title: T30177, an oligonucleotide stabilized by an intramolecular guanosine octet, is a potent inhibitor of laboratory strains and clinical isolates of human immunodeficiency virus type 1.
abstract: T30177, an oligonucleotide composed of only deoxyguanosine and thymidine, is 17 nucleotides in length and contains single phosphorothioate internucleoside linkages at its 5' and 3' ends for stability. This oligonucleotide does not share significant primary sequence homology with or possess any complementary (antisense) sequence motifs to the human immunodeficiency virus type 1 (HIV-1) genome. T30177 inhibited replication of multiple laboratory strains of HIV-1 in human T-cell lines, peripheral blood lymphocytes, and macrophages. T30177 was also found to be capable of inhibiting multiple clinical isolates of HIV-1 and preventing the cytopathic effect of HIV-1 in primary CD4+ T lymphocytes. In assays with human peripheral blood lymphocytes there was no observable toxicity associated with T30177 at the highest concentration tested (100 microM), while the median inhibitory concentration was determined to be in the range of 0.1 to 1.0 microM for the clinical isolates tested, resulting in a high therapeutic index for this drug. In temporal studies, the kinetics of addition of T30177 to infected cell cultures indicated that, like the known viral adsorption blocking agents dextran sulfate and Chicago sky blue, T30177 needed to be added to cells during or very soon after viral infection. However, analysis of nucleic acids extracted at 12 h postinfection from cells treated with T30177 at the time of virus infection established the presence of unintegrated viral cDNA, including circular proviral DNA, in the treated cells. In vitro analysis of viral enzymes revealed that T30177 was a potent inhibitor of HIV-1 integrase, reducing enzymatic activity by 50% at concentrations in the range of 0.050 to 0.09 microM. T30177 was also able to inhibit viral reverse transcriptase activity; however, the 50% inhibitory value obtained was in the range of 1 to 10 microM, depending on the template used in the enzymatic assay. No observable inhibition of viral protease was detected at the highest concentration of T30177 used (10 microM). In experiments in which T30177 was removed from infected cell cultures at 4 days post-HIV-1 infection, total suppression of virus production was observed for more than 27 days. PCR analysis of DNA extracted from cells treated in this fashion was unable to detect the presence of viral DNA 11 days after removal of the drug from the infected cell cultures. The ability of T30177 to inhibit both laboratory and clinical isolates of HIV-1 and the experimental data which suggest that T30177 represents a novel class of integrase inhibitors indicate that this compound is a viable candidate for evaluation as a therapeutic agent against HIV-1 in humans.
authors.affiliation: Triplex Pharmaceutical Corporation, The Woodlands, Texas 77380, USA.
authors.forename: J O
authors.initials: JO
authors.lastname: Ojwang
authors.forename: R W
authors.initials: RW
authors.lastname: Buckheit
authors.forename: Y
authors.initials: Y
authors.lastname: Pommier
authors.forename: A
authors.initials: A
authors.lastname: Mazumder
authors.forename: K
authors.initials: K
authors.lastname: De Vreese
authors.forename: J A
authors.initials: JA
authors.lastname: Esté
authors.forename: D
authors.initials: D
authors.lastname: Reymen
authors.forename: L A
authors.initials: LA
authors.lastname: Pallansch
authors.forename: C
authors.initials: C
authors.lastname: Lackman-Smith
authors.forename: T L
authors.initials: TL
authors.lastname: Wallace
mesh_terms: D000998:Antiviral Agents / Q000494:pharmacology*; D001483:Base Sequence; D002459:Cell Fusion / Q000187:drug effects; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D004254:DNA Nucleotidyltransferases / Q000037:antagonists & inhibitors; D004279:DNA, Viral / Q000096:biosynthesis; D005434:Flow Cytometry; D015658:HIV Infections / Q000821:virology*; D015497:HIV-1 / Q000187:drug effects* / Q000201:enzymology / Q000502:physiology; D006801:Humans; D019426:Integrases; D008214:Lymphocytes / Q000187:drug effects / Q000821:virology; D008264:Macrophages / Q000821:virology; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000494:pharmacology*; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology; D016176:T-Lymphocyte Subsets / Q000821:virology; D014779:Virus Replication / Q000187:drug effects
chemical_list: D000998:Antiviral Agents; D004279:DNA, Viral; D009841:Oligonucleotides; D018894:Reverse Transcriptase Inhibitors; C098172:T 30177; D004254:DNA Nucleotidyltransferases; D019426:Integrases