Variants of protein kinases

Abstract

The present invention concerns nucleic acid sequences and amino acid sequences of dominant negative variants of kinases, i.e. of sequences which inhibit activity of kinases in a dominant manner. The invention also concerns pharmaceutical compositions and detection methods using these sequences.

1 . An isolated nucleic acid sequence, selected from the group consisting of: 
 (i) the nucleic acid sequence depicted in anyone of SEQ ID NO: 1 to SEQ ID NO: 91;    (ii) nucleic acid sequences having at least 90% identity with the sequence of (i) with the proviso that each sequence is different than the original kinase nucleic acid sequence from which the sequences of (i) have been varied by alternative splicing; and    (iii) fragments of (i) or (ii) of at least 20 b.p., provided that said fragment contains a sequence which is not present, as a continuous stretch of nucleotides, in the original nucleic acid sequence from which the sequences of (i) have been varied by alternative splicing.    
     
     
         2 . An isolated nucleic acid sequence complementary to the nucleic acid sequence of  claim 1 .  
     
     
         3 . An isolated nucleic acid sequence coding for an amino acid sequences selected from the group consisting of: 
 (i) an amino acid sequence coded by the isolated nucleic acid sequence of alternative splice variants of  claim 1;     (ii) homologues of the amino acid sequences of (i) in which one or more amino acids has been added, deleted, replaced or chemically modified in the region, or adjacent to the region, where the amino acid sequences differs from the original amino acid sequence, coded by the original nucleic acid sequence from which the variant has been varied by alternative splicing; and    (iii) amino acid sequences as depicted in SEQ ID NO: 92 to SEQ ID NO: 182.    
     
     
         4 . A purified antibody which binds specifically to any of the amino acid sequences of  claim 3 .  
     
     
         5 . A purified antibody which binds to an amino acid sequence which is present only in the alternative splice variant depicted in the amino acid sequences of  claim 3 , but is not present in the original amino sequence.  
     
     
         6 . A purified antibody which binds to an amino acid sequence present in the original amino acid sequence, which amino acid sequence is not present in the amino acid sequences of  claim 3 .  
     
     
         7 . An expression vector comprising any one of the nucleic acid sequences of  claim 1  and control elements for the expression of the nucleic acid sequence in a suitable host.  
     
     
         8 . An expression vector comprising any one of the nucleic acid sequences of  claim 2 , and control elements for the expression of the nucleic acid sequences in a suitable host.  
     
     
         9 . A host cell transfected by the expression vector of  claim 8 .  
     
     
         10 . A host cell transfected by the expression vector of  claim 9 .  
     
     
         11 . A pharmaceutical composition comprising a pharmaceutically acceptable carrier and as an active ingredient an agent selected from the group consisting of: 
 (i) any one of the nucleic acid sequences of  claim 2;     (ii) the expression vector of  claim 8;  and    (iii) the purified antibody of  claim 4 .    
     
     
         12 . A pharmaceutical composition according to  claim 11 , for treatment of diseases which can be ameliorated or cured by decreasing the level of any one of the amino acid sequences of  claim 3 .  
     
     
         13 . A method for detecting a DNKV nucleic acid sequence in a biological sample, comprising the steps of: 
 (a) hybridizing to nucleic acid material of said biological sample any one of the nucleic acid sequences of  claim 1;  and    (b) detecting said hybridization complex;    
     
     
         14 . A method for detection of a nucleic acid sequence in a biological sample the method comprises: 
 (i) contacting the sample with probes for amplification of any one of SEQ ID NO: 1 to SEQ ID NO: 91;    (ii) proving reagents for amplification;    (iii) detecting the presence of amplified products,    said products indicating the presence of DNKV nucleic acid in the sample.    
     
     
         15 . A method for determining the level of DNKV nucleic acid sequences in a biological sample comprising the steps of: 
 (a) hybridizing to nucleic acid material of said biological sample any one of the nucleic acid sequences of  claim 1;  and    (b) determining the amount of hybridization complexes and normalizing said amount to provide the level of the variant nucleic acid sequences in the sample.    
     
     
         16 . A method for determining the ratio between the level of variant of the nucleic acid sequence in a first biological sample and the level of the original sequence from which the variant has been varied by alternative splicing in a second biological sample comprising: 
 (a) determining the level of the variant nucleic acid sequence in the first biological sample according to the method of  claim 15;     (b) determining the level of the original sequence in the second biological sample; and    (c) comprising the levels obtained in (a) and (b) to give said ratio.    
     
     
         17 . A method according to  claim 16 , wherein said first and said second biological samples are the same sample.  
     
     
         18 . A method according to  claim 13 , wherein the nucleic acid material of said biological sample are mRNA transcripts.  
     
     
         19 . A method according to  claim 13 , where the nucleic acid sequence is present in a nucleic acid chip.  
     
     
         20 . A method for detecting any one of the amino acid sequences of  claim 3  in a biological sample, comprising the steps of: 
 (a) contacting with said biological sample the antibody of  claim 4 , thereby forming an antibody-antigen complex; and    (b) detecting said antibody-antigen complex wherein the presence of said antibody-antigen complex correlates with the presence of the desired amino acid in said biological sample.    
     
     
         21 . A method for detecting the level of the amino acid sequence of any one of  claim 3  in a biological sample, comprising the steps of: 
 (a) contacting with said biological sample the antibody of  claim 4 , thereby forming an antibody-antigen complex; and    (b) detecting the amount of said antibody-antigen complex and normalizing said amount to provide the level of said amino acid sequence in the sample.    
     
     
         22 . A method for determining the ratio between the level of anyone of the amino acid sequence of  claim 3  present in a first biological sample and the level of the original amino acid sequences from which they were varied by alternative splicing, present in a second biological sample, the method comprising: 
 (a) determining the level of the amino acid sequences of  claim 3  into a first sample by the method of  claim 21;     (b) determining the level of the original amino acid sequence in the second sample; and    (c) comparing the level obtained in (a) and (b) to give said ratio.    
     
     
         23 . A method according to  claim 22 , wherein said first and said second biological samples are the same sample.  
     
     
         24 . A nucleic acid sequence according to  claim 1 , present on a data carrier.  
     
     
         25 . A data carrier comprising substantially all the nucleic acid sequences depicted in NV — 1 to NV — 91.  
     
     
         26 . A data carrier according to  claim 25  further comprising the amino acid sequences coded by anyone of NY — 92 to NY — 182.  
     
     
         27 . A data carrier according to  claim 26 , for use in an analysis of a nucleic acid sequence in which said nucleic acid sequence is compared to any of the sequences present on the data carrier.  
     
     
         28 . A software data combination comprising: 
 a nucleic acid sequence search and comparison software and data comprising substantially sequences depicted in NV — 1 to NY — 91; said combination when loaded into a computer can execute in the computer a search where a nucleotide sequence is compared to the sequences comprising said data.    
     
     
         29 . An inhibitor of kinase activity comprising any one of the isolated nucleic acid sequences of  claim 1 .  
     
     
         30 . An inhibitor of kinase activity comprising any one of the isolated nucleic acid sequences of  claim 2.