Evaluation of recombinant alphaviruses as vectors in gene therapy

Alphavirus vectors based on Sindbis virus and Semliki Forest virus (SFV) we re characterized as potential gene transfer vectors. Initial studies were p erformed using vectors engineered to transfer either lacZ or green fluoresc ent protein (GFP). High levels of gene transfer were achieved in human prim ary fibroblasts, BHK and 293T cells, with low levels of transduction observ ed in more than 20 other target cells. Alphavirus-based expression was gene rally very high, but transient in every cell type. Replication-competent al phavirus was never detected in SFV preparations but could be produced by Si ndbis-based vectors at a frequency of up to 3 x 10(-3) infectious units per mi. We constructed a human clotting factor IX (hFIX) cDNA-containing Sindb is virus and compared it with hFIX cDNA-harboring adenoviral and retroviral vectors. In most cases, hFIX levels obtained with Sindbis vector were init ially at least an order of magnitude higher than those obtained with other viral vectors. These data demonstrate that alphavirus vectors compare favor ably with adenovirus vectors as systems to promote high-level transient gen e expression and should be considered as an alternative vector for gene tra nsfer and potential gene therapy studies.