Vs-6063 (Defactinib) Targets Mesothelioma Cancer Stem Cells Directly and Through Inhibition of Tumor-Associated Macrophages and Cytokine Production

Abstract

ABSTRACT Aim: Focal adhesion kinase (FAK) is a non-receptor tyrosine kinase required for survival of cancer stem cells (CSCs). PYK2, the other FAK family member, shares 60% kinase domain homology. Based on the observation of reduced macrophage infiltration in PYK2 knockout mice, we investigated effects of PYK2 inhibition on tumor-associated macrophages (TAMs). Abundance of TAMs has been correlated with poor prognosis in multiple cancer types including mesothelioma. Defactinib is a potent, orally active FAK kinase inhibitor that has completed phase I and is currently in clinical development in several indications including mesothelioma. Methods: Cytokine release from human macrophages was measured in response to small molecule FAK kinase inhibitors. Serum IL-6 and F4/80-positive macrophages were assessed by ELISA and immunofluorescence respectively. Results: In addition to potent FAK inhibition, defactinib also inhibits PYK2 kinase activity with nanomolar potency. Using both FAK inhibitors and FAK siRNA, we demonstrated that FAK inhibition effectively reduces CSCs. Human monocyte-derived macrophages release IL-6 and IL-8, and these cytokines increased the proportion of ALDH-positive CSCs in mesothelioma and breast cancer cell lines. Defactinib dose-dependently inhibited production of IL-6 and IL-8, whereas a FAK-only reference inhibitor had no effect on these cytokines. Accordingly, PYK2 siRNA decreased IL-6 and IL-8 release from human macrophages. In a xenograft model in vivo, defactinib reduced serum IL-6 while the FAK-only reference compound did not. Furthermore, defactinib substantially reduced the number of F4/80-positive macrophages in the tumors. Conclusions: These data suggest that dual targeting of FAK and PYK2 by defactinib may more effectively suppress CSCs than inhibition of FAK alone. Given the sensitivity of mesothelioma CSCs to defactinib, the use of this agent in a maintenance setting after first line chemotherapy may be an attractive strategy to extend survival of patients with mesothelioma. These data support the ongoing COMMAND study with defactinib in mesothelioma (Clinical trial NCT01870609). Disclosure: M. Keegan: Employee of Verastem, Inc. Stock Ownership in Verastem, Inc.; J.E. Ring: Employee of Verastem, Inc. Stock Ownership in Verastem, Inc.; V.N. Kolev: Employee of Verastem, Inc. Stock Ownership in Verastem, Inc.; I.M. Shapiro: Employee of Verastem, Inc. Stock Ownership in Verastem, Inc.; M.V. Padval: Employee of Verastem, Inc. Stock Ownership in Verastem, Inc.; Q. Xu: Employee of Verastem, Inc. Stock Ownership in Verastem, Inc.; J.A. Pachter: Employee of Verastem, Inc. Stock Ownership in Verastem, Inc.