Pre-clinical humanized mouse models are a powerful tool to evaluate immunotherapies. NSG-SGM3 mice reconstituted with human stem cells (huSGM3) develop pronounced human myeloid cells due to transgenic expression of stem cell factor, granulocyte-macrophage colony-stimulating factor, and interleukin-3 (IL-3) compared with the widely used humanized NSG (huNSG) model. We assessed in vivo generation of CD19CAR T cells in huSGM3 mice upon single intravenous injection of the T cell-specific lentiviral vectors (LVs) CD4-LV and CD8LV. While in vivo CAR T cell generation was clearly detectable in individual mice, generation appeared less efficient than previously observed for huNSG mice. Especially for the CD4-LV group, this correlated with increased IL-15 and decreased GM-CSF levels, indicating activation of monocytes and macrophages. Co-culture assays identified macrophages as a potential barrier for gene transfer. Refining CD4-LV and CD8-LV with a less immunogenic surface by using modified packaging cells substantially improved the transduction of lymphocytes in vitro in the presence of macrophages, as well in vivo in huSGM3 mice. Notably, two mice that developed less CAR T cells showed high interferon-alpha or -beta levels before vector injection. Our data emphasize the relevance of innate immune responses for in vivo generation of CAR T cells, which can be overcome by vector surface engineering.

In vivo generation of CAR T cells in the presence of human myeloid cells

Milani, Michela;Cantore, Alessio;
2022

Abstract

Pre-clinical humanized mouse models are a powerful tool to evaluate immunotherapies. NSG-SGM3 mice reconstituted with human stem cells (huSGM3) develop pronounced human myeloid cells due to transgenic expression of stem cell factor, granulocyte-macrophage colony-stimulating factor, and interleukin-3 (IL-3) compared with the widely used humanized NSG (huNSG) model. We assessed in vivo generation of CD19CAR T cells in huSGM3 mice upon single intravenous injection of the T cell-specific lentiviral vectors (LVs) CD4-LV and CD8LV. While in vivo CAR T cell generation was clearly detectable in individual mice, generation appeared less efficient than previously observed for huNSG mice. Especially for the CD4-LV group, this correlated with increased IL-15 and decreased GM-CSF levels, indicating activation of monocytes and macrophages. Co-culture assays identified macrophages as a potential barrier for gene transfer. Refining CD4-LV and CD8-LV with a less immunogenic surface by using modified packaging cells substantially improved the transduction of lymphocytes in vitro in the presence of macrophages, as well in vivo in huSGM3 mice. Notably, two mice that developed less CAR T cells showed high interferon-alpha or -beta levels before vector injection. Our data emphasize the relevance of innate immune responses for in vivo generation of CAR T cells, which can be overcome by vector surface engineering.
CAR T cells
T cell targeted lentiviral vectors
humanized mouse model
innate immune response
in vivo gene delivery
macrophage
shielding
File in questo prodotto:
Non ci sono file associati a questo prodotto.

I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.

Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/20.500.11768/132671
Citazioni
  • ???jsp.display-item.citation.pmc??? ND
  • Scopus 1
  • ???jsp.display-item.citation.isi??? 1
social impact