[57] Optimized Lentiviral Vectors for HIV Gene Therapy: Multiplexed Expression of Small RNAs and Inclusion of MGMTP140K Drug Resistance Gene

Janet Chung, Lisa Scherer, Agnes Gardner, David DiGiusto, John Rossi. Molecular and Cell Biology, City of Hope, Duarte, CA; Laboratory for Cellular Medicine, City of Hope, Duarte, CA; Virology, City of Hope, Duarte, CA

Gene therapy with hematopoietic stem and progenitor cells (HSPCs) is a promising approach to engineering immunity to HIV and may lead to a functional cure for AIDS. In support of this approach, we created lentiviral vectors that utilized the MCM7 platform to express a diverse set of small anti-viral RNAs and a drug resistance gene (MGMTP140K). Multiple strategies for simultaneous expression of up to five RNA transgenes were tested. The placement and orientation of each transgene and its promoter were important determinants for optimal gene expression. RNA expression from the MCM7 platform with a U1 promoter was sufficient to provide protection from R5 tropic HIV in macrophages and resulted in reduced hematopoietic toxicity compared to constructs expressing RNA from independent Pol III promoters. Surprisingly, the addition of an HIV entry inhibitor and a nucleolar-localizing transcriptional inhibitor (TAR) did not enhance antiviral potency over constructs that targeted only viral RNA transcripts. We also demonstrated selective enrichment of gene modified HSPCs during in vitro expansion in the presence of BCNU. The use of these less toxic, potent anti-HIV vectors expressing a drug selection marker is likely to enhance the in vivo efficacy of our stem cell gene therapy approach to treating HIV/AIDS.
Keywords: Infectious Diseases; RNA Viral Vectors; RNAi and shRNA

Session: Poster Session: RNA Virus Vectors I (5:30 PM-7:30 PM)
Date/Time: Wednesday, May 21, 2014 - 5:30 PM
Room: Hall A and B South


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