title: Antisense-mediated inhibition of human immunodeficiency virus (HIV) replication by use of an HIV type 1-based vector results in severely attenuated mutants incapable of developing resistance.
abstract: We have constructed a human immunodeficiency virus type 1 (HIV-1)-based lentiviral vector expressing a 937-base antisense sequence against the HIV-1 envelope gene. Transduction of CD4(+) T lymphocytes with this vector results in expression of the therapeutic antisense sequence and subsequent inhibition of productive HIV-1 replication. In this report, we examined the effect of antisense-mediated suppression on the potential development of virus escape mutants using a permissive T-cell line cultured under conditions that over serial passages specifically allowed for generation and amplification of mutants selected for by antisense pressure. In the resulting virus clones, we found a significant increase in the number of deletions at the envelope target region (91% compared to 27.5% in wild-type HIV). Deletions were most often greater than 1 kb in length. These data demonstrate for the first time that during antisense-mediated suppression of HIV, mutants develop as a direct result of selective pressure on the HIV genomic RNA. Interestingly, in clones where deletions were not observed, there was a high rate of A-G transitions in mutants at the antisense target region but not outside this region, which is consistent with those mutations that are predicted as a result of antisense-mediated modification of double-stranded RNA by the enzyme double-stranded RNA-specific adenosine deaminase. These clones were not found to be escape mutants, as their replicative ability was severely attenuated, and they did not replicate in the presence of vector.
authors.affiliation: VIRxSYS Corporation, Gaithersburg, MD 20877, USA.
authors.forename: Xiaobin
authors.initials: X
authors.lastname: Lu
authors.forename: Qiao
authors.initials: Q
authors.lastname: Yu
authors.forename: Gwendolyn K
authors.initials: GK
authors.lastname: Binder
authors.forename: Ziping
authors.initials: Z
authors.lastname: Chen
authors.forename: Tatiana
authors.initials: T
authors.lastname: Slepushkina
authors.forename: John
authors.initials: J
authors.lastname: Rossi
authors.forename: Boro
authors.initials: B
authors.lastname: Dropulic
mesh_terms: D001483:Base Sequence; D015496:CD4-Positive T-Lymphocytes / Q000821:virology; D002460:Cell Line; D002478:Cells, Cultured; D015686:Gene Products, env / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D005822:Genetic Vectors; D015497:HIV-1 / Q000235:genetics / Q000502:physiology*; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D008969:Molecular Sequence Data; D009154:Mutation; D016372:RNA, Antisense / Q000378:metabolism / Q000494:pharmacology*; D017384:Sequence Deletion; D014161:Transduction, Genetic; D014779:Virus Replication / Q000187:drug effects*
chemical_list: D015686:Gene Products, env; D016372:RNA, Antisense