AS program is essential for HSC formation.

In the aorta-gonad-mesonephros (AGM) region of mouse embryos, pre-hematopoietic stem cells (pre-HSCs) are generated from rare and specialized hemogenic endothelial cells (HECs) via endothelial-to-hematopoietic transition, followed by maturation into bona fide hematopoietic stem cells (HSCs). As HECs also generate a lot of hematopoietic progenitors not fated to HSCs, powerful tools that are pre-HSC/HSC-specific become urgently critical. Here, using the gene knockin strategy, we firstly developed an Hlf-tdTomato reporter mouse model and detected Hlf-tdTomato expression exclusively in the hematopoietic cells including part of the immunophenotypic CD45– and CD45+ pre-HSCs in the embryonic day (E) 10.5 AGM region. By in vitro co-culture together with long-term transplantation assay stringent for HSC precursor identification, we further revealed that unlike the CD45– counterpart in which both Hlf-tdTomato-positive and negative sub-populations harbored HSC competence, the CD45+ E10.5 pre-HSCs existed exclusively in Hlf-tdTomato-positive cells. The result indicates that the cells should gain the expression of Hlf prior to or together with CD45 to give rise to functional HSCs. Furthermore, we constructed a novel Hlf-CreER mouse model and performed time-restricted genetic lineage tracing by a single dose induction at E9.5. We observed the labeling in E11.5 AGM precursors and their contribution to the immunophenotypic HSCs in fetal liver (FL). Importantly, these Hlf-labeled early cells contributed to and retained the size of the HSC pool in the bone marrow (BM), which continuously differentiated to maintain a balanced and long-term multi-lineage hematopoiesis in the adult. Therefore, we provided another valuable mouse model to specifically trace the fate of emerging HSCs during development.

Therapeutic antibodies have ushered in a new age of cancer immunotherapy. Historically, these therapies have targeted a limited subset of soluble and cell surface tumor-associated antigens (TAAs). T cell receptors (TCRs) on cytotoxic CD8+ T cells recognize peptide antigens bound to major histocompatibility class I (MHC-I) proteins, called HLA-A/B/C in humans. By this pathway, antigen is regularly sampled from the intracellular peptidome, processed, and presented to cytotoxic T cells. Expanding TCR-based recognition of soluble, intracellular TAAs presented on the surface of malignant cells by this mechanism is a propitious therapeutic strategy. Here we describe a novel platform for generating T cell receptor mimic (TCRm) antibodies using our humanized immunoglobulin (RenMabTM) mice engineered to express HLA. TCRm antibodies have the same binding properties as endogenous TCRs and recognize processed, HLA-bound peptides including intracellular tumor-associated antigens, viral oncoproteins, and cancer-testis antigen (CTA). TCRm antibodies bind peptide-HLA with high specificity and up to nanomolar affinity. Our optimized immunization protocols and high-throughput screening methods allow for one-step TCRm antibody generation. TCRm antibodies can also be used to assemble bispecific T cell engaging antibodies (BiTEs) to enhance tumor targeting of cytotoxic T cells. Biocytogen’s TCRm antibodies are a flexible and powerful tool for cancer immunotherapy. By enabling TCR-mediated recognition of an unrestricted repertoire of cancer neoantigens, TCRm antibodies may find broad clinical application. Citation Format: Jun Du, Taolin Liu, Wanbo Tang, Yue Zhang, Limin Zhao, Xin Jiao, Chao Sun, Pengfei Du, Yuqi Zhang, Baihong Liu, Qingcong Lin, Yi Yang. Targeting intracellular tumor antigens using fully human TCR mimic antibodies derived from HLA transgenic RenMiceTM [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2752.