Variants of human glycoprotein hormone alpha chain: compositions and uses thereof

Abstract

The present invention provides human glycoprotein hormone (hGPH) α-chain splice variants, including isolated nucleic acids encoding these variants and the encoded amino acid sequences, as well as antibodies, antisense oligonucleotides, expression vectors and host cells comprising these sequences. The present invention further discloses the use of these sequences in the diagnosis, prevention and treatment of symptoms, diseases and disorders related to glycoprotein hormones.

1 . An isolated polynucleotide comprising a nucleic acid sequence selected from the group consisting of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5 and SEQ ID NO:7.  
     
     
         2 . The polynucleotide according to  claim 1  encoding a polypeptide having at least one hGPH α-chain activity.  
     
     
         3 . An isolated polynucleotide comprising a nucleic acid sequence encoding a polypeptide having at least one hGPH α-chain activity having a sequence set forth in any one of SEQ ID NOs: 2, 4, 6 or fragments, variants or analogs thereof.  
     
     
         4 . The isolated polynucleotide according to  claim 3  wherein the nucleic acid sequence encodes a polypeptide having a sequence set forth in any one of SEQ ID NOs: 2, 4, 6 or fragments, variants or analogs thereof.  
     
     
         5 . The isolated polynucleotide according to  claim 3  wherein the nucleic acid sequence encodes a polypeptide comprising contiguous amino acids having at least 80% homology to amino acids 31 to 60 set forth in SEQ ID NO:2.  
     
     
         6 . The isolated polynucleotide according to  claim 3  wherein the nucleic acid sequence encodes a polypeptide comprising contiguous amino acids having at least 90% homology to amino acids 31 to 60 set forth in SEQ ID NO:2.  
     
     
         7 . The isolated polynucleotide according to  claim 3  wherein the nucleic acid sequence encodes a polypeptide comprising contiguous amino acids having at least 95% homology to amino acids 31 to 60 set forth in SEQ ID NO:2.  
     
     
         8 . The isolated polynucleotide according to  claim 3  wherein the nucleic acid sequence encodes a polypeptide comprising contiguous amino acids having 100% homology to amino acids 31 to 60 set forth in SEQ ID NO:2.  
     
     
         9 . The isolated polynucleotide according to  claim 3  wherein the nucleic acid sequence encodes a polypeptide comprising contiguous amino acids having at least 80% homology to amino acids 93-101 set forth in SEQ ID NO:4.  
     
     
         10 . The isolated polynucleotide according to  claim 3  wherein the nucleic acid sequence encodes a polypeptide comprising contiguous amino acids having at least 90% homology to amino acids 93-101 set forth in SEQ ID NO:4.  
     
     
         11 . The isolated polynucleotide according to  claim 3  wherein the nucleic acid sequence encodes a polypeptide comprising contiguous amino acids having at least 95% homology to amino acids 93-101 set forth in SEQ ID NO:4.  
     
     
         12 . The isolated polynucleotide according to  claim 3  wherein the nucleic acid sequence encodes a polypeptide comprising contiguous amino acids having 100% homology to amino acids 93-101 set forth in SEQ ID NO:4.  
     
     
         13 . The isolated polynucleotide according to  claim 3  wherein the nucleic acid sequence encodes a polypeptide comprising contiguous amino acids having at least 80% homology to amino acids 30-72 set forth in SEQ ID NO:6.  
     
     
         14 . The isolated polynucleotide according to  claim 3  wherein the nucleic acid sequence encodes a polypeptide comprising contiguous amino acids having at least 90% homology to amino acids 30-72 set forth in SEQ ID NO:6.  
     
     
         15 . The isolated polynucleotide according to  claim 3  wherein the nucleic acid sequence encodes a polypeptide comprising contiguous amino acids having at least 95% homology to amino acids 30-72 set forth in SEQ ID NO:6.  
     
     
         16 . The isolated polynucleotide according to  claim 3  wherein the nucleic acid sequence encodes a polypeptide comprising contiguous amino acids having 100% homology to amino acids 30-72 set forth in SEQ ID NO:6.  
     
     
         17 . An isolated polynucleotide comprising a nucleic acid sequence complementary to the nucleic acid sequence according to  claim 3 .  
     
     
         18 . An isolated polynucleotide comprising a nucleic acid sequence that hybridizes under stringent conditions to a polynucleotide sequence according to  claim 1 .  
     
     
         19 . A polypeptide having at least one hGPH α-chain activity having an amino acid sequence set forth in any one of SEQ ID NOs: 2, 4, 6 or fragments, analogs or derivatives thereof.  
     
     
         20 . The polypeptide according to  claim 19 , comprising contiguous amino acids having at least 80% homology to amino acids 31 to 60 set forth in SEQ ID NO:2.  
     
     
         21 . The polypeptide according to  claim 19 , comprising contiguous amino acids having at least 80% homology to amino acids 93 to 101 set forth in SEQ ID NO:4.  
     
     
         22 . The polypeptide according to  claim 19 , comprising contiguous amino acids having at least 80% homology to amino acids 30 to 72 set forth in SEQ ID NO:6.  
     
     
         23 . A polypeptide having at least one hGPH α-chain activity comprising (a) a first portion having an amino acid sequence being at least about 90% homologous to positions 1-29 of the amino acid sequence of WT hGPH α-chain (SEQ ID NO:9), which are also amino acids 1-29 set forth in SEQ ID NO:2; a second portion, having an amino acid sequence being at least about 80% homologous to an amino acid sequence ETGFHHVAQAALKLLSSSNPPTKASQSARIT (positions 30-60 of SEQ ID NO:2); and a third portion, having an amino acid sequence being at least about 90% homologous to positions 30-116 of the amino acid sequence of WT hGPH α-chain (SEQ ID NO:9), which are also amino acids 61-147 set forth in SEQ ID NO:2; wherein the first, the second and the third portions are contiguous and in sequential order, or (b) a first portion having an amino acid sequence being at least about 90% homologous to positions 1-92 of the amino acid sequence of WT hGPH α-chain (SEQ ID NO:9), which are also amino acids 1-92 set forth in SEQ ID NO:4; and a second portion, having an amino acid sequence being at least about 80% homologous to an amino acid sequence RTSRSPEAF (positions 93-101 of SEQ ID NO:4); wherein the first and the second portions are contiguous and in sequential order, or (c) a first portion having an amino acid sequence being at least about 90% homologous to positions 1-29 of the amino acid sequence of WT hGPH α-chain (SEQ ID NO:9), which are also amino acids 1-29 set forth in SEQ ID NO:6; and a second portion, having an amino acid sequence being at least about 80% homologous to an amino acid sequence ETGFHHVAQAALKLLSSSNPPTKASQSARITGVSYCAGLIDSF (positions 30-72 of SEQ ID NO:6); wherein the first and the second portions are contiguous and in sequential order.  
     
     
         24 . An insertion portion of a polypeptide having at least one hGPH α-chain activity having an amino acid sequence set forth in SEQ ID NO:2, wherein the insertion comprises an amino acid sequence being at least about 80% homologous to an amino acid sequence ETGFHHVAQAALKLLSSSNPPTKASQSARIT (positions 30-60 of SEQ ID NO:2).  
     
     
         25 . A tail portion of a polypeptide having at least one hGPH α-chain activity having an amino acid sequence (a) as set forth in SEQ ID NO:4, wherein the tail portion comprises an amino acid sequence at least about 80% homologous to an amino acid sequence RTSRSPEAF (positions 93-101 of SEQ ID NO:4), or (b) as set forth in SEQ ID NO:6, wherein the tail portion comprises an amino acid sequence at least about 80% homologous to an amino acid sequence ETGFHHVAQAALKLLSSSNPPTKASQSARITGVSYCAGLIDSF (positions 30-72 of SEQ ID NO:6).  
     
     
         26 . A bridge portion of SEQ ID NO:2, comprising a polypeptide having a length “n”, wherein “n” is at least about 10 amino acids in length, wherein (a) at least two amino acids comprise QE, having a structure as follows (numbering according to SEQ ID NO:2): a sequence starting from any of amino acid numbers 29−x to 29; and ending at any of amino acid numbers 30+((n−2)−x), in which x varies from 0 to n−2; wherein the total amino acid number does not exceed 147, or (b) at least two amino acids comprise TD, having a structure as follows (numbering according to SEQ ID NO:2): a sequence starting from any of amino acid numbers 60−x to 60; and ending at any of amino acid numbers 61+((n−2)−x), in which x varies from 0 to n−2; wherein the total amino acid number does not exceed 147, or (c) at least two amino acids comprise VR, having a structure as follows (numbering according to SEQ ID NO:4): a sequence starting from any of amino acid numbers 92−x to 92; and ending at any of amino acid numbers 93+((n−2)−x), in which x varies from 0 to n−2; wherein the total amino acid number does not exceed 101, or (d) at least two amino acids comprise QE, having a structure as follows (numbering according to SEQ ID NO:6): a sequence starting from any of amino acid numbers 29−x to 29; and ending at any of amino acid numbers 30+((n−2)−x), in which x varies from 0 to n−2; wherein the total amino acid number does not exceed 72.  
     
     
         27 . The bridge portion according to  claim 26 , wherein n is between 4 to 9 amino acids in length or is at least about 20 amino acids in length.  
     
     
         28 . The bridge portion according to  claim 26 , comprising a homologous polypeptide having at least 80% homology to said polypeptide.  
     
     
         29 . An antibody which binds to at least one epitope of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NOS: 2, 4, 6 or fragments, variants or analogs thereof.  
     
     
         30 . An antibody-antigen complex formed with antibodies and epitopes according to  claim 29 .  
     
     
         31 . A method for detecting a polynucleotide which encodes hGPH α-chain in a biological sample comprising: 
 a. hybridizing the polynucleotide sequence according to  claim 17  to nucleic acid material of a biological sample, thereby forming a hybridization complex; and    b. detecting the hybridization complex, wherein the presence of the complex correlates with the presence of a polynucleotide encoding hGPH α-chain in the biological sample.    
     
     
         32 . A pharmaceutical composition comprising as an active ingredient a polynucleotide sequence according to  claim 1  and a pharmaceutically acceptable diluent or carrier.  
     
     
         33 . A pharmaceutical composition comprising as an active ingredient a polypeptide according to  claim 19  and a pharmaceutically acceptable diluent or carrier.  
     
     
         34 . A method for preventing, treating or ameliorating a GPH related diseases or disorder, comprising administering to a subject in need thereof a pharmaceutical composition comprising as an active ingredient a polypeptide according to  claim 19 .  
     
     
         35 . A method for diagnosing a GPH related conditions, diseases or disorders, comprising detecting specific expression of a polynucleotide according to  claim 1 .  
     
     
         36 . The method according to  claim 35 , wherein detection of the expression is performed with a NAT-based assay.  
     
     
         37 . The method according to  claim 35 , wherein detection of the expression is performed with an immunoassay.  
     
     
         38 . The method according to  claim 35  wherein the condition, disease or disorder is selected from the group consisting of pregnancy (normal and ectopic); trophoblastic tumors; tumors of the testes; other cancers including breast, colorectal, lung and ovary cancer; benign conditions including cirrhosis, duodenal ulcer, and inflammatory bowel disease; fertility and infertility; pituitary diseases; general hormonal imbalance; precocious puberty; amenorrhea; hypogonadism; ovarian cysts; thyroid diseases, including hyperthyroidism and hypothyroidism; and under active thyroid, specifically in newborns.  
     
     
         39 . A method for targeting a therapeutic agent to a cell bearing a GPH receptor, comprising administering to a subject in need thereof a pharmaceutical composition comprising as an active ingredient an hGPH-α-chain according to  claim 19  conjugated to the therapeutic agent.  
     
     
         40 . A method for targeting a diagnostic agent to a cell bearing a GPH receptor, comprising administering to a subject in need thereof a pharmaceutical composition comprising as an active ingredient an hGPH-α-chain according to  claim 19  conjugated to the diagnostic agent.