EPEN-26. Chemokine receptor blockade reverses CCL2 mediated immunosuppression and restores CAR-T cell function in posterior fossa ependymoma

Abstract

Trastuzumab-based HER2 CAR-T constructs have demonstrated preclinical efficacy in medulloblastoma and are being evaluated for use in children and young adults with recurrent or refractory CNS tumors. Preliminary results demonstrate immune activation but no objective tumor response in three patients, including two patients with posterior fossa (PF)-EPN. A key finding in the serum and CSF of all three patients was very high levels of the inflammatory chemokine CCL2 following treatment with CAR-T cells. Preclinical studies suggest that high levels of CCL2 may impede T cell mediated anti-tumor activity in CNS tumors. The role of CCL2 to enhance or diminish CAR-T cell efficacy for CNS tumors is unknown. We evaluated a second generation trastuzumab-based HER2 CAR construct with a 4-1BB co-stimulatory domain in two ultra-high-risk patient-derived xenograft (PDX) models that faithfully recapitulate PFA-EPN. In contrast to preclinical studies in other cancers, treatment with trastuzumab-based HER2 CAR-T cell alone causes only partial regression of tumors and robust infiltration of immunosuppressive monocytes in PFA-EPN PDX mouse models. We studied constitutive NF-kB activation because it is a hallmark of PFA-EPN that drives dysregulation of inflammatory genes and forms an immunosuppressive tumor microenvironment. Upon tumor recognition, CAR-T cells produce high amounts of the cytokine tumor necrosis factor-alpha, which is an extracellular stimulus that propagates NF-kB activation in PFA-EPN. We show that HER2 CAR-T cell treatment causes increased nuclear translocation of the RELA NF-kB subunit, which induces CCL2 gene transcription and chemokine release. This results in CCL2-CCR2 ligand/receptor mediated influx of inflammatory monocytes and regulatory T cells, impairing CAR-T cell effector function. Inhibition of CCR2 restores anti-tumor CAR-T cytotoxicity against bulky orthotopic tumors by decreasing the infiltration of inflammatory monocytes and regulatory T cells. Combinatorial strategies addressing tumor mediated immunosuppression should be evaluated in upcoming CAR-T cell trials for patients with high-risk CNS tumors.