Evacuated blood collection tubes containing protease inhibitors for the assessment of contact system activation

Patent metadata

Field	Value
Publication number	US-11826758-B2
Application number	US-202217821681-A
Country	US
Kind code	B2
Filing date	Aug 23, 2022
Priority date	Aug 13, 2015
Publication date	Nov 28, 2023
Grant date	Nov 28, 2023

Abstract

Disclosed herein are evacuated blood collection tubes comprising protease inhibitor cocktails in liquid form and uses thereof for assessing features associated with the contact system in a subject, including the endogenous level of contact system activation, the endogenous level of a drug that targets a component of contact system during treatment, and/or the immunogenicity of such a drug.

First claim

What is claimed is:

1. A method for assessing the endogenous level of contact system activation in a subject, the method comprising:
(i) collecting blood from a subject to an evacuated blood collection tube;
(ii) processing the blood to produce a plasma sample; and
(iii) measuring the level of contact system activation in the plasma sample;
wherein the evacuated blood collection tube comprises a liquid formulation that comprises a mixture of protease inhibitors, wherein the mixture of protease inhibitors comprises EPI-KAL2 provided by SEQ ID NO: 1 and leupeptin.

2. The method of claim 1 , wherein in step (i), the evacuated blood collection tube is not the first tube filled with blood from the subject.

3. The method of claim 1 , wherein step (ii) is performed within one hour after step (i).

4. The method of claim 1 , wherein the subject is a human subject.

5. The method of claim 4 , wherein the human subject is a human patient having a disease associated with the contact system.

6. The method of claim 5 , wherein the disease is hereditary angioedema (HAE) or idiopathic angioedema.

7. The method of claim 6 , wherein the human patient has HAE with normal C1 esterase inhibitor (C1-INH).

8. The method of claim 4 , wherein the human subject is treated with a drug targeting a component of the contact system.

9. The method of claim 8 , wherein the component of the contact system is plasma kallikrein; optionally wherein the drug inhibits active plasma kallikrein.

10. The method of claim 9 , wherein the drug is an antibody that binds active plasma kallikrein.

11. The method of claim 1 , wherein step (iii) is performed by measuring the level of one or more biomarkers indicative of contact system activation.

12. The method of claim 11 , wherein the one or more biomarkers are selected from the group consisting of prekallikrein, active plasma kallikrein (pKal), α2M-pKal complex, active factor XII, active factor XI, high molecular weight kininogen (HMWK), and a bradykinin metabolite; optionally wherein the one or more biomarkers comprise cleaved HMWK and/or intact HMWK.

13. A method for assessing the level of a drug targeting the contact system in a subject, the method comprising:
(i) collecting blood from a subject to an evacuated blood collection tube; wherein the subject is administered with a drug that targets a component of the contact system;
(ii) processing the blood to produce a plasma sample; and
(iii) measuring the level of the drug in the plasma sample
wherein the evacuated blood collection tube comprises a liquid formulation that comprises a mixture of protease inhibitors, wherein the mixture of protease inhibitors comprises EPI-KAL2 provided by SEQ ID NO: 1 and leupeptin.

14. A method for assessing immunogenicity of a drug targeting the contact system, the method comprising:
(i) collecting blood from a subject to an evacuated blood collection tube; wherein the subject is administered with a drug that targets a component of the contact system;
(ii) processing the blood to produce a plasma sample; and
(iii) measuring the level of antibodies that bind to the drug in the plasma sample;
wherein the evacuated blood collection tube comprises a liquid formulation that comprises a mixture of protease inhibitors, wherein the mixture of protease inhibitors comprises EPI-KAL2 provided by SEQ ID NO: 1 and leupeptin.

15. The method of claim 14 , wherein the method further comprises, prior to step (iii), isolating antibodies that bind the drug from the plasma sample; optionally wherein the antibodies are isolated by a solid phase extraction and acid dissociation (SPEAD) assay.

16. The method of claim 1 , wherein the liquid formulation comprises 5-15 μM EPI-KAL2 and 200-300 μM leupeptin.

17. The method of claim 1 , wherein the EPI-KAL2 is biotinylated.

18. The method of claim 1 , wherein the liquid formulation further comprises polybrene and EDTA.

19. The method of claim 1 , wherein the liquid formulation has a pH of 4-6.

20. The method of claim 1 , wherein the mixture of protease inhibitors is substantially free of protease inhibitors that are unstable in an aqueous solution.

Assignees

• Takeda Pharmaceuticals Co

Inventors

• Sexton Daniel J
• Faucette Ryan

Classifications

• B01L2200/16

  Reagents, handling or storing thereof · CPC title

• B01L3/5082 Primary

  Test tubes per se · CPC title

• A61B5/150755

  Blood sample preparation for further analysis, e.g. by separating blood components or by mixing · CPC title

• A61B5/15003

  for venous or arterial blood · CPC title

• A61B5/154 Primary

  {Devices} using pre-evacuated means · CPC title