title: Potent 2'-amino-, and 2'-fluoro-2'-deoxyribonucleotide RNA inhibitors of keratinocyte growth factor.
abstract: Reiterative in vitro selection-amplification from random oligonucleotide libraries allows the identification of molecules with specific functions such as binding to specific proteins. The therapeutic usefulness of such molecules depends on their high affinity and nuclease resistance. Libraries of RNA molecules containing 2'amino-(2'NH2)- or 2'fluoro-(2'F)-2'-deoxypyrimidines could yield ligands with similar nuclease resistance but not necessarily with similar affinities. This is because the intramolecular helices containing 2'NH2 have lower melting temperatures (Tm) compared with helices containing 2'F, giving them thermodynamically less stable structures and possibly weaker affinities. We tested these ideas by isolating high-affinity ligands to human keratinocyte growth factor from libraries containing modified RNA molecules with either 2'NH2 or 2'F pyrimidines. We demonstrated that 2'F RNA ligands have affinities (Kd approximately 0.3-3 pM) and bioactivities (Ki approximately 34 pM) superior to 2'NH2 ligands (Kd approximately 400 pM and Ki approximately 10 nM). In addition, 2'F ligands have extreme thermo-stabilities (Tm approximately 78 degrees C in low salt, and specificities).
authors.affiliation: NeXstar Pharmaceuticals, Inc., Boulder, CO 80301, USA.
authors.forename: N C
authors.initials: NC
authors.lastname: Pagratis
authors.forename: C
authors.initials: C
authors.lastname: Bell
authors.forename: Y F
authors.initials: YF
authors.lastname: Chang
authors.forename: S
authors.initials: S
authors.lastname: Jennings
authors.forename: T
authors.initials: T
authors.lastname: Fitzwater
authors.forename: D
authors.initials: D
authors.lastname: Jellinek
authors.forename: C
authors.initials: C
authors.lastname: Dang
mesh_terms: D016475:3T3 Cells / Q000187:drug effects / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D003854:Deoxyribonucleotides / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology*; D004355:Drug Stability; D051526:Fibroblast Growth Factor 10; D051523:Fibroblast Growth Factor 7; D005346:Fibroblast Growth Factors; D006133:Growth Substances / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D012313:RNA / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology*; D012260:Ribonucleases / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D003854:Deoxyribonucleotides; C496453:FGF7 protein, human; C496454:Fgf7 protein, mouse; D051526:Fibroblast Growth Factor 10; D006133:Growth Substances; D051523:Fibroblast Growth Factor 7; D005346:Fibroblast Growth Factors; D012313:RNA; D012260:Ribonucleases