Inhibition of autophagy sensitizes gastrointestinal stromal tumor cells to TKI/Bcl-2 inhibitors-induced apoptosis.

Abstract

11036 Background: Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumor of the GI tract. Most GISTs are driven by mutations in KIT or platelet-derived growth factor receptor-α (PDGFRA), which responds well to imatinib, a tyrosine kinase inhibitor (TKI) that blocks KIT and PDGFR-α signaling. Bcl-2 family plays a critical role in the regulation of cell apoptosis in GISTs. ABT-737 as an inhibitor of Bcl-2/Bcl-xL can result in a time and dose-dependent activation of apoptosis. Autophagy is a key mechanism to promote tumor cells survival, inhibition of which can induce the cell death in GISTs. Chloroquine, an antimalarial drug, has been also identified as an autophagy inhibitor. In this study, we assessed the combinational effects of imatinib, ABT-737 and chloroquine in GIST cells. Methods: Human GIST cell lines, GIST-T1 and GIST-882, were employed in our study. Cells were treated with imatinib, ABT-737 and chloroquine either separately or in different combinations. Cell viability was tested by means of MTS and synergistic effects were analyzed by isobologram software. The levels of related proteins of apoptosis (PARP, Caspase-3) and autophagy (LC3-II, beclin-1) were measured by western blot. Cell apoptosis and cell cycle were tested by flow cytometry. Results: Cell viability assay indicated cell survival percentage of double or triple drug combinations ( < 5%) dramatically decreased compared to single drug treatments (42%, 36% or 12%) ( P< 0.05). Isobologram analysis revealed triple drugs combination had stronger synergistic effects than double drugs combinations (CI = 0.204 vs 0.309 or 0.356, P< 0.05). Cell apoptosis percentage of double (32.9% or 36.6%) or triple drugs combinations (66.5%) significantly increased compared to single treatments (6.1%, 6.1% or 13.1%) ( P< 0.05). Western blot showed drugs combinations increased cleavage of PARP and Caspase-3 levels, but inhibited autophagy. Conclusions: The combination of imatinib, ABT-737 and chloroquine has collaborative effects on the treatment of GISTs in vitro. The combined strategy may enhance the clinical efficacy, which provides a rationale for the clinical evaluation of these drug combinations in GISTs treatment.