The Cerebroventricular Environment Reprograms Locally Infused CAR T Cells for Superior Activity Against Both CNS and Systemic B Cell Lymphoma
Authors: Wang Xiuli , Huynh Christian , Urak Ryan , Walter Miriam , Weng Lihong , Lim Laura , Vyas Vibhuti , Aguilar Brenda , Brito Alfonso , Sarkissian Aniee , Bandara Achini , Budde Elizabeth E , Popplewell Leslie , Siddiqi Tanya , Brown Christine , Forman Stephen J 
Published: October 2019

Abstract

Central nervous system lymphoma (CNSL) is a lymphoid malignancy in which tumors from lymph tissue start in the brain, spinal cord, eyes, and/or meninges (primary CNSL) or present as a result of metastasis from initial systemic sites to the CNS (secondary CNSL). The incidence of primary CNS lymphoma has been increasing over the past 20 years. CNS lymphomas carry a worse prognosis than systemic lymphoma, and therefore, effective treatment is urgently needed for CNS disease.
T cells that are genetically engineered with chimeric antigen receptors (CAR) targeting CD19 have broad applications in adoptive therapy of B cell malignancies and have shown tremendous potential in the treatment of systemic lymphoma. During the early phase of CD19-CAR T cell studies, most if not all protocols excluded patients with active CNS involvement. In all CD19-CAR T cell trials, T cell products are administrated intravenously. Systemic CD19-CAR T administration for ALL and DLBCL has resulted in complete remission of concurrent CNS disease. CD19-CAR T cell trafficking to the cerebrospinal fluid (CSF) is frequently reported; however, there has been no evidence thus far to indicate that CAR T cells in CSF are related to neurotoxicity. Therefore, an increasing number of CD19-CAR T cell trial protocols no longer exclude patients with active CNS lymphoma involvement. Based on the success of CD19-CAR T cell therapy in ALL and lymphoma, we aimed to translate this strategy toward a more effective therapy for CNS B cell disease.
Methods and Results: Isolated naïve and central memory T cells were genetically modified with CD19-CAR lentivirus and expanded in vitro for 14 days. A mouse model with both CNS and systemic lymphoma in the same animal was established by simultaneously engrafting Daudi cells (human B cell lymphoma) intracranially and subcutaneously into NSG mice. We then administered 2x10^6 CD19-CAR T cells via two delivery routes: intracerebroventricular (i.c.v.) to bypass the blood-brain barrier and target tumor throughout the entire CNS, and intravenous injection (i.v.). We repeatedly observed that a single i.c.v. infusion was capable of completely eradicating CNS lymphoma and systemic lymphoma in all mice by day 14 post CAR T cell infusion and 100% of mice remained tumor free for 300 days until the termination of the experiment. In contrast, a single delivery of CD19-CAR T cells via i.v. infusion resulted in a noticeably delayed antitumor activity with complete remissions only observed approximately 40 days post CAR T cell treatment. Eventually, the tumors relapsed and all i.v. treated mice died before day 180 (Figure 1).
T cell trafficking experiments demonstrated that i.c.v. CAR T cells are able to efficiently migrate to the periphery, home to systemic tumor locations, and dramatically expand outside the CNS. We were able to detect CAR T cells in the blood, bone marrow, and spleens of mice that received i.c.v. therapy at 300 days post CAR T cell treatment. These persisting T cells are CD4 dominant and express high levels of CD28 with a broad TCR repertoire. The persisting T cells also maintain anti-tumor functionality and are able to resist tumor re-challenge. Further mechanistic studies indicate that factors within the CSF are able to reprogram i.c.v. infused CAR T cells and upregulate genes that are related to memory function.
In conclusion, our studies suggest that CAR T cells administrated via i.c.v. and nurtured by CSF exhibit better efficacy, expansion, and persistence, resulting in disease elimination. More interestingly, i.c.v. delivered CAR T cells efficiently traffic beyond the CNS to the periphery and completely eradicate systemic tumors in the same mouse. This study is the first to demonstrate that locally delivered CAR T cells are capable of efficiently treating both systemic lymphoma and concurrent CNS disease, which can lower the risk of cytokine release syndrome and avoid toxicities derived from lymphodepletion and systemic infusion of CAR T cells.

Disclosures
Wang: Mustang Therapeutics: Other: Licensing Agreement, Patents & Royalties, Research Funding. Budde:Mustang Therapeutics: Consultancy, Other: Licensing Agreement, Patents & Royalties, Research Funding. Brown:Mustang Therapeutics: Consultancy, Other: Licensing Agreement, Patents & Royalties, Research Funding. Forman:Mustang Therapeutics: Other: Licensing Agreement, Patents & Royalties, Research Funding.