Rna aptamers and methods for identifying the same

Abstract

RNA aptamers and methods for identifying the same are disclosed. The RNA aptamers selectively bind coagulation factors, E2F family members, Ang1 or Ang2, and therapeutic and other uses for the RNA aptamers are also disclosed.

1 . A method of identifying a ligand which binds to a target, the method comprising:
 (a) preparing a candidate oligonucleotide mixture;   (b) contacting the candidate oligonucleotide mixture with the target in a low stringency buffer in the presence of a partitioning matrix to produce a binding mixture which comprises a first plurality of ligands which bind the target and a second plurality of ligands which do not bind the target, wherein the first plurality of ligands has increased affinity to the target relative to the second plurality of ligands and/or relative to the partitioning matrix;   (c) removing the second plurality of ligands;   (d) collecting the ligands that are bound to the target to produce a first collected ligand mixture.   
     
     
         2 . The method of  claim 1 , wherein steps (a)-(d) are repeated one or more times. 
     
     
         3 . The method of  claim 1 , further comprising:
 (e) repeating steps (a)-(d) one or more times;   (f) contacting the first plurality of ligands with the target in a higher stringency buffer, wherein the first plurality of ligands has increased affinity to the target relative to the second plurality of ligands;   (g) removing the second plurality of ligands; and   (h) collecting the first plurality of ligands.   
     
     
         4 . The method of  claim 3 , wherein steps (f), (g) and (h) are repeated one or more times. 
     
     
         5 . The method of  claim 1 , further comprising amplifying the first collected ligand mixture or the second collected ligand mixture, or both, to yield a ligand enriched mixture, whereby at least one desired ligand to the target is identified. 
     
     
         6 . The method of  claim 1 , wherein the candidate oligonucleotide mixture comprises single strand nucleic acids. 
     
     
         7 . The method of  claim 6 , wherein the single stranded nucleic acids are ribonucleic acids. 
     
     
         8 . The method of  claim 6 , wherein the single stranded nucleic acids are deoxyribonucleic acids. 
     
     
         9 . The method of  claim 6 , wherein said candidate oligonucleotide mixture comprises 2′-modified ribonucleic acids. 
     
     
         10 . The method of  claim 9 , wherein said 2′-modified ribonucleic acids comprise 2′-fluoro (2′-F) modified nucleic acids. 
     
     
         11 . The method of  claim 1 , wherein the lower stringency buffer has a pH of about 7-8. 
     
     
         12 . The method of  claim 1 , wherein the lower stringency buffer has a NaCl concentration of about 10 mM. 
     
     
         13 . The method of  claim 3 , wherein the higher stringency buffer comprises a physiological buffer. 
     
     
         14 . The method of  claim 13 , wherein the physiological buffer has a pH of about 7.4 and a NaCl concentration of about 150 mM NaCl. 
     
     
         15 . A product identified by the process of  claim 1 .