Clinical-scale lentiviral vector transduction of PBL for TCR gene therapy and potential for expression in less-differentiated cells.
In human gene therapy applications, lentiviral vectors may have advantages over gamma-retroviral vectors because of their ability to transduce nondividing cells, their resistance to gene silencing, and a lack of integration site preference. In this study, we used VSV-G pseudotype third generation lentiviral vectors harboring specific antitumor T-cell receptor (TCR) to establish clinical-scale lentiviral transduction of peripheral blood lymphocyte (PBL). Spinoculation (1000g, 32 degrees C for 2 h) in the presence of protamine sulfate represents the most efficient and economical approach to transduce a large number of PBLs compared with RetroNectin-based methods. Up to 20 million cells per well of a 6-well plate were efficiently transduced and underwent an average 50-fold expansion in 2 weeks. TCR transduced PBL-mediated specific antitumor activities including interferon-gamma release and cell lysis. Compared with gamma-retroviral vectors, the TCR transgene could be preferentially expressed on a less-differentiated cell population.
Yang
Shicheng
S
Surgery Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Rosenberg
Steven A
SA
Morgan
Richard A
RA
D024221:Antineoplastic Protocols; D002454:Cell Differentiation; D045744:Cell Line, Tumor; D003131:Combined Modality Therapy; D003602:Cytotoxicity, Immunologic; D005822:Genetic Vectors; D006801:Humans; D007111:Immunity, Cellular / Q000235:genetics; D007371:Interferon-gamma / Q000378:metabolism; D016086:Lentivirus / Q000276:immunology*; D008214:Lymphocytes / Q000378:metabolism*; D040542:Mechanotransduction, Cellular / Q000276:immunology; D008545:Melanoma / Q000473:pathology / Q000628:therapy*; D008863:Microspheres; D011479:Protamines / Q000378:metabolism; D011948:Receptors, Antigen, T-Cell / Q000235:genetics* / Q000378:metabolism; D016176:T-Lymphocyte Subsets / Q000378:metabolism* / Q000473:pathology; D013601:T-Lymphocytes / Q000378:metabolism* / Q000473:pathology; D014161:Transduction, Genetic; D019076:Transgenes
D011479:Protamines; D011948:Receptors, Antigen, T-Cell; D007371:Interferon-gamma