Abstract 6117: Development of preclinical models for the evaluation of conventional and <i>in vivo</i> CAR-T therapies

Conventional CAR-T manufacturing involves ex vivo expansion and engineering of T cells either from patients (autologous) or healthy donors (allogenic), which is time- and cost-consuming. The recent advancement of in vivo gene editing using lentiviral vectors allows CAR-T generation in vivo, which is time- and cost-effective compared to the conventional approaches. To evaluate their in vivo anti-tumor efficacy and toxicity, genetically engineered mouse models which express the human-specific target represent a viable choice for preclinical testing. While immunodeficient mouse models are crucial for preclinical evaluation, their utility is often compromised by the development of graft-versus-host disease (GvHD). To overcome this limitation, GemPharmatech has developed a novel NCG-MHC-dKO mouse model, which combined severe immunodeficiency with MHC class I and II molecules knockout to minimize GvHD occurrence and extend treatment window. We assessed the efficacy of an in vivo CAR-T vector using this newly developed model. NCG-MHC-dKO mice bearing luciferase-tagged Nalm6 tumors received intraperitoneal injection of 2×107 PBMCs, followed by administration of a single dose of the lentiviral vector encoding an anti-CD19 CAR transgene. Successful in vivo gene editing was confirmed by flow cytometry analysis using antibodies specific for CAR in reconstituted human T cells. Tumor burden was monitored longitudinally using an In Vivo bioluminescence Imaging System (IVIS®), quantified as total bioluminescent flux. The in vivo CAR-T vector treatment demonstrated robust growth inhibition of Nalm6-Luc tumor in huPBMC-NCG-MHC-dKO mice, with minimal GvHD occurrence throughout the course of the study, while mock-treated groups did not show any obvious tumor growth inhibition, confirming the specificity of the CAR-T vector. These findings established the NCG-MHC-dKO mouse model with PBMC engraftment as a suitable platform for preclinical evaluation of both conventional and in vivo CAR-T therapeutic strategies. Citation Format: Hongyan Sun, Xiaoliu Yang, Shiying Guo, Jianming Xu, Xiang Gao, Jun Wang. Development of preclinical models for the evaluation of conventional and in vivo CAR-T therapies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6117.