Mutated structural protein of a parvovirus
The present invention is related to a structural protein of a parvovirus with an amino acid insertion at the insertion site I-453, a library comprising the protein, a multimeric structure comprising the protein, a nucleic acid encoding the protein, a vector, virus, or cell comprising the nucleic acid, a process for the preparation of the protein, a medicament comprising the protein, nucleic acid, or multimeric structure as well as methods and uses involving the protein, nucleic acid, or multimeric structure.
1. A structural protein of a parvovirus which comprises a first amino acid insertion of one or more amino acids into I-453, and a second amino acid insertion of one or more amino acids into I-587, wherein the first amino acid insertion is directly C-terminal of amino acid G 453 and the second amino acid insertion is directly C-terminal of amino acid N587 in the sequence of AAV-2 or the corresponding amino acids of any other parvovirus, wherein the first amino acid insertion and the second amino acid insertion are each a tolerogen-derived epitope.
2. The structural protein of claim 1 , wherein the insertion is located on the surface of the capsid formed by the structural proteins.
3. The structural protein of claim 1 , wherein the structural protein with the insertion is capable of particle formation.
4. The structural protein of claim 1 , wherein the parvovirus is an adeno-associated virus.
5. The structural protein of claim 1 , wherein the parvovirus is selected from the group consisting of AAV-1, AAV-2, AAV-3b, AAV-4, AAV-5, AAV-6, AAV-7, AAV-8, AAV-9, AAV-10, AAV-11, AAV-12, b-AAV, feline panleukopenia virus (FPV), canine parvovirus (CPV), B19, goose parvovirus (GPV) and minute virus of mice (MVM).
6. The structural protein of claim 1 , wherein the amino acid insertion has a length of about 4 to about 30 amino acids.
7. The structural protein of claim 1 , wherein the tolerogen-derived epitope is derived from a protein from the group consisting of CETP, CD20, acetylcholine receptors, IL13R, EGFR, IgE, Melan A, HMW MAA, CA125, Her2/NEU, CCR5, L1 cell adhesion molecule, VEGF, EGFR, CD20, TNF-α, IL-6, IL9, IL-13, IL-17, and β-amyloid.
8. The structural protein of claim 7 , wherein the tolerogen-derived epitope is selected from the group consisting of VNLTWSRASG (SEQ ID NO: 50), EFCINHRGYWVCGD (SEQ ID NO:55), EDGQVMDVDLS (SEQ ID NO: 85), EKQRNGTLT (SEQ ID NO: 86), TYQCRVTHPHLPRALMR (SEQ ID NO: 87), RHSTTQPRKTKGSG (SEQ ID NO: 88), DSNPRGVSAYLSR (SEQ ID NO: 89), TITCLWDLAPSK (SEQ ID NO: 90), KTKGSGFFVF (SEQ ID NO: 91), THPHLPRALMRS (SEQ ID NO: 92), GETYQCRVTHPHLPRALMRSTTK (SEQ ID NO: 93), LPRALMRS (SEQ ID NO: 94), INHRGYWV (SEQ ID NO: 95), CDAGSVRTNAPD (SEQ ID NO: 60), AKAVSNLTESRSESLQS (SEQ ID NO: 96), SLTGDEFKKVLET (SEQ ID NO: 97), REAVAYRFEED (SEQ ID NO: 98), INPEIITLDG (SEQ ID NO: 99), DISVTGAPVITATYL (SEQ ID NO: 100), DISVTGAPVITA (SEQ ID NO: 101), PKTVSNLTESSSESVQS (SEQ ID NO: 102), SLMGDEFKAVLET (SEQ ID NO: 103), QHSVAYTFEED (SEQ ID NO: 104), INPEIITRDG (SEQ ID NO: 105), DISLTGDPVITASYL (SEQ ID NO: 106), DISLTGDPVITA (SEQ ID NO: 107), DQSIDFEIDSA (SEQ ID NO: 108), KNVSEDLPLPTFSPTLLGDS (SEQ ID NO: 109), KNVSEDLPLPT (SEQ ID NO: 110), CDSGRVRTDAPD (SEQ ID NO: 111), FPEHLLVDFLQSLS (SEQ ID NO: 112), DAEFRHDSG (SEQ ID NO: 65), HYAAAQWDFGNTMCQL (SEQ ID NO: 113), YAAQWDFGNTMCQ (SEQ ID NO: 114), RSQKEGLHYT (SEQ ID NO: 115), SSRTPSDKPVAHWANPQAE (SEQ ID NO: 116), SRTPSDKPVAHWANP (SEQ ID NO: 117), SSRTPSDKP (SEQ ID NO: 118), NADGNVDYHMNSVP (SEQ ID NO: 119), DGNVDYHMNSV (SEQ ID NO: 120), RSFKEFLQSSLRALRQ (SEQ ID NO: 121); FKEFLQSSLRA (SEQ ID NO: 122), and QMWAPQWGPD (SEQ ID NO: 123).
9. The structural protein of claim 1 , wherein the amino acid insertion brings about an alteration in a chromatographic property of the structural protein.
10. The structural protein of claim 5 , wherein the parvovirus structural protein comprises one or more further mutation(s) at a site different to I-453 and I-587 independently selected from a point mutation, an internal deletion, an N-terminal deletion, an insertion and a substitution.
11. The structural protein of claim 10 , wherein the further mutation is an insertion of a B-cell epitope or a cytotoxic T-cell epitope.
12. The structural protein of claim 10 , wherein the further mutation reduces the transducing activity of the particle for a given target cell by at least 50%.
13. The structural protein of claim 10 , wherein the further mutation reduces the ability to induce a B-cell response against a parvovirus-specific epitope and/or mimotope.
14. An isolated nucleic acid molecule encoding a structural protein of claim 1 .
15. A vector comprising the nucleic acid molecule of claim 14 .
16. A composition comprising at least one parvovirus structural protein according to claim 1 or a nucleic acid molecule according to claim 14 .