@article{f70c060d9c7b46d3b01a76c18a4799c9,
title = "Foamy virus vectors expressing anti-HIV transgenes efficiently block HIV-1 replication",
abstract = "Gene therapy has the potential to control human immunodeficiency virus (HIV) in patients who do not respond to traditional antiviral therapy. In this study, we tested foamy virus (FV) vectors expressing three anti-HIV transgenes, both individually and in a combination vector. The transgenes tested in this study are RevM10, a dominant negative version of the viral rev protein, Sh1, a short hairpin RNA directed against a conserved overlapping sequence of tat and rev, and membrane-associated C46 (maC46), a membrane-attached peptide that blocks HIV cell entry. FV vectors efficiently transduce hematopoietic stem cells and, unlike lentivirus (LV) vectors, do not share viral proteins with HIV. The titers of the FV vectors described in this study were not affected by anti-HIV transgenes. On a direct comparison of FV vectors expressing the individual transgenes, entry inhibition using the maC46 transgene was found to be the most effective at blocking HIV replication. A clinically relevant FV vector expressing three anti-HIV transgenes effectively blocked HIV infection in primary macrophages derived from transduced, peripheral blood CD34-selected cells and in a cell line used for propagating HIV, CEM×174. These results suggest that there are potential benefits of using FV vectors in HIV gene therapy.",
author = "Taylor, {Jason A.} and Lucia Vojtech and Ingrid Bahner and Kohn, {Donald B.} and Laer, {Dorothee Von} and Russell, {David W.} and Richard, {Robert E.}",
note = "Funding Information: John Rossi provided critically important plasmids as well as technical advice and encouragement. Erik Olson and James Allen of the University of Washington (UW) Vector Core assisted J.A.T. with vector production and development of vector storage methods. Jennifer Hemplemann and Kevin Otto provided technical assistance. Reagents including the 2F5 antibody and TZM-bl and CEMx174 cells were obtained from the NIH AIDS Research & Reference Reagent Program. D.W.R. was supported by grants HL085107 and HL53750. J.A.T. was supported by K08 AI65389 from National Institute of Allergy and Infectious Diseases. R.R. was supported by K23 RR16509 from the National Center for Research Resources, a pilot project grant from the General Clinical Research Center at the UW, supported by the NIH, grant M01RR-00037, and a pilot grant from the Fred Hutchinson Cancer Research Center. RR also received support from the Core Center for Experimental Hematology at the Fred Hutchinson Cancer Research Center (DK56465). Funding Information: Hematopoietic cell culture, transduction, and differentiation. Human CD34– selected cells were isolated from apheresed granulocyte colony-stimulating factor-mobilized healthy donors using a CliniMacs system (Miltenyi, Auburn, CA), and processed by the Cell Processing Core, Fred Hutchinson Cancer Research Center, sponsored by the National Institutes of Health National Heart, Lung, and Blood Institute Program for Excellence in Gene Therapy. Transductions were performed for 12 hours in CH296-coated (10 μg/cm 2 , Takara Shuzo, Otsu, Japan) 48-well plates with 1 × 10 5 cells per well in 200 μl volume containing concentrated FV vector stock at an MOI of 3 (as determined on HT1080 cells) in Iscove's modified Dulbecco's medium (IMDM) supplemented with 20% (vol/vol) FBS (HyClone), 100 ng/ml of flt-3 ligand (FL), 100 ng/ml stem-cell factor (SCF), and 50 ng/ml of thrombopoietin. The following day, cells were washed and re-plated in IMDM with 20% (vol/vol) FBS, 50 ng/ml of FL, 50 ng/ml SCF, 50 ng/ml interleukin-3 (IL-3), and 25 ng/ml thrombopoietin. At day 4 cells were FACS sorted (FACSVantage SE cell sorter, Becton Dickinson, Franklin Lakes, NJ) for GFP or maC46 expression (using the 2F5 antibody against GP-41 obtained from the NIH AIDS Research & Reagent Program). Sorted cells were re-plated in IMDM with 20% (vol/vol) FBS, 50 ng/ml of SCF, 50 ng/ml IL-3, and 50 ng/ml IL-6 for 8 days, followed by re-plating in IMDM with 20% (vol/vol) FBS, 50 ng/ml of granulocyte macrophage colony-stimulating factor, 25 ng/ml SCF, 25 ng/ml IL-3, and 25 ng/ml IL-6 for 22 days of further expansion and differentiation. Adherent cells were re-plated in 48-well plates at 40,000 cells per well and maintained in IMDM with 20% (vol/vol) FBS, 50 ng/ml of granulocyte–macrophage colony-stimulating factor, 25 ng/ml SCF, 25 ng/ml IL-3, and 25 ng/ml IL-6. The following day the media were replaced with 100 μl of media containing HIV JR-FL , at an MOI of 0.25. After 2 hours the media were removed, cells washed and 200 μl of new media added. Every 4 days media were changed, and p24 levels were determined using enzyme-linked immunosorbent assay (ZeptoMetrix, Buffalo, NY). ",
year = "2008",
month = jan,
doi = "10.1038/sj.mt.6300335",
language = "English (US)",
volume = "16",
pages = "46--51",
journal = "Molecular Therapy",
issn = "1525-0016",
publisher = "Nature Publishing Group",
number = "1",
}