Synergistic action of a RAF inhibitor and a dual PI-3 Kinase / mTOR inhibitor in thyroid cancer

Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Abstract Purpose: In thyroid cancer clinical trials, agents targeting VEGF receptors and RET, among other kinases, have led to partial responses but few complete or durable responses. The RAF-MEK-ERK and PI3K-AKT-mTOR signaling pathways are frequently activated in differentiated and medullary thyroid cancer (DTC and MTC) and may provide therapeutic targets for these diseases. We tested a novel drug combination targeting RAF, PI3K and mTOR, plus VEGFR2 and RET, in thyroid cancer preclinical models with defined genetic backgrounds. Experimental Design RAF265, an ATP-competitive pan-RAF inhibitor active against VEGFR2, and BEZ-235, a PI3K inhibitor also active against Torc1 and Torc2, were tested alone and in combination in a panel of thyroid cancer lines. We tested RAF265 and BEZ-235 for kinase inhibition, growth inhibition and cell cycle alterations, and inhibition of signaling targets and tumor growth in xenograft models. Results Both drugs potently inhibited their kinase targets in the ERK and PI3K pathways. In addition, RAF265 had significant RET inhibitory activity (IC50 25-50 nM for RET C634W). The combination strongly inhibited proliferation of DTC and MTC lines with mutations in RAS, BRAF, PTEN, and RET. Synergy was demonstrated for B-CPAP (BRAF V600E) and TT cells (RET C634W). The combination of both drugs significantly inhibited growth of CAL62 (KRAS G12R/G12R) and TT xenografts, thoroughly inhibiting ERK and PI3K pathway signaling. Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Conclusions Combined blockade of ERK and PI3K signaling potently inhibits growth in preclinical models representing the key genotypes seen in refractory thyroid cancer. These targets and therapies are promising for further development in both DTC and MTC. The submitted manuscript is translational-relevant for several reasons. First, the clinical problem-advanced thyroid cancer results in approximately 1700 deaths per year in the U.S. There are no approved therapies for the majority of cases, although vandetanib recently was approved for advanced medullary thyroid cancer. Our preclinical development in thyroid cancer is aimed at this unmet need. Second, the signaling pathways that we examined are the two pathways most commonly activated by mutation in thyroid cancer. Third, the model systems that we chose are standard preclinical models for this field. Fourth, the drugs that we studied have clinical activity in other solid tumor types and are proceeding with development. Fifth, the preclinical activity that we observed …