Inhibition of the PI3K and mTOR survival pathways enhances apoptosis in human mesothelioma tumor fragment spheroids

Abstract

Malignant mesothelioma exhibits significant resistance to apoptosis. We have characterized a novel in vitro model of human mesothelioma tissue grown in culture as tumor fragment spheroids to identify the mechanisms of resistance. Spheroids maintained many features of the original tumors, including the presence of viable proliferating mesothelioma cells, for up to 3 months. Mesothelioma cell monolayers or 2-week old human mesothelioma spheroids were exposed to the death receptor ligand TRAIL and cycloheximide (CHX) for 48 h and studied for apoptosis by expression of cleaved caspase 3 in mesothelioma (cytokeratin-positive) cells. Unlike the nearly 100% apoptosis in the monolayers, TRAIL plus CHX induced apoptosis in only 32 ± 4% (mean ± SEM) of mesothelioma cells within spheroids (versus 7 ± 1% in untreated spheroids). Apoptosis of mesothelioma cells in spheroids increased to 47 ± 6% by inhibition of the PI3K pathway with LY294002 and to 50 ± 17% by inhibition of the mTOR pathway with rapamycin. Inhibition of the pathways was confirmed with staining for p-Akt or p70-S6K, targets of the PI3K and mTOR pathways, respectively. Targeting these survival pathways therefore presents a promising approach for mesothelioma and perhaps other tumors resistant to apoptosis. This research was supported by an NIH R01 NCI-095671 to VCB.