Research Article abstract
Nature Biotechnology 15, 68 - 73 (1997)
doi:10.1038/nbt0197-68
Potent 2'-amino-, and 2'-fluoro-2'- deoxyribonucleotide RNA inhibitors of keratinocyte growth factor
Nikos C. Pagratis*, 1, Carol Bell1, Ying-Fon Chang1, Susan Jennings1, Tim Fitzwater1, Derek Jellinek2 & Chinh Dang1
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 thermostabilities (Tm approximately 78°C in low salt, and specificities).
- 1NeXstar Pharmaceuticals, Inc., 2860 Wilderness PL, Boulder, CO 80301.
- 2Present Address: Children's Medical Research Institute, Locked Bag 23, Wentworthville, 2145 NSW, Australia.
- *e-mail: pagratis@nexstar.com
