Effects of sterol-O-acyl transferase 1 (SOAT1) inhibitors in canine adrenocortical cells: in vitro investigation

Abstract

Current medical treatment options in dogs with cortisol-secreting adrenocortical adenomas (ACAs) and adrenocortical carcinomas (ACCs) are limited. Therefore, there is a strong need of new medical therapies. Recently, sterol-O-acyl-transferase 1 (SOAT1) inhibitors Sandoz 58-035 (S035) and ATR-101 regained attention. By inhibiting SOAT1, free cholesterol and fatty acids accumulate, leading to endoplasmic reticulum stress (ER-stress). ER-stress activates the unfolded protein response (UPR), ultimately leading to apoptosis and thereby decreased steroidogenesis. Inhibition of SOAT1 could thus be an interesting therapy option in dogs with cortisol-secreting adrenocortical tumors (ATs), but can only be useful if canine ATs express SOAT1.

The aim of this study was to determine whether the use of SOAT1 inhibitors could have potential in the treatment of canine ATs. Quantitative real-time polymerase chain reaction (RT-qPCR) and immunohistochemistry (IHC) revealed SOAT1 expression at the mRNA and protein level in all ACAs and ACCs. Expression did not significantly differ between the groups, although expression was more variable in ACCs. After 24 hours, the cortisol concentration, determined by radioimmunoassay (RIA), of ACTH-stimulated normal adrenocortical cells (n=4) incubated with mitotane or ATR-101 decreased. The cortisol concentrations at the maximum compound concentration (100 uM mitotane, 100 nM ATR- 101) were 32.9% ± 54.2 and 28.8% ± 31.2, respectively. S035 did not decrease the cortisol concentration of ACTH-stimulated normal adrenocortical cells. The cortisol concentrations of mitotane and ATR-101 incubated ACC cells (n=1) were decreased (8.5% and 65.5%, respectively). S035 did not reduce the cortisol concentration of ACC cells. Mitotane and ATR-101 increased apoptosis of non- stimulated normal adrenocortical cells (n=2), determined by the Caspase-Glo® 3/7 Assay, with 476.0% ± 132.5 and 433.5% ± 19.7, respectively. S035 had little effect on apoptosis of normal adrenocortical cells. In non-stimulated ACC cells (n=1), mitotane, S035 and ATR-101 increased apoptosis (842.7%, 372.8% and 571.0%, respectively). After 6 hours, the UPR seemed to be activated in mitotane incubated non-stimulated and ACTH-stimulated ACC cells. The UPR was activated to some extent in S035 and ATR-101 incubated non-stimulated and ACTH-stimulated ACC cells (n=1).

Current study has brought important preliminary data on SOAT1 inhibitors in dogs. It will serve as a solid base for further studies about SOAT1 inhibitors in humans and dogs, which we are setting up in cooperation with the Endocrine research group from the Medical Center from Würzburg, Germany.