CAR-T Cells Targeting BAFF-Receptor for B-Cell Malignancies: A Potential Alternative to CD19

Background: Chimeric antigen receptor (CAR) T-cell adoptive therapies have the potential to revolutionize the treatment of cancers. Particularly in hematological malignancies, there are reports of promising clinical outcomes in advanced non-Hodgkin lymphomas (NHLs) with CD19-CAR T-cell therapy. However, disease relapse is problematic, and is thought to be caused by poor long-term persistence of the CAR T-cells, and loss of the CD19 target on tumors. Thus, there is an urgent need for improved novel CAR T-cell therapies directed at alternative targets.

The CAR T-cell platform relies on antibody-derived single chain fragments (scFv), which are genetically engineered into chimeric T-cell receptors, and that recognize target cell surface proteins on tumors. One potential target is B-cell activating factor receptor (BAFF-R), a tumor necrosis factor receptor superfamily protein (TNFRSF13C) specifically involved in B lymphocyte development and mature B-cell survival, that is primarily expressed on B cells and various subtypes of B-cell NHLs and ALL. We have recently developed a humanized therapeutic BAFF-R antibody with strong anti B-cell tumor activity.

Method and Results: We adapted a scFv based on our humanized anti-BAFF-R antibody onto a second generation CAR platform containing CD3ζ and 4-1BB intracellular signaling domains. In response to BAFF-R-expressing malignant human B cells (NHLs, acute lymphoblastic leukemias, and chronic lymphocytic leukemias), our CAR T cells readily proliferated and secreted cytotoxic cytokines. We demonstrated both significant levels of BAFF-R CAR T-cell activation and malignant B-cell killing in vitro .

Established human NHL tumors in xenogeneic models were eliminated following BAFF-R CAR T-cell treatments in vivo . Remarkable tumor-free survival was repeatedly observed in human lymphoma xenograft models including JeKo-1 (mantle cell lymphoma) and Raji (Burkitt lymphoma) in NSG mice. Moreover, upon tumor re-challenge and without any further treatment tumors failed to develop, due to persisting CAR T cells conferring continued anti-lymphoma activity. We pursued optimization of CAR T-cell persistence by comparing three subsets of early stage T cells (central memory, T_CM; memory stem, T_SCM; and naïve, T_N) as potential starting material for CAR T cell generation. Our in vivo studies show CAR T cells from the T_N population retained highest potency eradicating established tumors at a minimal therapeutic dose compared to other subtypes; 80% of T_N CAR treated mice achieved long-term survival compared to 20% of T_CM and 40% of T_SCM CAR treated mice. We also observed the long term anti-tumor effects conferred by CD8+ CAR T cells required the addition of CD4+ CAR T cells. Finally, we performed a head-to-head comparison of BAFF-R CAR T cells with CD19 CAR T cells in the Raji model (Figure). Comparable expression levels of BAFF-R and CD19 were observed on the tumor cells (A), and both treatments elicited significant antitumor effects compared to T-cell and saline controls (B-C). However, the BAFF-R CAR T-cell treatment demonstrated long-term tumor free survival in all treated mice compared to the CD19 CAR T-cell treated cohort, which only showed delayed tumor growth (P<0.01, B-C).

Conclusion: BAFF-R CAR T cells demonstrate remarkable efficacy against B-cell malignancies. Unexpectedly, we found that naïve T cells were superior to T_CM or T_SCM subpopulations as starting material in vivo . Naïve T-cell starting products, after transduction and expansion, generated sufficient numbers of T_CM and T_SCM CAR T cells for activity. Targeting BAFF-R combined with our strategy to improve CAR T-cell persistence potentially addresses unmet clinical needs in B-cell NHLs and ALL, particularly in the setting of CD19 CAR-T-resistance or CD19-negative relapse.

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