Vγ2Vδ2 T cell-based immunotherapy has benefited some patients in clinical trials, but the overall efficacy is low for solid tumor patients. In this study, a bispecific antibody against both PD-L1 and CD3 (PD-L1 x CD3), Y111, could efficiently bridge T cells and PD-L1 expressing tumor cells. The Y111 prompted fresh CD8+ T cell-mediated lysis of H358 cells, but spared this effect on the fresh Vδ2+ T cells enriched from the same donors, which suggested that Y111 could bypass the anti-tumor capacity of the fresh Vγ2Vδ2 T cells. As the adoptive transfer of the expanded Vγ2Vδ2 T cells was approved to be safe and well-tolerated in clinical trials, we hypothesized that the combination of the expanded Vγ2Vδ2 T cells with the Y111 would provide an alternative approach of immunotherapy. Y111 induced the activation of the expanded Vγ2Vδ2 T cells in a dose-dependent fashion in the presence of PD-L1 positive tumor cells. Moreover, Y111 increased the cytotoxicity of the expanded Vγ2Vδ2 T cells against various NSCLC-derived tumor cell lines with the releases of granzyme B, IFNγ, and TNFα in vitro. Meanwhile, the adoptive transferred Vγ2Vδ2 T cells together with the Y111 inhibited the growth of the established xenografts in NPG mice. Taken together, our data suggested a clinical potential for the adoptive transferring the Vγ2Vδ2 T cells with the Y111 to treat PD-L1 positive solid tumors.

Background A T cell-redirecting bispecific antibody consisting of a tumor-binding unit and a T cell-binding unit is a large group of antibody-based biologics against death-causing cancer diseases. The anti-CD38 × anti-CD3 bispecific antibody (Y150) is potential for treating multiple myeloma (MM). When developing a cell-based reporter gene bioassay to assess the activities of Y150, it was found that the expression of CD38 on the human T lymphocyte cells (Jurkat) caused the nonspecific activation which interfered with the specific T cells activation of mediated by the Y150 and CD38(+) tumor cells. Methods Here we first knocked-out the CD38 expression on Jurkat T cell line by CRISPR-Cas9 technology, then developed a stable monoclonal CD38(−) Jurkat T cell line with an NFAT-RE driving luciferase expressing system. Further based on the CD38(−) Jurkat cell, we developed a reporter gene method to assess the bioactivity of the anti-CD38 × anti-CD3 bsAb. Results Knocking out CD38 expression abolished the nonspecific self-activation of the Jurkat cells. The selected stable monoclonal CD38(−) Jurkat T cell line assured the robustness of the report genes assay for the anti-CD38 × anti-CD3 bsAb. The relative potencies of the Y150 measured by the developed reporter gene assay were correlated with those by the flow-cytometry-based cell cytotoxicity assay and by the ELISA-based binding assay. Conclusions The developed reporter gene assay was Mechanism of Action (MOA)-reflective for the bioactivity of anti-CD38 × anti-CD3 antibody, and suitable for the quality control for the bsAb product.

Background: Anti-cancer immunotherapy based on the adoptive transfer of Vγ2Vδ2 T cells has benefited to some patients in clinical trials, but the overall responses are inconsistent. Therefore, new strategies are urgently needed to improve the current therapy.Methods: In this study, a designed bispecific antibody Y111, which binds to both CD3 and PD-L1, is applied to optimally potentiate Vγ2Vδ2 T cell-based killing of cancer cells. The binding activities of Y111 was determined by Flow cytometry. CFSE/PI-based flow cytometry was applied to check the re-directed killing ability induced by Y111 in the condition of using T cell subsets, or expanded- and purified- Vγ2Vδ2 T cells as effector cells. Moreover, expanded- and purified- Vγ2Vδ2 T cells were co-cultured with tumor cells in the presence/absence of Y111 to assess the activation, degranulation, and cytokine production by intracellular cytokine staining, and CBA method. Finally, NPG-based subcutaneous tumor mouse models were used to check the in vivo therapeutic efficacy of the combination of transfused Vγ2Vδ2 T cells and Y111.Results: Due to its binding activities, Y111 apparently prompts fresh αβ-mediated lysis of tumor cell line H358 cells, but spare the effect on the fresh enriched Vγ2Vδ2 T cells from the same donors. However, Y111 increases cytotoxicity of expanded and purified Vγ2Vδ2 T cells against various NSCLC-derived tumor cell lines in a tumor cell dependent fashion. Y111 also prompted the releases of granzyme B, IFNγ and TNFα. Supporting to these observations in vitro, a combination of adoptive transferring Vγ2Vδ2 T cell and Y111 into the tumor-bearing NPG mice inhibited the growth of the established tumors in the mice.Conclusions: Taken together, our data suggest clinical potential for adoptive transferring the bispecific antibody-armored Vγ2Vδ2 T cells to treat solid tumors, such as NSCLC.