Cytolytic T lymphocytes play an important role in host defense against viral infections, including human immunodeficiency virus (HIV). In a phase I clinical trial (protocol 080 of the AIDS Clinical Trials Group), generation of CD8+ effector cells from peripheral blood of patients with acquired immunodeficiency syndrome (AIDS)-related complex (ARC) or AIDS and safety of autologous adoptive transfer of these cells were evaluated. For therapeutic infusions, CD8+ T cells were purified by positive selection on anti-CD8 monoclonal antibody-coated flasks from leukapheresed peripheral blood of seven patients. These CD8+ T cells were cultured in the presence of interleukin-2 and phytohemagglutinin for up to 3 weeks to obtain cells sufficient for therapeutic infusions (10(8) to 10(10)). All 31 cell cultures established from the seven patients and used for therapy were highly enriched in CD8+ (mean, 97%), CD8+HLA-DR+ (50%), cytotoxic CD8+CD11b- (82%), and memory CD29+ (78%) T lymphocytes. In vitro expanded CD8+ cells had excellent cytotoxic function at the time they were used for therapy, including HIV-specific activity against autologous targets infected with vaccinia vectors expressing HIV-IIIb antigens, gag, pol, and env. Anti-HIV activity of cultured CD8+ cells was significantly higher than that of autologous fresh peripheral blood lymphocytes. Our results show that CD8+ T lymphocytes obtained from peripheral blood of symptomatic HIV-infected patients can be purified, cultured to obtain large numbers of cells with enhanced anti-HIV activity, and safely infused into patients with AIDS as a form of immunotherapy.
There is increasing interest in adeno-associated virus (AAV) vectors for a wide variety of gene therapy applications. AAV is a nonpathogenic human parvovirus that can mediate long-term transduction of a number of cell types without provoking a significant immune response. These properties make AAV especially attractive for use in gene therapy of rheumatoid arthritis (RA), a chronic inflammatory disease. To investigate the potential of AAV in gene therapy of arthritis, the ability of AAV to infect synovium in vitro and in vivo was tested. Three human RA synovial fibroblast cell lines and two murine (one DBA/1J and one DBA1J×C3H F1) synovial fibroblast cell lines were used to test AAV transduction in vitro. The cell lines (2 × 10 5 cells) were infected with 10 4 particles/cell of a murine IL-10-encoding vector (AAV-mIL-10) alone or with the addition of a low titer (100 particles/cell) of an E1-, E3-deleted recombinant adenovirus to provide E4orf6 activity to enhance second-strand synthesis. The supernatants were harvested from the wells at various time points and assayed for mIL-10 expression by ELISA. Both human synovial cell lines infected with AAV alone demonstrated low-level transgene expression throughout the course of the study. However, by day 10, all human cultures coinfected with adenovirus showed a 16-to 56-fold increase in mIL-10 compared to cultures infected with AAV-mIL10 alone. By day 30, a 31-to 135-fold increase was observed. No such increase was observed in any of the mouse cell lines. To determine the AAV transduction efficiency for synovium in vivo, human RA synovial tissues obtained from patients undergoing joint-replacement surgery were implanted subcutaneously on the backs of NOD.CB17-Prkdc SCID mice. After allowing a 2-week period for engraftment, tissues were injected with 3.4 × 10 11 particles of AAV-luciferase alone or in combination with 1.0 × 10 11 particles of adenovirus. Two weeks following AAV administration, the tissues were homogenized and assayed for expression of luciferase. Only the tissues coinfected with adenovirus had luciferase levels above background. A similar experiment with AAV-LacZ demonstrated X-gal staining only of synovial tissues coinfected with adenovirus. These findings demonstrate a preferential ability of AAV to transduce human, compared to mouse, synovial tissue and suggest that second strand synthesis may be a limiting factor in gene transduction. Further studies to elucidate the mechanisms limiting gene transduction in human synovium may allow optimization of this vector for the treatment of arthritis. Research of the last years has demonstrated clearly the role of rheumatoid arthritis synovial fibroblasts (RA-SF) in the destruction of articular cartilage. It has been understood that RA-SF not only exhibit features of activation and altered apoptosis, but following attachment to cartilage secrete large amounts of matrix degrading enzymes that mediate the destruction of extracellular matrix. Given recent advances in the field of gene transfer, we have been wor...
Cytolytic T lymphocytes play an important role in host defense against viral infections, including human immunodeficiency virus (HIV). In a phase I clinical trial (protocol 080 of the AIDS Clinical Trials Group), generation of CD8+ effector cells from peripheral blood of patients with acquired immunodeficiency syndrome (AIDS)-related complex (ARC) or AIDS and safety of autologous adoptive transfer of these cells were evaluated. For therapeutic infusions, CD8+ T cells were purified by positive selection on anti-CD8 monoclonal antibody-coated flasks from leukapheresed peripheral blood of seven patients. These CD8+ T cells were cultured in the presence of interleukin-2 and phytohemagglutinin for up to 3 weeks to obtain cells sufficient for therapeutic infusions (10(8) to 10(10)). All 31 cell cultures established from the seven patients and used for therapy were highly enriched in CD8+ (mean, 97%), CD8+HLA-DR+ (50%), cytotoxic CD8+CD11b- (82%), and memory CD29+ (78%) T lymphocytes. In vitro expanded CD8+ cells had excellent cytotoxic function at the time they were used for therapy, including HIV-specific activity against autologous targets infected with vaccinia vectors expressing HIV-IIIb antigens, gag, pol, and env. Anti-HIV activity of cultured CD8+ cells was significantly higher than that of autologous fresh peripheral blood lymphocytes. Our results show that CD8+ T lymphocytes obtained from peripheral blood of symptomatic HIV-infected patients can be purified, cultured to obtain large numbers of cells with enhanced anti-HIV activity, and safely infused into patients with AIDS as a form of immunotherapy.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
customersupport@researchsolutions.com
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.