Delivery of antisense constructs and ribozymes to inhibit cartilage destruction in the SCID mouse model of RA

Abstract

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 working on specific strategies to interfere with the expression of disease relevant matrix degrading enzymes using the complementary approaches of ribozymes and antisense expression constructs. Ribozymes are short RNA molecules that have catalytic activity and are capable of cleaving mRNA thus inhibiting its translation. We have used retroviral gene transfer of ribozymes against MMP-1 as well as cathepsins B and L to inhibit the expression of these enzymes in RA-SF both in vitro and when implanted together with normal articular cartilage into severe combined immunodeficient (SCID) mice. As demonstrated in vitro, gene transfer of such ribozymes results in a sustained, up to 60% decrease of enzyme production in RA-SF over 60 days. Currently, SCID mouse experiments are underway to study the effect of gene transfer with these ribozymes on cartilage degradation in vivo. To evaluate the potential of antisense constructs as an alternative approach, we have generated antisense constructs against the novel MT1-MMP and transduced RA-SF using a retroviral system. First data indicate a high efficacy of MT1-MMP antisense in inhibiting the production of MT1-MMP. However, constantly high levels of MT1-MMP antisense RNA are needed for a sustained effect in RA-SF. The focus of our current efforts is to optimize the construct both in terms of specificity and its level of expression.