% nitrogen). Cell vitality and proliferation were determined using the microculture tetrazolium assay. Apoptotic cells were quantified by flow cytometry. Enzyme-linked immunosorbent assay was used to visualize inhibition of the mammalian target of rapamycin (mTOR) by rapamycin and synthesis of vascular endothelial growth factor. Rapamycin delayed proliferation of cancer cell lines but did not induce apoptosis. No direct hypoxia interactions of rapamycin were found. Enzyme-linked immunosorbent assay gave evidence of mTOR inhibition in all cell lines and a significant decrease of hypoxia-induced vascular endothelial growth factor synthesis. Because proliferation in bladder cancer cells was delayed, but no apoptosis was induced, cell cycle arrest caused by rapamycin was feasible. Rapamycin decreased hypoxia-induced synthesis of vascular endothelial growth factor. These findings should be substantiated in an animal model to verify the antiangiogenetic effects of rapamycin.