ABSTRACT
Unmatched human leukocyte antigens (HLA) expressed by allogeneic donor cells are the major target for immunological rejection. In order to reduce the immunogenicity of allograft cells, we have developed lentiviral vectors for delivery of short hairpin ribonucleic acid (shRNA) against Class I HLA. This approach was evaluated in both an established human embryonic kidney cell line and primary human CD34+ hematopoietic stem/progenitor cells. Target cells transduced with lentiviral vectors expressing either HLA-A*0201 allele-specific or HLA-A, -B, -C consensus sequence-specific shRNA showed effective knockdown of cell surface HLA expression. Mixed lymphocyte-target cell reactions showed significantly reduced interferon-gamma production from alloreactive cytotoxic T lymphocytes and significantly reduced levels of target cell apoptosis after shRNA-mediated knockdown of HLA expression and target cell survival correlated with vector transduction efficiency. Furthermore, increasing resistance to complement-dependent cytotoxicity mediated by anti-HLA antibodies was observed to correlate with increasing levels of shRNA vector transduction in primary human CD34+ cells. Notably, non-HLA restricted killing by lymphokine-activated killer cells was not incurred after HLA knockdown. These data demonstrate the potential for genetic engineering strategies targeting incompatible HLA alleles to reduce both cellular and humoral responses and enable graft survival after transplantation of allogeneic cells and tissues.
Subject(s)
Genetic Therapy/methods , Graft Rejection/genetics , Graft Rejection/immunology , Histocompatibility Antigens Class I/genetics , Histocompatibility Antigens Class I/immunology , RNA Interference/immunology , Antibody Specificity , Antigens, CD34/metabolism , Cell Line, Transformed , Fetus/cytology , Fetus/metabolism , HEK293 Cells , Humans , Isoantigens/immunology , Killer Cells, Lymphokine-Activated/immunology , Lentivirus/genetics , Primary Cell CultureABSTRACT
PMDC05, a leukemic plasmacytoid dendritic cell (pDC) line which was established in our laboratory, showed a capacity of generating antigen-specific cytotoxic T lymphocytes (CTLs). In order to enhance an antigen presenting ability of PMDC05, PMDC05 was transduced with CD80 gene by lentiviral vector, which was named as PMDC11. PMDC11 displayed a strong antigen presenting ability even without any stimulation, and by culturing with stimulators such as calcium ionophore PMDC11 gained a more potent antigen presenting ability. Our data suggested PMDC11 could be applied as antigen presenting cells more efficiently in adoptive cellular immunotherapy for tumors and severe infections in comparison with PMDC05.
