Silencing mutated β-catenin inhibits cell proliferation and stimulates apoptosis in the adrenocortical cancer cell line H295R

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