Oncogene-targeted therapies are initially effective in most patients with BRAF-mutated melanoma. However, patients frequently develop resistance during therapy. At the time of therapy resistance, they are also less likely to respond to immune checkpoint inhibitors (ICI) with response rates ranging from 0-12%, compared to 40-60% when used in the first line. This may be in part be due to rapid tumor growth that outpaces pharmacodynamics of ICI. New drugs that directly control tumor growth and effectively enhance ICI activity are therefore needed. We report that the small molecule CX-6258 has potent activity against both RAF/MEK inhibitor (RMS) and resistant (RMR) BRAF-mutant melanoma cell lines. CX-6258 is annotated as a pan-PIM kinase inhibitor, with an EC50 corrected for growth rate (GR50) of ~200 nM; in contrast, other PIM inhibitors had no in vitro activity, suggesting that PIMs may not be the true target in melanoma - we therefore sought to systematically study the targets of this compound. Using an unbiased kinase profiling approach, we identified Haspin Kinase (HASPIN) as the key target of CX-6258 with an IC50~10nM. Haspin kinase (Haspin) is a poorly understood mitotic kinase, whose only well-defined function is phosphorylation of Histon H3 at Thr3 (pH3T3). Treatment with CX-6258 resulted in reduced proliferation, reduced phosphorylation of pH3T3, frequent formation of micronuclei (MN), recruitment of cGAS and activation of the cGAS-STING-pathway. By using CRISPR/CAS9 knock outs of PIM kinase we show that these targets are not required for CX-6258 function. Furthermore, siRNA mediated knock-down of HASPIN phenocopied CX-6258. In murine models, CX-6258 induces a potent cGAS dependent type-I-interferon response in tumor cells, increases IFNγ-producing CD8+ T-cells, reduces Tregs, and enhances responses to ICIs in vivo. HASPIN is more strongly expressed in malignant compared to healthy tissue and its inhibition by CX-6258 has minimal toxicity in ex vivo expanded human tumor-infiltrating-lymphocytes (TILs) and in vitro differentiated human neurons, suggesting a potential therapeutic index for anti-cancer therapy. CX-6258 also shows activity in several Ewing sarcoma and multiple myeloma cell lines with. In summary, HASPIN inhibition may overcome drug resistance in melanoma, synergize with ICIs and target a vulnerability in different cancer lineages.
Citation Format: Johannes C. Melms, Sreeram Vallabhaneni, Caitlin E. Mills, Kai Wucherpfennig, Peter K. Sorger, Benjamin Izar. Inhibition of Haspin kinase to promote cell-intrinsic and extrinsic anticancer therapy [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr B132. doi:10.1158/1535-7163.TARG-19-B132