Abstract
Background: Chimeric antigen receptor (CAR) T cell therapy has transformed outcomes in advanced B-cell lymphomas, achieving remarkable responses in a subset of patients with advanced disease. However, relapse is commonly observed. We have demonstrated that durable responses are associated with activation of the non-CAR T cell repertoire potentially overcoming therapeutic resistance arising from antigen escape and limited CAR T persistence (Cheloni et al., 2025). We have developed a personalized dendritic cell (DC)-based cancer vaccine capable of eliciting a polyclonal T cell response targeting a broad array of tumor antigens (Rosenblatt et al., 2016). We hypothesized that combining CAR T cells with vaccine-primed T cells (Vac T) would enhance therapeutic efficacy by pairing the potent cytotoxic activity of CAR T cells with a sustained polyclonal anti-tumor T cell response.
Methods: We evaluated the in vivo efficacy of this combination strategy in an immunocompetent Balb/c murine model of aggressive lymphoma (A20). Vac T were generated in vivo by vaccinating healthy mice with DC/A20 fusion cells followed by ex vivo T cell expansion with 4-1BB agonist and CD3/CD28 Dynabeads. Antigen specificity was confirmed by MHC-I tetramer staining. Mice bearing parental CD19⁺ A20 tumors or mixed CD19⁺/CD19⁻ A20 tumors were treated with Vac T, CD19-directed CAR T cells, or the combination. Tumor burden was evaluated by bioluminescence imaging of luciferase-tagged A20 cells.
Results: We previously showed that DC/A20 vaccine elicits a robust anti-lymphoma T cell response in vitro characterized by upregulation of activation markers and recognition of MHC-I-restricted tumor epitopes. Additionally, ex vivo vaccine priming enhanced CAR T activation and persistence, resulting in heightened tumor killing. These findings support the immunogenicity and functional potency of the vaccine platform and provide the foundation for the in vivo combinatorial strategy described here.
In mice bearing CD19⁺ A20 tumors, both CAR T and Vac T monotherapies reduced tumor burden compared to controls, though Vac T monotherapy did not confer survival benefit. Strikingly, the combination of CAR T and Vac T demonstrated a synergistic anti-tumor activity, with significantly lower tumor burden and a median survival not reached at 9 months post-tumor inoculation (vs 21, 22, and 46 days in mice treated with naïve T, Vac T, or CAR T cells, respectively). Similarly, in a model of limited CAR T cell efficacy characterized by high disease burden and suboptimal lymphodepletion, while neither monotherapy achieved statistical benefit over controls, the combination therapy demonstrated a survival advantage (p≤0.05 vs control) with some mice remaining tumor-free beyond 220 days. In an antigen-loss model, where tumors included a mixture of CD19⁺ and CD19⁻ A20 cells, CAR T monotherapy failed to improve survival, confirming immune escape. In contrast, Vac T significantly reduced tumor burden and improved survival, with the combination therapy efficiently targeting both tumor populations (median survival 40 days vs 22-34 in CAR T or Vac T alone; p vs naïve T≤0.001; p vs CAR T≤0.05).
Conclusions: Vac T synergize with CAR T cells to enhance tumor clearance, overcome antigen-loss escape, and improve therapeutic efficacy, even in resistant disease settings. These findings provide a strong preclinical rationale for clinical translation. A clinical trial combining DC/tumor fusion vaccination and CAR T therapy is planned.
