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US 12,662,503 B2Granted
Reagents and methods for screening MPS I, II, IIIA, IIIB, IVA, VI, AND VII
AssigneeUniversity of Washington through its Center for Commercialization
InventorsMichael H. Gelb, Arun Babu Kumar, Frances Hocutt, Zdenek Spacil, Mariana Natali Barcenas Rodriguez, Frantisek Turecek, C. Ronald Scott
Life of this patent
Estimating the term…
Abstract
Reagents, methods, and kits for assaying enzymes associated with lysosomal storage diseases MPS-I, MPS-II, MPS-IIIA, MPS-IIIB, MPS-IVA, MPS-VI, and MPS VII.
Claims33
A method for assaying enzymatic activities of one or more lysosomal enzymes, comprising:
- (a) contacting a sample with a first solution to provide a solution comprising one or more lysosomal enzymes;
- (b) contacting the one or more lysosomal enzymes in solution with an enzyme substrate for each lysosomal enzyme to be analyzed and incubating the substrates with the enzymes for a time sufficient to provide a solution comprising a first enzyme product for each lysosomal enzyme present in the sample;
- (c) subjecting the first enzyme products to a glycohydrolase to provide a second enzyme product for each first enzyme product susceptible to further enzymatic action by the glycohydrolase;
- and
- (d) determining the quantities of one or more of the first enzyme products and/or one or more of the second enzyme products
- wherein the substrate has a carbohydrate moiety and an aglycone moiety and has a formula:
- or a salt thereof,
- wherein
- L2 is a linker comprising 1-20 carbon atoms in which one or more carbon atoms may be replaced with a heteroatom selected from N, O, and S, and/or one or more of carbon atoms may be substituted with a C1-C6 alkyl group or halogen;
- L3 is a linker comprising 1-20 carbon atoms in which one or more carbon atoms may be replaced with a heteroatom selected from N, O, or S, and/or one or more of carbon atoms may be substituted with a C1-C6 alkyl group or halogen;
- L4 is optional and when present is a linker comprising 1-20 carbon atoms in which one or more carbon atoms may be replaced with a heteroatom selected from N, O, or S), and/or one or more of carbon atoms may be substituted with a C1-C6 alkyl group or halogen,
- R1 is a C1-C10 alkyl group;
- R2 at each occurrence is independently selected from a C1-C10 alkyl group, a C1-C10 alkoxy group, halogen, nitro, —C(═O)NHR, or —C(═O)OR, where R is C1-C8 alkyl group;
- R3 is a C1-C10 alkyl group or a substituted or unsubstituted C6-C10 aryl group;
- n is 0, 1, 2, 3, or 4;
- and
- S is
14 more elements
A method for assaying enzymatic activities of one or more lysosomal enzymes, comprising:
- (a) contacting a sample with a first solution to provide a solution comprising one or more lysosomal enzymes;
- (b) contacting the one or more lysosomal enzymes in solution with an enzyme substrate for each lysosomal enzyme to be analyzed and incubating the substrates with the enzymes for a time sufficient to provide a solution comprising a first enzyme product for each lysosomal enzyme present in the sample;
- (c) subjecting the first enzyme products to reaction with an exogenous glycohydrolase to provide a second enzyme product for each first enzyme product susceptible to further enzymatic action by the glycohydrolase;
- and
- (d) determining the quantities of one or more of the second enzyme products.
2 more elements
Dependencies are inferred from the claim text.