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Abstract

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. Autologous cultured chondrocyte transplantation was introduced in Sweden in 1987 for the treatment of full thickness chondral defects of the knee. The results from the first 219 patients with a follow up of 2-10 years treated using this technique are reported.Patients were assessed with three types of endpoints: patient and physician derived clinical rating scales, arthroscopy assessments of cartilage fill, integration, and surface hardness and histology of graft biopsies. There was a high percentage of good to excellent results (84-90%) in patients with different types of single femoral condyle lesions, while other types of lesions had a lower degree of success (mean 74%). Furthermore, the long-term durability of the grafted area has been evaluated in a group of 61 patients with femoral condyle and patellar lesions who where followed for a mean of 7.4 (range 5-11) years and where the durability was calculated as the comparison between the long-term follow up and clinical status at 2 years post surgery. At 2 ...