Work

Engineered Human Hematopoietic Stem Cells Generated in Developing Chimeric Animals from a New Class of Blastomere-like Human Totipotent Stem Cells

Year: 2026

Type: article

Abstract: Introduction Although immune-deficient mice engrafted with human CD34 + HSC are widely employed in human immunology studies, these models have limitations. Generation of a fully diverse human lympho-hematopoietic system in research animals would radically change the landscape of biotechnology. Objectives/Hypothesis Human HSC generated within developing murine AGM, fetal liver, and thymic niches of chimeric animal embryos can give rise to a full repertoire of human lymphocytes, myeloid cells, and HLA-expressing APC. Methods We have reported Tankyrase/PARP1 Inhibitor-Regulated Naïve Stem Cells (TIRN-SC) by chemically reverting conventional hiPSC to an improved stem cell state that contributes differentiating human cells into developing mouse embryos. TIRN-SC are a new class of human blastomere-like totipotent stem cells with high epigenetic plasticity. Results In directed differentiation protocols, TIRN-SC generated 5-to-50-fold greater numbers of granulocyte-macrophage progenitors per culture volume and input cell than conventional hiPSC. TIRN-SC hematopoietic cultures produced two-fold greater purities of cultures of CD34 + CD45 + progenitors and with >90% CD11b + and >85% CD33 + myeloid progeny. To evaluate in vivo developmental potential, TIRN-SC were injected into murine embryos to develop human-murine chimeras. TIRN-SC i njection into 8C-16C-staged murine embryos resulted in efficient integration of human cells to both extra-embryonic (placenta) and embryonic (neural, hematopoietic) lineages of blastocyst and fetal chimeras , including human CD34 + CD45 + hematopoietic progenitors that developed within murine fetal livers. TIRN-SC chimerism efficiencies could be further optimized to high levels of 20-50% using immunosuppression or ectopic expression of a murine-E-Cadherin (mECad) transgene; the latter strategy promoted enhanced human cell targeting to murine hematopoietic organs ( e.g ., AGM and fetal liver; Fig. 1 ). Transgenic mECad-TIRN-SC achieved human cell chimerism rates of 2%-80% and 2%-18% in whole murine E14.5 feti and fetal livers, respectively. Toward our goal of generating humanized pigs that can produce unlimited amounts of engineered cells in their peripheral blood, we prepared CAR-expressing TIRN-SC lines recognizing the glioblastoma-specific disialoganglioside surface tumor antigen GD2 . Direct and adaptor-based anti-GD2 CAR-expressing TIRN-SC-derived myeloid cells were evaluated for tumor cell-dependent activation, cellular killing, and phagocytosis. Conclusions TIRN-SC will allow the development of animals with humanized immune systems and cellular immunotherapy animal biofactories. For example, chimeric animals with human lymphohematopoietic systems from engineered TIRN-SC could provide unlimited sources of patient-specific or transgenic CAR-expressing NK,T, and myeloid cells in their peripheral blood.

Source: Transplantation and Cellular Therapy

DOI: 10.1016/j.jtct.2025.12.985

Authors: Ludovic Zimmerlin, Willem Buys, Dr. Elias T. Zambidis

Institutions: Johns Hopkins University, Johns Hopkins Medicine, Sidney Kimmel Comprehensive Cancer Center

Language: English

Topic: Pluripotent Stem Cells Research

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

SDG: Good health and well-being

Open Access status: hybrid

Estimated APC paid: $3,650