Organic compounds

Abstract

The present invention relates to isolated polypeptides which comprise an amino acid sequence consisting of a mutated functional Abl kinase domain, said mutated functional kinase domain being resistant to inhibition of its tyrosine kinase activity by N-[4-methyl-3-(4-pyridin-3-yl-pyrimidin-2-ylamino)-phenyl]-4-(4-methyl-piperazin-1-ylmethyl)-benzamide or a salt thereof, to the use of such polypeptides to screen for compounds which inhibit the tyrosine kinase activity of such polypeptides, to nucleic acid molecules encoding such polypeptides, to recombinant vectors and host cells comprising such nucleic acid molecules and to the use of such nucleic acid molecules in the production of such polypeptides for use in screening for compounds which inhibit the tyrosine kinase activity of such polypeptides.

1 . An isolated polypeptide which comprises a mutated functional Abl kinase domain that is resistant to inhibition of its tyrosine kinase activity by N-[4-methyl-3-(4-pyridin-3-yl-pyrimidin-2-ylamino)-phenyl]-4-(4-methyl-piperazin-1-ylmethyl)-benzamide or a salt thereof. 
     
     
         2 . An isolated polypeptide according to  claim 1 , wherein the mutated functional Abl kinase domain comprises the amino acid sequence of the native human Abl kinase domain or an essentially similar sequence thereof in which at least one amino acid is replaced by another amino acid. 
     
     
         3 . An isolated polypeptide according to  claim 2 , wherein in the amino acid sequence of the native human Abl kinase domain or an essentially similar sequence thereof at least one amino acid selected from Leu248, Glu255, Lys271, Glu286, Met290, Thr315, Tyr320, Asn322, Glu373, His375 and Ala380 is replaced by another amino acid. 
     
     
         4 . An isolated polypeptide according to  claim 3 , wherein in the amino acid sequence of the native human Abl kinase domain or an essentially similar sequence thereof at least one amino acid selected from Leu248, Glu255, Lys271, Glu286, Met290, Tyr320, Asn322, Glu373, His375 and Ala380 is replaced by another amino acid. 
     
     
         5 . An isolated polypeptide according to  claim 4 , wherein in the amino acid sequence of the native human Abl kinase domain or an essentially similar sequence thereof at least one amino acid selected from Leu248, Lys271, Glu286, Met290, Tyr320, Asn322, Glu373, His375 and Ala380 is replaced by another amino acid. 
     
     
         6 . An isolated polypeptide according to  claim 3 , wherein in the amino acid sequence of the native human Abl kinase domain or an essentially similar sequence thereof at least one amino acid selected from Glu255, Thr315 and Ala380 is replaced by another amino acid. 
     
     
         7 . An isolated polypeptide according to  claim 6 , wherein in the amino acid sequence of the native human Abl kinase domain or an essentially similar sequence thereof at least one amino acid selected from Glu255 and Ala380 is replaced by another amino acid. 
     
     
         8 . An isolated polypeptide according to  claim 2 , wherein in the amino acid sequence of the native human Abl kinase domain or an essentially similar sequence thereof a single amino acid is replaced by another amino acid. 
     
     
         9 . An isolated polypeptide according to  claim 7 , wherein in the amino acid sequence of the native human Abl kinase domain or an essentially similar sequence thereof Glu255 is replaced by another amino acid. 
     
     
         10 . An isolated polypeptide according to  claim 3 , wherein the amino acid sequence of the native human Abl kinase domain or an essentially similar sequence thereof contains at least one amino acid mutation selected from Glu255Val, Glu255Lys, Thr315Val, Thr315Leu, Thr315Met, Thr315Gln, Thr315Phe and Ala380Thr. 
     
     
         11 . An isolated polypeptide according to  claim 10 , wherein the amino acid sequence of the native human Abl kinase domain or an essentially similar sequence thereof contains at least one amino acid mutation selected from Glu255Val, Thr315Val, Thr315Leu, Thr315Met, Thr315Gln, Thr315Phe and Ala380Thr. 
     
     
         12 . An isolated polypeptide according to  claim 11 , wherein the amino acid sequence of the native human Abl kinase domain or an essentially similar sequence thereof contains at least one amino acid mutation selected from Thr315Leu, Thr315Met, Thr315Gln and Thr315Phe. 
     
     
         13 . An isolated polypeptide according to  claim 10 , wherein the amino acid sequence of the native human Abl kinase domain or an essentially similar sequence thereof contains a single amino acid mutation. 
     
     
         14 . An isolated polypeptide according to  claim 11 , wherein the amino acid sequence of the native human Abl kinase domain or an essentially similar sequence thereof contains the amino acid mutation Glu255Val. 
     
     
         15 . An isolated polypeptide according to  claim 2 , wherein the amino acid sequence of the native human Abl kinase domain consists of amino acids 229-500 of SEQ ID NO:2. 
     
     
         16 . An isolated polypeptide according to  claim 2 , which is a Bcr-Abl tyrosine kinase. 
     
     
         17 . Use of a polypeptide according to  claim 2  to screen for compounds which inhibit the tyrosine kinase activity of said polypeptide. 
     
     
         18 . An isolated nucleic acid molecule comprising a nucleotide sequence that encodes a polypeptide according to  claim 2 . 
     
     
         19 . Use of a nucleic acid molecule according to  claim 18  in the production of a polypeptide according to  claim 2  for use in screening for compounds which inhibit the tyrosine kinase activity of said polypeptide. 
     
     
         20 . A recombinant vector comprising a nucleic acid molecule according to  claim 18 . 
     
     
         21 . A recombinant vector according to  claim 20 , which is a recombinant expression vector. 
     
     
         22 . A host cell comprising a recombinant vector according to  claim 20 .