Patent application EP19173440A (EP), filed 2019-05-09
Publication number: EP3736286A1
Title: SINGLE CHAIN FACTOR VIII MOLECULE
Abstract: [0001]    The present invention relates to a recombinant Factor VIII protein comprising, in a single chain, a heavy chain portion comprising an A1 and an A2 domain and a light chain portion comprising an A3, C1 and C2 domain of Factor VIII, wherein the B-domain is partially deleted in two deletions, the first leading to the presence of a defined processing sequence cleavable by thrombin, and the second leading to absence of the furin cleavage recognition site at position R1664-R1667. An internal fragment of the B-domain is maintained. Nucleic acids encoding said protein, host cells and methods of preparing the protein are also provided, as well as a pharmaceutical composition comprising the protein, nucleic acid or host cell, which may be used for treatment of hemophilia A.
IPC codes: A61K  38/    37            A I
C07K  14/   755            A I
C12N  15/    00            A I
Claims: number: 1
text: A recombinant Factor VIII protein comprising, in a single chain, a heavy chain portion comprising an A1 and an A2 domain and a light chain portion comprising an A3, C1 and C2 domain of Factor VIII, wherein,
number: 2
text: A recombinant Factor VIII protein comprising, in a single chain, a heavy chain portion comprising an A1 and an A2 domain and a light chain portion comprising an A3, C1 and C2 domain of Factor VIII, wherein,
number: 3
text: The recombinant Factor VIII protein of any one of the preceding claims, wherein the processing sequence is SEQ ID NO: 4 or a sequence having at most one amino acid substitution in said sequence, wherein, optionally, the F, the S C-terminal to the F, the Q or the N are substituted.
number: 4
text: The recombinant Factor VIII protein of any one of the preceding claims, wherein the processing sequence is selected from the group consisting of SEQ ID NO: 4, 6, 7 or 8.
number: 5
text: The recombinant Factor VIII protein of any one of the preceding claims, wherein the processing sequence is SEQ ID NO: 4.
number: 6
text: The recombinant Factor VIII protein of any one of the preceding claims, wherein the amino acids corresponding to amino acids K1663 to L1674 of wild type Factor VIII are deleted, leading to a second deletion.
number: 7
text: The recombinant Factor VIII protein of any one of the preceding claims, not comprising a furin cleavage recognition site, and not comprising a sequence having more than 75% sequence identity to the furin cleavage recognition site RHQR between the processing sequence and the merging sequence,
preferably, not comprising a sequence having more than 40% sequence identity to SEQ ID NO: 15.
number: 8
text: The recombinant Factor VIII protein of any one of the preceding claims, comprising a processing sequence and, C-terminal to said processing sequence, a merging sequence, wherein said sequences are selected from the group consisting of
number: 9
text: The recombinant Factor VIII protein of any one of the preceding claims, further comprising a third thrombin cleavage site between the A1 and A2 domain.
number: 10
text: The recombinant Factor VIII protein of any one of the preceding claims that is a fusion protein with at least one heterologous fusion partner selected from the group consisting of an Fc region, an albumin-binding sequence, albumin, PAS polypeptides, HAP polypeptides, the C-terminal peptide of the beta subunit of chorionic gonadotropin, albumin-binding small molecules, polyethylenglycol, hydroxyethyl starch,
wherein, optionally, said heterologous fusion partner is inserted directly C-terminal to said processing sequence.
number: 11
text: A nucleic acid encoding a recombinant Factor VIII protein of any one of the preceding claims, wherein said polynucleotide optionally is an expression vector suitable for expression of said recombinant Factor VIII protein in a mammalian cell selected from the group comprising a human cell.
number: 12
text: A host cell comprising a nucleic acid of claim 11, wherein preferably the host cell is a mammalian cell comprising an expression vector suitable for expression of said recombinant Factor VIII protein in said cell.
number: 13
text: A method for preparing a Factor VIII protein, comprising culturing the host cell of claim 12 under conditions suitable for expression of the Factor VIII protein and isolating the Factor VIII protein, wherein the method optionally comprises formulating the Factor VIII protein as a pharmaceutical composition.
number: 14
text: A pharmaceutical composition comprising the recombinant Factor VIII protein of any of claims 1-10, the nucleic acid of claim 11, or the host cell of claim 12.
number: 15
text: The pharmaceutical composition of claim 14 for use in treatment of hemophilia A,
wherein, optionally, the treatment is immune tolerance induction.
Office action: search_report, dated 2020-11-11
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ref_id: - DATABASE Geneseq [online] 12 July 2007 (2007-07-12), "Human B-domain deleted factor VIII protein, SEQ ID : 5.", XP002795192, retrieved from EBI accession no. GSP:AGA99685 Database accession no. AGA99685
* sequence . *
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ref_id: [ ]     - "Human B-domain deleted factor VIII protein, SEQ ID : 5.", Geneseq, (20070712), Database accession no. AGA99685, URL: EBI, XP002795192 [ ] * sequence . *
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Final disposition: pending (2020-11-11)