Empirical advantages of adeno associated viral vectors for in vivo gene therapy for arthritis

Objective, To evaluate the utility of the adeno associated viral (AAV) vect or for gene delivery to joint cells in vivo and in vitro, and to assess its potential as a vector for arthritis gene therapy. Methods. A recombinant AAV (rAAV) vector expressing the bacterial beta-gala ctosidase (beta-gal) gene (rAAV-CMV-LacZ) was directly introduced into heal thy-normal mouse knees, or arthritic knees in mice overexpressing tumor nec rosis factor-alpha (hTNF alpha-Tg). beta-gal expression levels were determi ned by immunohistochemistry and chemiluminescence. The transduction efficie ncy of this vector on primary. fibroblast-like synoviocytes (FLS) in vitro was determined by FAGS. The effects of UV and gamma-irradiation as well as TNF-alpha on transduction efficiency were determined using the same methods . Results, We found little evidence of rAAV transduction in the joint cells o f healthy mice. Target gene expression was detected in all animals at Day 3 , and peaked at Day 7 before returning to baseline levels 21 days after inj ection. In contrast, synoviocytes, articular chondrocytes, and meniscal cel ls of diseased mice were transduced by rAAV-CMV-LacZ in hTNF alpha-Tg anima ls. Transduction efficiencies correlated with joint damage, and target gene expression was up to 10-fold greater than that seen in the normal mice. In vitro, we found that rAAV transduction of FLS can be enhanced by pretreatm ent with UV or gamma-irradiation and TNF-alpha stimulation. Conclusion, We find that rAAV vectors have several empirical advantages for in vivo gene therapy for arthritis: (1) rAAV preferentially transduces art hritic joint cells in vivo. (2) rAAV can transduce both FLS and chondrocyte s in vivo. (3) rAAV transduction of FLS can be augmented by pretreatment wi th agents that induce DNA repair enzymes.