Endocrine Abstracts

Background: Adrenocortical carcinoma (ACC) is a rare endocrine-related cancer with limited treatment options. The current therapeutic standard is mitotane, which is adrenolytic via unknown mechanisms. Many chemotherapeutic drugs are known to induce apoptosis and autophagy. The role of autophagy is of particular interest as it can promote either tumor cell survival or death.

Objective: We characterized the autophagic responses in H295R, CU-ACC1, and CU-ACC2 cell lines, hypothesizing that mitotane induces both apoptosis and autophagy and might synergize with autophagy inhibitors such as chloroquine (CQ) or bafilomycin (BAF).

Methods: Proliferation, MTS, and colony formation assays were performed to characterize ACC cell line sensitivity to mitotane. Western blots assessed cleaved PARP and LC3-II accumulation. Incucyte monitoring of caspase 3/7 activity determined mitotane-induced apoptosis with genetic silencing of critical autophagic regulator, ATG5, with or without autophagy inhibitors CQ and BAF.

Results: H295R, CU-ACC1, and CU-ACC2 show differential dose-dependent apoptotic and autophagic responses to mitotane. Silencing of ATG5 in mitotane-treated cells increased cell death in CU-ACC2, but not H295R. CQ and BAF synergized with mitotane in CU-ACC1 and CU-ACC2, but less in H295R. Genetic knockdown of ATG5 in CQ-treated cells did not further augment cell death. In addition, treatment with other autophagy inhibitors, VPS34-IN1 or SBI-0206965, did not increase caspase 3/7 expression.

Discussion: Mitotane induces autophagy differentially in ACC cell lines, correlating to their sensitivities to CQ. Autophagy is a targetable pathway in ACC. CQ, especially in combination with mitotane, is an effective treatment to induce tumor cell death. CQ induces cell death via autophagic and/or non-autophagic mechanisms. These data support the potential use of CQ in patients with ACC. Funded by VA Merit Review to MEW/KKV and laboratory funds