title: CRISPR/Cas9-mediated PD-1 disruption enhances anti-tumor efficacy of human chimeric antigen receptor T cells.
abstract: Immunotherapies with chimeric antigen receptor (CAR) T cells and checkpoint inhibitors (including antibodies that antagonize programmed cell death protein 1 [PD-1]) have both opened new avenues for cancer treatment, but the clinical potential of combined disruption of inhibitory checkpoints and CAR T cell therapy remains incompletely explored. Here we show that programmed death ligand 1 (PD-L1) expression on tumor cells can render human CAR T cells (anti-CD19 4-1BBζ) hypo-functional, resulting in impaired tumor clearance in a sub-cutaneous xenograft model. To overcome this suppressed anti-tumor response, we developed a protocol for combined Cas9 ribonucleoprotein (Cas9 RNP)-mediated gene editing and lentiviral transduction to generate PD-1 deficient anti-CD19 CAR T cells. Pdcd1 (PD-1) disruption augmented CAR T cell mediated killing of tumor cells in vitro and enhanced clearance of PD-L1+ tumor xenografts in vivo. This study demonstrates improved therapeutic efficacy of Cas9-edited CAR T cells and highlights the potential of precision genome engineering to enhance next-generation cell therapies.
authors.affiliation: Department of Cellular & Molecular Pharmacology, San Francisco, CA, USA.|Center for Systems and Synthetic Biology, University of California San Francisco, San Francisco, 94158, CA, USA.|Helen Diller Family Comprehensive Cancer Center, University of California San Francisco, San Francisco, CA, 94158, USA.|Howard Hughes Medical Institute, University of California San Francisco, San Francisco, 94158, CA, USA.
authors.forename: Levi J
authors.initials: LJ
authors.lastname: Rupp
authors.affiliation: Diabetes Center, University of California San Francisco, San Francisco, CA, 94143, USA.|Department of Microbiology and Immunology, University of California San Francisco, San Francisco, CA, 94143, USA.
authors.forename: Kathrin
authors.initials: K
authors.lastname: Schumann
authors.affiliation: Department of Cellular & Molecular Pharmacology, San Francisco, CA, USA.|Center for Systems and Synthetic Biology, University of California San Francisco, San Francisco, 94158, CA, USA.|Helen Diller Family Comprehensive Cancer Center, University of California San Francisco, San Francisco, CA, 94158, USA.
authors.forename: Kole T
authors.initials: KT
authors.lastname: Roybal
authors.affiliation: Department of Epidemiology and Biostatistics, Department of Bioengineering and Therapeutic Sciences, Institute for Human Genetics, University of California, San Francisco, CA, 94143, USA.|Biological and Medical Informatics Graduate Program, University of California, San Francisco, CA, 94158, USA.
authors.forename: Rachel E
authors.initials: RE
authors.lastname: Gate
authors.affiliation: Department of Epidemiology and Biostatistics, Department of Bioengineering and Therapeutic Sciences, Institute for Human Genetics, University of California, San Francisco, CA, 94143, USA.
authors.forename: Chun J
authors.initials: CJ
authors.lastname: Ye
authors.affiliation: Department of Cellular & Molecular Pharmacology, San Francisco, CA, USA. Wendell.lim@ucsf.edu.|Center for Systems and Synthetic Biology, University of California San Francisco, San Francisco, 94158, CA, USA. Wendell.lim@ucsf.edu.|Helen Diller Family Comprehensive Cancer Center, University of California San Francisco, San Francisco, CA, 94158, USA. Wendell.lim@ucsf.edu.|Howard Hughes Medical Institute, University of California San Francisco, San Francisco, 94158, CA, USA. Wendell.lim@ucsf.edu.
authors.forename: Wendell A
authors.initials: WA
authors.lastname: Lim
authors.affiliation: Helen Diller Family Comprehensive Cancer Center, University of California San Francisco, San Francisco, CA, 94158, USA.|Diabetes Center, University of California San Francisco, San Francisco, CA, 94143, USA.|Department of Microbiology and Immunology, University of California San Francisco, San Francisco, CA, 94143, USA.|Division of Infectious Diseases and Rheumatology, Department of Medicine, University of California San Francisco, San Francisco, CA, 94143, USA.|Innovative Genomics Institute, University of California Berkeley, Berkeley, CA, 94720, USA.
authors.forename: Alexander
authors.initials: A
authors.lastname: Marson
mesh_terms: D000818:Animals; D018941:Antigens, CD19 / Q000235:genetics / Q000276:immunology; D000951:Antigens, Neoplasm / Q000235:genetics / Q000276:immunology; D060890:B7-H1 Antigen / Q000378:metabolism; D015415:Biomarkers; D064113:CRISPR-Cas Systems; D045744:Cell Line, Tumor; D005260:Female; D015870:Gene Expression; D055785:Gene Knockdown Techniques; D018390:Gene Targeting; D006801:Humans; D016130:Immunophenotyping; D051379:Mice; D018345:Mice, Knockout; D009369:Neoplasms / Q000235:genetics* / Q000276:immunology* / Q000473:pathology / Q000628:therapy; D061026:Programmed Cell Death 1 Receptor / Q000235:genetics* / Q000378:metabolism; D011948:Receptors, Antigen, T-Cell / Q000235:genetics* / Q000378:metabolism*; D013601:T-Lymphocytes / Q000276:immunology* / Q000378:metabolism*
chemical_list: D018941:Antigens, CD19; D000951:Antigens, Neoplasm; D060890:B7-H1 Antigen; D015415:Biomarkers; C423236:CD274 protein, human; C105992:PDCD1 protein, human; D061026:Programmed Cell Death 1 Receptor; D011948:Receptors, Antigen, T-Cell