Development of a nonintegrating Rev-dependent lentiviral vector carrying diphtheria toxin A chain and human TRAF6 to target HIV reservoirs.
Persistence of human immunodeficiency virus (HIV) despite highly active antiretroviral therapy (HAART) is a lasting challenge to virus eradication. To develop a strategy complementary to HAART, we constructed a series of Rev-dependent lentiviral vectors carrying diphtheria toxin A chain (DT-A) and its attenuated mutants, as well as human tumor necrosis factor receptor-associated factor 6 (TRAF6). Expression of these suicide genes following delivery through viral particles is dependent on Rev, which exists only in infected cells. Among these toxins, DT-A has been known to trigger cell death with as little as a single molecule, whereas two of the attenuated mutants in this study, DT-A(176) and DT-A(Delta N), were well tolerated by cells at low levels. TRAF6 induced apoptosis only with persistent overexpression. Thus, these suicide genes, which induce cell death at different expression levels, offer a balance between efficacy and safety. To minimize possible mutagenesis introduced by retroviral integration in nontarget cells, we further developed a nonintegrating Rev-dependent (NIRD) lentiviral vector to deliver these genes. In addition, we constructed a DT-A-resistant human cell line by introducing a human elongation factor 2 mutant into HEK293T cells. This allowed us to manufacture the first high-titer NIRD lentiviral particles carrying DT-A to target HIV-positive cells.
Wang
Z
Z
Transplantation Biology Research Center, Massachusetts General Hospital, Harvard Medical School, Harvard University, Boston, MA, USA.
Tang
Z
Z
Zheng
Y
Y
Yu
D
D
Spear
M
M
Iyer
S R
SR
Bishop
B
B
Wu
Y
Y
D017209:Apoptosis; D016923:Cell Death; D002460:Cell Line; D004279:DNA, Viral / Q000378:metabolism; D004167:Diphtheria Toxin / Q000235:genetics* / Q000378:metabolism; D015200:Fluoroimmunoassay; D015694:Gene Products, rev / Q000235:genetics*; D015316:Genetic Therapy / Q000379:methods; D005822:Genetic Vectors / Q000235:genetics* / Q000378:metabolism; D015658:HIV Infections / Q000235:genetics; D015497:HIV-1 / Q000235:genetics / Q000502:physiology*; D006801:Humans; D016086:Lentivirus / Q000235:genetics* / Q000378:metabolism; D010446:Peptide Fragments / Q000235:genetics* / Q000378:metabolism; D048029:TNF Receptor-Associated Factor 6 / Q000235:genetics* / Q000378:metabolism; D014779:Virus Replication / Q000502:physiology
D004279:DNA, Viral; D004167:Diphtheria Toxin; D015694:Gene Products, rev; D010446:Peptide Fragments; D048029:TNF Receptor-Associated Factor 6; C029795:diphtheria toxin fragment A
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