Methods of determination of activation or inactivation of atrial natriuretic peptide (anp) and brain natriuretic peptide (bnp) hormonal systems

Abstract

An in vivo method of determining activation or inactivation of the atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) hormonal systems, the method comprising simultaneously detecting the presence or amount of atrial and brain natriuretic peptide prohormones (proANP and proBNP) or fragments thereof in a sample.

1 . An agent which comprises:
 (a) (i) proANP (SEQ ID NO:1), ANP (SEQ ID NO:2), or NT-proANP (SEQ ID NO:3);
 (ii) a homologous sequence having at least 70% identity to (a)(i); or 
 (iii) a fragment of (a)(i) or (a)(ii) which is at least 6 amino acids in length; and 
   (b) (i) pro-BNP (SEQ ID NO:4), BNP (SEQ ID NO:5), or NT-proBNP (SEQ ID NO:6);
 (ii) a homologous sequence having at least 70% identity to (b)(i); or 
 (iii) a fragment of (b)(i) or (b)(ii) which is at least 6 amino acids in length. 
   
     
     
         2 . An agent according to  claim 1  which comprises or consists of:
 (a) proBNP 15-24  and proANP 82-96 ; 
 (b) proBNP 1-37  and proANP 29-98 ; 
 (c) proBNP 10-29  and proANP 20-80 ; 
 (d) proBNP 1-76  and proANP 1-98 ; 
 (e) proBNP 10-29  and proANP 60-80 ; 
 (f) proBNP 1-108  and proANP 1-126 ; or 
 (g) proBNP 77-92  and proANP 112-126 . 
 
     
     
         3 . An agent according to  claim 2  which comprises any one of SEQ ID NOs: 13, 14, 15, 17, 18, 19, or 20. 
     
     
         4 . An agent according to  claim 1  which is labelled with a detectable label. 
     
     
         5 . A polypeptide agent according to  claim 1 . 
     
     
         6 . A polynucleotide comprising sequence which encodes a polypeptide agent according to  claim 5  or sequence which is complementary to said coding sequence. 
     
     
         7 . A polynucleotide according to  claim 6  which comprises:
 (a) (i) SEQ ID NOs: 7, 8, or 9;
 (ii) a sequence complementary to (a)(i); 
 (iii) a sequence which hybridizes under medium or high stringent conditions to (a)(i) or (a)(ii); 
 (iv) a sequence which is degenerate as a result of the genetic code to (a)(i), (a)(ii), or (a)(iii); 
 (v) a sequence having at least 70% identity to any of the sequences in (a)(i) to (a)(iv); or 
 (vi) a fragment of any of the sequences in (a)(i) to (a)(v); and 
 
 (b) (i) SEQ ID NOs: 10, 11, or 12;
 (ii) a sequence complementary to (b)(i); 
 (iii) a sequence which hybridizes under medium or high stringent conditions to (b)(i) or (b)(ii); 
 (iv) a sequence which is degenerate as a result of the genetic code to (b)(i), (b)(ii), or (b)(iii); 
 (v) a sequence having at least 70% identity to any of the sequences in (b)(i) to (b)(iv); or 
 (vi) a fragment of any of the sequences in (b)(i) to (b)(v). 
 
 
     
     
         8 . An expression vector comprising a polynucleotide according to  claim 6 . 
     
     
         9 . A host cell comprising a polynucleotide according to  claim 6 . 
     
     
         10 . A process for producing a polypeptide agent according to  claim 5  which process comprises:
 (I) cultivating a host cell comprising a polynucleotide, or its complement, which encodes a polypeptide which comprises:
 (a) (i) proANP (SEQ ID NO:1), ANP (SEQ ID NO:2), or NT-proANP (SEQ ID NO:3);
 (ii) a homologous sequence having at least 70% identity to (a)(i); or 
 (iii) a fragment of (a)(i) or (a)(ii) which is at least 6 amino acids in length; and 
 
 (b) (i) pro-BNP (SEQ ID NO:4), BNP (SEQ ID NO:5), or NT-proBNP (SEQ ID NO:6);
 (ii) a homologous sequence having at least 70% identity to (b)(i); or 
 (iii) a fragment of (b)(i) or (b)(ii) which is at least 6 amino acids in length. under conditions to provide for expression of the polypeptide; and optionally 
 
 
 (II) recovering the expressed polypeptide. 
 
     
     
         11 . A process for producing a polypeptide agent according to  claim 5  which comprises chemical synthesis. 
     
     
         12 . A diagnostic kit comprising:
 (a) a bi or oligo specific first binding substance that is able to bind to both
 (I) (i) proANP (SEQ ID NO: 1), ANP (SEQ ID NO: 2), or NT-proANP (SEQ ID NO: 3);
 (ii) a homologous sequence having at least 70% identity to (i); or 
 (iii) a fragment of (i) or (ii) which is at least 6 amino acids in length; and 
 
 (II) (i) proBNP (SEQ ID NO: 4), BNP (SEQ ID NO: 5), or NT-proBNP (SEQ ID NO: 6);
 (ii) a homologous sequence having at least 70% identity to (i); or 
 (iii) a fragment of (i) or (ii) which is at least 6 amino acids in length; or 
 
   (b) said first binding substance and a fusion polypeptide agent comprising both:
 (I) proANP (SEQ ID NO:1), ANP (SEQ ID NO:2), or NT-proANP (SEQ ID NO:3); and 
 (II) proBNP (SEQ ID NO:4), BNP (SEQ ID NO:5), or NT-proBNP (SEQ ID NO:6), wherein said fusion polypeptide agent can be bound by said first binding substance, and optionally the binding substance and/or the agent is labelled. 
   
     
     
         13 . A kit according to  claim 12  wherein the first binding substance comprises
 (a) bi- or oligo-specific binding substance; 
 (b) a mixture of mono-specific binding substances; 
 (c) natriuretic receptor GC-A (SEQ ID NO: 33); 
 (d) homologous sequence having at least 70% identity to (c); 
 (e) a fragment of (c) or (d) which is at least 400 amino acids in length; 
 (f) an extracellular binding domain of the natriuretic receptor GC-A (SEQ ID NO: 34); 
 and/or 
 (g) is present on a solid support comprising a bi- or oligo-specific antibody, fragment or derivative thereof which is able to bind to both:
 (a) (i) proANP (SEQ ID NO: 1), ANP (SEQ ID NO: 2), or NT-proANP (SEQ ID NO: 3);
 (ii) a homologous sequence having at least 70% identity to (i); or 
 (iii) a fragment of (i) or (ii) which is at least 6 amino acids in length; and 
 
 (b) (i) proBNP (SEQ ID NO: 4), BNP (SEQ ID NO: 5), or NT-proBNP (SEQ ID NO: 6);
 (ii) a homologous sequence having at least 70% identity to (i); or 
 (iii) a fragment of (i) or (ii) which is at least 6 amino acids in length. 
 
 
 
     
     
         14 . A kit according to  claim 12  wherein the agent comprises
 (a) (i) proANP (SEQ ID NO: 1), ANP (SEQ ID NO: 2), or NT-proANP (SEQ ID NO: 3);
 (ii) a homologous sequence having at least 70% identity to (i); or 
 (iii) a fragment of (i) or (ii) which is at least 6 amino acids in length; and 
 
 (b) (i) proBNP (SEQ ID NO: 4), BNP (SEQ ID NO: 5), or NT-proBNP (SEQ ID NO: 6);
 (ii) a homologous sequence having at least 70% identity to (i); or 
 (iii) a fragment of (i) or (ii) which is at least 6 amino acids in length.