Abstract 123: Establishment of humanized CD3EDG/4-1BB mice for testing novel T cell engagers

Abstract Agonism of the 4-1BB receptor is an attractive therapeutic strategy for tumor cell therapy, due to the ability of this receptor to stimulate activity of T and NK cells for tumor cell killing. Many classes of biologics targeting 4-1BB have been generated to date, including bispecific or multispecific engagers designed to stimulate 4-1BB and CD3 activity on the T cell surface. To provide a preclinical platform to test such therapies, Biocytogen generated a humanized B-hCD3EDG/h4-1BB mouse model. This was achieved by crossing humanized mice expressing chimeric CD3E, CD3D and CD3G with mice expressing the human extracellular region of 4-1BB. Previously, we showed that these humanized CD3EDG mice and humanized 4-1BB mice can be used for validating efficacy of anti-human CD3 and 4-1BB antibody-based therapies, respectively. Here, we confirmed expression of human CD3 and 4-1BB on splenocytes in homozygous humanized B-hCD3EDG/h4-1BB mice by flow cytometry, which also demonstrated lack of murine CD3 and 4-1BB expression. Assessment of leukocyte subpopulations in the spleen and peripheral blood confirmed that humanization of the mice did not significantly alter the development or frequency of blood cell populations. Further evaluation of lymphocyte frequency was also performed, which demonstrates no significant alterations in the CD4/CD8 T cell ratio in spleen, lymph node, or peripheral blood of the animals. Presence of Treg and NK cells was also confirmed. Taken together, these data indicate that the humanized B-hCD3EDG/h4-1BB model is a novel, robust tool for testing novel T cell engagers. Citation Format: Chengzhang Shang, Chong Li, Lei Zhao, Qingqing Xu, James Jin, Menglian Ren. Establishment of humanized CD3EDG/4-1BB mice for testing novel T cell engagers abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts) ; 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84 (6Suppl): Abstract nr 123.