Clinical trials using
kinase inhibitors have demonstrated transient partial responses and disease control in patients with progressive
medullary thyroid cancer (MTC). The goal of this study was to identify potential combinatorial strategies to improve on these results using
sorafenib, a multikinase inhibitor with activity in MTC, as a base compound to explore signaling that might predict synergystic interactions. Two human MTC cell lines, TT and MZ-CRC-1, which harbor endogenous C634W or M918T RET mutations, respectively, were exposed to
sorafenib,
everolimus, and
AZD6244 alone and in combination. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl tetrasodium
bromide (MTT) and
poly (ADP-ribose) polymerase (PARP) cleavage assays were performed to measure cell survival and apoptosis. Western blots were performed to confirm activity of the compounds and to determine possible mechanisms of resistance and predictors of synergy. As a solitary agent,
sorafenib was the most active compound on MTT assay. Western blots confirmed that
sorafenib,
everolimus, and
AZD6244 inhibited their anticipated targets. At concentrations below its IC(50),
sorafenib-treated TT and MZ-CRC-1 cells demonstrated transient inhibition and then re-activation of Erk over 6 h. In concordance, synergistic effects were only identified using
sorafenib in combination with the
Mek inhibitor
AZD6244 (P<0.001 for each cell line). Cells treated with
everolimus demonstrated activation of Akt and Ret via
TORC2 complex-dependent and
TORC2 complex-independent mechanisms respectively.
Everolimus was neither additive nor syngergistic in combination with
sorafenib or
AZD6244. In conclusion,
sorafenib combined with a
Mek inhibitor demonstrated synergy in MTC cells in vitro. Mechanisms of resistance to
everolimus in MTC cells likely involved TORC2-dependent and TORC2-independent pathways.