Nuclease-resistant nucleic acid ligands to vascular permeability factor/vascular endothelial growth factor.
| Autor: | Green LS; NeXstar Pharmaceuticals, Boulder, Colorado 80301, USA., Jellinek D, Bell C, Beebe LA, Feistner BD, Gill SC, Jucker FM, Janjić N |
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| Jazyk: | angličtina |
| Zdroj: | Chemistry & biology [Chem Biol] 1995 Oct; Vol. 2 (10), pp. 683-95. |
| DOI: | 10.1016/1074-5521(95)90032-2 |
| Abstrakt: | Background: Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) is a potent inducer of new blood vessel growth (angiogenesis) that contributes to the pathology of many angiogenesis-associated disease states such as psoriasis, rheumatoid arthritis and cancer. Few molecular entities capable of binding to VPF/VEGF with high affinity and specificity have been described to date. Results: Nuclease-resistant 2'-amino-2'-deoxypyrimidine nucleotide RNA (2'-aminopyrimidine RNA) ligands that bind to VPF/VEGF with high affinity have been identified by iterative rounds of affinity-selection/amplification from two independent random libraries. The sequence information that confers high affinity binding to VPF/VEGF is contained in a contiguous stretch of 24 nucleotides, 5'-CCCUGAUGGUAGACGCCGGGGUG-3' (2'-aminopyrimidine nucleotides are designated with italic letters). Of the 14 ribopurines in this minimal ligand, 10 can be substituted with the corresponding 2'-O-methylpurine nucleotides without a reduction in binding affinity to VPF/VEGF. In fact, the 2'-O-methyl substitution at permissive positions leads to a approximately 17-fold improvement in the binding affinity to VPF/VEGF. The higher affinity results from the reduction in the dissociation rate constant of the 2'-O-methyl-substituted RNA ligand from the protein compared to the unsubstituted ligand. The 2'-O-methyl-substituted minimal ligand, which folds into a bulged hairpin motif, is also more thermally stable than the unsubstituted ligand. Nuclease resistance of the ligand is further improved by the 2'-O-methyl substitutions and the addition of short phosphorothioate caps to the 3'- and 5'-ends. Conclusions: We have used the SELEX (systematic evolution of ligands by exponential enrichment) process in conjunction with post-SELEX modifications to define a highly nuclease-resistant oligonucleotide that binds to VPF/VEGF with high affinity and specificity. |
| Databáze: | MEDLINE |
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