Reactivation of p53 mutants by prima-1 [corrected] in thyroid cancer cells

Abstract

Most undifferentiated thyroid carcinomas express p53 mutants and thereafter, are very resistant to chemotherapy. p53 reactivation and induction of massive apoptosis (Prima-1) is a compound restoring the tumor-suppressor activity of p53 mutants. We tested the effect of Prima-1 in thyroid cancer cells harboring p53 mutations. Increasing doses of Prima-1 reduced viability of thyroid cancer cells at a variable extent (range 20-80%). Prima-1 up-regulated p53 target genes (p21(WAF1) , BCL2-associated X protein (Bax), and murine double minute 2 (MDM2)), in BC-PAP and Hth-74 cells (expressing D259Y/K286E and K286E p53 mutants) but had no effect in SW1736 (p53 null) and TPC-1 (expressing wild-type p53) thyroid cancer cells. Prima-1 also increased the cytotoxic effects of either doxorubicin or cisplatin in thyroid cancer cells, including the chemo-resistant 8305C, Hth-74 and BC-PAP cells. Moreover, real-time PCR and Western blot indicated that Prima-1 increases the mRNA of thyroid-specific differentiation markers in thyroid cancer cells. Fluorescence-activated cell sorting analysis revealed that Prima-1 effect on thyroid cancer cells occurs via the enhancement of both cell cycle arrest and apoptosis. Small interfering RNA experiments indicated that Prima-1 effect is mediated by p53 mutants but not by the p53 paralog p73. Moreover, in C-643 thyroid cancer cells, forced to ectopically express wild-type p53, Prima-1 prevented the dominant negative effect of double K248Q/K286E p53 mutant. Finally, co-IP experiments indicated that in Hth-74 cells Prima-1 prevents the ability of p53 mutants to sequestrate the p53 paralog TAp73. These in vitro studies imply that p53 mutant reactivation by small compounds may become a novel anticancer therapy in undifferentiated thyroid carcinomas.