HLA DR Genome Editing with TALENs in Human iPSCs Produced Immune-Tolerant Dendritic Cells

Abstract

Although human induced pluripotent stem cells (iPSCs) can serve as a universal cell source for regenerative medicine, the use of iPSCs in clinical applications is limited by prohibitive costs and prolonged generation time. Moreover, allogeneic iPSC transplantation requires preclusion of mismatches between the donor and recipient human leukocyte antigen (HLA). We, therefore, generated universally compatible immune nonresponsive human iPSCs by gene editing. Transcription activator-like effector nucleases (TALENs) were designed for selective elimination of HLA DR expression. The engineered nucleases completely disrupted the expression of HLA DR on human dermal fibroblast cells (HDF) that did not express HLA DR even after stimulation with IFN-γ. Teratomas formed by HLA DR knockout iPSCs did not express HLA DR, and dendritic cells differentiated from HLA DR knockout iPSCs reduced CD4+ T cell activation. These engineered iPSCs might provide a novel translational approach to treat multiple recipients from a limited number of cell donors.

CAR-neutrophil mediated delivery of tumor-microenvironment responsive nanodrugs for glioblastoma chemo-immunotherapy

“…Particularly, off-target toxicity profiling of CAR-neutrophils with or without loading nanodrugs in infused animals, including cytokine release syndrome, neurotoxicity, and on-target off-tumor toxicities observed in CAR-T cells 57 , are needed, despite the short lifespan of neutrophils. While feasible approaches, such as engineering hypoimmunogenic universal donor hPSCs [58][59][60][61] and banking human leukocyte antigen (HLA)-homozygous hPSC libraries 62 , are readily available to avoid the potential risk of graft-versus-host disease (GvHD), preclinical animal models with an intact immune system is still needed to assess the translational potential of our neutrophil therapeutics. Finally, limited anti-tumor cytotoxicity and extension of animal lifespan of CAR-neutrophil nanodrug therapeutics were observed.…”

Genetic engineering strategies to enhance antitumor reactivity and reduce alloreactivity for allogeneic cell-based cancer therapy

“…After 13 days of quadruple gRNAs transfection, Lee et al observed that the gene-edited CD3 + T cells composed 62.1% of HLA-I/II-double-negative cells, while retaining surface phenotype and functionality. However, for generation of universally compatible iPSCs ( 142 ), results concluded that TALEN restriction enzymes had the greatest consistency; they were more maneuverable than ZFNs and had fewer off-target events than CRISPR/Cas 9. Even when stimulated with high concentrations of IFN-γ, TALEN-treated fibroblasts lacked HLA-DR expression, which was maintained within iPSCs generation ( 142 ).…”

“…However, for generation of universally compatible iPSCs ( 142 ), results concluded that TALEN restriction enzymes had the greatest consistency; they were more maneuverable than ZFNs and had fewer off-target events than CRISPR/Cas 9. Even when stimulated with high concentrations of IFN-γ, TALEN-treated fibroblasts lacked HLA-DR expression, which was maintained within iPSCs generation ( 142 ). HLA-DR knockout remained effective and safe in these therapies, suggesting their potential to modify effector cells to prevent HvG effect in cancer immunotherapy ( 143 ).…”

Reprogramming the tumor microenvironment by genome editing for precision cancer therapy

“…T cells engineered with TALENs enhance the antitumor efficacy of adoptive immunotherapy [ 9 ]. TALENs have also been applied to edit the genome of human induced pluripotent stem cells (iPSCs), making these cells differentiate into immune cells with potential antitumor activity [ 10 ]. The emergence of the CRISPR/Cas system produced a revolution in gene editing technology.…”