HCV clade and prototype sequences thereof

• US 20050069870A1
• Filed: 08/27/2004
• Published: 03/31/2005
• Est. Priority Date: 08/29/2003
• Status: Active Grant

• Current Assignee: INNOGENETICS N (H.U. Group Holdings, Inc.)
• Original Assignee: INNOGENETICS N (H.U. Group Holdings, Inc.)
• Inventors: Quint, Wim, Van Doorn, Leen-Jan, Sablon, Erwin
• Granted Patent: US 8,124,747 B2
• Time in Patent Office: Days
• US Class Current: 435/5
• 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 31/14 for RNA viruses
• CPC Class Codes: C07K 14/005 from viruses
• CPC Class Codes: C12N 2770/24222 New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
• CPC Class Codes: C12N 2770/24251 Methods of production or purification of viral material
• CPC Class Codes: C12N 7/00 Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof preparing medicinal viral antigen or antibody compositions, e.g. virus vaccines, A61K39/00
• CPC Class Codes: C12Q 1/707 non-A, non-B Hepatitis, excluding hepatitis D
• CPC Class Codes: C12Q 2525/117 incorporating modified base

Abstract

The current invention relates to a previously unrecognized clade of HCV genotypes as well as to diagnostic, prophylactic and therapeutic applications of nucleic acids, proteins, and antibodies to said protein, derived of or based on the newly characterized hepatitis C viruses.

Claims

1-42. -42. (canceled)

43. An isolated nucleic acid comprising an HCV nucleic acid sequence of an HCV lade

represented by any of the following HCV nucleic acid sequences which are prototype

sequences of said HCV lade:

(i) a nucleic acid sequence defined by SEQ ID NOs;

1, 2, 9, 10 or 16;

(ii) a nucleic acid sequence encoding a protein defined by SEQ ID NOs;

3, 4 or 17;

(iii) a nucleic acid sequence which is the complement of a nucleic acid sequence of (i) or (ii);

(iv) a nucleic acid sequence hybridizing to a nucleic acid sequence of (i), (ii) or (iii) under stringent conditions;

wherein said nucleic acid is DNA, cDNA or a synthetic nucleic acid, or wherein said nucleic acid is RNA and wherein “

T”

is replaced by “

U”

.

45. An oligonucleotide comprising at least 8 contiguous nucleotides taken from an HCV nucleic acid sequence according to claim 43 and wherein said oligonucleotide comprises at least one nucleotide that is different from any HCV nucleic acid sequence of an HCV of clades 1 to 6 or different from any HCV nucleic acid sequence of an HCV genotype or subtype of clades 1 to 6.

46. The oligonucleotide primer comprising at least of contiguous nucleotides taken from an HCV nucleic acid sequence of an HCV lade represented by any of the following HCV nucleic acid sequences which are prototype sequences of said HCV lade:

(i) a nucleic acid sequence defined by SEQ ID NOs;

1, 2, 9, 10 or 16;

(ii) a nucleic acid sequence encoding a protein defined by SEQ ID NOs;

3, 4 or 17;

(iii) a nucleic acid sequence which is the complement of a nucleic acid sequence of (i) or (ii);

(iv) a nucleic acid sequence hybridizing to a nucleic acid sequence of (i), (ii) or (iii) under stringent conditions;

wherein said nucleic acid is DNA, cDNA or a synthetic nucleic acid, or wherein said nucleic acid is RNA and wherein “

T”

is replaced by “

U”

;

wherein said oligonucleotide comprises at least one nucleotide that is different from any HCV nucleic acid sequence of an HCV of clades 1 to 6 or different from any HCV nucleic acid sequence of an HCV genotype or subtype of clades 1 to 6;

said primer being capable of directing specific amplification of an HCV nucleic acid sequence according to claim 43.

47. The oligonucleotide probe comprising at least 8 contiguous nucleotides taken from an HCV nucleic acid sequence of an HCV lade represented by any of the following HCV nucleic acid sequences which are prototype sequences of said HCV lade:

(i) a nucleic acid sequence defined by SEQ ID NOs;

1, 2, 9, 10 or 16;

(ii) a nucleic acid sequence encoding a protein defined by SEQ ID NOs;

3, 4 or 17;

(iii) a nucleic acid sequence which is the complement of a nucleic acid sequence of (i) or (ii);

(iv) a nucleic acid sequence hybridizing to a nucleic acid sequence of (i), (ii) or (iii) under stringent conditions;

wherein said nucleic acid is DNA, cDNA or a synthetic nucleic acid, or wherein said nucleic acid is RNA and wherein “

T”

is replaced by “

U”

;

wherein said oligonucleotide comprises at least one nucleotide that is different from any HCV nucleic acid sequence of an HCV of clades 1 to 6 or different from any HCV nucleic acid sequence of an HCV genotype or subtype of clades 1 to 6;

said probe being capable of specifically hybridizing to an HCV nucleic acid sequence according to claim 43.

48. The oligonucleotide by any of the following HCV nucleic acid sequences which are prototype sequences of said HCV lade:

(i) a nucleic acid sequence defined by SEQ ID NOs;

1, 2, 9, 10 or 16;

(ii) a nucleic acid sequence encoding a protein defined by SEQ ID NOs;

3, 4 or 17;

(iii) a nucleic acid sequence which is the complement of a nucleic acid sequence of (i) or (ii);

(iv) a nucleic acid sequence hybridizing to a nucleic acid sequence of (i), (ii) or (iii) under stringent conditions;

wherein said nucleic acid is DNA, cDNA or a synthetic nucleic acid, or wherein said nucleic acid is RNA and wherein “

T”

is replaced by “

U”

;

wherein said oligonucleotide comprises at least one nucleotide that is different from any HCV nucleic acid sequence of an HCV of clades 1 to 6 or different from any HCV nucleic acid sequence of an HCV genotype or subtype of clades 1 to 6;

said oligonucleotide being capable of specifically detecting an HCV nucleic acid sequence according to claim 43.

49. The nucleic acid sequence according to claim 43, further comprising

one or more modified nucleotide bases;

one or more labeled nucleotides;

one or more peptide nucleic acid monomers;

one or more locked nucleic acid monomers; and

/or

wherein the backbone of said nucleic acid is modified.

50. The oligonucleotide sequence of claim 45, further comprising

one or more modified nucleotide bases;

one or more labeled nucleotides;

one or more peptide nucleic acid monomers;

one or more locked nucleic acid monomers; and

/or

wherein the backbone of said oligonucleotide is modified.

51. An isolated protein comprising an HCV amino acid sequence encoded by an HCV nucleic acid sequence according to claim 43.

52. The isolated protein according to claim 51 wherein said HCV amino acid sequence is defined by SEQ ID NOs:

3, 4 or 17.

53. A peptide of at least 5 contiguous amino acids taken from an HCV amino acid sequence according to claim 51 and wherein said peptide comprises at least one amino acid that is different from any HCV amino acid sequence of an HCV of clades 1 to 6 or different from any HCV amino acid sequence of an HCV genotype or subtype of clades 1 to 6.

54. The protein according to claim 51, wherein if said HCV amino acid sequence is comprising at least one cysteine, said cysteine is reversibly or irreversibly protected.

55. A recombinant vector comprising a nucleic acid or oligonucleotide according to claim 43.

56. The recombinant vector according to claim 55 which is an expression vector capable of directing expression of an HCV protein encoded by the HCV nucleic acid sequence comprised in said vector.

57. A host cell transformed with a nucleic acid or oligonucleotide according to claim 43 or with a recombinant vector comprising said nucleic acid or oligonucleotide.

58. A method for detecting the presence of an HCV virus in a biological sample comprising the step of detecting the presence of a nucleic acid or oligonucleotide according to claim 43.

59. A method for determining the genotype of an HCV virus in a biological sample comprising the step of detecting the presence of a nucleic acid or oligonucleotide according to claim 43.

60. The method according to claim 58 wherein the detection of the presence of said HCV nucleic acid sequence is based on at least one of an amplification reaction, a sequencing reaction, a hybridization reaction or a reverse hybridization reaction.

61. The method according to claim 58 wherein the detection of the presence of said HCV nucleic acid sequence comprises:

amplification of said HCV nucleic acid sequence with an oligonucleotide;

hybridization or reverse hybridization of said HCV nucleic acid with an oligonucleotide;

or

detection of said HCV nucleic acid with an oligonucleotide.

62. A method for detecting the presence of an HCV virus in a biological sample comprising the steps of:

(i) obtaining a target HCV nucleic acid from a biological sample suspected to contain an HCV nucleic acid sequence according to claim 43;

(ii) obtaining the nucleic acid sequence of the target HCV nucleic acid of (i);

(iii) inferring from the nucleic acid sequence obtained in (ii) the presence of an HCV nucleic acid sequence according to claim 43 and, therefrom, the presence of an HCV virus in said biological sample.

63. A method for detecting the presence of an HCV virus in a biological sample comprising the steps of:

(i) obtaining a target HCV nucleic acid from a biological sample suspected to contain an HCV nucleic acid sequence according to claim 43;

(ii) contacting the target HCV nucleic acid of (i) with an oligonucleotide, and said contacting generating a discriminatory signal;

(iii) inferring from the discriminatory signal obtained in (ii) the presence of the HCV nucleic acid sequence and, therefrom, the presence of an HCV virus in said biological sample.

64. A method for determining the genotype of an HCV virus in a biological sample comprising the steps of:

(i) obtaining a target HCV nucleic acid from a biological sample suspected to contain an HCV nucleic acid sequence according to claim 43;

(ii) obtaining the nucleic acid sequence of the target HCV nucleic acid of (i);

(iii) inferring from the nucleic acid sequence obtained in (ii) the presence of the HCV nucleic acid sequence and, therefrom, the genotype of said HCV virus in said biological sample.

65. A method for detecting the genotype of an HCV virus in a biological sample comprising the steps of:

(i) obtaining a target HCV nucleic acid from a biological sample suspected to contain an HCV nucleic acid sequence according to claim 43;

(ii) contacting the target HCV nucleic acid of (i) with an oligonucleotide, and said contacting generating a discriminatory signal;

(iii) inferring from the discriminatory signal obtained in (ii) the presence of the HCV nucleic acid sequence and, therefrom, the genotype of said HCV virus in said biological sample.

66. A diagnostic kit for detecting the presence and/or for determining the genotype of an HCV virus in a biological sample, said kit comprising at least one of a nucleic acid according to claim 43 or an oligonucleotide relating thereto.

67. The diagnostic kit according to claim 66 wherein said nucleic acid or oligonucleotide is attached to a solid support.

68. A method for detecting antibodies to an HCV virus present in a biological sample comprising the step of detecting said antibodies with at least one protein or peptide according to claim 51.

70. A method for typing of an HCV virus present in a biological sample comprising the steps of:

(i) contacting said biological sample with a protein or peptide according to claim 51;

(ii) detecting the immunological complex formed in (i) between the antibodies to HCV present in said biological sample and said protein or peptide;

(iii) inferring from the immunological complex detected in (ii) the type of HCV virus present in said biological sample.

71. A diagnostic kit for detecting antibodies to an HCV virus or for typing of an HCV virus, said kit comprising at least one protein or peptide according to claim 51.

72. The diagnostic kit according to claim 71 wherein said protein or peptide are bound to a solid support.

73. An antibody raised upon immunization of a mammal with at least one protein or peptide according to claim 51.

74. The antibody according to claim 73 which is a monoclonal antibody or a humanized antibody.

75. A method for detecting the presence of HCV antigens in a biological sample comprising the steps of:

(i) contacting said biological sample with an antibody according to claim 73 under conditions allowing the formation of an immunological complex between HCV antigens and said antibody;

(ii) detecting the immunological complex formed in (i);

(iii) inferring from the immunological complex detected in (ii) the presence of HCV antigens present in said biological sample.

76. A method for typing of an HCV virus present in a biological sample comprising the steps of:

(i) contacting said biological sample with an antibody according to claim 73;

(ii) detecting the immunological complex formed in (i) between the HCV antigens present in said biological sample and said antibody;

(iii) inferring from the immunological complex detected in (ii) the type of HCV virus present in said biological sample.

77. A method for detecting antibodies to an HCV virus present in a biological sample comprising contacting an antigen with said antibodies in the presence of an isolated antibody according to claim 73 as competitor of binding of said antigen to said antibody.

78. A method for detecting the presence of HCV antigens in a biological sample comprising contacting said antigen with an antibody to said antigen in the presence of an isolated protein or peptide according to claim 51 as competitor of binding of said antigen to said antibody.

79. A diagnostic kit for detecting the presence of HCV antigens, said kit comprising at least an antibody according to claim 73.

80. A diagnostic kit for detecting the presence of HCV antigens, said kit comprising at least one isolated protein or peptide according to claim 51.

81. A composition comprising at least one of an nucleic acid according to claim 43;

and at least one of a suitable carrier, adjuvant or vehicle.

82. A composition comprising a protein of claim 51 and at least one carrier.

83. A composition comprising a vector of claim 55 and at least one carrier.

84. A composition comprising an antibody of claim 73 and at least one carrier.

85. A composition of claim 81, said composition being immunogenic.

86. A composition of claim 82, said composition being immunogenic.

87. A composition of claim 83, said composition being immunogenic.

88. A composition of claim 84, said composition being immunogenic.

89. A vaccine comprising a composition of claim 81.

90. A vaccine comprising a composition of claim 82.

91. A vaccine comprising a composition of claim 83.

92. A vaccine comprising a composition of claim 84.

93. A method for the recombinant production of a protein or peptide according to claim 51 comprising the steps of:

(i) transformation of an appropriate cellular host with a recombinant vector encoding said protein or peptide;

(ii) culturing the transformed cellular host of (i) under conditions enabling the expression of said protein;

(iii) harvesting the protein expressed in (ii).

44. An isolated nucleic acid comprising an HCV nucleic acid sequence of an HCV genotype represented by any of the following HCV nucleic acid sequences which are prototype sequences of said HCV genotype:

(i) a nucleic acid sequence defined by SEQ ID NOs;

1, 2, 9, 10, or 16 wherein SEQ ID NOs;

1 and 9 and SEQ ID NOs;

2, 10 and 16 each represent a different subtype of said HCV genotype;

(ii) a nucleic acid sequence encoding a protein defined by SEQ ID NOs;

3, 4 or 17;

(iii) a nucleic acid sequence which is the complement of a nucleic acid sequence of (i) or (ii);

(iv) a nucleic acid sequence hybridizing to a nucleic acid sequence of (i), (ii) or (iii) under stringent conditions;

wherein said nucleic acid is DNA, cDNA or a synthetic nucleic acid, or wherein said nucleic acid is RNA and wherein “

T”

is replaced by “

U”

.

69. A method for detecting antibodies to an HCV virus present in a biological sample comprising the steps of:

(i) contacting said biological sample with an isolated protein or part thereof or derivative of any thereof according to the invention;

(ii) detecting the immunological complex formed in (i) between said antibodies and said protein or part thereof or derivative of any thereof.