Abstract A34: Inhibition of autophagy sensitizes gastrointestinal stromal tumor cells to TKI/Bcl-2 inhibitors-induced apoptosis
Authors: Zhang Shuchao , Hu Guozhi , Paz-mejia Ana cristina , Li Luyuan , Trent Jonathan C. 
Published: August 2018

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 that the cell survival percentage of double- or triple-drug combinations dramatically decreased (less than 5%) compared to that of single-drug treatment (42%, 36%, or 12%) (P&lt;0.05). Isobologram analysis revealed a strongly synergistic drug interaction, with combination indices (CI) &lt; 0.5 in most of double and triple combinations. Importantly, triple-drugs combination showed even more strongly synergistic effects compared to double-drugs combinations (CI =0.204 vs. 0.309 or 0.356, P&lt;0.05). Cell apoptosis assay showed that the cell apoptosis percentage of double- (32.9% or 36.6%) or triple-drugs combinations (66.5%) also significantly increased over that of single-drug treatments (6.1%, 6.1%, or 13.1%) (P&lt;0.05). Finally, Western blot discovered that double- or triple-drugs combinations increased the cleavage of PARP as well as expression of Caspase-3, but inhibited autophagy in comparison to single-drug treatments.

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.

Citation Format: Shuchao Zhang, Guozhi Hu, Ana cristina Paz-mejia, Luyuan Li, Jonathan C. Trent. Inhibition of autophagy sensitizes gastrointestinal stromal tumor cells to TKI/Bcl-2 inhibitors-induced apoptosis [abstract]. In: Proceedings of the AACR Conference on Advances in Sarcomas: From Basic Science to Clinical Translation; May 16-19, 2017; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(2_Suppl):Abstract nr A34.