US2008145379A1PendingUtilityA1
Recombinant DNA molecules encoding aminopeptidase enzymes and their use in the preparation of vaccines against helminth infections
Est. expiryMay 8, 2012(expired)· nominal 20-yr term from priority
Inventors:Margaret GrahamTrevor S. SmithEdward Albert MunnDavid KnoxJoanna Jane OliverSusan Elizabeth Newton
C12N 9/48C12N 2799/026C07K 14/4354C07K 2319/02C07K 9/00C07K 9/001C07H 13/04A61P 33/10A61P 33/00A61K 2039/55566A61K 39/0003A61K 39/00
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Claims
Abstract
The present invention provides nucleic acid molecules containing nucleotide sequences encoding helminth aminopeptidase enzymes, and antigenic fragments and functionally-equivalent variants thereof, their use in the preparation of vaccines, for use against helminth parasites, and synthetic polypeptides encoded by them.
Claims
exact text as granted — not AI-modified1 . A synthetic polypeptide comprising an amino acid sequence encoded by a nucleotide sequence which encodes a helminth aminopeptidase or an antigenic portion thereof, said nucleic acid selected from the group consisting of i) any sequence of SEQ ID NOs:1-15 or 19-21, ii) a sequence fully complementary to said sequence of (i), iii) a portion of said sequence of (i) wherein said portion of said sequence encodes an antigenic peptide of said helminth aminopeptidase, and (iv) a sequence which hybridizes with said sequence of (i) or (ii) under conditions of 2 times SSC, 65 C (where SSC=0.15 M NaCl, 0.015 M sodium citrate, pH 7.2) and which has at least 60% identity with said sequence of (i), other than a synthetic polypeptide corresponding to the protein doublet H110D or a synthetic polypeptide selected from:
(SEQ ID NO: 25)
(a)
Met Gly Tyr Pro Val Val Lys Val Glu Glu Phe,
(SEQ ID NO: 26)
(b)
Met Gly Phe Pro Val Leu Thr Val Glu Ser,
(SEQ ID NO: 27)
(c)
Met Gly/Phe Asn Phe Lys Ile Glu/Val Thr/Glu
Ala Gly,
(SEQ ID NO: 28)
(d)
Met Lys Pro/Glu Thr/Val Lys Asp/Ala Thr/Lys
Leu - Ile Thr,
(SEQ ID NO: 29)
(e)
Met Leu Ala Leu Asp Tyr His Ser - Phe Val,
(SEQ ID NO: 30)
(f)
Met Leu Ala Glu/Tyr Asp Gln/Ala Glu Asp Val,
(SEQ ID NO: 31)
(g)
Met Gly Phe Pro Leu Val Thr Val Glu Ala Phe
Tyr,
(SEQ ID NO: 32)
(h)
Met Lys Thr Pro Glu Phe Ala Val/Leu Gln Ala
Phe/Thr Ala Thr Ser/Gly Phe Pro,
(SEQ ID NO: 33)
(i)
Lys His/Tyr Asn/Val Ser Pro Ala Ala Glu Asn/
Leu Leu Asn/Gly,
(SEQ ID NO: 34)
(j)
Lys - Thr Ser Val Ala Glu Ala Phe Asn,
(SEQ ID NO: 35)
(k)
Lys Ala Ala Glu Val Ala Glu Ala Phe Asp - Ile
- - - Lys Gly,
(SEQ ID NO: 36)
(l)
Lys Ala Val Glu Val/Pro Ala Glu Ala Phe Asp
Asp Ile Thr? Tyr - - Gly Pro Ser,
(SEQ ID NO: 37)
(m)
Lys - Glu Glu Thr Glu Ile Phe Asn Met,
(SEQ ID NO: 38)
(n)
Lys - - - Pro Phe Asn/Asp Ile Glu Ala Leu,
(SEQ ID NO: 39)
(o)
Asp Gln Ala Phe Ser Thr Asp Ala Lys,
(SEQ ID NO: 40)
(p)
Met Gly Tyr Pro Val Val Lys Val Glu Glu Phe -
Ala Thr Ala Leu,
(SEQ ID NO: 41)
(q)
Met Gly Phe Pro Val Leu Thr Val Glu Ser -
Tyr? - Thr,
(SEQ ID NO: 42)
(r)
Met Glu/Phe Asn Phe Leu Ile Glu/Val Thr/Glu
Ala Gly - Ile Thr,
(SEQ ID NO: 43)
(s)
Met Gly Phe Leu Val Thr Val Glu Ala Phe Tyr -
Thr Ser,
(SEQ ID NO: 44)
(t)
Met Lys Thr Pro Glu Phe Ala Val/Leu Gln Ala
Phe/Thr Ala Thr Ser/Gly Phe Pro
(SEQ ID NO: 45)
(u)
Met Lys Pro/Glu Thr/Val Leu Asp/Ala Thr/Lys
Leu - Ile Thr - Gly,
(SEQ ID NO: 46)
(v)
Met Leu Ala Leu Asp Tyr His Ser - Phe Val
Gly?,
(SEQ ID NO: 47)
(w)
Met Leu Ala Glu/Tyr Asp Gln/Ala Glu Asp Val,
(SEQ ID NO: 48)
(x)
Lys His/Tyr Asn/Val Ser Pro Ala Ala Glu Asn/
Leu Leu Asn/Gly,
(SEQ ID NO: 49)
(y)
Lys - Thr Ser Val Ala Glu Ala Phe Asn,
(SEQ ID NO: 50)
(z)
Lys Ala Ala Glu Val Ala Glu Ala Phe Asp -
lIe - - - Lys Gly,
(SEQ ID NO: 51)
(aa)
Lys Ala Val Glu Val/Pro Ala Glu Ala Phe Asp
Asp Ile Thr? Tyr - - Gly Pro Ser,
(SEQ ID NO: 52)
(bb)
Lys - Glu Gln Thr Glu Ile Phe Asn Met,
(SEQ ID NO: 53)
(cc)
Lys - - - Pro Phe Asn/Asp Ile Glu Ala Leu
and
(SEQ ID NO: 54)
(dd)
Asp Gln Ala Phe Ser Thr Asp Ala Lys.
2 . A synthetic polypeptide comprising an amino acid sequence encoded by a nucleotide sequence which encodes a polypeptide which raises protective antibodies against helminth parasites, which nucleotide sequence incorporates at least one aminopeptidase-encoding sequence, selected from the group consisting of i) any sequence of SEQ ID NOS: 1-15 or 19-21, ii) a sequence fully complementary to said sequence of (i), iii) a portion of said sequence of (i), and iv) a sequence which hybridizes with said sequence of (i) or (ii) under conditions of 2 times SSC, 65 C (where SSC=0.15 M NaCl, 0.015 M sodium citrate, pH 7.2) and which has at least 60% identity with said sequence of (i), other than a synthetic polypeptide corresponding to the protein doublet H110D or a synthetic polypeptide as defined in (a) to (dd) in claim 1 .
3 . A synthetic polypeptide comprising an amino acid sequence encoded by a nucleotide sequence which corresponds to or which is complementary to one or more sequences selected from the group of sequences consisting of i) M1(SEQ ID NO:1), ii) B1A (SEQ ID NO:2), iii) B1A-3′ (SEQ ID NO:3), iv) B2 (SEQ ID NO:4), v) M1AUS (SEQ ID NO:5), vi) AusB1 (SEQ ID NO:6), vii) 014-015 (2.5 PCR) (SEQ ID NO:7), viii) 014-872 (3.5 PCR clone 2) (SEQ ID NO:8), ix) A-648 (5′ END of B1) (SEQ ID NO:9), x) A-650 (5′-end of 2.5 PCR) (SEQ ID NO:10), xi) α-649 (5′ end of 3.5 PCR) (SEQ ID NO:11), xii) 014-178 (3′ end of AustB1 clone 2) (SEQ ID NO:12), xiii) 014-178 (3′ end of AustB1 clones 3 and 6) (SEQ ID NO:13) xiv) 014-872 (3.5 PCR clone 10) (SEQ ID NO:14), xv) 014-872 (3.5 PCR clone 19) (SEQ ID NO:15), xvi) H11-1 (SEQ ID NO:19), xvii) H11-2 (SEQ ID NO:20), xviii) H11-3 (SEQ ID NO:21), and xix) a sequence which hybridizes with any of said sequences of (i)-(xviii) under conditions of 2 times SSC, 65 C (where SSC=0.15 M NaCl, 0.015 M sodium citrate, pH 7.2) and which has at least 60% identity with said sequence of (i)-(xviii), other than a synthetic polypeptide corresponding to the protein doublet H110D or a synthetic polypeptide as defined in (a) to (dd) in claim 1 .
4 . A synthetic polypeptide as claimed in claim 1 , 2 or 3 substantially free from other Haemonchus contortus components.
5 . A synthetic polypeptide as claimed in claim 1 , 2 or 3 in the form of a fusion polypeptide, comprising an additional polypeptide fused to said amino acid sequence as defined in any one of claims 1 to 3 .
6 . A synthetic polypeptide as claimed in claim 1 , 2 or 3 linked to an oligosaccharide having the structure:
7 . A synthetic polypeptide as claimed in claim 6 wherein said oligosaccharide has the structure:
8 . A synthetic polypeptide as claimed in claim 1 , 2 or 3 wherein said synthetic polypeptide is a recombinant polypeptide.
9 . A synthetic polypeptide as claimed in claim 1 , 2 or 3 prepared by a method comprising the steps:
a) culturing a prokaryotic or eukaryotic cell containing a nucleic acid molecule comprising at least one nucleotide sequence which encodes a helminth aminopeptidase or an antigenic portion thereof, said nucleic acid selected from the group consisting of i) any sequence of SEQ ID NOs:1-15 or 19-21, ii) a sequence fully complementary to said sequence of (i), iii) a portion of said sequence of (i) wherein said portion of said sequence encodes an antigenic peptide of said helminth aminopeptidase, and (iv) a sequence which hybridizes with said sequence of (i) or (ii) under conditions of 2 times SSC, 65 C (where SSC=0.15 M NaCl, 0.015 M sodium citrate, pH 7.2) and which has at least 60% identity with said sequence of (i), under conditions whereby said synthetic polypeptide is expressed, and b) recovering said synthetic polypeptide thus produced.
10 . A vaccine composition for stimulating immune responses against helminth parasites in a human or non-human animal, comprising at least one synthetic polypeptide as defined in claim 1 , 2 or 3 , or a virus or host cell having inserted therein a nucleic acid molecule encoding said polypeptide, for stimulation of an immune response to polypeptides encoded by the inserted nucleic acid molecule, together with a pharmaceutically acceptable carrier.
11 . A method of stimulating an immune response against helminth parasites in a human or non-human animal comprising administering to said animal a vaccine composition as defined in claim 10 .Join the waitlist — get patent alerts
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