Nitric oxide S-nitrosylates CSF1R to augment the action 1 of CSF1R inhibition against castration resistant prostate cancer

During progression of prostate cancer, sustained oxidative overload in cancer cells potentiates the overall tumor microenvironment (TME). Targeting the TME using colony-stimulating factor 1 receptor (CSF1R) inhibition is a promising therapy for castration-resistant prostate cancer (CRPC). However, the therapeutic response to sustained CSF1R blockade therapy (CSF1Ri) is limited as a monotherapy. We postulated that one of the causative agents for reduced efficacy of CSF1Ri and increased oxidation in CRPC is endothelial nitric oxide syntheses (eNOS). Results showed that in high grade PCa human specimens, eNOS is positively correlated with CSF1-CSF1R signaling and remains in an un-coupled state. The uncoupling disables eNOS to generate sufficient Nitric oxide (NO) that are required for inducing effective S-nitrosylation of CSF1R molecule at specific cysteine sites (Cys 224, Cys 278 and Cys 830). Importantly, we found that S-nitrosylation of CSF1R molecule at Cys 224, Cys 278 and Cys 830 sites is necessary for effective inhibition of tumor promoting cytokines (which are downstream of CSF1-CSF1R signaling) by CSF1R blockade. In this context, we studied if exogenous NO treatment could rescue the side effects of eNOS uncoupling. Results showed that exogenous NO treatment (using S-nitrosoglutathione (GSNO)) is effective in not only inducing S-Nitrosylation of CSF1R molecule, but it helps in rescuing the excess oxidation in tumor regions, reducing overall tumor burden, suppresses the tumor promoting cytokines which are ineffectively suppressed by CSF1R blockade. Together these results postulated that NO therapy could act as an effective combinatorial partner with CSF1R blockade against CRPC. In this context, results demonstrated that exogenous NO treatment successfully augment the anti-tumor ability of CSF1Ri in murine models of CRPC. Importantly, the overall tumor reduction was most effective in NO-CSF1Ri therapy compared to NO or CSF1Ri mono therapies. Moreover, Immunophenotyping of tumor grafts showed that the NO-CSF1Ri combination significantly decreased intratumoral percentage of anti-inflammatory macrophages, myeloid derived progenitor cells and increased the percentage of pro-inflammatory macrophages, cytotoxic T lymphocytes, and effector T cells respectively. Together, our study suggests that the NO-CSF1Ri combination has the potential to act as a therapeutic agent that restore control over TME and improve the outcomes of PCa patients.