8537 Background: The advent of targeted therapy has revolutionized the treatment of cancer. The mutant BRAFV600E protein is found in over 50% of melanomas and thyroid carcinomas, resulting in elevated kinase activity, increased mitogen-activated protein kinase (MAPK) pathway signaling, and cell proliferation. Vemurafenib and PLX4720 were designed to selectively inhibit the BRAF kinase, and clinical trials of vemurafenib in metastatic melanoma have demonstrated a response rate of over 50% and an overall survival advantage over standard dacarbazine therapy. Approximately 20-30% of individuals treated with vemurafenib develop cutaneous squamous cell carcinoma (cSCC) highlighting the importance of understanding toxicities associated with this drug. Paradoxical ERK activation in BRAF wild-type, RAS-mutant cells is thought to be the major mechanism by which this occurs, as evidenced by the presence of RAS mutations in 60% of such lesions. Methods: Using a combination of BRAF-wild-type cSCC cell lines, primary human keratinocytes, as well as a UV mouse model of cSCC and human cSCC samples, we identified novel effects of BRAFi on apoptosis. Results: Here we show an unexpected and novel effect of vemurafenib and PLX4720 in suppressing apoptosis through the inhibition of multiple off-target kinases. JNK signaling and apoptosis are suppressed in cSCC lesions arising in vemurafenib-treated patients as well as in irradiated mouse skin. This occurs independently of paradoxical ERK signaling and in the presence of MEK inhibitor. Treatment with PLX4720 greatly accelerates the development of UV-induced cSCC in mice without Ras mutations. Kinome screening identified ZAK and MKK4 (MEK4 / MAP2K4) kinases as inhibited by vemurafenib, leading to suppression of MKK4 and MKK7 (MAP2K7) phosphorylation. Knockdown of inhibited off-target kinases recapitulates these anti-apoptotic effects of vemurafenib. Conclusions: Our results implicate suppression of JNK signaling, independent of ERK activation, as an additional, complementary mechanism of adverse effects of vemurafenib. This has broad implications for combination therapies with other modalities that induce apoptosis and for the long-term use of vemurafenib in the adjuvant setting.
An Engrailed1 enhancer underlies human thermoregulatory evolution
