Delivery of Human Interferon-γ via Gene TransferIn Vitro:Prolonged Expression and Induction of Macrophage Antimicrobial Activity

Abstract

Daily parenteral administration of exogenous interferon-gamma (IFN-gamma) induces or accelerates recovery in experimental and human infections. To develop an alternative delivery system, a replication-defective recombinant adenovirus expressing human IFN-gamma was constructed. The complete coding region of IFN-gamma was amplified by RT-PCR and inserted into an adenovirus cloning vector under the control of a human cytomegalovirus promoter. Recombinant adenovirus containing the IFN-gamma minigene (dAv-IFN-gamma) was isolated from 293 cells co-transfected with the linearized plasmid and an E1 region-deleted fragment of adenovirus genome. Following in vitro infection with dAv-IFN-gamma, dose-dependent and time-dependent expression of IFN-gamma, mRNA and production of soluble protein were demonstrated in human diploid fibroblat and HeLa cell cultures by Northern blot and ELISA, respectively. Extracellular protein secretion persisted for > = 4 weeks following initial transfection, and secreted IFN-gamma induced both antiviral activity (8000-25,000 U/ml) and macrophage activation with killing of intracellular Toxoplasma gondii and leishmania donovani. These results establish that dAv-IFN-gamma generates long-term secretion of biologically active IFN-gamma in vitro and suggest that this vector may be a useful delivery system for cytokine therapy.

Altered serum cytokine and immunoglobulin levels in the workers exposed to antimony

“…As in the case of berylliosis, the chronic lung in¯ammation is related to HLA-DP genetics and not to the extent to which workers were exposed to beryllium. 40 The present study suggests that Sb exposure may suppress host defense against infectious microorganisms, because IFNg, a major cytokine mediating cell-mediated immune responses for the clearance of infectious pathogens, 41 was signi®cantly lowered in the Group A Sb-exposed workers (Figure 3). Furthermore, the Group B subjects also showed lower IFNg levels than Group C. According to the urine Sb levels (Table 1), Group B intra-factory control subjects were indirectly exposed to Sb although to a signi®cantly lesser degree than the Group A workers.…”

Development of DNA-Designed Avian IgY Antibodies for Quantitative Determination of Bovine Interferon-Gamma

“…2). Stoeckle et al (1996) [16] have demonstrated that human IFN-γ can be secreted in vitro by transfected HeLa cells and fibroblasts using a replication defective adenovirus for >4 weeks. To express caprine IFN-γ protein in COS-7 cells, Beyer et al (1998) [17] have used vectors containing CMV and caprine arthritis encephalitis virus long terminal repeat (CAEV LTR) promoters and employed Lipofectamine transfection method.…”

Interferon‐γ Therapy: Evaluation of Routes of Administration and Delivery Systems

“…This process, termed transfection, can be accomplished using a number of vectors, which include viral vectors, polymers, and liposomes, and can be accomplished in vivo or in vitro 116. This approach has been used in numerous studies to obtain a sustained local IFN‐γ concentration 117–123. For example, Stoeckle et al have demonstrated in vitro that IFN‐γ can be secreted by transfected HeLa cells and fibroblasts using a replication‐defective adenovirus for > 4 weeks 120.…”

“…This approach has been used in numerous studies to obtain a sustained local IFN‐γ concentration 117–123. For example, Stoeckle et al have demonstrated in vitro that IFN‐γ can be secreted by transfected HeLa cells and fibroblasts using a replication‐defective adenovirus for > 4 weeks 120. The possible utility of the approach has also been demonstrated in Phase I clinical trials.…”