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CD7-specific Single Chain Antibody-mediated Delivery of siRNA to T Cells Inhibits HIV Replication in a Humanized Mouse Model
Priti Kumar*1, H Ban2, S Kim1, M Peipp3, G Fey3, L Schultz4, N Manjunath1, S Lee2, and P Shankar1
1Immune Disease Inst, Boston, MA, US; 2Hanyang Univ, Seoul, Korea; 3Univ of Erlangen, Germany; and 4Jackson Lab, Bar Harbor, ME, US
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 scFvCD7in 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 intravenously administered
scFv/9R complexed host and antiviral siRNA 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-PBL mice
resulted in reduced levels of surface CD4 expression on T cells. Mice infected
with HIV-1 and treated weekly with scFvCD7-9R/siRNA complexes targeting a
combination of viral genes and the host co-receptor molecule CCR5 successfully
maintained CD4/CD3 T cell ratios as long as 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.
Conclusions: scFvCD7-9R/antiviral siRNA treatment helped
maintain CD4 T cell numbers and 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.
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