The tyrosine kinase inhibitor nilotinib is more efficient than mitotane in decreasing cell viability in spheroids prepared from adrenocortical carcinoma cells

Abstract\ud \ud Background\ud New drugs for adrenocortical carcinoma (ACC) are needed because most patients undergo rapid disease progression despite surgery and adjuvant therapy with mitotane. In this study, we aimed to investigate the in vitro effects of different chemotherapy drugs, alone or combined with mitotane, on the viability of adrenocortical carcinoma cells.\ud \ud \ud Methods\ud Everolimus, sunitinib, zoledronic acid, imatinib and nilotinib cytotoxicity, alone or combined with mitotane were tested on ACC H295R cells in monolayer or spheroid cultures using MTS assays and confocal microscopy. Moreover, the nilotinib effects were investigated in spheroids cultured from patient tumor-derived ACC-T36 cells.\ud \ud \ud Results\ud Morphological characterization of H295R cell spheroids using histochemistry was performed and showed that dense, homogenously sized, multicellular spheroids were obtained. We observed that sunitinib and nilotinib alone were equally effective in a monolayer preparation, whereas mitotane was the most effective even at a low dose. A combination of sunitinib and mitotane was the most effective treatment, with only 23.8% of cells in the monolayer remaining viable. Spheroid preparations showed resistance to different drugs, although the poor effect produced by mitotane alone was surprising, with a cell viability of 84.6% in comparison with 13.1% in monolayer cells. The most ineffective drugs in spheroid preparations were everolimus, zoledronic acid and imatinib. In both cell types, nilotinib, either alone or in combination with mitotane induced more significant cell viability inhibition in monolayer and spheroid preparations. In addition, the mechanism of nilotinib activity involves the ERK1/2 pathway.\ud \ud \ud Conclusion\ud Taken together, our data identified nilotinib as a cytotoxic drug that combined with ERK inhibitors deserves to be tested as a novel therapy for adrenocortical carcinoma.This work was supported by FAPESP, the São Paulo State Research Foundation (nº 2011/07656-3 and 2015/014199-9). CFPL received funding from Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) and ES and IPC were recipients of scholarships from Capes (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior)