Compositions and methods for enhancing coagulation factor VIII function

• US 9,670,267 B2
• Filed: 08/25/2014
• Issued: 06/06/2017
• Est. Priority Date: 10/02/2009
• Status: Active Grant

• Current Assignee: Children's Hospital of Philadelphia (Children's Hospital of Philadelphia Foundation)
• Original Assignee: Children's Hospital of Philadelphia (Children's Hospital of Philadelphia Foundation)
• Inventors: Camire, Rodney, Arruda, Valder, Iacobelli, Nicholas
• Primary Examiner(s): TSAY, MARSHA M
• Application Number:
  US14/468,108
• Publication Number:
  US 20150056271A1
• Time in Patent Office: 1,016 Days
• Field of Search: None
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61P 7/04 Antihaemorrhagics; Procoagulants; Haemostatic agents; Antifibrinolytic agents
• CPC Class Codes: C07K 14/755 Factors VIII , e.g. factor VIII C (AHF), factor VIII Ag (VWF)

Abstract

Factor VIII variants and methods of use thereof are disclosed.

Claims

1. An isolated recombinant nucleic acid encoding a FVIII variant having a modified PACE/furin cleavage site and a B domain deletion (BDD) for modulating hemostasis, said human PACE/furin cleavage site consisting of amino acids RHQR, said FVIII variant being a human variant selected from the group consisting essentially of:

i) a FVIII variant with said BDD and wherein R is substituted with an H in the PACE/furin cleavage site;

ii a FVIII variant with said BDD and wherein one amino acid at said PACE/furin cleavage site is deleted;

iii) a FVIII variant with said BDD and wherein R at said cleavage site is substituted with an amino acid selected from the group consisting of a serine, lysine, methionine, cysteine, proline and tyrosine; and

iv) a FVIII variant wherein one or more amino acids at said PACE/furin cleavage site and a B domain are deleted,

each of i), ii), iii), and iv) exhibiting increased specific activity and stability relative to human FVIII-BDD lacking said substitution and deletions.

2. An expression vector comprising the nucleic acid of claim 1.

3. The vector of claim 2, selected from the group consisting of an adenoviral vector, an adeno-associated virus (AAV) vector, a retroviral vector, a plasmid, and a lentiviral vector.

4. A method for treatment of a hemostasis related disorder in a patient in need thereof comprising administration of a therapeutically effective amount of the vector of claim 3 in a biologically acceptable carrier, wherein said vector is an adeno associated virus (AAV) vector.

5. The method of claim 4, wherein said FVIII variant is a pro-coagulant and said disorder is selected from the group consisting of hemophilia A, von Willebrand diseases and bleeding associated with trauma, injury, thrombosis, thrombocytopenia, stroke, coagulopathy, disseminated intravascular coagulation (DIC) and over-anticoagulation treatment disorders.

6. The method of claim 4 or claim 5, wherein said FVIII variant is encapsulated in a liposome or mixed with phospholipids or micelles.

7. A host cell expressing the FVIII variant encoded by the nucleic acid of claim 1.

8. The nucleic acid encoding the FVIII variant as claimed in claim 1, wherein R at said cleavage site is substituted with an amino acid selected from the group consisting of a serine, lysine, methionine, cysteine, proline and tyrosine.

9. The nucleic acid encoding the FVIII variant as claimed in claim 1, wherein at least one amino acid at said PACE/furin cleavage site is deleted.

10. The nucleic acid encoding the FVIII variant of claim 1, wherein two of the amino acids at the PACE/Furin cleavage site are deleted.

11. The nucleic acid encoding the FVIII variant of claim 1, wherein three of the amino acids at the PACE/Furin cleavage site are deleted.

12. The nucleic acid encoding the FVIII variant of claim 1, wherein all four of the amino acids at the PACE/Furin cleavage site are deleted.

13. An adeno associated virus (AAV) vector comprising the nucleic acid of claim 1.

14. The adeno associated virus (AAV) vector of claim 13, further comprising ITR (inverted terminal repeat) elements.