Peptide quantitation assay for differentiating full-length high molecular weight kininogen (HMWK) and cleaved HMWK

Patent metadata

Field	Value
Publication number	US-12135329-B2
Application number	US-202318308813-A
Country	US
Kind code	B2
Filing date	Apr 28, 2023
Priority date	Dec 15, 2015
Publication date	Nov 5, 2024
Grant date	Nov 5, 2024

Abstract

Methods for differentiating full-length high molecular weight kininogen (HMWK) and cleaved HMWK in a sample are provided herein. Such methods may comprise treating a biological sample with a protease to generate a plurality of digested peptides, and measuring one or more signature peptides, which are indicative of cleaved HMWK and/or full-length HMWK.

First claim

What is claimed is:

1. A method for identifying a subject having or at risk for hereditary angioedema (HAE), the method comprising:
(i) providing a biological sample of a subject,
(ii) contacting the sample with a protease to generate a plurality of digested peptides;
(iii) measuring the level of a signature peptide in the plurality of digested peptides, wherein the signature peptide is indicative of the 46 kDa light chain and is KHNLGHGH (SEQ ID NO: 1), or the 56 kDa light chain and is SSRIGE (SEQ ID NO: 5); and
(iv) identifying the subject as having HAE or being at risk for HAE attack based on the level of cleaved HMWK, wherein an elevated level of cleaved HMWK in the biological sample as compared with a predetermined reference value indicates that the subject has HAE or is at risk for HAE attack.

2. The method of claim 1 , wherein the protease is chymotrypsin.

3. The method of claim 1 , wherein the protease is endoproteinase Glu-C or cathepsin.

4. The method of claim 1 , further comprising (v) administering to the subject identified as having HAE an effective amount of a therapeutic agent.

5. The method of claim 1 , wherein the measuring is performed by liquid chromatography-mass spectrometry (LC-MS).

6. The method of claim 1 , wherein the biological sample is a blood sample or a plasma sample.

7. The method of claim 4 , wherein the biological sample is a plasma sample collected in an evacuated blood collection tube, which comprises a liquid formulation that comprises a mixture of protease inhibitors.

8. The method of claim 1 , wherein step (ii) is performed in the presence of a reducing agent.

9. The method of claim 1 , wherein step (ii) is performed in the absence of a protease inhibitor, the absence of an anticoagulant, or both the absence of a protease inhibitor and an anticoagulant.

10. The method of claim 1 , wherein the subject is a human patient having a symptom of HAE.

11. The method of claim 1 , wherein the subject is resistant to an anti-histamine therapy, a corticosteroid therapy, or both.

12. A method for identifying a subject having or at risk for hereditary angioedema (HAE), the method comprising:
(i) providing a biological sample of a subject,
(ii) contacting the sample with a protease to generate a plurality of digested peptides;
(iii) measuring the level of a first signature peptide and a second signature peptide in the plurality of digested peptides, wherein the first signature peptide is indicative of cleaved high molecular weight kininogen (HMWK) and the second signature peptide is indicative of intact HMWK;
(iv) determining the ratio between the first signature peptide and the second signature peptide,
(v) identifying the subject as having HAE or being at risk for HAE attack based on the ratio between the first signature peptide and the second signature peptide, wherein an elevated ratio of cleaved HMWK to intact HMWK in the biological sample as compared with a predetermined reference value indicates that the subject has HAE or is at risk for HAE attack wherein the first signature peptide is indicative of the 56 kDa light chain and is SSRIGE (SEQ ID NO: 5) and the second signature peptide indicative of full-length HMWK is SYYFDLTDGLS (SEQ ID NO: 10).

13. The method of claim 12 , wherein the protease is endoproteinase Glu-C or cathepsin.

14. The method of claim 12 , wherein step (ii) is performed in the absence of a protease inhibitor, an anticoagulant, or both the absence of a protease inhibitor and an anticoagulant.

15. The method of claim 12 , further comprising (vi) administering to the subject identified as having HAE an effective amount of a therapeutic agent.

16. The method of claim 12 , wherein the measuring is performed by liquid chromatography-mass spectrometry (LC-MS).

17. The method of claim 12 , wherein the biological sample is a blood sample or a plasma sample, and is optionally collected in an evacuated blood collection tube, which comprises a liquid formulation that comprises a mixture of protease inhibitors.

Assignees

• Takeda Pharmaceuticals Co

Inventors

• Wu Jiang
• Zhang Guodong
• Sexton Daniel J
• Faucette Ryan
• Mustafa Gul M
• Szewc Mark

Classifications

• G01N2560/00

  Chemical aspects of mass spectrometric analysis of biological material · CPC title

• G01N2800/224

  Haemostasis or coagulation · CPC title

• G01N2030/8831

  involving peptides or proteins · CPC title

• G01N33/6848 Primary

  Methods of protein analysis involving mass spectrometry · CPC title

• G01N30/7233

  interfaced to liquid or supercritical fluid chromatograph (interfaces in general for introducing or extracting samples to be analysed with specially adapted mass spectrometer, see H01J49/04) · CPC title