Patent application EP19165681A (EP), filed 2019-03-28
Publication number: EP3715358A1
Title: MUTATED ADENO-ASSOCIATED VIRUS CAPSID PROTEINS, AAV PARTICLE COMPRISING THE SAME AND LIVER DIRECTED AAV VECTOR GENE THERAPY
Abstract: [0001]    The present invention relates in a first aspect to a mutated adeno-associated virus (AAV) capsid protein or fragment thereof having an insert composed of an oligopeptide optionally having further amino acids representing linker sequences flanking both sites of said oligopeptide. These mutated AAV capsid proteins are more efficient in targeting hepatic tissue, hepatocytes, hepatic cells and cell lines or hepatocellular carcinoma (HCC) with partially higher specificity. Further, a mutated AAV particle comprising the mutated AAV capsid protein according to the present invention is provided. In addition, a nucleic acid encoding the mutated AAV capsid protein according to the present invention is identified together with a corresponding nucleic acid vector, in particular, a plasmid. In addition, a host cell containing the nucleic acid vector or the nucleic acid molecule according to the present invention. Further, a use of the AAV particle or the nucleic acid or the nucleic acid vector according to the present invention in the manufacture of an AAV particle, or in the manufacture of a medicament for gene therapy is described. Moreover, the described protein, particle molecules as well as nucleic acid vectors for use in targeting hepatocytes and/or HCC are described. In particular, said components for use in treating diseases involving hepatocytes or for treating HCC are disclosed, in particular, for use in gene therapy, e.g. for use in a transfer of a gene of interest into hepatocytes, heptatic tissue or HCC.
IPC codes: C07K  14/   005            A I
C07K  14/   015            A I
C12N  15/    10            A I
C12N  15/    35            A I
C12N  15/    86            A I
C12N   5/    10            A I
Claims: number: 1
text: A mutated adeno-associated virus (AAV) capsid protein or a fragment thereof wherein an insert is inserted after at least one the amino acids having the amino acid numbers 139, 161, 261, 381, 447, 453, 459, 534, 570, 573, 584, 585, 586, 587, 588, 589 of SEQ ID No. 2 corresponding to the capsid protein of AAV type 2 or the homolog capsid protein of the other serotypes of AAV, namely, AAV1, AAV3b, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10,AAV11, and AAV bovine, the insert comprises an oligopeptide of at least four amino acids and at most thirty amino acids, like consisting of five to twelve amino acids, in particular, seven amino acids, said insert may optionally has further amino acids representing a linker sequence at both sites of said oligopeptide, said linker sequence having independently from one another a size of 1 to 3 amino acids, if present; wherein the mutated AAV capsid protein or a fragment thereof or its homologs being more efficient in targeting hepatic tissue, hepatocytes, hepatic cells and cell lines or hepatocellular carcinoma (HCC) with higher specificity.
number: 2
text: The mutated AAV capsid protein, a fragment thereof or a homolog thereof according to claim 1 wherein the AAV is type 1, 2, 3b, 5, 6, 7, 8, 9, 10, 4, 11, and bovine.
number: 3
text: The mutated AAV capsid protein, a fragment thereof or a homolog thereof according to claim 1 wherein the insert are at the positions following at least one of the amino acids of 261, 453, 534, 570, 573, 587 and 588, more preferably 453 and 587.
number: 4
text: The mutated AAV capsid protein, a fragment thereof or a homolog thereof according to any one of the preceding claims wherein the mutated AAV capsid protein, the fragment thereof, or the homolog thereof comprise at least one further mutation selected from a point mutation, an internal or terminal deletion, a second insertion and a substitution.
number: 5
text: The mutated AAV capsid protein, a fragment thereof, or a homolog thereof according to any one of the preceding claims when the linker sequence is an amino acid sequence containing or consisting of the amino acids G, S and/or A, in particular, wherein the linker consists of at least one of amino acids A or S, in particular, having amino acids AAA N-terminally and AA C-terminally, or ASA N-terminally and AA C-terminally, flanking the oligopeptide of at least four to at most thirty amino acids, like at least five amino acids and at most twelve amino acids, like seven amino acids.
number: 6
text: The mutated AAV capsid protein, a fragment thereof, or a homolog thereof according to any one of the preceding claims wherein the oligopeptide having a size of seven amino acids is a sequence according to any one of SEQ ID No. 13 to 275.
number: 7
text: The mutated AAV capsid protein, a fragment thereof, or a homolog thereof according to any one of the preceding claims wherein the insert is any one of SEQ ID No. 276 to 538.
number: 8
text: A mutated adeno-associated virus (AAV) particle comprising the mutated AAV capsid protein, a fragment thereof or a homolog thereof according to any one of claims 1 to 7.
number: 9
text: A nucleic acid molecule encoding the mutated AAV capsid protein, a fragment thereof or a homolog thereof according to any one of claims 1 to 8.
number: 10
text: A nucleic acid vector, in particular, a plasmid comprising the nucleic acid molecule according to claim 9.
number: 11
text: A host cell containing the nucleic acid vector according to claim 10 or a nucleic acid molecule according to claim 9.
number: 12
text: A use of the AAV particle according to claim 8 or the nucleic acid molecule according to claim 9 or the nucleic acid vector according to claim 10 in the manufacture of an AAV particle, or in manufacture of a medicament for gene therapy.
number: 13
text: The mutated AAV capsid protein, a fragment thereof or a homolog according to any one of claims 1 to 7, the mutated AAV particle according to claim 8, the nucleic acid molecule according to claim 9, nucleic acid vector according to claim 10, or the nucleic acid vector according to claim 11 for use in transducing hepatocytes and/or HCC in vivo or ex vivo.
number: 14
text: The mutated AAV capsid protein, a fragment thereof or a homolog according to any one of claims 1 to 7, the mutated AAV particle according to claim 8, the nucleic acid molecule according to claim 9, or the nucleic acid vector according to claim 10 for use in treating diseases involving hepatocytes: bleeding disorders including Hemophilia A, Hemophilia B, Von Willebrand disease, metabolic disorders including Alpha-1-antitrypsin deficiency, familial hypercholesterolemia, ornithine transcarbamylase deficiency, crigler-najjar syndrome, acute intermittent porphyria, glycogen storage disease type 1a, liver cancer including hepatocellular carcinoma, hepatoblastoma, collangiocarcinoma, autoimmune disorders including multiple sclerosis, latent autoimmune diabetes, induction of tolerance towards allogenic transplants including liver, kidneys, heart, stem cells, surrogate liver-directed gene therapy including lipoproteinlipase deficiency, diabetes and ornithine transcarbamylase deficiency; or for treating HCC
number: 15
text: The mutated AAV capsid protein, a fragment thereof or a homolog according to any one of claims 1 to 7, the mutated AAV particle according to claim 8, the nucleic acid molecule according to claim 9, he nucleic acid vector according to claim 10, or the nucleic acid vector according to claim 11 for use in transfer of a gene of interest into hepatocytes or HCC, in particular, for use in gene therapy.
Office action: search_report, dated 2020-09-30
Rejection reasons: novelty_epc_54
Prior art cited: ref_id: EP2158211A2
ref_type: patent
severity: background
ref_id: EP1456383A2
ref_type: patent
severity: background
ref_id: EP2363487A2
ref_type: patent
severity: background
ref_id: US2002192823A1
ref_type: patent
severity: background
ref_id: WO2012149160A2
ref_type: patent
severity: background
ref_id: - FITZPATRICK ET AL.: "Molecular Therapy: Methods & Clinical Development", ELSEVIER LTD., pages: 119 - 129
ref_type: npl
severity: background
ref_id: - WAGHRAY, A. ET AL., WJH, vol. 7, no. 8, 2015, pages 1020 - 1029
ref_type: npl
severity: background
ref_id: - BUNING, H. ET AL., CURRENT OPINION IN PHARMACOLOGY, vol. 24, 2015, pages 94 - 104
ref_type: npl
severity: background
ref_id: - BIINING; SVRIVASTAVA, MOL THER METHODS CLIN DEV, vol. 12, 2019, pages 248 - 265
ref_type: npl
severity: background
ref_id: - MULLER, O. J. ET AL., NATURE BIOTECHNOLOGY, vol. 21, no. 9, 2003, pages 1040 - 1046
ref_type: npl
severity: background
ref_id: - PERABO, L. ET AL., MOLECULAR THERAPY, vol. 8, no. 1, 2003, pages 151 - 157
ref_type: npl
severity: background
ref_id: - SCHMITZ ET AL., J. HEPATOL., 2004, pages 787 - 797
ref_type: npl
severity: background
ref_id: - HAUPENTHAL J. ET AL., NEOPLASIA, vol. 14, no. 5, 2012, pages 410 - 419
ref_type: npl
severity: background
ref_id: - LESZEK LISOWSKI ET AL: "Selection and evaluation of clinically relevant AAV variants in a xenograft liver model", NATURE, vol. 506, no. 7488, 25 December 2013 (2013-12-25), London, pages 382 - 386, XP055573596, ISSN: 0028-0836, DOI: 10.1038/nature12875
ref_type: npl
severity: background
ref_id: - C. RAUPP ET AL: "The Threefold Protrusions of Adeno-Associated Virus Type 8 Are Involved in Cell Surface Targeting as Well as Postattachment Processing", JOURNAL OF VIROLOGY., vol. 86, no. 17, 20 June 2012 (2012-06-20), US, pages 9396 - 9408, XP055269892, ISSN: 0022-538X, DOI: 10.1128/JVI.00209-12
ref_type: npl
severity: background
ref_id: WO2018189244A1
ref_type: patent
severity: background
ref_id: WO2017143100A1
ref_type: patent
severity: background
ref_id: WO2013029030A1
ref_type: patent
severity: background
ref_id: EP1456383A2
ref_type: patent
severity: background
ref_id: - NADJA MEUMANN: "Identification and characterisation of novel liver-directed AAV capsid variants exhibiting highly improved liver transduction features", HUMAN GENE THERAPY, vol. 28, no. 12, 1 December 2017 (2017-12-01), pages A104, XP055593867
ref_type: npl
severity: background
ref_id: - NADJA MEUMANN: "Novel liver-directed adeno-associated viral capsid variants with striking transduction efficiency in murine and human hepatocytes", MOLECULAR THERAPY, vol. 26, no. 5 Suppl 1, 1 May 2018 (2018-05-01), pages 180 - 181, XP055593874
ref_type: npl
severity: background
ref_id: - CHRISTINA RAUPP: "Analysis of Infection Relevant Protein Domains of the Adeno-Associated Virus Serotype 8 in Comparison to Serotype 2 - INAUGURAL-DISSERTATION zur Erlangung der Doktorw[upsilon]rde der Naturwissenschaftlich-Mathematischen Gesamtfakultät der Ruprecht-Karls-Universität Heidelberg", 18 June 2010 (2010-06-18), XP055593764, Retrieved from the Internet <URL:http://archiv.ub.uni-heidelberg.de/volltextserver/10925/1/complete_CRAUPPdiss.pdf> [retrieved on 20190604]
ref_type: npl
severity: background
ref_id: - STEFAN MICHELFELDER ET AL: "Successful Expansion but Not Complete Restriction of Tropism of Adeno-Associated Virus by In Vivo Biopanning of Random Virus Display Peptide Libraries", PLOS ONE, vol. 4, no. 4, 9 April 2009 (2009-04-09), pages e5122, XP055147203, DOI: 10.1371/journal.pone.0005122
ref_type: npl
severity: background
ref_id: - K VARADI ET AL: "Novel random peptide libraries displayed on AAV serotype 9 for selection of endothelial cell-directed gene transfer vectors", GENE THERAPY, vol. 19, no. 8, 1 August 2012 (2012-08-01), pages 800 - 809, XP055129104, ISSN: 0969-7128, DOI: 10.1038/gt.2011.143
ref_type: npl
severity: background
ref_id: EP1456383
ref_type: patent
severity: background
ref_id: WO2013029030
ref_type: patent
severity: background
ref_id: WO2017143100
ref_type: patent
severity: background
ref_id: WO2018189244
ref_type: patent
severity: novelty_destroying
ref_id: - NADJA MEUMANN, "Identification and characterisation of novel liver-directed AAV capsid variants exhibiting highly improved liver transduction features", HUMAN GENE THERAPY, (20171201), vol. 28, no. 12, page A104, XP055593867 [X] 1-3,8-13,15 * abstract * [I] 1-5,8-15
ref_type: npl
severity: novelty_destroying
ref_id: - NADJA MEUMANN, "Novel liver-directed adeno-associated viral capsid variants with striking transduction efficiency in murine and human hepatocytes", MOLECULAR THERAPY, (20180501), vol. 26, no. 5 Suppl 1, pages 180 - 181, XP055593874 [X] 1-3,8-13,15 * abstract * [I] 1-5,8-15
ref_type: npl
severity: novelty_destroying
ref_id: - Christina Raupp, "Analysis of Infection Relevant Protein Domains of the Adeno-Associated Virus Serotype 8 in Comparison to Serotype 2 - INAUGURAL-DISSERTATION zur Erlangung der Doktorw[upsilon]rde der Naturwissenschaftlich-Mathematischen Gesamtfakultät der Ruprecht-Karls-Universität Heidelberg", (20100618), URL: http://archiv.ub.uni-heidelberg.de/volltextserver/10925/1/complete_CRAUPPdiss.pdf, (20190604), XP055593764 [A] 1-15 * see the entire section 3.4; pages  36-37; figures 3-19,3-21; tables 3-3 *
ref_type: npl
severity: background
ref_id: - STEFAN MICHELFELDER ET AL, "Successful Expansion but Not Complete Restriction of Tropism of Adeno-Associated Virus by In Vivo Biopanning of Random Virus Display Peptide Libraries", PLOS ONE, (20090409), vol. 4, no. 4, doi:10.1371/journal.pone.0005122, page e5122, XP055147203 [A] 1-15 * the whole document *
ref_type: npl
severity: background
ref_id: - K VARADI ET AL, "Novel random peptide libraries displayed on AAV serotype 9 for selection of endothelial cell-directed gene transfer vectors", GENE THERAPY, (20120801), vol. 19, no. 8, doi:10.1038/gt.2011.143, ISSN 0969-7128, pages 800 - 809, XP055129104 [A] 1-15 * the whole document *
ref_type: npl
severity: background
Final disposition: pending (2020-09-30)