title: Safe engineering of CAR T cells for adoptive cell therapy of cancer using long-term episomal gene transfer.
abstract: Chimeric antigen receptor (CAR) T-cell therapy is a new successful treatment for refractory B-cell leukemia. Successful therapeutic outcome depends on long-term expression of CAR transgene in T cells, which is achieved by delivering transgene using integrating gamma retrovirus (RV) or lentivirus (LV). However, uncontrolled RV/LV integration in host cell genomes has the potential risk of causing insertional mutagenesis. Herein, we describe a novel episomal long-term cell engineering method using non-integrating lentiviral (NILV) vector containing a scaffold/matrix attachment region (S/MAR) element, for either expression of transgenes or silencing of target genes. The insertional events of this vector into the genome of host cells are below detection level. CD19 CAR T cells engineered with a NILV-S/MAR vector have similar levels of CAR expression as T cells engineered with an integrating LV vector, even after numerous rounds of cell division. NILV-S/MAR-engineered CD19 CAR T cells exhibited similar cytotoxic capacity upon CD19(+) target cell recognition as LV-engineered T cells and are as effective in controlling tumor growth in vivo We propose that NILV-S/MAR vectors are superior to current options as they enable long-term transgene expression without the risk of insertional mutagenesis and genotoxicity.
authors.affiliation: Department of Immunology, Genetics and Pathology, Science for Life Laboratory, Uppsala University, Uppsala, Sweden.
authors.forename: Chuan
authors.initials: C
authors.lastname: Jin
authors.affiliation: Department of Immunology, Genetics and Pathology, Science for Life Laboratory, Uppsala University, Uppsala, Sweden.
authors.forename: Grammatiki
authors.initials: G
authors.lastname: Fotaki
authors.affiliation: Department of Immunology, Genetics and Pathology, Science for Life Laboratory, Uppsala University, Uppsala, Sweden.
authors.forename: Mohanraj
authors.initials: M
authors.lastname: Ramachandran
authors.affiliation: Department of Immunology, Genetics and Pathology, Science for Life Laboratory, Uppsala University, Uppsala, Sweden.
authors.forename: Berith
authors.initials: B
authors.lastname: Nilsson
authors.affiliation: Department of Immunology, Genetics and Pathology, Science for Life Laboratory, Uppsala University, Uppsala, Sweden magnus.essand@igp.uu.se di.yu@igp.uu.se.
authors.forename: Magnus
authors.identifier: 0000-0002-9725-0422
authors.initials: M
authors.lastname: Essand
authors.affiliation: Department of Immunology, Genetics and Pathology, Science for Life Laboratory, Uppsala University, Uppsala, Sweden magnus.essand@igp.uu.se di.yu@igp.uu.se.
authors.forename: Di
authors.initials: D
authors.lastname: Yu
mesh_terms: D019264:Adoptive Transfer / Q000379:methods*; D000818:Animals; D018941:Antigens, CD19 / Q000276:immunology; D060846:Cell Engineering / Q000379:methods*; D064987:Cell- and Tissue-Based Therapy / Q000379:methods*; D002478:Cells, Cultured; D004195:Disease Models, Animal; D015870:Gene Expression; D005822:Genetic Vectors; D042822:Genomic Instability; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D009369:Neoplasms / Q000628:therapy*; D010957:Plasmids; D011946:Receptors, Antigen / Q000096:biosynthesis / Q000235:genetics; D013601:T-Lymphocytes / Q000276:immunology*; D019076:Transgenes; D016896:Treatment Outcome
chemical_list: D018941:Antigens, CD19; C000604811:CD19 molecule, human; D011946:Receptors, Antigen
keywords: CAR T cells; episomal cell engineering; non‐integrating lentivirus (NILV); scaffold/matrix attachment region (S/MAR) element; self‐replicating DNA