Silencing Mutated β-Catenin Inhibits Cell Proliferation and Stimulates Apoptosis in the Adrenocortical Cancer Cell Line H295R

Context: Adrenocortical carcinoma (ACC) is a rare and highly aggressive endocrine neoplasm, with limited therapeutic options. Activating beta-catenin somatic mutations are found in ACC and have been associated with a poor clinical outcome. In fact, activation of the Wnt/beta-catenin signaling pathway seems to play a major role in ACC aggressiveness, and might, thus, represent a promising therapeutic target. Objective: Similar to patient tumor specimen the H295 cell line derived from an ACC harbors a natural activating beta-catenin mutation. We herein assess the in vitro and in vivo effect of beta-catenin inactivation using a doxycyclin (dox) inducible shRNA plasmid in H295R adrenocortical cancer cells line (clone named sh beta). Results: Following dox treatment a profound reduction in beta-catenin expression was detectable in sh beta clones in comparison to control clones (Ctr). Accordingly, we observed a decrease in Wnt/beta catenin-dependent luciferase reporter activity as well as a decreased expression of AXIN2 representing an endogenous beta-catenin target gene. Concomitantly, beta-catenin silencing resulted in a decreased cell proliferation, cell cycle alterations with cell accumulation in the G1 phase and increased apoptosis in vitro. In vivo, on established tumor xenografts in athymic nude mice, 9 days of beta-catenin silencing resulted in a significant reduction of CTNNB1 and AXIN2 expression. Moreover, continous beta-catenin silencing, starting 3 days after tumor cell inoculation, was associated with a complete absence of tumor growth in the sh beta group while tumors were present in all animals of the control group. Conclusion: In summary, these experiments provide evidences that Wnt/beta-catenin pathway inhibition in ACC is a promising therapeutic target.