Interleukin-4 adenoviral gene therapy reduces production of inflammatory cytokines and prostaglandin E2 by rheumatoid arthritis synovium ex vivo.

BACKGROUND: The rheumatoid arthritis (RA) joint is characterized by an inflammatory synovial pannus which mediates tissue destruction. Interleukin (IL)-4 reduces the production of many proinflammatory cytokines, particularly by activated macrophages. Therefore, we examined the ability of adenovirally delivered IL-4 for the treatment of human RA to reduce the secretion of proinflammatory molecules. METHODS: Adenoviral vectors encoding the genes for human IL-4 (AxCAIL-4) and bacterial beta-galactosidase (AxCAlacZ) were generated and examined for appropriate production and biological activity. RA synovial tissue (ST) explants or fibroblasts were infected with AxCAIL-4 or a beta-galactosidase producing vector, as a control, and conditioned medium (CM) was collected for ELISA analysis. RESULTS: AxCAIL-4 decreased the production of the inflammatory cytokines IL-1 beta and tumor necrosis factor-alpha in RA ST explant CM. IL-8 levels were significantly reduced by 71%, 88%, and 82% at 24, 48, and 72 hours, respectively, in RA ST explant CM. In the same CM, monocyte chemotactic protein-1 (MCP-1) levels decreased 60% at 48 hours. In contrast, RA synovial fibroblast CM levels of MCP-1 were increased by AxCAIL-4. Epithelial neutrophil activating peptide-78 levels produced by RA ST explants were significantly decreased by AxCAIL-4 by 88%, 92%, and 93% at 24, 48, and 72 hours, respectively. Growth related gene product-alpha levels were likewise decreased in RA ST explant CM. In ST explants as well as RA synovial fibroblasts, IL-4 treatment decreased prostaglandin E2 (PGE2) production. CONCLUSIONS: Increased expression of IL-4 via gene therapy may decrease RA-associated inflammation by reducing proinflammatory cytokines and PGE2.

Differential structure-activity relationships of phosphoramidon analogues for inhibition of three metalloproteases: endothelin-converting enzyme, neutral endopeptidase, and angiotensin-converting enzyme.

The structure-activity relationships of phosphoramidon analogues for inhibition of endothelin-converting enzyme (ECE), neutral endopeptidase 24.11 (NEP), and angiotensin-converting enzyme (ACE) were compared. Phosphoramidon inhibited ECE, NEP, and ACE activities with IC50 values of 3.5, 0.034, and 78 microM, respectively. Removal of the rhamnose moiety of phosphoramidon (dipeptide 3) reduced the potency for ECE (IC50 = 70 microM), whereas the potencies for NEP (0.003 microM) and ACE (0.20 microM) were increased. Addition of 2-(2-naphthyl)ethyl to dipeptide 3 improved the potency for ECE (0.55 microM) but weakened the potency for NEP (0.02 microM), and had no significant change for ACE. Interchange between Leu and Trp abolished the inhibitory activities for ECE and NEP, but the compound remained active for ACE. These results suggest that a hydrophobic group in the P1 position of phosphoramidon analogues increases the potency for ECE significantly, whereas compounds containing a free phosphonic acid are optimal for inhibition of NEP and ACE. Furthermore, an aromatic group in the P'2 position is essential for the inhibition of ECE and NEP, but not ACE.