title: Generation of robust cytotoxic T lymphocytes against prostate specific antigen by transduction of dendritic cells using protein and recombinant adeno-associated virus.
abstract: Prostate cancer is the most common male cancer and there is an urgent need for adjuvant therapy such as immunotherapy. Recombinant adeno-associated virus type 2 (rAAV) vectors are useful for antigen gene-loading of human dendritic cells (DC) and for the rapid generation of cytotoxic T lymphocytes (CTL). In this study, we report a protocol for AAV-loading of DC with the AAV-loading of self-antigen prostate specific antigen (PSA) resulting in generation of CTL. PSA and cytokine expression, Cell surface marker analysis of DC and CTL cells were done using a FACScalibur flow cytometer. Chromium-51 release assay was used to analyze the killing activity of CTL. It was found that AAV-loading of DC with the PSA gene is superior to PSA protein loading of the same antigen for generating effective CTL. AAV/PSA-loading of DC was found to result in: (1) strong, rapid PSA-specific, MHC Class I-restricted CTL, (2) PSA expression in DC, (3) high CD80, CD83, and CD86 expression on DC, (4) high level of IL-12 and low level of IL-10 in DC, (5) T cell populations with significant interferon gamma (IFNgamma) expression, but low IL-4 expression, (6) high proliferation of T cell populations, (7) high CD8:CD4 and CD8:CD56 T cell ratios. The reason for generation of robust CTL is partly explained by the characteristics of DC and CTL described. This protocol may be useful for adoptive immunotherapy against self antigens such as PSA for prostate cancer.
authors.affiliation: Gene Therapy Program, University of Arkansas for Medical Sciences, Little Rock, AR 72205, USA, and Department of Oncology, Nanfang Hospital, Guangzhou, China. Mahadevanmahendran@uams.edu
authors.forename: Mahendran
authors.initials: M
authors.lastname: Mahadevan
authors.forename: Yong
authors.initials: Y
authors.lastname: Liu
authors.forename: Changxuan
authors.initials: C
authors.lastname: You
authors.forename: Rongcheng
authors.initials: R
authors.lastname: Luo
authors.forename: Hong
authors.initials: H
authors.lastname: You
authors.forename: Jawahar L
authors.initials: JL
authors.lastname: Mehta
authors.forename: Paul L
authors.initials: PL
authors.lastname: Hermonat
mesh_terms: D015703:Antigens, CD / Q000032:analysis; D002478:Cells, Cultured; D016207:Cytokines / Q000378:metabolism; D003713:Dendritic Cells / Q000276:immunology*; D000229:Dependovirus / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D015395:Histocompatibility Antigens Class I / Q000276:immunology; D006801:Humans; D016219:Immunotherapy, Adoptive / Q000379:methods; D008297:Male; D017430:Prostate-Specific Antigen / Q000235:genetics / Q000276:immunology*; D011471:Prostatic Neoplasms / Q000628:therapy; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology* / Q000637:transplantation; D014161:Transduction, Genetic
chemical_list: D015703:Antigens, CD; D016207:Cytokines; D015395:Histocompatibility Antigens Class I; D017430:Prostate-Specific Antigen