CD7-specific Single Chain Antibody Mediated Delivery of siRNA to T Cells Inhibits HIV Replication in a Humanized Mouse Model

초록

Background: A serious hurdle to the development of RNA interference as a therapy for AIDS is the delivery of siRNA to T lymphocytes, the major target of HIV infection, which are difficult cells to transfect even in vitro. We have employed a single-chain antibody (scFv) to the pan T cell surface antigen CD7 to deliver antiviral siRNA into T cells and tested this method as a potential therapy for HIV infection in a humanized mouse model. Methods: We expressed CD7scFv in E.coli using the bacterial expression vector pET26b and conjugated the purified scFv to an oligo-9R peptide (scFvCD7-9R) that can bind nucleic acids by charge interactions. To test the ability of the chimeric protein to deliver siRNA to T cells in vivo, we reconstituted NOD/SCIDIL2rγc-/- mice with human peripheral blood lymphocytes (Hu/PBL) from normal or HIV seropositive donors. Mice reconstituted with PBL from normal donors were infected ip with HIVBaL. The ability of iv administered scFv/9R complexed host and/or antiviral siRNAs to suppress viral replication was evaluated by serial analyses of CD4/CD3 T cell ratios (ratio used to normalize for reconstitution levels), p24 antigen levels and HIV-1 RNA levels in sera of experimental mice. Results: scFvCD7-9R efficiently delivered CD4 siRNA into human T cells in vitro. In vivo administration to Hu/PBLmice resulted in reduced levels of surface CD4 expression on T cells. Mice infected with HIV-1 and treated on a weekly basis with scFvCD7-9R/ siRNA complexes targeting a combination of viral genes and the host coreceptor molecule CCR5 successfully maintained CD4/CD3 T cell ratios up to 5 weeks after infection in contrast to control mice that displayed a marked reduction in CD4 T cell numbers. p24 antigen levels were undetectable in 3 of the 4 protected mice. scFvCD7-9R/antiviral siRNA treatment also helped maintain CD4 T cell numbers with reduced plasma viral loads in Hu-PBL mice reconstituted with PBMC from donors seropositive for HIV, indicating that this method can contain viral replication even in established HIV infections. Conclusions: Our results show that scFvCD7-9R could be further developed as a potential therapeutic for HIV-1 infection. Keywords: RNAi, CD7, humanized mice