Structural motifs determining the high anti-thrombin activity of NU172, an aptamer currently in Phase II clinical trials

Human α-thrombin is a trypsin-like serine protease that has the unique ability to convert soluble fibrinogen in insoluble fibrin clot. In addition to the active site, this enzyme owns two electropositive regions, (exosite I and II), located at opposite sides of its globular shape and playing an essential role in the recognition of many cofactors and substrates.1 A special class of thrombin synthetic ligands is represented by aptamers, which are short single stranded DNA or RNA oligonucleotides that bind their targets with very high affinity and specificity by adopting defined and stable three dimensional structural motifs.2 In particular, anti-thrombin aptamers adopt a G-quadruplex structure.3-5 Recently, the researchers interest has moved on a new class of oligonucleotides in which the addition of a duplex-forming sequences to a G-quadruplex module results in an improvement of the antithrombotic action. One of these oligonucleotides is NU172 that possesses an high anticoagulant activity with an IC50 value of 5–10 μg/mL in plasma6 and is currently the only thrombin binding aptamer evaluated in Phase II clinical trials (ClinicalTrials.gov identifier NCT00808964) for anticoagulation in heart disease treatments by ARCA Biopharma, Inc.7 The crystallographic characterization of thrombin-NU172 complex in the presence of either potassium or sodium ions provides together with conformational solution studies on NU172 variants and anticoagulant assays have provided a reasonable interpretation of the peculiar antithrombotic properties of NU172. Details on our results will be discussed at the Meeting. References: 1. Di Cera, E. J Thromb Haemost. 2007, 5 Suppl 1, 196 – 202. 2. Woodruff, R.S.; Sullenger, B.A. Arterioscler Thromb Vasc Biol. 2015, 35, 2083 – 2091. 3. Russo Krauss, I; Pica, A.; Merlino, A.; Mazzarella, L.; Sica, F. Acta Crystallogr D Biol Crystallogr. 2013, 69, 2403 – 2411. 4. Russo Krauss, I.; Spiridonova, V.; Pica, A.; Napolitano, V.; Sica, F. Nucleic Acids Res. 2016, 44, 983 – 991. 5. Russo Krauss, I.; Napolitano, V.; Petraccone, L.; Troisi, R.; Spiridonava, V.; Mattia, C.A.; Sica, F. Int J Biol Macromol. 2018, 107, 1697 – 1705. 6. Keefe, A.D.; Pai, S.; Ellington, A. Nat Rev Drug Discov. 2010, 9, 537 – 550. 7. Zhou, J.; Rossi, J. Nat Rev Drug Discov. 2017, 16, 181 – 202.