Aminoglycoside-arginine conjugates that bind TAR RNA: synthesis, characterization, and antiviral activity.
Regulation of HIV gene expression is crucially dependent on binding of the trans-activator protein, Tat, to the trans-activation response RNA element, TAR, found at the 5' end of all HIV-1 transcripts. Tat-TAR interaction is mediated by a short arginine-rich domain of the protein. Disruption of this interaction could, in theory, create a state of complete viral latency. A new class of small-molecule peptidomimetic TAR RNA binders, conjugates of aminoglycosides and arginine, was recently designed [Litovchick, A., Evdokimov, A. G., and Lapidot, A. (1999) FEBS Lett. 445, 73-79]. Two of these compounds, the tri-arginine derivative of gentamicin C (R3G) and the tetra-arginine derivative of kanamycin A (R4K), bind efficiently and specifically to TAR RNA. These compounds display negligible toxicity while being transported and accumulated in cell nuclei. Here we present a detailed synthesis and chemical characterization of the aminoglycoside-arginine conjugates R3G and R4K as well as GB4K, the tetra-gamma-guanidinobutyric derivative of kanamycin A. Their binding sites on TAR RNA were assigned by RNase A, uranyl nitrate, and lead acetate footprinting. The conjugates interact with TAR RNA in the widened major groove, formed by the UCU bulge and the neighboring base pairs of the upper stem portion of TAR, the binding site of Tat protein, and Tat-derived peptides (e.g., R52). Our results suggest an additional binding site of R4K and R3G compounds, in the lower stem-bulge region of TAR. The antiviral activity of the conjugates in cultured equine dermal fibroblasts infected with equine infectious anemia virus, used as a model system of HIV-infected cells, is also presented.
Litovchick
A
A
Departments of Organic Chemistry and Structural Biology, The Weizmann Institute of Science, 76100 Rehovot, Israel.
Evdokimov
A G
AG
Lapidot
A
A
D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D001120:Arginine / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D001665:Binding Sites; D002478:Cells, Cultured; D018983:DNA Footprinting; D005839:Gentamicins / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D016325:HIV Long Terminal Repeat; D006736:Horses; D006801:Humans; D004860:Infectious Anemia Virus, Equine / Q000187:drug effects*; D007424:Intracellular Fluid / Q000378:metabolism / Q000821:virology; D007612:Kanamycin / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D007854:Lead / Q000378:metabolism; D019906:Nuclear Magnetic Resonance, Biomolecular; D009942:Organometallic Compounds / Q000378:metabolism; D012367:RNA, Viral / Q000378:metabolism; D051381:Rats; D012259:Ribonuclease, Pancreatic; D014502:Uranyl Nitrate / Q000378:metabolism
D000998:Antiviral Agents; D005839:Gentamicins; D009942:Organometallic Compounds; D012367:RNA, Viral; D014502:Uranyl Nitrate; C100093:gentamicin C; D007854:Lead; D007612:Kanamycin; D001120:Arginine; D012259:Ribonuclease, Pancreatic; C008261:lead acetate