PL2.1 Exploiting the DNAM-1 system for chimeric antigen receptor (CAR) T cell therapy of glioblastoma

Abstract

Abstract BACKGROUND Cancer immunotherapy with genetically engineered T cells that express a chimeric antigen receptor (CAR) has led to impressive responses in extracranial malignancies and is also explored against glioblastoma. However, CAR T cell strategies that are currently being explored against glioblastoma target single tumor antigens, which are non-homogeneously expressed and are prone to antigen escape. Furthermore, the immunosuppressive brain tumor microenvironment hampers anti-tumor efficacy. METHODS By immunohistochemistry and flow cytometry, we investigated the expression of CD155 and CD112, which are ligands to the activating immune cell receptor DNAX accessory molecule-1 (DNAM-1), in human and mouse glioma cell lines as well as in human glioblastoma samples. To understand their functional role, we generated CD155 or CD112 knock-out glioma cell lines using CRISPR/Cas9 and studied proliferation, sensitivity to irradiation or temozolomide as well as migration. To exploit the promiscuous binding features of DNAM-1, we generated different first or second-generation CAR T cells that use DNAM-1 as a tumor-binding domain. Subsequently, we investigated their anti-tumor activity in vitro in co-culture assays and in vivo in syngeneic orthotopic murine glioma models. RESULTS CD155 and CD112 are homogenously expressed in human and mouse glioma cell lines and human glioblastoma tissues. Knock-out of these ligands affected the migration of tumor cells, but did not affect proliferation or sensitivity to irradition or temozolomide. DNAM-1-based CAR T cells demonstrated high cytolytic activity and effector cytokine secretion in vitro. In vivo, DNAM-1 based CAR T cells reached to the tumor site in the brain upon intravenous administration, prolonged survival of orthotopic glioma-bearing mice and led to a durable anti-tumor response in a fraction of mice. The treatment was tolerated without toxicities. CONCLUSION We elucidated the tumor-intrinisic role of CD155 and CD112 and provide the first systematical preclincal assessment of DNAM-1 CAR T cells against glioma. These findings provide a rationale to test this immunotherapeutic strategy also in human glioma patients.