Aptamer

By similar logic, protein affinity Hbraries have been constmcted to identify protein—protein combining sites, as in antibody—antigen interaction (19) and recombinant Hbraries have been made which produce a repertoire of antibodies in E. coli (20). In another case, a potential DNA-based therapeutic strategy has been studied (21). DNAs from a partially randomized Hbrary were selected to bind thrombin in vitro. Oligonucleotides, termed aptamers that bound thrombin shared a conserved sequence 14—17 nucleotides long. 

Combinatorial Hbraries are limited by the number of sequences that can be synthesized. For example, a Hbrary consisting of one molecule each of a 60-nucleotide sequence randomized at each position, would have a mass of >10 g, weU beyond the capacity for synthesis and manipulation. Thus, even if nucleotide addition is random at all the steps during synthesis of the oligonucleotide only a minority of the sequences can be present in the output from a laboratory-scale chemical DNA synthesis reaction. In analyzing these random but incomplete Hbraries, the protocol is efficient enough to allow selection of aptamers of lowest dissociation constants (K ) from the mixture after a small number of repetitive selection and amplification cycles. Once a smaller population of oligonucleotides is amplified, the aptamer sequences can be used as the basis for constmcting a less complex Hbrary for further selection. 

VEGF inhibitors Humanized neutralizing antibodies, antisense oligonucleotides, siRNA, aptamers... 

Oligonucleic acid or peptide molecules that bind a specific target molecule. Nucleic acid aptamer species can be engineered through treated rounds of in vitro selection to bind to various molecular targets such as... 

Pegaptanib (Macugen) Aptamer VEGF-165 Inhibition of ligand binding AMD... 

Bunka DH, Stockley PG (2006) Aptamers come of age - at last, Nat Rev 4 588-596 Damm EM, Pelkmans L (2006) Systems biology of virus entry in mammalian cells. Cell Microbiol 8 1219-1227... 

Townsend L, Devivar R, Turk S, Nassiri M, Drach J (1995) Design, synthesis, and antiviral activity of certain 2,5,6-trihalo-l-(beta-d-ribofuranosyl)benzimidazoles. J Med Chem 38 4098 105 Turlure F, Devroe E, Silver PA, Engelman A (2004) Human cell proteins and human immunodeficiency virus DNA integration. Front Biosd 9 3187-3208 Umehara T, Fukuda K, Nishikawa F, Kohara M, Hasegawa T, NisUkawa S (2005) Rational design of dual-functional aptamers that inhibit the protease and helicase activities of HCV NS3. J Biochem 137 339-347... 

Kleimnan et al. 2008). In addition, synthetic siRNAs are also subject to degradation in vivo by nuclease activity. Besides side effects and instability, the efficient and specific delivery of the RNAi indncers to the target cell still requires optimization. Here we snmmarize the cnrrent statns of nncleic acid-based antiviral therapentics. The focns will be on antiviral strategies nsing antisense and RNAi technology. Additionally, antiviral ribozymes and aptamers will be discussed briefly, with a focus on recent studies. Gene therapy approaches and delivery systems are the subject of Chapter 11 of this book. 

Matsugami A, Kobayashi S, Ouhashi K, Uesugi S, Yamamoto R, Taira K, Nishikawa S, Kumar PK, Katahira M (2003) Structural basis of the highly efficient trapping of the HIV Tat protein by an RNA aptamer. Structure 11 533-545... 

Bridonneau. R, Bunch, S., Tengler, R., Hill, K., Carter, J., Pieken, W., Tinner-meier, D., Lehrman, R., and Drolet, D. W., Purification of a highly modified RNA-aptamer. Effect of complete denaturation during chromatography on product recovery and specific activity, /. Chromatogr. B, 726, 237, 1999. 

Bracht F., Schror K. Isolation and identification of aptamers from defibrotide that act as thrombin antagonists in vitro. Biochem Biophys Res Commun 1994 200,933-7. 

Van den Bossche, J. et al. (2010) Efficient receptor-independent intracellular translocation of aptamers mediated by conjugation to carbon nanotubes. Chemical Communications, 7379-7381. Mu, Q.X., Broughton, D.L. and Yan, B. 

Holeman LA, Robinson SL, Szostak JW, Wilson C (1998) Isolation and characterization of fluorophore-binding RNA aptamers. Fold Des 3 423 -31... 

Gouda, H. Kuntz, I.D. Case, D.A. Kollman, P.A., Free energy calculations for theophylline binding to an RNA aptamer Comparison of MM-PBSA and thermodynamic integration methods, Biopolymers 2003, 68,16-34. 

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