IP Library › Granted Patent US 12,295,972
Granted Patent B2
US 12,295,972 · App. 18/810,381 · Granted May 13, 2025

Methods using freeze-dried platelet derivative compositions for restoring hemostasis in a subject

Inventors: Keith Andrew Moskowitz (Westfield, IN); Braden Carl Ishler (Montgomery Village, MD); William Matthew Dickerson (Washington, DC); Narendra Nath Tandon (Gaithersburg, MD); Amber Nicole Lee (Montgomery Village, MD); Stephen Edward Amos (Buckeystown, MD); Rafael Jorda (Merignac, FR); Michael Alexander Mathews (Arlington, VA)
Assignee: Cellphire, Inc.
A61K35/19A61K9/19A61P7/04
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Quick Facts
Patent No.
US 12,295,972
App. No.
18/810,381
Granted
May 13, 2025
Kind
B2
Abstract

Provided herein are methods and compositions for treating a coagulopathy in a subject. Such methods can include administering to the subject in need thereof, for example because they have been administered an anticoagulant agent, an effective amount of a composition including platelets, or in illustrative embodiments platelet derivatives, and in further illustrative embodiments freeze-dried platelet derivatives (FDPDs). Various properties of exemplary embodiments of such methods and platelet derivatives used therein, as well as numerous additional aspects and embodiments are provided herein.

Claims (68)

1. A method for restoring hemostasis in a subject, wherein the method comprises:

rehydrating a platelet derivative composition in the form of a powder, said composition comprising an effective amount of the platelet derivatives and an incubating agent comprising one or more saccharides, one or more salts, and a buffer to form a rehydrated platelet derivative composition, and

administering the rehydrated platelet derivative composition comprising the effective amount of the platelet derivatives to the subject,

wherein the subject has been treated or is being treated with at least one antiplatelet agent,

wherein at least 70% of the platelet derivatives in the rehydrated platelet derivative composition are CD41 positive platelet derivatives when measured using flow cytometry, and less than 5% of the CD41 positive platelet derivatives are microparticles,

wherein at least 70% of the platelet derivatives in the rehydrated platelet derivative composition are CD42 positive platelet derivatives, when measured using flow cytometry,

wherein at least 70% of the platelet derivatives in the rehydrated platelet derivative composition are CD62 positive platelet derivatives, when measured using flow cytometry,

wherein the platelet derivatives are capable of generating thrombin in an in vitro thrombin formation assay,

wherein the platelet derivatives have in vitro occlusion activity, and

wherein the platelet derivatives show an inability to increase expression of a platelet activation marker in the presence of an agonist as compared to the expression of the platelet activation marker in the absence of the agonist, and wherein the agonist is thrombin receptor activator peptide 6 (TRAP-6) and the platelet activation marker can be detected by binding of Annexin V to the platelet derivatives.

2. The method of claim 1 , wherein the subject is being treated with an antiplatelet agent.

3. The method of claim 2 , wherein the one or more saccharides comprise trehalose.

4. The method of claim 3 , wherein the platelet derivatives have the in vitro occlusion activity, such that when forced through a collagen-coated microchannel at a concentration of at least 255×10 3 particles/μL they are capable of attaining a pressure of 80 kPa in less than 14 minutes in platelet-reduced citrated whole blood in an in vitro total thrombus-formation analysis system (T-TAS) assay.

5. The method of claim 3 , wherein less than 3.5% of the CD41 positive platelet derivatives are microparticles.

6. The method of claim 3 , wherein the one or more saccharides further comprise polysucrose, and wherein the one or more salts are selected from the group consisting of phosphate salts, sodium salts, potassium salts, calcium salts, magnesium salts, and a combination of two or more thereof.

7. The method of claim 6 , wherein the antiplatelet agent is selected from the group consisting of aspirin, cangrelor, ticagrelor, clopidogrel, prasugrel, eptifibatide, tirofiban, abciximab, terutroban, picotamide, elinogrel, ticlopidine, ibuprofen, vorapaxar, atopaxar, cilostazol, prostaglandin E1, epoprostenol, dipyridamole, treprostinil sodium, and sarpogrelate.

8. The method of claim 6 , wherein the administering comprises administering intravenously the rehydrated platelet derivative composition comprising the effective amount of the platelet derivatives.

9. The method of claim 6 , wherein the antiplatelet agent is selected from the group consisting of cangrelor, ticagrelor, clopidogrel, prasugrel, eptifibatide, tirofiban, abciximab, terutroban, picotamide, elinogrel, ticlopidine, ibuprofen, vorapaxar, atopaxar, cilostazol, prostaglandin E1, epoprostenol, dipyridamole, treprostinil sodium, and sarpogrelate.

10. The method of claim 9 , wherein the subject is being treated with aspirin in addition to the antiplatelet agent.

11. The method of claim 6 , wherein the platelet derivatives are capable of generating thrombin in the in vitro thrombin formation assay, such that the platelet derivatives at a concentration of at least 4.8×10 3 particles/μl generate a thrombin peak height of at least 25 nM in the presence of a reagent containing tissue factor and phospholipids.

12. The method of claim 6 , wherein less than 3.5% of the CD41 positive platelet derivatives are microparticles, and wherein the rehydrated platelet derivative composition comprises trehalose in an amount from 50 mM to 200 mM, and polysucrose in amount from 3% to 10%.

13. The method of claim 1 , wherein the method is for preparing a subject for surgery.

14. A method for restoring hemostasis in a subject,

wherein the method comprises:

rehydrating a platelet derivative composition in the form of a powder, said composition comprising an effective amount of the platelet derivatives and an incubating agent comprising one or more saccharides, one or more salts, and a buffer to form a rehydrated platelet derivative composition, and

administering the rehydrated platelet derivative composition comprising the effective amount of the platelet derivatives to the subject,

wherein the rehydrated platelet derivative composition comprises a population of platelet derivatives comprising CD 41 positive platelet derivatives, wherein less than 5% of the CD 41 positive platelet derivatives are microparticles,

wherein at least 70% of the platelet derivatives in the rehydrated platelet derivative composition are CD42 positive platelet derivatives, when measured using flow cytometry,

wherein at least 70% of the platelet derivatives in the rehydrated platelet derivative composition are CD 41 positive platelet derivatives, when measured using flow cytometry,

wherein at least 70% of the platelet derivatives in the rehydrated platelet derivative composition are CD62 positive platelet derivatives, when measured using flow cytometry,

wherein the platelet derivatives are capable of generating thrombin in an in vitro thrombin formation assay,

wherein the platelet derivatives have in vitro occlusion activity, and

wherein the platelet derivatives show an inability to increase expression of a platelet activation marker in the presence of an agonist as compared to the expression of the platelet activation marker in the absence of the agonist, and wherein the agonist is thrombin receptor activator peptide 6 (TRAP-6) and the platelet activation marker can be detected by binding of Annexin V to the platelet derivatives.

15. The method of claim 14 , wherein the subject has a clotting-related disorder selected from the group consisting of Von Willebrand Disease, hemophilia, thrombasthenia, and thrombocytopenia.

16. The method of claim 15 , wherein the one or more saccharides comprise trehalose and polysucrose.

17. The method of claim 16 , wherein the trehalose in the rehydrated platelet derivative composition is in an amount from 50 mM to 200 mM, and wherein the polysucrose in the rehydrated platelet derivative composition in amount from 3% to 10%.

18. The method of claim 16 , wherein the one or more salts are selected from the group consisting of phosphate salts, sodium salts, potassium salts, calcium salts, magnesium salts, and a combination of two or more thereof.

19. The method of claim 18 , wherein the buffer comprises 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES).

20. The method of claim 16 , wherein less than 3.5% of the CD 41 positive platelet derivatives are microparticles.

21. The method of claim 16 , wherein the platelet derivatives have the in vitro occlusion activity, such that when forced through a collagen-coated microchannel at a concentration of at least 255×10 3 particles/μL they are capable of attaining a pressure of 80 kPa in less than 14 minutes in platelet-reduced citrated whole blood in an in vitro total thrombus-formation analysis system (T-TAS) assay.

22. The method of claim 16 , wherein the administering comprises administering intravenously the rehydrated platelet derivative composition comprising the effective amount of the platelet derivatives.

23. A method for administering platelet derivatives to a subject,

wherein the method comprises:

rehydrating a platelet derivative composition in the form of a powder, said composition comprising an effective amount of the platelet derivatives and an incubating agent comprising one or more saccharides comprising 10-60% trehalose by weight, one or more salts, and a buffer to form a rehydrated platelet derivative composition, and

administering intravenously the rehydrated platelet derivative composition comprising the effective amount of the platelet derivatives to the subject,

wherein the rehydrated platelet derivative composition comprises a population of platelet derivatives comprising CD 41 positive platelet derivatives, wherein less than 5% of the CD 41 positive platelet derivatives are microparticles,

wherein at least 70% of the platelet derivatives in the rehydrated platelet derivative composition are CD42 positive platelet derivatives, when measured using flow cytometry,

wherein at least 70% of the platelet derivatives in the rehydrated platelet derivative composition are CD41 positive platelet derivatives, when measured using flow cytometry,

wherein at least 70% of the platelet derivatives in the rehydrated platelet derivative composition are CD62 positive platelet derivatives, when measured using flow cytometry,

wherein the platelet derivatives in the rehydrated platelet derivative composition have thrombospondin (TSP) on their surface at a level that is greater than a level of TSP on the surface of resting platelets,

wherein the platelet derivatives in the rehydrated platelet derivative composition have von Willebrand factor (vWF) on their surface at a level that is greater than a level of vWF on the surface of resting platelets,

wherein the platelet derivatives are capable of generating thrombin in an in vitro thrombin formation assay,

wherein the platelet derivatives have in vitro occlusion activity,

wherein the platelet derivatives show an inability to increase expression of a platelet activation marker in the presence of an agonist as compared to the expression of the platelet activation marker in the absence of the agonist, and wherein the agonist is thrombin receptor activator peptide 6 (TRAP-6) and the platelet activation marker can be detected by binding of Annexin V to the platelet derivatives, and

wherein the administering the rehydrated platelet derivative composition restores hemostasis in the subject.

24. The method of claim 23 , wherein the one or more saccharides further comprises polysucrose.

25. The method of claim 24 , wherein the composition in the form of the powder comprises the trehalose in a range of 20-35% by weight, and the polysucrose in a range of 45-60% by weight.

26. The method of claim 25 , wherein the platelet derivative composition in the form of the powder is in a container,

wherein the container is at least one container from a plurality of containers,

wherein the plurality of containers comprise the platelet derivatives from at least 2 different lots in separate containers, and

wherein the amount of microparticles in the powder of any two containers chosen from different lots, differs by less than 10%.

27. The method of claim 23 , wherein the platelet derivative composition in the form of the powder is prepared by a process, comprising:

performing tangential flow filtration (TFF) of a platelet composition comprising platelets in a preparation agent comprising a buffering agent, trehalose in an amount in the range of 10 mM to 500 mM, and polysucrose in an amount in the range of 3% to 7%, thereby preparing a TFF-treated composition comprising at least 1000×10 3 platelets/μl in an aqueous medium having less than or equal to 7.5% plasma protein and having less than 5.0% microparticles by scattering intensity;

freeze drying the TFF-treated composition comprising platelets in the aqueous medium to form a freeze-dried platelet derivative composition comprising platelet derivatives; and

heating the freeze-dried platelet derivative composition at a temperature in the range of 60° C. to 90° C. for at least 1 hour to not more than 36 hours to thermally treat the platelet derivatives in the freeze-dried platelet derivative composition to form the platelet derivative composition in the form of the powder.

28. The method of claim 17 , wherein the platelet derivatives are capable of generating thrombin in the in vitro thrombin formation assay, such that the platelet derivatives at a concentration of at least 4.8×10 3 particles/μl generate a thrombin peak height of at least 25 nM in the presence of a reagent containing tissue factor and phospholipids.

29. The method of claim 17 , wherein less than 3.5% of the CD 41 positive platelet derivatives are microparticles.

30. The method of claim 14 , wherein less than 3.5% of the CD 41 positive platelet derivatives are microparticles.

Assignments (1)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Aug 20, 2024
From: MOSKOWITZ, KEITH ANDREW; ISHLER, BRADEN CARL; DICKERSON, WILLIAM MATTHEW; TANDON, NARENDRA NATH; LEE, AMBER NICOLE; AMOS, STEPHEN EDWARD; JORDA, RAFAEL; MATHEWS, MICHAEL ALEXANDER
To: CELLPHIRE, INC.
Reel/Frame 068346/0967 →
Continuity (6)
Continuation 17674770 · Feb 17, 2022
Provisional Application 63264227 · Nov 17, 2021
Provisional Application 63276420 · Nov 5, 2021
Provisional Application 63275937 · Nov 4, 2021
Provisional Application 63150338 · Feb 17, 2021
Related Publication 20240408141A1 · Dec 12, 2024
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