New polypeptides and peptides, nucleic acids coding for them, and their use in the field of tumor therapy, inflammation or immunology

Abstract

The invention relates: to a polypeptide containing in its peptidic chain the amino acid sequence of 311 amino acids of FIG. 3, or a fragment of this sequence, with said fragment being such that it is liable to produce antibodies capable of forming a complex with the amino acid sequence of FIG. 3, or an amino acid sequence having a percentage of homology of at least 50%, preferably 75%, and advantageously 90% with the amino acid sequence of FIG. 3, and to pharmaceutical compositions containing, as active substance, at least one of the polypeptides of the invention or of the antagonists of the polypeptides of the invention as antitumor compounds, or antiinflammatory compounds or as growth activators of T-cells and B-cells, as bone repair compounds as inducer of immunosuppressive cells, as inhibitors of anti-colony stimulating factor; or as trypoanocidal agents; or part of the polypeptides of the invention, capable of binding to the above-defined receptor.

1 . Polypeptide containing in its peptidic chain 
 the amino acid sequence of 311 amino acids of FIG. 3,    or a fragment of this sequence, with said fragment being such that it is liable to produce antibodies capable of forming a complex with the amino acid sequence of FIG. 3,    or an amino acid sequence having a percentage of homology of at least 50%, preferably 75%, and advantageously 90% with the amino acid sequence of FIG. 3,    or a sequence liable to form a complex with antibodies raised 
 against the amino acid sequence of FIG. 3  
 or against pep1(m)  
 or against pep2(m)  
 or against pep3(m).  
   
     
     
         2 . Polypeptide according to  claim 1  containing in its peptidic chain 
 the amino acid sequence of 311 amino acids of FIG. 2,  
 or a fragment of this sequence, with said fragment being such that it is liable to produce antibodies capable of forming a complex with the amino acid sequence of FIG. 2  
 or an amino acid sequence having a percentage of homology of at least 50%, preferably 75%, and advantageously 90% with the amino acid sequence of FIG. 2,  
 or a sequence liable to form a complex with antibodies raised 
 against the amino acid sequence of FIG. 2  
 or against pep1(h)  
 or against pep2(h)  
 or against pep3(h).  
 
 
     
     
         3 . Polypeptide according to  claim 1 , characterized by the fact that it is constituted by the sequence represented in FIG. 3, extending from the extremity constituted by amino acid at position (1) to the extremity constituted by amino acid at position (311), or that it contains at 
 least one of the following peptides: 
 Cys-Ser-Trp-Lys-Gly-Ser-Gly-Leu-Thr  
 Val-Glu-Trp-Met-Tyr-Pro-Thr-Gly-Ala-Leu-Ile-Val-Asn-Leu-Arg-Pro-Asn-Thr-Phe-Ser-Pro-Ala  
 Asp-Ser-Ser-Gly-Ala-Asn-Ile-Tyr-Leu-Glu-Lys-Thr-Gly-Glu-Leu-Arg-Leu-Leu-Val  
 Leu-Glu-Glii-Gly-Gly-Leu-Phe-Val-Glu-Ala-Thr-Pro-Gln-Gln-Asp-Ile  
 Arg-Arg-Thr-Thr-Gly-Phe-Gln-Tyr-Glu-Leu  
 Leu-Ser-Ala-Pro-Cys-Arg-Pro-Cys-Ser-Asp-Thr-Glu-Val-Leu-Leu-Ala  
 Arg-Gln-Lys-Ser-Arg-Val-Phe  
 Cys-Gly-Val-Arg-Pro-Gly-His- Gly  
 Phe-Leu-Phe-Thr-Gly-His  
 Leu-Gly-Cys-Ala-Pro-Arg-Phe  
 Asp-Phe-Gln-Arg-Met-Tyr-Arg  
   
     
     
         4 . Polypeptide according to  claim 2 , characterized by the fact that it is constituted by the sequence represented in FIG. 2, extending from the extremity constituted by amino acid at position (1) to the extremity constituted by amino acid at position (311).  
     
     
         5 . Muteins deriving from anyone of the polypeptides of  claims 1  to  4 , containing modifications consisting of substitutions and/or deletions and/or additions of one or several amino acids, insofar that said modifications do not alter the hydropathicity profile such as defined by Kyte and Doolitle (1982) and such as represented in FIG. 6 a  and  6   b.    
     
     
         6 . Polypeptide according to any one of  claims 1  to  5  presenting at least one of the following properties: 
 promoting the incorporation of  3 H-thymidine in rat femur pre- and osteoblasts cells, in 3-week-old mice thymocytes, in splenic cells or lymph node cells, advantageously upon co-stimulation with IFN-γ,  
 promoting the incorporation of  3 H-thymidine in thymocytes, advantageously upon co-stimulation with IL4,  
 promoting the activation or cytotoxicity or mobility of LAK cells.  
 promoting the recruitment of suppressive peritoneal exudate cells upon injection in vivo,  
 promoting the generation of immunocompetent lymph node cells, preferantially after ConA, PHA or LPS induction, upon in vivo intrafootpath injection,  
 exerting a trypanocidal or trypanolytical activity on the pleomorph bloodstream trypanosomes in vitro.  
 
     
     
         7 . Amino acid sequence constituted by a polypeptide according to anyone of  claims 1  to  6 , and a protein or a heterologous sequence with respect to said polypeptide, with said protein or heterologous sequence comprising, for instance, from about 10 to about 100 amino acids.  
     
     
         8 . Nucleic acid characterized by the fact that it comprises or is constituted by: 
 a nucleotide sequence which is effectively homologous with any of the nucleotide sequence coding for the polypeptides according to any one of  claims 1  to  6 ,    a nucleotide sequence liable to hybridize with anyone of the nucleotide sequence coding for the polypeptides according to anyone of  claims 1  to  6 ,    or a nucleotide sequence which, further to translation or further to transcription and to translation, leads to anyone of the polypetide according to  claims 1  to  6 ,    or the complementary sequences of the above-mentioned nucleotide sequences.    
     
     
         9 . Nucleic acid according to  claim 8 , which comprises or is constituted by: 
 a nucleotide sequence which is effectively homologous with any of the nucleotide sequences of FIG. 1,    a nucleotide sequence liable to hybridize with the complementary strand of the nucleotide sequence of FIG. 1,    the nucleotide sequence of FIG. 1,    the complementary sequences of the above-mentioned sequences.    
     
     
         10 . Nucleic acid according to  claim 8 , which comprises or is constituted by: 
 a nucleotide sequence which is effectively homologous with any of the nucleotide sequences of FIG. 2,    a nucleotide sequence liable to hybridize with the complementary strand of the nucleotide sequence of FIG. 2,    the nucleotide sequence of FIG. 2,    the complementary sequences of the above-mentioned sequences.    
     
     
         11 . Nucleic acid according to  claim 8 , which comprises or is constituted by: 
 a nucleotide sequence which is effectively homologous with any of the nucleotide sequences of FIG. 3,    a nucleotide sequence liable to hybridize with the complementary strand of the nucleotide sequence of FIG. 3,    the nucleotide sequence of FIG. 3,    the complementary sequences of the above-mentioned sequences.    
     
     
         12 . Recombinant nucleic acid containing at least one of the nucleotide sequences of anyone of  claims 8  to  11  combined with or inserted in a heterologous nucleic acid.  
     
     
         13 . Recombinant vector particularly for cloning and/or expression, comprising a vector sequence, notably of the type plasmid, cosmid, phage, or virus DNA and a recombinant nucleic acid according to anyone of  claims 8  to  12 , inserted in one of the nonessential sites for its replication.  
     
     
         14 . Recombinant vector according to  claim 13 , containing necessary elements to promote the expression in a cellular host of polypeptides coded by nucleic acids according to anyone of  claims 8  to  12 , inserted in said vector and notably a promoter recognized by the RNA polymerase of the cellular host, particularly an inducible promoter and possibly a sequence coding for transcription, termination, and possibly a signal sequence and/or an anchoring sequence.  
     
     
         15 . Recombinant vector according to  claim 13 , containing the elements enabling the expression of a nucleotide sequence according to anyone of  claims 8  to  11 , as a mature protein or as part of a fusion protein, the fusion moiety which is used in the fusion protein being part of a nonhomologous protein (such as mTNF) chosen to optimize the expression of a fusion protein.  
     
     
         16 . Cellular host chosen from among bacteria such as  E. coli  or chosen from among eukaryotic organisms, such as COS cells, baculovirus or vaccinia virus, which is transformed by a recombinant vector according to anyone of  claims 13  to  15  and containing the regulatory elements enabling the expression of the nucleotide sequence coding for the polypeptide according to anyone of  claims 1  to  7  in this host.  
     
     
         17 . Expression product of a nucleic acid expressed by a transformed cellular host according to  claim 16 .  
     
     
         18 . Antibody characterized by being specifically directed against a polypeptide according to anyone of  claim 1  to  7 .  
     
     
         19 . Nucleotidic probe, hybridizing with any of the nucleic sequences according to anyone of  claims 8  to  11 .  
     
     
         20 . Process for preparing a recombinant polypeptide according to anyone of  claims 1  to  7 , comprising the following steps: 
 the culture in an appropriate medium of a cellular host which has previously been transformed by an appropriate vector containing a nucleic acid according to anyone of  claims 8  to  11 , and  
 the recovery of the polypeptide produced by the above-said transformed cellular host from the above-said culture medium or from the cellular host.  
 
     
     
         21 . Process for detecting the capacity of a molecule to behave as a ligand or as a receptor with respect to a polypeptide according to anyone of  claims 1  to  6 , characterized by: 
 contacting the molecule with a cellular host which has previously been transformed by a vector itself modified by an insert coding for said polypeptide, this host carrying on its surface one or several specific sites of this polypeptide, possibly after induction of the expresion of this insert, with said contacting being carried out under conditions enabling binding between at least one of these specific sites and said molecule to be formed if it happens to present an affinity for said polypeptide,  
 detecting the possible formation of a complex of the type ligand-polypeptide or receptor-polypeptide.  
 
     
     
         22 . Immunogenic composition containing, as active substance, at least one of the polypetides of FIG. 2, or anyone of the peptides pep1(h), pep2(h) or pep3(h).  
     
     
         23 . Pharmaceutical compositions containing, as active substance, at least anyone of the polypeptides of  claims 1  to  6  or of the antagonists of the polypeptides above-defined as antitumor compounds, or antiinflammatory compounds, as growth activators of T-cells and B-cells, as bone repair compounds, as inducer of immunosuppressive cells as inhibitors of anti-colony stimulating factor; or as trypanocidal agents; or part of the polypeptides of the invention, capable of binding to the receptor defined above.  
     
     
         24 . Antisense oligonucleotides or antisense mRNA derived from the nucleotide sequences of anyone of  claims 8  to  11 .  
     
     
         25 . Nonhuman mammalian transgenic animal which contains, in its genome, a nucleic acid sequence according to anyone of  claims 8  to  11 , and which can be used to study the effects of pharmacological compositions and to prepare different cell types from transgenic animals which express the nucleotide sequence according to anyone of  claims 8  to  11  in a constitutive or inducible way.  
     
     
         26 . “Knock-out” nonhuman mammalian transgenic animal in which the natural gene, effectively homologous with any one of the nucleotide sequences according to anyone of  claims 8  to  11 , is rendered nonfunctional, for instance by homologous recombination, with said animal being suitable for the study of the possible loss of functions or the possible restoration effects caused by the reintroduction into the animals of the polypeptides according to anyone of  claims 8  to  11 .