Targeting the Autophagy Pathway Using Ectopic Expression of Beclin 1 in Combination with Rapamycin in Drug-Resistant v-Ha-ras-Transformed NIH 3T3 Cells

摘要

The effectiveness of an apoptosis-targeting therapy may be limited in tumor cells with defects in apoptosis. Recen-tly, considerable attention in the field of cancer therapy has been focused on the mammalian rapamycin target (mTOR), inhibition of which results in autophagic cell death. In our study using multidrug-resistant v-Ha-ras-transformed NIH3T3 (Ras-NIH 3T3/Mdr) cells, we demon-strated that rapamycin-induced cell death may result from 2 different mechanisms. At high rapamycin concentrations (≥100 nM), cell death may occur via an autophagy-depen-dent pathway, whereas at lower concentrations (≤ 10 nM), cell death may occur after G1-phase cell cycle arrest. This effect was accompanied by upregulation of p21Cip1 and p27Kip1 expression via an autophagy-independent pathway. We also tested whether inhibition of mTOR with low con-centrations of rapamycin and ectopic Beclin-1 expression would further sensitize multidrug resistance (MDR)-posi-tive cancer cells by upregulating autophagy. Rapamycin at low concentrations might be insufficient to initiate auto-phagosome formation in autophagy but Beclin-1 overex-pression triggered additional processes downstream of mTOR during G1 cell cycle arrest by rapamycin. Our find-ings suggest that these combination strategies targeting autophagic cell death may yield significant benefits for cancer patients, because lowering rapamycin concentra-tion for cancer treatment minimizes its side effects in pa-tients undergoing chemotherapy.