AbstractIn contrast to its limited expression in normal tissues, the B7 homolog 3 protein (B7-H3) is over-expressed in many human tumors, making it an attractive target for chimeric antigen receptor (CAR) T cell therapy.To address the technical challenges limiting broader patient access to autologous CAR T cell therapy, we armoured Epstein-Barr Virus Specific T Cells (EBVSTs) with CARs as an off-the-shelf therapy against solid tumors. We identified and utilised a nanobody as the domain specifically targeting B7-H3 in our CAR construct to reduce the stability issues associated with scFv-based domains. Our B7H3.CAR armored EBVSTs demonstrated potent in-vitro and in-vivo activity against multiple B7-H3 positive tumor and patient-derived xenografts models. Analysis of human healthy tissues and tumors microarrays indicated that B7-H3 expression, while elevated in patient tumors, was differentially low in healthy tissues. Consistent with this, mouse T cells expressing a murine equivalent of our B7H3.CAR displayed good on-target off-tumor safety in immunocompetent mice bearing syngeneic tumors. Further in-vitro evaluation revealed that while human T, B and NK cells were unaffected by B7H3.CAR EBVSTs targeting, monocytes were readily targeted due to the upregulation of B7-H3. Consequent to this targeting and depletion of B7-H3 positive human myeloid cells which are key mediators of CRS, a low incidence of cytokine release syndrome was observed in humanized mice after B7H3.CAR EBVST treatment. Lastly, we showed that B7H3.CAR EBVSTs can target myeloid-derived suppressor cells (MDSCs) that express B7-H3 and in doing so, alleviate MDSC-driven immune suppression.Our findings demonstrate that our nanobody-based B7H3.CAR EBVSTs present not only an effective and safe off-the-shelf approach for targeting any B7-H3-positive solid tumor while also addressing challenges that hindered the clinical development of CAR T cells in such indications.Citation Format: Siok Ping Yeo, Lindsay Kua, Jin Wei Tan, Joanna Kristyn Lim, Fiona Wong, May Delos Santos, Chek Meng Poh, Angeline Goh XH, Xin Yu Koh, Xiao Hua Zhou, Rajarethinam R, Qing Feng Chen, Zhisheng Her, Ivan D Horak, Lionel Low, Kar Wai Tan. Off-The-Shelf Epstein Barr Virus Specific T Cells Armored with B7-H3-Targeting Chimeric Antigen Receptors Is an Effective and Safe Tumor- Agnostic Therapy For B7-H3 Positive Solid Malignancies [abstract]. In: Proceedings of Frontiers in Cancer Science; 2023 Nov 6-8; Singapore. Philadelphia (PA): AACR; Cancer Res 2024;84(8_Suppl):Abstract nr P66.
