Novel isoforms of human pregnancy-associated protein-E

• US 20040063134A1
• Filed: 09/30/2003
• Published: 04/01/2004
• Est. Priority Date: 05/26/2000
• Status: Abandoned Application

• Current Assignee: Amersham PLC (General Electric Company)
• Original Assignee: Amersham PLC (General Electric Company)
• Inventors: Gu, Yizhong, Shannon, Mark E.
• Application Number:
  US10/675,685
• Publication Number:
  US 20040063134A1
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: A01K 2217/05 Animals comprising random inserted nucleic acids (transgenic)
• CPC Class Codes: A01K 2217/075 inducing loss of function, i.e. knock out
• 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: C07K 14/47 from mammals
• CPC Class Codes: C07K 14/4748 Tumour specific antigens; Tumour rejection antigen precursors [TRAP], e.g. MAGE
• CPC Class Codes: C07K 14/705 Receptors; Cell surface antigens; Cell surface determinants tumour specific antigens C07K14/4748
• CPC Class Codes: C07K 2319/00 Fusion polypeptide
• CPC Class Codes: C07K 2319/02 containing a signal sequence
• CPC Class Codes: C07K 2319/40 containing a tag for immunodetection, or an epitope for immunisation
• CPC Class Codes: C07K 2319/60 containing spectroscopic/fluorescent detection, e.g. green fluorescent protein [GFP]
• CPC Class Codes: C12N 15/1034 Isolating an individual clone by screening libraries
• CPC Class Codes: C12N 15/66 General methods for inserting a gene into a vector to form a recombinant vector using cleavage and ligation; Use of non-functional linkers or adaptors, e.g. linkers containing the sequence for a restriction endonuclease
• CPC Class Codes: C12Q 1/6837 using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C12Q 1/6883 for diseases caused by alterations of genetic material
• CPC Class Codes: C12Q 1/6886 for cancer immunoassay for cancer G01N33/574
• CPC Class Codes: C12Q 2600/156 Polymorphic or mutational markers
• CPC Class Codes: C12Q 2600/158 Expression markers
• CPC Class Codes: G16B 25/00 ICT specially adapted for hybridisation; ICT specially adapted for gene or protein expression
• CPC Class Codes: G16B 30/00 ICT specially adapted for sequence analysis involving nucleotides or amino acids
• CPC Class Codes: Y02A 90/10 Information and communication technologies [ICT] supporting adaptation to climate change, e.g. for weather forecasting or climate simulation

Abstract

The invention provides isolated nucleic acids that encode three novel isoforms of human pregnancy associated plasma protein E, hPAPP-E, and fragments thereof, vectors for propagating and expressing PAPP-E nucleic acids, host cells comprising the nucleic acids and vectors of the present invention, proteins, protein fragments, and protein fusions of the novel PAPP-E isoforms, and antibodies thereto. The invention further provides transgenic cells and non-human organisms comprising human PAPP-E isoform nucleic acids, and transgenic cells and non-human organisms with targeted disruption of the endogenous orthologue of the human PAPP-E gene. The invention further provides pharmaceutical formulations of the nucleic acids, proteins, and antibodies of the present invention, and diagnostic, investigational, and therapeutic methods based on the PAPP-E nucleic acids, proteins, and antibodies of the present invention.

Claims

1. An isolated nucleic acid comprising a nucleotide sequence selected from the group consisting of:

(i) the nucleotide sequence of SEQ ID NO;

1, (ii) the complement of the nucleotide sequence of SEQ ID NO;

1;

(iii) the nucleotide sequence of SEQ ID NO;

2, (iv) a degenerate variant of the nucleotide sequence of SEQ ID NO;

2, or (v) the complement (iii) or (iv), (vi) a nucleotide sequence that encodes a polypeptide having the sequence of SEQ ID NO;

3, (vii) a nucleotide seuqence that encodes a polypeptide having the sequence of SEQ ID NO;

3 with conservative amino acid substitutions, and (viii) a nucleotide sequence that is the complement of (vii) or (viii).

24. The isolated nucleic acid of any of claims 1-23, wherein said isolated nucleic acid is detectably labeled.

25. The isolated nucleic acid of any one of claims 1-23, wherein said nucleic acid is attached to a substrate.

27. A recombinant nucleic acid vector, comprising:

the isolated nucleic acid of any one of claims 1-23.

28. The vector of claim 27, wherein said isolated nucleic acid is operably linked to genetic elements capable of driving expression of said nucleic acid in a host cell.

29. A host cell comprising the isolated nucleic acid of any one of claims 1-23 or the vector of either of claims 27-28.

34. A pharmaceutical composition comprising:

the nucleic acid of any one of claims 1-23, the polypeptide of any one of claims 30-31, or the antibody of claim 32;

and

a pharmaceutically acceptable excipient.

36. A method of preventing or abrogating pregnancy, comprising:

vaccinating a human subject with the nucleic acid of any one of claims 1-23 or the polypeptide of any one of claims 30-31, or administering an effective amount of the antibody of claim 32.

2. An isolated nucleic acid comprising a nucleotide sequence selected from the group consisting of:

(i) the nucleotide sequence of SEQ ID NO;

4, (ii) the complement of the nucleotide sequence of SEQ ID NO;

4, (iii) a nucleotide sequence that encodes a polypeptide having the sequence of SEQ ID NO;

7, (iv) a nucleotide sequence that encodes a polypeptide having the sequence of SEQ ID NO;

7 with conservative amino acid substitutions, and (iii) the complement of (i) or (ii),

wherein said isolated nucleic acid is no more than 50 kb in length.

3. An isolated nucleic acid comprising a sequence that hybridizes under high stringency conditions to a hybridization probe the nucleotide sequence of which (i) encodes a polypeptide having the sequence of SEQ ID NO:

7, (ii) encodes a polypeptide having the sequence of SEQ ID NO;

7 with conservative amino acid substitutions, or (iii) is the complement of (i) or (ii),

wherein said isolated nucleic acid is no more than 50 kb in length.

4. The isolated nucleic acid of claim 3, wherein said nucleic acid, or the complement of said nucleic acid, encodes a polypeptide having metalloproteinase activity.

5. The isolated nucleic acid of claim 3, wherein said nucleic acid, or the complement of said nucleic acid, is expressed in placental tissue.

6. An isolated nucleic acid comprising at least 17 nucleotides of (i) SEQ ID NO:

4 or (ii) the complement of SEQ ID NO;

4,

wherein said isolated nucleic acid is no more than 50 kb in length.

7. An isolated nucleic acid comprising (i) a nucleotide sequence that encodes a peptide of at least 8 contiguous amino acids of SEQ ID NO:

7, or (ii) the complement of a nucleotide sequence that encodes a peptide of at least 8 contiguous amino acids of SEQ ID NO;

7,

wherein said isolated nucleic is no more than 50 kb in length.

8. An isolated nucleic acid comprising a nucleotide sequence selected from the group consisting of:

(i) the nucleotide sequence of SEQ ID NO;

8, (ii) the complement of the nucleotide sequence of SEQ ID NO;

8, (iii) the nucleotide sequence of SEQ ID NO;

9, (iv) a degenerate variant of the nucleotide sequence of SEQ ID NO;

9, or (v) the complement (iii) or (iv), (vi) a nucleotide sequence that encodes a polypeptide having the sequence of SEQ ID NO;

10, (vii) a nucleotide sequence that encodes a polypeptide having the sequence of SEQ ID NO;

10 with conservative amino acid substitutions, and (viii) a nucleotide sequence that is the complement of (vi) or (vii).

9. An isolated nucleic acid comprising a nucleotide sequence selected from the group consisting of:

(i) the nucleotide sequence of SEQ ID NO;

11,

(ii) a degenerate variant of SEQ ID NO;

11, (iii) the complement of (i) or (ii), (iv) a nucleotide sequence that encodes SEQ ID NO;

12, (v) a nucleotide sequence that encodes SEQ ID NO;

12 with conservative substititions, (vi) the complement of (iv) or (v),

wherein said isolated nucleic acid is no more than 50 kb in length.

10. An isolated nucleic acid comprising a nucleotide sequence (i) that encodes a polypeptide having the sequence of SEQ ID NO:

14, (ii) that encodes a polypeptide having the sequence of SEQ ID NO;

14 with conservative amino acid substitutions, or (iii) the complement of a nucleotide sequence that encodes a polypeptide with the amino acid sequence of SEQ ID NO;

14,

wherein wherein said isolated nucleic acid is no more than 50 kb in length.

11. An isolated nucleic acid comprising a sequence that hybridizes under high stringency conditions to a probe the nucleotide sequence of which consists of SEQ ID NO:

11 or the complement of SEQ ID NO;

11,

wherein said isolated nucleic acid is no more than 50 kb in length.

12. The isolated nucleic acid of claim 11, wherein said nucleic acid, or the complement of said nucleic acid, encodes a polypeptide having metalloproteinase activity.

13. The isolated nucleic acid of claim 11, wherein said nucleic acid, or the complement of said nucleic acid, is expressed in placental tissue.

14. An isolated nucleic acid comprising at least 17 nucleotides of (i) SEQ ID NO:

11, (ii) a degenerate variant of SEQ ID NO;

11, or (iii) the complement of (i) or (ii),

wherein said isolated nucleic acid is no more than 50 kb in length.

15. An isolated nucleic acid comprising a nucleotide sequence that (i) encodes a polypeptide having the sequence of at least 8 contiguous amino acids of SEQ ID NO:

12, (ii) encodes a polypeptide having the sequence of at least 8 contiguous amino acids of SEQ ID NO;

12 with conservative amino acid substitutions, or (iii) is the complement of (i) or (ii).

16. An isolated nucleic acid comprising a nucleotide sequence at least 90% identical to SEQ ID NO:

11.

17. An isolated nucleic acid comprising a nucleotide sequence selected from the group consisting of:

(i) the nucleotide sequence of SEQ ID NO;

15, (ii) a degenerate variant of SEQ ID NO;

15, (iii) the complement of (i) or (ii), (iv) a nucleotide sequence that encodes a polypeptide having the sequence of SEQ ID NO;

16, (v) a nucleotide sequence that encodes a polypeptide having the sequence of SEQ ID NO;

16 with conservative amino acid substitutions, and (vi) the complement of (iv) or (v).

18. An isolated nucleic acid comprising a sequence selected from the group consisting of:

(i) the nucleotide sequence of SEQ ID NO;

17, (ii) a degenerate variant of SEQ ID NO;

17, (iii) the complement of (i) or (ii), (iv) a nucleotide sequence that encodes SEQ ID NO;

18, (v) a nucleotide sequence that encodes SEQ ID NO;

18 with conservative amino acid substititions, and (vi) the complement of (i) or (ii).

19. The isolated nucleic acid of claim 18 wherein said nucleic acid, or the complement of said nucleic acid, encodes a polypeptide having metalloproteinase activity.

20. The isolated nucleic acid of claim 18 wherein said nucleic acid, or the complement of said nucleic acid, is expressed in placental tissue.

21. An isolated nucleic acid comprising a nucleotide sequence of no more than one fragment of SEQ ID NOs:

19 to 41 or the complement of SEQ ID NOs;

19 to 41, wherein said fragment comprises at least 17 contiguous nucleotides of said SEQ ID NO;

or its complement and hybridizes under high stringency conditions to a nucleic acid expressed in human placenta.

22. The isolated nucleic acid of claim 68, wherein said fragment comprises at least 50 contiguous nucleotides of said SEQ ID NO:

or its complement.

23. An isolated nucleic acid comprising the nucleotide sequence of SEQ ID NO:

65 or its complement,

wherein said isolated nucleic acid is no more than 50 kb in length.

26. A microarray, comprising:

a plurality of nucleic acids addressably disposed upon a substrate,

wherein at least one of said nucleic acids is the isolated nucleic acid of any one of 1-23.

30. An isolated polypeptide, said polypeptide comprising the amino acid sequence of SEQ ID NO:

3, SEQ ID NO;

10, or SEQ ID NO;

16.

31. An isolated polypeptide comprising a sequence of at least 8 contiguous amino acids of SEQ ID NO:

7, SEQ ID NO;

12, SEQ ID NO;

13, SEQ ID NO;

14, or SEQ ID NO;

18.

32. An antibody, said antibody selected from the group consisting of:

(i) antibodies capable of binding specifically to polypeptides comprising an amino acid sequence as provided in SEQ ID NO;

7, and the binding of which can be competitively inhibited by a polypeptide the sequence of which is given in SEQ ID NO;

7;

(ii) antibodies capable of binding specifically to polypeptides comprising an amino acid sequence as provided in SEQ ID NO;

14, and the binding of which can be competitively inhibited by a polypeptide the sequence of which is given in SEQ ID NO;

14; and

(iii) antibodies capable of binding specifically to polypeptides comprising an amino acid sequence as provided in SEQ ID NO;

18, and the binding of which can be competitively inhibited by a polypeptide the amino acid sequence of which is given in SEQ ID NO;

18.

33. The antibody of claim 32, which antibody is detectably labeled.

35. A method of antenatal diagnosis of dysgenetic pregnancies, comprising:

quantifying levels of PAPP-E protein circulating in maternal serum during first or second trimester of pregnancy, and comparing to a standard, decreased levels of circulating PAPP-E as compared to standard indicating increased likelihood of a dysgenetic pregnancy.