Antisense oligodeoxynucleotide: inhibitor of splicing of mRNA of human immunodeficiency virus.
Antisense oligodeoxynucleotide (ODN), which are directed against the splice acceptor site of exon II of the regulatory gene tat of the human immunodeficiency virus type 1 (HIV-1), have been described. These 20-mer ODN's displayed moderate anti-HIV activity in vitro. Using the same antisense ODN (termed ODN-2), which was additionally modified and protected both at the 3'- and the 5'-terminus by two phosphorothioate internucleotide linkages, a strong anti-HIV activity (EC50: 2.7 micrograms/ml) could be measured in the HIV-1/CEM- and HIV-1/HeLa-T4+ cell system. The analogous ODNs which were protected only at one end were either inactive (up to 10 micrograms/ml) or displayed a low antiviral activity. Time kinetic studies revealed that the antisense ODN-2 reduced the release of HIV-1 already after an incubation time of 1 h. By applying S1 nuclease protection procedures, it could be established that the antisense ODN-2 inhibited splicing of high molecular weight transcript to the 2-kb tat mRNA in HIV-1-infected CEM cells. Transfection experiments with pU3R-III chloramphenicol acetyltransferase expression vector in HeLa-T4+ cells revealed that the antisense ODN-2 blocked the Tat protein-mediated transactivation process. In co-transfection experiments using pSV2tat72 or scrape loading studies with purified Tat, the transactivation was restored. These data indicate that the selected antisense ODN-2 displays its anti-HIV effect by blocking the splicing process leading to the functional 2-kb tat mRNA.
Daum
T
T
Institut für Physiologische Chemie, Johannes Gutenburg-Universität, Mainz, BRD.
Engels
J W
JW
Mag
M
M
Muth
J
J
Lücking
S
S
Schröder
H C
HC
Matthes
E
E
Müller
W E
WE
D000998:Antiviral Agents / Q000494:pharmacology*; D001483:Base Sequence; D002478:Cells, Cultured; D004279:DNA, Viral; D015756:Genes, tat; D015497:HIV-1 / Q000187:drug effects* / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D012326:RNA Splicing / Q000187:drug effects*; D012333:RNA, Messenger / Q000378:metabolism*; D015533:Transcriptional Activation / Q000187:drug effects
D000998:Antiviral Agents; D004279:DNA, Viral; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger