Gene knock-outs in human CD34+ hematopoietic stem and progenitor cells and in the human immune system of mice
Year: 2023
Type: article
Abstract: Human CD34+ hematopoietic stem and progenitor cells (HSPCs) are a standard source of cells for clinical HSC transplantations as well as experimental xenotransplantation to generate "humanized mice". To further extend the range of applications of these humanized mice, we developed a protocol to efficiently edit the genomes of human CD34+ HSPCs before transplantation. In the past, manipulating HSPCs has been complicated by the fact that they are inherently difficult to transduce with lentivectors, and rapidly lose their stemness and engraftment potential during in vitro culture. However, with optimized nucleofection of sgRNA:Cas9 ribonucleoprotein complexes, we are now able to edit a candidate gene in CD34+ HSPCs with almost 100% efficiency, and transplant these modified cells in immunodeficient mice with high engraftment levels and multilineage hematopoietic differentiation. The result is a humanized mouse from which we knocked out a gene of interest from their human immune system.
Source: PLoS ONE
Authors: Daniel A. Kuppers, Jonathan Linton, Sergio Ortiz Espinosa, Kelly M. McKenna, Anthony Rongvaux, Patrick J. Paddison
Institutions: Fred Hutch Cancer Center, University of Washington
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $1,805
Funders: Bill and Melinda Gates Foundation, Yale University, Fred Hutchinson Cancer Research Center, Regeneron Pharmaceuticals, National Institutes of Health, National Heart, Lung, and Blood Institute, National Cancer Institute, National Institute of General Medical Sciences, National Institute of Allergy and Infectious Diseases, National Institute of Diabetes and Digestive and Kidney Diseases
Awards: G1305388156, G191830569, G1971433595, G23584040, G2362813460, G2594373096, Progenitor Cell Biology Consortium Administrative Coordinating Center, Core Center of Excellence in Hematology, G378552126, G4266498966, G4430470024, G4945730252, G5003652954, G6482580321, G6612846274, G6845504420, G7113653239, G7200330922, G7792392908, G7949248321, G8571044719, G863835797, G8800477068, G9000085113, G9000122962, G9000269946, Cancer Center Support Grant, Stem Cell and Transplantation Biology
OpenAlex ID: W4382468026