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Details for Patent: 7,176,295

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Details for Patent: 7,176,295

Title:Systematic evolution of ligands by exponential enrichment: blended SELEX
Abstract: A method is described for generating blended nucleic acid ligands containing non-nucleic acid functional units. Specifically, a SELEX identified RNA ligand to the integrin gpIIbIIIa is conjugated to the peptide Gly-Arg-Gly-Asp-Thr-Pro (SEQ ID NO:1). This blended RNA ligand inhibits the biological activity of gpIIbIIIa with high specificity. Also described is a single-stranded DNA ligand to elastase coupled to N-methoxysuccinyl-Ala-Ala-Pro-Val-chloromethyl ketone (SEQ ID NO:2). This elastase blended nucleic acid ligand inhibits the biological activity of elastase.
Inventor(s): Biesecker; Gregory (Boulder, CO), Jayasena; Sumeda (Boulder, CO), Gold; Larry (Boulder, CO), Smith; Drew (Boulder, CO), Kirschenheuter; Gary P. (Arvada, CO)
Assignee: Gilead Sciences, Inc. (Foster City, CA)
Filing Date:Oct 02, 2002
Application Number:10/263,456
Claims:1. A blended nucleic acid ligand of a target compound prepared by a method comprising the steps of: a) identifying a nucleic acid ligand of a target compound from a candidate mixture comprised of nucleic acids each having a region of randomized sequence by a method comprising: i) contacting the candidate mixture with the target, wherein nucleic acids having an increased affinity to the target relative to the candidate mixture may be partitioned from the remainder of the candidate mixture; ii) partitioning the increased affinity nucleic acids from the remainder of the candidate mixture, and iii) amplifying the increased affinity nucleic acids to yield a ligand-enriched mixture of nucleic acids, whereby nucleic acid ligands of the target may be identified; and b) attaching at least one functional unit to said nucleic acid ligand to yield a blended nucleic acid ligand of the target compound, wherein the functional unit is selected from the group consisting of proteins, peptides, photoreactive groups, chemically-reactive groups, active site directed compounds, lipids, biotin, and fluorescent compounds.

2. The blended nucleic acid ligand of claim 1 wherein said at least one functional unit is attached to an oligonucleotide capable of hybridizing to said nucleic acid ligand, and wherein step b) is accomplished by hybridizing said oligonucleotide to said nucleic acid ligand.

3. A diagnostic reagent comprising a blended nucleic acid ligand of a target compound and a reporter molecule, wherein said diagnostic reagent is prepared by a method comprising the steps of: a) identifying a nucleic acid ligand of a target compound from a candidate mixture comprised of nucleic acids each having a region of randomized sequence by a method comprising: i) contacting the candidate mixture with the target, wherein nucleic acids having an increased affinity to the target relative to the candidate mixture; ii) partitioning the increased affinity nucleic acids from the remainder of the candidate mixture, and iii) amplifying the increased affinity nucleic acids to yield a ligand-enriched mixture of nucleic acids, whereby a nucleic acid ligand of the target compound may be identified; b) attaching at least one functional unit to said nucleic acid ligand to yield a blended nucleic acid ligand of the target compound, wherein the functional unit is selected from the group consisting of proteins, peptides, photoreactive groups, chemically-reactive groups, active site directed compounds, lipids, biotin, and fluorescent compounds; and c) attaching at least one reporter molecule to said blended nucleic acid ligand, whereby a diagnostic reagent is prepared.

4. The diagnostic reagent of claim 3 wherein said at least one functional unit is attached to an oligonucleotide capable of hybridizing to said nucleic acid ligand, and wherein step b) is accomplished by hybridizing said oligonucleotide to said nucleic acid ligand.
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