Lna oligonucleotides and the treatment of cancer

Abstract

The present disclosure concerns LNA oligonucleotides having a (sub)sequence of the general formula 5′-( Me C x )(T x ) Me C x A s A s t s C s C s a s t s g s g s Me C x A x ( G x )(c)-3′, and preferably of the general formula 5′- Me C x T x Me C x A s a s t s c s c s a s t s g s g s Me C x A x G x c-3′, wherein capital letters designate an LNA nucleotide analogue selected from β-D-oxy-LNA, β-D-thio-LNA, β-D-amino-LNA and α-L-oxy-LNA, small letters designate a deoxynucleotide, and underline designates either an LNA nucleotide analogue as defined above or a deoxynucleotide. Such LNA oligonucleotides exhibit surprisingly good properties with respect to inhibition of the expression of Survivin by means of an anti-sense mechanism, and thereby lead to reduction or inhibition of tumour development in vivo. The LNA oligonucleotides are superior to other LNA oligonucletides targeting Surviving mRNA measured by functional read outs such as apoptosis induction and proliferation inhibition, and is potent in down-regulating Survivin mRNA and protein in transfected cancer cell lines, and induce apoptosis in combination with Taxol superior compared to other LNA oligonucleotides.

1 . A liquid pharmaceutical composition comprising an LNA oligonucleotide in an aqueous carrier, said LNA oligonucleotide having a total of 12-20 nucleotides/LNA nucleotide analogues and comprising the following (sub)sequence: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO. 28) 
                 
                     
                   5'-( Me   C   x )(T x ) Me C x A s   A   s t s c s c s a s t s g s g s   Me C x A x ( G   x )(c)-3'. 
                 
             
                
                
               
            
           
         
         wherein capital letters designate an LNA nucleotide analogue selected from β-D-oxy-LNA, β-D-thio-LNA, β-D-amino-LNA and α-L-oxy-LNA, small letters designate a deoxynucleotide, underline designates either an LNA nucleotide analogue as defined above or a deoxynucleotide, subscript “s” designates a phosphorothioate link between neighbouring nucleotides/LNA nucleotide analogues, and subscript “x” designates either a phosphorothioate link or a phosphorodiester link between neighbouring nucleotides/LNA nucleotide analogues; and where the nucleotide units in bracket each represent an optional unit, and 
         said aqueous carrier comprising a buffer for keeping the pH in the range of 4.0-8.5, and having an ionic strength of 20-2000 mM. 
       
     
     
         2 . The composition according to  claim 1 , wherein the LNA oligonucleotide has a total of 16-20 nucleotides/LNA nucleotide analogues and comprises the following (sub)sequence: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO. 1) 
                 
                     
                   5'- Me   C   x T x   Me C x A s a s t s c s c s a s t s g s g s   Me C x A x   G   x c-3'. 
                 
             
                
                
               
            
           
         
       
     
     
         3 . The composition according to  claim 1 , wherein all LNA nucleotide analogues are β-D-oxy-LNA. 
     
     
         4 . The composition according to  claim 1 , wherein the LNA oligonucleotide is a compound selected from the group consisting of SEQ ID NOS. 2, 3, 4, 5, 6, 7, 8, 9, and 10. 
     
     
         5 . The composition according to  claim 4 , wherein the LNA oligonucleotide is the compound with SEQ ID NO. 2. 
     
     
         6 . A liquid pharmaceutical composition comprising a conjugate in an aqueous carrier, said conjugate consisting of an LNA oligonucleotide as defined in  claim 1  and at least one non-nucleotide/non-polynucleotide moiety covalently attached to said oligonucleotide; and
 said aqueous carrier comprising a buffer for keeping the pH in the range of 4.0-8.5, and having an ionic strength of 20-2000 mM. 
 
     
     
         7 . The composition according to  claim 6 , wherein the LNA oligonucleotide has a total of 16-20 nucleotides/LNA nucleotide analogues and comprises the following (sub)sequence: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO. 1) 
                 
                     
                   5'- Me   C   x T x   Me C x A s a s t s c s c s a s t s g s g s   Me C x A x   G   x c-3'. 
                 
             
                
                
               
            
           
         
       
     
     
         8 . A pharmaceutical composition comprising at least one taxane compound and an LNA oligonucleotide as defined in  claim 1  in a pharmaceutically acceptable carrier, and wherein the weight ratio between the taxane compound(s) and the LNA oligonucleotide in said composition is in the range of 50:1 to 1:25. 
     
     
         9 . The composition according to  claim 8 , wherein the LNA oligonucleotide has a total of 16-20 nucleotides/LNA nucleotide analogues and comprises the following (sub)sequence: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO. 1) 
                 
                     
                   5'- Me   C   x T x   Me C x A s a s t s c s c s a s t s g s g s   Me C x A x   G   x c-3'. 
                 
             
                
                
               
            
           
         
       
     
     
         10 . A pharmaceutical composition comprising at least one taxane compound and a conjugate in a pharmaceutically acceptable carrier, said conjugate consisting of an LNA oligonucleotide as defined in  claim 1  and at least one non-nucleotide/non-polynucleotide moiety covalently attached to said oligonucleotide, and
 wherein the weight ratio between the taxane compound(s) and the LNA oligonucleotide part of the conjugate in said composition is in the range of 50:1 to 1:25. 
 
     
     
         11 . The composition according to  claim 10 , wherein the LNA oligonucleotide has a total of 16-20 nucleotides/LNA nucleotide analogues and comprises the following (sub)sequence: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO. 1) 
                 
                     
                   5'- Me   C   x T x   Me C x A s a s t s c s c s a s t s g s g s   Me C x A x   G   x c-3'. 
                 
             
                
                
               
            
           
         
       
     
     
         12 . A method of treating a mammal, in particular a human, suffering from or susceptible to a cancer disease, the method comprising the step of administering to the mammal one or more therapeutically effective doses of a first pharmaceutical composition comprising an LNA oligonucleotide as define in  claim 1 . 
     
     
         13 . The method according to  claim 12 , wherein the LNA oligonucleotide has a total of 16-20 nucleotides/LNA nucleotide analogues and comprises the following (sub)sequence: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO. 1) 
                 
                     
                   5'- Me   C   x T x   Me C x A s a s t s c s c s a s t s g s g s   Me C x A x   G   x c-3'. 
                 
             
                
                
               
            
           
         
       
     
     
         14 . A method of treating a mammal, in particular a human, suffering from or susceptible to a cancer disease, the method comprising the step of administering to the mammal one or more therapeutically effective doses of a first pharmaceutical composition comprising a conjugate, said conjugate consisting of an LNA oligonucleotide as defined in  claim 1  and at least one non-nucleotide/non-polynucleotide moiety covalently attached to said oligonucleotide. 
     
     
         15 . The method according to  claim 14 , wherein the LNA oligonucleotide has a total of 16-20 nucleotides/LNA nucleotide analogues and comprises the following (sub)sequence: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO. 1) 
                 
                     
                   5'- Me   C   x T x   Me C x A s a s t s c s c s a s t s g s g s   Me C x A x   G   x c-3'. 
                 
             
                
                
               
            
           
         
       
     
     
         16 . The method according to  claim 13 , wherein all LNA nucleotide analogues are β-D-oxy-LNA. 
     
     
         17 . The method according to  claim 13 , wherein the LNA oligonucleotide is a compound selected from the group consisting of SEQ ID NOS. 2, 3, 4, 5, 6, 7, 8, 9, and 10. 
     
     
         18 . The method according to  claim 17 , wherein the LNA oligonucleotide is the compound with SEQ ID NO. 2. 
     
     
         19 . The method according to  claim 13 , wherein the cancer disease is selected from the group consisting of acute myelocytic leukemia, diffuse B-cell lymphoma, acute lymphocytic leukemia, hepatic cancer, renal cancer, urinary tract cancer, and colorectal cancer. 
     
     
         20 . A kit comprising
 (a) a first component containing one or more injectable solution doses of an LNA oligonucleotide as defined in  claim 1 , and   (b) a second component containing one or more injectable solutions of one or more taxane compounds; and   wherein the weight ratio between the at least one taxane compound in one solution of the second component and the at least one LNA oligonucleotide in one solution dose of the first component is in the range of 50:1 to 1:25, and/or   wherein the injectable solution doses of the LNA oligonucleotide comprise a buffer for keeping the pH in the range of 4.0-8.5, and having an ionic strength of 20-2000 mM.   
     
     
         21 . The kit according to  claim 20 , wherein the LNA oligonucleotide has a total of 16-20 nucleotides/LNA nucleotide analogues and comprises the following (sub)sequence: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO. 1) 
                 
                     
                   5'- Me   C   x T x   Me C x A s a s t s c s c s a s t s g s g s   Me C x A x   G   x c-3'. 
                 
             
                
                
               
            
           
         
       
     
     
         22 . A kit comprising
 (a) a first component containing an LNA oligonucleotide as defined in  claim 1  in solid form, and   (b) a second component containing saline or a buffer solution adapted for reconstitution of said LNA oligonucleotide.   
     
     
         23 . The kit according to  claim 22 , wherein the LNA oligonucleotide has a total of 16-20 nucleotides/LNA nucleotide analogues and comprises the following (sub)sequence: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO. 1) 
                 
                     
                   5'- Me   C   x T x   Me C x A s a s t s c s c s a s t s g s g s   Me C x A x   G   x c-3'.