Sequence-specific inhibition of human immunodeficiency virus (HIV) reverse transcription by antisense oligonucleotides: comparative study in cell-free assays and in HIV-infected cells.
We have investigated two regions of the viral RNA of human immunodeficiency virus type 1 (HIV-1) as potential targets for antisense oligonucleotides. An oligodeoxynucleotide targeted to the U5 region of the viral genome was shown to block the elongation of cDNA synthesized by HIV-1 reverse transcriptase in vitro. This arrest of reverse transcription was independent of the presence of RNase H activity associated with the reverse transcriptase enzyme. A second oligodeoxynucleotide targeted to a site adjacent to the primer binding site inhibited reverse transcription in an RNase H-dependent manner. These two oligonucleotides were covalently linked to a poly(L-lysine) carrier and tested for their ability to inhibit HIV-1 infection in cell cultures. Both oligonucleotides inhibited virus production in a sequence- and dose-dependent manner. PCR analysis showed that they inhibited proviral DNA synthesis in infected cells. In contrast, an antisense oligonucleotide targeted to the tat sequence did not inhibit proviral DNA synthesis but inhibited viral production at a later step of virus development. These experiments show that antisense oligonucleotides targeted to two regions of HIV-1 viral RNA can inhibit the first step of viral infection--i.e., reverse transcription--and prevent the synthesis of proviral DNA in cell cultures.
Bordier
B
B
Institut de Biochimie Cellulaire et Neurochimie, Centre National de la Recherche Scientifique, Bordeaux, France.
Perala-Heape
M
M
Degols
G
G
Lebleu
B
B
Litvak
S
S
Sarih-Cottin
L
L
Hélène
C
C
D001483:Base Sequence; D002460:Cell Line; D002474:Cell-Free System; D017931:DNA Primers; D004261:DNA Replication; D054303:HIV Reverse Transcriptase; D015497:HIV-1 / Q000201:enzymology*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D011107:Polylysine; D016133:Polymerase Chain Reaction; D011533:Proviruses / Q000201:enzymology; D012367:RNA, Viral / Q000737:chemistry / Q000378:metabolism; D012194:RNA-Directed DNA Polymerase / Q000187:drug effects*; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology*; D016914:Ribonuclease H / Q000378:metabolism
D017931:DNA Primers; D016376:Oligonucleotides, Antisense; D012367:RNA, Viral; D018894:Reverse Transcriptase Inhibitors; D011107:Polylysine; D054303:HIV Reverse Transcriptase; D012194:RNA-Directed DNA Polymerase; D016914:Ribonuclease H