Development of Novel Second Generation DC/Tumor Fusion Vaccine in Lymphoma

Abstract

Our personalized cancer vaccine comprises patient tumor cells fused with autologous dendritic cells (DCs) and limits the risk of antigen escape by presenting a broad array of tumor antigens in the context of DC mediated co-stimulation. In clinical trials of hematologic malignancies patients, vaccination induced expansion of tumor-specific T cells (Tc) and led to prolonged remission in a subset of patients. Currently, we are developing a novel second-generation (2G) vaccine. Fusions are presented in the context of a unique biomatrix expressing high levels of 41BB ligand (41BBL), to further accentuate Tc activation. In this study, we show the efficacy of vaccination in a preclinical lymphoma model and enhanced potency of the 2G vaccine. We first showed the fusions potency in generating anti-tumor immunity in the A20 lymphoma murine model. Bone marrow derived mononuclear cells cultured with GM-CSF and IL-4 were fused to A20 cells. Fusions effectively induced tumor specific immunity as manifested by potent lysis of A20 Tc in vitro as compared to unstimulated Tc in a CTL assay. Consistently, vaccine effectively induced tumor specific immunity in an immunocompetent murine model. Balb/C mice (30) underwent IV inoculation with 750,000, luciferase transduced, A20 cells. 24h post inoculation, 15 mice were vaccinated with 105 fusions. 10 days post inoculation, in the untreated cohort all mice had detectable tumor whereas in the treated group, 5 mice didn't show disease and 5 mice showed minimal disease as detected by BLI imaging. We further showed that patient autologous vaccine stimulated Tc mediated lysis of primary lymphoma cells. DC, generated from patient's peripheral blood mononuclear cells cultured with GM-CSF and IL-4 and matured with TNFa were fused to primary lymphoma cells isolated from resected tumor. In a standard CTL assay, fusion stimulated Tc potently lysed autologous tumor cells as compared to unstimulated Tc (25.7% as compared to 12.66%). To enhance vaccine potency, we developed a biomatrix expressing 41BBL. By carbodiimide chemistry we covalently bonded 41BBL protein to an alginate (Alg)-based scaffold, promoting a supporting microenvironment for Tc and fusions co-culture. We cultured syngeneic Tc with fusions within a scaffold with or without bound 41BBL and examined Tc cytotoxicity by a CTL assay. Tc stimulated by vaccine within the Alg/41BBL scaffold showed higher levels of tumor lysis compared to the percent killed by Tc cultured within an Alg scaffold (22.95% and 13.95 respectively). In this study we succeeded in demonstrating the fusions capacity to generate tumor specific Tc cytotoxicity. We presented patient derived tumor results supporting the applicable nature of the vaccine. In addition, we developed a 2G vaccine comprised of the original vaccine presented to the Tc in an Alg/41BBL scaffold acting as a nurturing microenvironment for Tc tumor specific immune response.