Rapid generation of combined CMV-specific CD4+ and CD8+ T-cell lines for adoptive transfer into recipients of allogeneic stem cell transplants.
Adoptive transfer of cytomegalovirus (CMV)-specific T cells can restore long-lasting, virus-specific immunity and clear CMV viremia in recipients of allogeneic stem cell transplants if CD4(+) and CD8(+) CMV-specific T cells are detected in the recipient after transfer. Current protocols for generating virus-specific T cells use live virus, require leukapheresis of the donor, and are time consuming. To circumvent these limitations, a clinical-scale protocol was developed to generate CMV-specific T cells by using autologous cellular and serum components derived from a single 500-mL blood draw. CMV-specific T cells were stimulated simultaneously with CMV-specific major histocompatibility complex class I (MHC I)- restricted peptides and CMV antigen. Activated T cells were isolated with the interferon-gamma (IFN-gamma) secretion assay and expanded for 10 days. In 8 randomly selected, CMV-seropositive donors, 1.34 x 10(8) combined CD4(+) and CD8(+) CMV-specific T cells, on average, were generated, as determined by antigen-triggered IFN-gamma production. CMV-infected fibroblasts were efficiently lysed by the generated T cells, and CMV-specific CD4(+) and CD8(+) T cells expanded if they were stimulated with natural processed antigen. On the other hand, CD4(+) and CD8(+) T cell-mediated alloreactivity of generated CMV-specific T-cell lines was reduced compared with that of the starting population. In conclusion, the culture system developed allowed the rapid generation of allodepleted, highly enriched, combined CD4(+) and CD8(+) CMV-specific T cells under conditions mimicking good manufacturing practice.
Rauser
Georg
G
Medizinische Klinik II, Eberhard-Karls-Universität, Tuebingen, Germany.
Einsele
Hermann
H
Sinzger
Christian
C
Wernet
Dorothee
D
Kuntz
Gabriele
G
Assenmacher
Mario
M
Campbell
John D M
JD
Topp
Max S
MS
D019264:Adoptive Transfer / Q000379:methods*; D000956:Antigens, Viral / Q000276:immunology; D001782:Blood Donors; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology* / Q000637:transplantation; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology* / Q000637:transplantation; D018929:Cell Culture Techniques / Q000379:methods; D003587:Cytomegalovirus / Q000276:immunology*; D018380:Hematopoietic Stem Cell Transplantation / Q000379:methods*; D006801:Humans; D008213:Lymphocyte Activation; D010455:Peptides / Q000276:immunology; D014184:Transplantation, Homologous
D000956:Antigens, Viral; D010455:Peptides