title: Harnessing autophagy for cell fate control gene therapy.
abstract: We hypothesized that rapamycin, through induction of autophagy and promotion of an antiapoptotic phenotype, would permit lentiviral (LV)-based transgene delivery to human T-Rapa cells, which are being tested in phase II clinical trials in the setting of allogeneic hematopoietic cell transplantation. Manufactured T-Rapa cells were exposed to supernatant enriched for a LV vector encoding a fusion protein consisting of truncated CD19 (for cell surface marking) and DTYMK/TMPKΔ, which provides "cell-fate control" due to its ability to phosphorylate (activate) AZT prodrug. LV-transduction in rapamycin-treated T-Rapa cells: (1) resulted in mitochondrial autophagy and a resultant antiapoptotic phenotype, which was reversed by the autophagy inhibitor 3-MA; (2) yielded changes in MAP1LC3B and SQSTM1 expression, which were reversed by 3-MA; and (3) increased T-Rapa cell expression of the CD19-DTYMKΔ fusion protein, despite their reduced proliferative status. Importantly, although the transgene-expressing T-Rapa cells expressed an antiapoptotic phenotype, they were highly susceptible to cell death via AZT exposure both in vitro and in vivo (in a human-into-mouse xenogeneic transplantation model). Therefore, rapamycin induction of T cell autophagy can be used for gene therapy applications, including the CD19-DTYMKΔ cell-fate control axis to improve the safety of T cell immuno-gene therapy.
authors: Experimental Transplantation and Immunology Branch; National Cancer Institute; National Institutes of Health; Bethesda, MD USA.
authors: Tania C
authors: TC
authors: Felizardo
authors: Jason
authors: J
authors: Foley
authors: Kevin
authors: K
authors: Steed
authors: Boro
authors: B
authors: Dropulic
authors: Shoba
authors: S
authors: Amarnath
authors: Jeffrey A
authors: JA
authors: Medin
authors: Daniel H
authors: DH
authors: Fowler
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D018941:Antigens, CD19 / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects; D001343:Autophagy / Q000187:drug effects; D015496:CD4-Positive T-Lymphocytes / Q000166:cytology / Q000187:drug effects; D019070:Cell Lineage / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D015316:Genetic Therapy; D006801:Humans; D016086:Lentivirus / Q000378:metabolism; D051379:Mice; D009703:Nucleoside-Phosphate Kinase / Q000378:metabolism; D010641:Phenotype; D020123:Sirolimus / Q000494:pharmacology; D014161:Transduction, Genetic; D019076:Transgenes / Q000235:genetics; D015215:Zidovudine / Q000494:pharmacology
chemical_list: D018941:Antigens, CD19; D015215:Zidovudine; C025946:3-methyladenine; D009703:Nucleoside-Phosphate Kinase; C020093:dTMP kinase; D000225:Adenine; D020123:Sirolimus
keywords: DTYMK/TMPK; autophagy; cell-fate control; rapamycin; suicide gene
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