Protein present at the surface of hematopoietic stem cells of the lymphoid line and of nk cells, and uses thereof

Abstract

The invention concerns a protein present at the surface of hematopoietic stem cells of the lymphoid cell line and mature NK cells, the corresponding isolated cDNA sequence and their uses as marker of said cells and for preparing antibodies directed against said protein. The invention also concerns the uses of said antibodies for selecting cells expressing at their surface said protein. Said isolated protein has a structure (a) comprising an extracellular domain located between positions 21 and 152, five transmembrane domains located between positions 153 and 295 and a cytoplasmic domain located between positions 296 and 350, with reference to the sequence SEQ ID NO:2; an apparent molecular weight of about 36 to 38 kDa, and its precursor, which comprises in N-terminal of structure (a), a signal sequence of 20 amino acids, is selected in the group consisting of the protein of SEQ ID NO:2, and the proteins having an amino acid sequence having at least 70% identity or at least 85% similarity and preferably at least 95% identity or at least 99% similarity with the sequence SEQ ID NO:2.

1 . An antibody,characterized in that it is directed against a surface protein having a structure comprising an extracellular domain, five transmembrane domains and a cytoplasmic domain, located, respectively, between positions 21 and 152, 153 and 295, and 296 and 350, with reference to the sequence SEQ ID NO:2, which surface protein has an apparent molecular weight of approximately 36-38 kD, is expressed at the surface of all lymphoid progenitor cells and all mature NK cells, has an amino acid sequence selected from the group consisting of the sequence SEQ ID NO:2 and the sequences which have at least 70% identity or at least 85% similarity with said sequence SEQ ID NO:2, over the entire said sequence SEQ ID NO:2, excluding the sequences which have the accession numbers SWALL EBI Q9Y3U9, GENESEQ W 63688, and GENESEQ W 74904 and the 321 amino acid C-terminal fragment of the sequence GENESEQ W 74904.  
     
     
         2 . An antibody, characterized in that it is directed against a fragment of the surface protein as claimed in  claim 1  which is selected from the group consisting of: 
 the fragments defined by the sequences SEQ ID NO:4, 6, 7, 8, 9 and 24,    the fragments of at least 7 amino acids of the sequence corresponding to that located between positions 153 and 295 with reference to the sequence SEQ ID NO:2, and    the fragments of at least 7 amino acids of the sequence corresponding to that located between positions 296 and 350 with reference to the sequence SEQ ID NO:2.    
     
     
         3 . The antibody as claimed in  claim 2 , characterized in that it is directed against the fragment of sequence SEQ ID NO:7.  
     
     
         4 . The antibody as claimed in  claim 1 , characterized in that it is selected from monoclonal antibodies and polyclonal antibodies.  
     
     
         5 . An antibody, characterized in that it is directed against a human cell surface protein wherein said protein: 
 consists of the sequence SEQ ID NO:25,    is derived from a human precursor of the sequence SEQ ID NO:2 that comprises a 20 amino acid N-terminal signal sequence,    is expressed at the surface of human lymphoid progenitor cells and mature NK cells,    has an apparent molecular weight of approximately 36-38 kDa, and    has a structure comprising an extracellular domain, five transmembrane domains and a cytoplasmic domain, located, respectively, between positions 21 and 152, 153 and 295, and 296 and 350, with reference to the sequence SEQ ID NO:2.    
     
     
         6 . An isolated and purified cDNA, which encodes a surface protein having a structure comprising an extracellular domain, five transmembrane domains and a cytoplasmic domain, located, respectively, between positions 21 and 152, 153 and 295, and 296 and 350, with reference to the sequence SEQ ID NO:2, which surface protein has an apparent molecular weight of approximately 36-38 kD, is expressed at the surface of all lymphoid progenitor cells and all mature NK cells, has an amino acid sequence from the group consisting of the sequence SEQ ID NO:2 and the sequences which have a least 70% identity or at least 85% similarity with said sequence SEQ ID NO:2, over the entire said sequence SEQ ID NO:2, excluding the sequences which have the accession numbers SWALL EBI Q9Y3U9, GENESEQ W 63688, and GENESEQ W 74904 and the 321 amino acid C-terminal fragment of the sequence GENESEQ W 74904, or a fragment of this protein.  
     
     
         7 . The isolated and purified cDNA as claimed in  claim 6 , which is selected from the group consisting of the sequences SEQ ID NO:1, 3, 5, 10, 11, 12, 13 and 23 and SEQ ID NO:16-20.  
     
     
         8 . A method for selectively removing the cells expressing a surface protein from a biological sample comprising a heterogeneous population of cells, which method comprises: 
 bringing said biological sample into contact with an antibody as claimed in  claim 1 , and    removing the cells which bind to said antibody.    
     
     
         9 . The removal method as claimed in  claim 8 , wherein said antibody is attached to a solid support.  
     
     
         10 . A method for detecting and/or quantifying and/or isolating the NK cells and/or the lymphoid cell progenitors in a heterogeneous population of cells, said method comprising: 
 bringing a sample comprising a heterogeneous population of cells into contact with an antibody as claimed in  claim 1 , and    detecting the formation of the cell-antibody complex.    
     
     
         11 . The method as claimed in  claim 10 , wherein, when said sample consists of bone marrow, of umbilical cord blood, of fetal liver or of any other fetal or adult hematopoietic tissue, the method also comprises bringing the cells which have formed a complex with said antibody into contact with an antibody specific for pre-B cells (anti-cD19 antibody or anti-CD10 antibody) and/or for pre-T cells (anti-CD3 antibody, anti-CD4 antibody or anti-CD8 antibody).  
     
     
         12 . A medicinal product for the treatment of autoimmune diseases containing the antibody as claimed in  claim 1 .  
     
     
         13 . A method for preparing a product which can be transplanted, which comprises using the cells of bone marrow, of umbilical cord blood, of fetal liver or of any other fetal or adult hematopoietic tissue, depleted of NK cells and/or of lymphoid progenitors, and obtained using the method as claimed in  claim 11 .  
     
     
         14 . A method for diagnosing malignant and benign hemopathies, characterized in that it comprises detecting a nucleic acid sequence as claimed in  claim 6  in a biological sample.  
     
     
         15 . A kit for detecting and/or quantifying and/or isolating the NK cells and/or the lymphoid cell progenitors (pre-B, pre-T and pre-NK), characterized in that it comprises, besides useful amounts of buffers suitable for carrying out said detection, suitable doses of an antibody as claimed in  claim 1 , and suitable doses of a reagent for revealing the cell-antibody complex possibly formed.  
     
     
         16 . An isolated and purified cDNA, which encodes a fragment of the surface protein as defined in  claim 6  which is selected from the group consisting of: 
 the fragments defined by the sequences SEQ ID NO:4, 6, 7, 8, 9 and 24,    the fragments of at least 7 amino acids of the sequence corresponding to that located between positions 153 and 295 with reference to the sequence SEQ ID NO:2, and    the fragments of at least 7 amino acids of the sequence corresponding to that located between positions 296 and 350 with reference to the sequence SEQ ID NO:2.    
     
     
         17 . A medicinal product for the treatment of autoimmune diseases containing the antibody as claimed in  claim 2 .  
     
     
         18 . A reagent for detecting NK cells which includes fragments of a surface protein as defined in  claim 1 , comprising at least 7 consecutive amino acids of the extracellular domain.