Enhancing mitotane efficacy in adrenocortical carcinoma by calcineurin inhibition with cyclosporine A

Abstract

Aims: The aim of this study is to determine the effect of calcineurin (CaN) in adrenocortical cancer (ACC) cells, which is a rare but aggressive type of cancer resistant to mitotane therapy. The intracellular calcium signaling pathway is one of the most important mechanisms for cells. The effect of intracellular calcium concentration [(Ca2+i)] on the function of cancer cells is also known. CaN, activated by the binding of calmodulin and Ca2+, is critical in this pathway. Methods: H295 adrenocortical cancer cells were treated with mitotane, cyclosporine A (CsA), and a combination of both. Cell viability, apoptosis, cell cycle, and gene expression levels of apoptosis-related genes (BCL2, BAX, TP53) were analyzed. Western blotting was used to measure CaN protein levels, and wound healing assays assessed cell migration. Results: CsA significantly suppressed CaN protein levels in a dose-dependent manner, reducing cell viability and increasing apoptosis in H295 cells. Mitotane alone also suppressed CaN protein, but the combination of mitotane and CsA had a synergistic effect, further decreasing cell viability and increasing apoptosis. The combination treatment led to significant suppression of the BCL2 gene and upregulation of TP53. Cell cycle analysis showed increased arrest in the G0/G1 phase with combination treatment. Conclusion: Suppression of CaN by CsA enhances the cytotoxic effects of mitotane on ACC cells, suggesting a potential therapeutic strategy to improve ACC treatment outcomes. This study highlights the importance of targeting intracellular calcium signaling pathways to overcome resistance and enhance the efficacy of existing cancer therapies.