title: Inhibition of PAI-1 antiproteolytic activity against tPA by RNA aptamers.
abstract: Plasminogen activator inhibitor-1 (PAI-1; SERPINE1) inhibits the plasminogen activators: tissue-type plasminogen activator (tPA) and urokinase-type plasminogen activator (uPA). Elevated levels of PAI-1 have been correlated with an increased risk for cardiovascular disease. Pharmacologically suppressing PAI-1 might prevent, or successfully treat PAI-1 related vascular diseases. This can potentially be accomplished by using small RNA molecules (aptamers). This study's goal is to develop RNA aptamers to a region of PAI-1 that will prevent the ability of PAI-1 to interact with the plasminogen activators. The aptamers were generated through a systematic evolution of ligands via exponential enrichment approach that ensures the creation of RNA molecules that bind to our target protein, PAI-1. In vitro assays were used to determine the effect of these aptamers on PAI-1's inhibitory activity. Three aptamers that bind to PAI-1 with affinities in the nanomolar range were isolated. The aptamer clones R10-4 and R10-2 inhibited PAI-1's antiproteolytic activity against tPA and disrupted PAI-1's ability to form a stable covalent complex with tPA. Increasing aptamer concentrations correlated positively with an increase in cleaved PAI-1. To the best of our knowledge, this is the first report of RNA molecules that inhibit the antiproteolytic activity of PAI-1.
authors.affiliation: 1 Department of Pediatric Hematology, Johns Hopkins University School of Medicine , Baltimore, Maryland.
authors.forename: Jared
authors.initials: J
authors.lastname: Damare
authors.forename: Stephanie
authors.initials: S
authors.lastname: Brandal
authors.forename: Yolanda M
authors.initials: YM
authors.lastname: Fortenberry
mesh_terms: D052157:Aptamers, Nucleotide / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D017395:Plasminogen Activator Inhibitor 1 / Q000737:chemistry* / Q000378:metabolism; D011480:Protease Inhibitors / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D059748:Proteolysis; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D052156:SELEX Aptamer Technique; D010959:Tissue Plasminogen Activator / Q000737:chemistry* / Q000378:metabolism; D014568:Urokinase-Type Plasminogen Activator / Q000737:chemistry / Q000378:metabolism
chemical_list: D052157:Aptamers, Nucleotide; D017395:Plasminogen Activator Inhibitor 1; D011480:Protease Inhibitors; D011994:Recombinant Proteins; C051321:SERPINE1 protein, human; D010959:Tissue Plasminogen Activator; D014568:Urokinase-Type Plasminogen Activator