Non-Clinical Cell Therapy Development Using the NCG Mouse Model as a Test System
Viktorija Smutova1, Camila Pará1, Morgan K Foret1
1Charles River Laboratories, Laval, QC, Canada.
Abstract:
The NCG triple immunodeficient mice on a NOD/Nju background lack functional/mature T, B, and NK cells, and have reduced macrophage and dendritic cell function. This study characterized the NCG mouse model for toxicity, engraftment and tumorigenicity assessments of cell therapies, using CD34+ hHSPC adult mobilized cells with two myeloablation regimens.Mice received sub-lethal irradiation or busulfan and were then injected intravenously with CD34+ hHSPCs (1.0 x 106 cells/mouse) or PBS (control), while positive control animals received 2 x 106 HL-60 cells/mouse. hCD34+ cell donors were treated with the mobilizing agent G-CSF prior to leukapheresis. Following injections, mouse blood samples were collected to assess engraftment rates by flow cytometry with body weights recorded periodically up to 20 weeks post-cell injection. No significant clinical signs or body weight changes were observed. At week 10 post-cell injection, the peripheral blood chimerism of hCD45+ cells was above 20%. While mCD45+ concentration was constant between week 10 and 17 in whole blood samples, hCD45+ concentration and chimerism slightly decreased at week 17. However, chimerism remained above 10%, with busulfan-treated mice presenting higher values. Chimerism was further assessed by quantifying human Alu sequences in blood and multiple organs using qPCR. Alu sequences were most abundant in the spleen and bone marrow, while lowest in the testes. In the positive control group, expected mortalities due to tumorigenesis were observed between days 27 and 40 post-cell injection. Overall, study results may be used to inform study design and potential toxicological endpoints relevant to non-clinical cell therapy development.
Insights
NCG mice support human hematopoietic stem cell engraftment for cell therapy studies. Busulfan preconditioning resulted in higher sustained chimerism, with no observed toxicity in this immunodeficient mouse model.
Area of Science:
- Immunology
- Hematology
- Preclinical Research
Background:
- NCG mice are triple immunodeficient, lacking T, B, and NK cells, with impaired myeloid function.
- This model is suitable for evaluating cell therapies due to its immune deficiencies.
Purpose of the Study:
- To characterize the NCG mouse model for cell therapy assessments.
- To evaluate toxicity, engraftment, and tumorigenicity of CD34+ human hematopoietic stem progenitor cells (hHSPCs).
Main Methods:
- NCG mice received sub-lethal irradiation or busulfan myeloablation before intravenous injection of CD34+ hHSPCs.
- Engraftment was assessed via flow cytometry (hCD45+ cells) and qPCR (human Alu sequences) up to 20 weeks.
- Tumorigenicity was evaluated using HL-60 cells as a positive control.
Main Results:
- No significant clinical signs or body weight changes were observed in hHSPC-injected mice.
- Peripheral blood chimerism of hCD45+ cells reached over 20% by week 10 and remained above 10% at week 17.
- Busulfan treatment led to higher sustained chimerism compared to irradiation.
- Human Alu sequences were detected in multiple organs, highest in spleen and bone marrow.
- Positive control group showed expected mortality due to tumorigenesis.
Conclusions:
- The NCG mouse model is well-suited for preclinical cell therapy development.
- Busulfan is an effective myeloablative regimen for achieving robust hHSPC engraftment in NCG mice.
- The study provides data to inform the design of future cell therapy studies and identify toxicological endpoints.
