Xenotransplantation of Human Hematopoietic Stem Cells into NBSGW Mice: A Basic Model for Preclinical Development of Gene Therapy Approaches

Abstract

Background. The gene therapy based on hematopoietic cell xenotransplantation is becoming a powerful and universally applied therapeutic strategy in an ever-expanding range of human diseases. One of the current issues in implementing the techniques of genome modification in hematopoietic stem cells (HSCs) into clinical practice is to assure the quality and safety of gene and cell therapy products for human use. This is achieved by animal model testing at the stage of preclinical studies. With this purpose in view, NBSGW mice seem to be a unique and promising model for human HSC engraftment without pre-conditioning.
 Aim. To test the NBSGW mouse model for human HSC engraftment, to optimize the methods of assessing the state of the animals and monitoring the chimerism level for translational preclinical development of HSC-based products for gene and cell therapy.
 Materials & Methods. The xenograft models of NBSGW mice were generated using the samples of the selected peripheral blood CD34+ HSCs from a healthy donor. Serial transplantation was performed by intravenous injection of bone marrow cells from primary recipients with a high chimerism level. Engraftment efficiency was evaluated by flow cytofluorometry (FCF) and droplet digital PCR (ddPCR). Subpopulation pattern of human cell engraftment was assessed by FCF.
 Results. The tested HSC transplantation regimen is characterized by favorable toxicity profile. In the entire study sample of mice, the FCF analysis showed a long-term engraftment of human cells with a high chimerism level (23.5–93.6 %) in the bone marrow of the animals, also after serial transplantation, which was confirmed by ddPCR. The B-lineage differentiation cells predominated in all tested samples (of peripheral blood, bone marrow, and spleen) from mice after primary and serial transplantation. The ddPCR assay can be used as an additional tool for validating the level of human cell engraftment determined by FCF.
 Conclusion. NBSGW mice present a promising reference model for preclinical development of gene and cell therapy products based on human primary HSCs with a modified genome.