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CRISPR-Cas9 disruption of PD-1 enhances activity of universal EGFRvIII CAR T cells in a preclinical model of human glioblastoma.

ABSTRACT: Despite remarkable success in the treatment of hematological malignancies, CAR T-cell therapies for solid tumors have floundered, in large part due to local immune suppression and the effects of prolonged stimulation leading to T-cell dysfunction and exhaustion. One mechanism by which gliomas and other cancers can hamper CAR T cells is through surface expression of inhibitory ligands such as programmed cell death ligand 1 (PD-L1). Using the CRIPSR-Cas9 system, we created universal CAR T cells resistant to PD-1 inhibition through multiplexed gene disruption of endogenous T-cell receptor (TRAC), beta-2 microglobulin (B2M) and PD-1 (PDCD1). Triple gene-edited CAR T cells demonstrated enhanced activity in preclinical glioma models. Prolonged survival in mice bearing intracranial tumors was achieved after intracerebral, but not intravenous administration. CRISPR-Cas9 gene-editing not only provides a potential source of allogeneic, universal donor cells, but also enables simultaneous disruption of checkpoint signaling that otherwise impedes maximal antitumor functionality.

SUBMITTER: Choi BD

PROVIDER: S-EPMC6857271 | biostudies-literature | 2019 Nov

REPOSITORIES: biostudies-literature

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CRISPR-Cas9 disruption of PD-1 enhances activity of universal EGFRvIII CAR T cells in a preclinical model of human glioblastoma.

Choi Bryan D BD Yu Xiaoling X Castano Ana P AP Darr Henia H Henderson Daniel B DB Bouffard Amanda A AA Larson Rebecca C RC Scarfò Irene I Bailey Stefanie R SR Gerhard Genevieve M GM Frigault Matthew J MJ Leick Mark B MB Schmidts Andrea A Sagert Jason G JG Curry William T WT Carter Bob S BS Maus Marcela V MV

Journal for immunotherapy of cancer 20191114 1

Despite remarkable success in the treatment of hematological malignancies, CAR T-cell therapies for solid tumors have floundered, in large part due to local immune suppression and the effects of prolonged stimulation leading to T-cell dysfunction and exhaustion. One mechanism by which gliomas and other cancers can hamper CAR T cells is through surface expression of inhibitory ligands such as programmed cell death ligand 1 (PD-L1). Using the CRIPSR-Cas9 system, we created universal CAR T cells re ...[more]

PMID: 31727131

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Project description:BackgroundGlioblastoma (GBM) is the most common malignant brain tumor and has "immunologically cold" features. Changing GBM to an "immunologically hot" tumor requires a strong trigger that induces initial immune responses in GBM. Allogeneic natural killer cells (NKCs) have gained considerable attention as promising immunotherapeutic tools against cancer, where gene-edited NKCs would result in effective anti-cancer treatment. The present study focused on the immune checkpoint molecule cytokine-inducible SH2-containing protein (CISH, or CIS) as a critical negative regulator in NKCs.MethodsThe GBM tumor environment featured with immunological aspect was analyzed with Cancer immunogram and GlioVis. We generated human primary CIS-deleted NKCs (NK dCIS) using clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) with single guide RNA targeting genome sites on CIS coding exons. The genome-edited NKCs underwent microarray with differential expression analysis and gene set enrichment analysis (GSEA). The anti-GBM activity of the genome-edited NKCs was evaluated by apoptosis induction effects against allogeneic GBM cells and spheroids. We further detected in vivo antitumor effects using xenograft brain tumor mice.ResultsWe successfully induced human CIS-deleted NKCs (NK dCIS) by combining our specific human NKC expansion method available for clinical application and genome editing technology. CIS gene-specific guide RNA/Cas9 protein complex suppressed CIS expression in the expanded NKCs with high expansion efficacy. Comprehensive gene expression analysis demonstrated increased expression of 265 genes and decreased expression of 86 genes in the NK dCIS. Gene set enrichment analysis revealed that the enriched genes were involved in NKC effector functions. Functional analysis revealed that the NK dCIS had increased interferon (IFN)ɤ and tumor necrosis factor (TNF) production. CIS deletion enhanced NKC-mediated apoptosis induction against allogeneic GBM cells and spheroids. Intracranial administration of the allogeneic NKCs prolonged the overall survival of xenograft brain tumor mice. Furthermore, the NK dCIS extended the overall survival of the mice.ConclusionThe findings demonstrated the successful induction of human primary NK dCIS with CRISPR/Cas9 with efficient expansion. CIS deletion enhanced the NKC-mediated anti-tumor effects in allogeneic GBM and could be a promising immunotherapeutic alternative for patients with GBM.

Dataset Information

CRISPR-Cas9 disruption of PD-1 enhances activity of universal EGFRvIII CAR T cells in a preclinical model of human glioblastoma.

Publications

CRISPR-Cas9 disruption of PD-1 enhances activity of universal EGFRvIII CAR T cells in a preclinical model of human glioblastoma.

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