title: Antisense oligodeoxynucleotide: inhibitor of splicing of mRNA of human immunodeficiency virus.
abstract: 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.
authors.affiliation: Institut für Physiologische Chemie, Johannes Gutenburg-Universität, Mainz, BRD.
authors.forename: T
authors.initials: T
authors.lastname: Daum
authors.forename: J W
authors.initials: JW
authors.lastname: Engels
authors.forename: M
authors.initials: M
authors.lastname: Mag
authors.forename: J
authors.initials: J
authors.lastname: Muth
authors.forename: S
authors.initials: S
authors.lastname: Lücking
authors.forename: H C
authors.initials: HC
authors.lastname: Schröder
authors.forename: E
authors.initials: E
authors.lastname: Matthes
authors.forename: W E
authors.initials: WE
authors.lastname: Müller
mesh_terms: 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
chemical_list: D000998:Antiviral Agents; D004279:DNA, Viral; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger