RNAi modulation of the BCR-ABL fusion gene and uses thereof

Abstract

The invention relates to compositions and methods for modulating the expression of Bcr-Abl, and more particularly to the down-regulation of Bcr-Abl mRNA and Bcr-Abl protein levels by oligonucleotides via RNA interference, e.g., chemically modified oligonucleotides.

1 . An iRNA agent comprising an antisense strand which differs at no more than 1, 2, or 3 nucleotides from a sequence of 15 or more contiguous nucleotides of one of: SEQ ID No. 2, 6, 12, 14, 16,or20.  
     
     
         2 . The iRNA agent of  claim 1 , wherein the antisense strand has at least 15 contiguous nucleotides of one of: SEQ ID No. 2, 6, 12, 14, 16, or 20.  
     
     
         3 . The iRNA agent of  claim 1 , further comprising a sense strand which differs at no more than 1, 2, or 3 nucleotides from a sequence of 15 or more contiguous nucleotides of one of: SEQ ID No. 1, 5, 11, 13, 15, or 19.  
     
     
         4 . The iRNA agent of  claim 1 , further comprising a sense strand having a sequence of 15 contiguous nucleotides of one of: SEQ ID No. 1, 5, 11, 13, 15, or 19.  
     
     
         5 . The iRNA agent of  claim 1 , wherein the agent is one of: agent no. 3, 4 or 5.  
     
     
         6 . The iRNA agent of  claim 1 , wherein the sense strand has at least 15 contiguous nucleotides of the sense sequences of agent no. 3, 4 or 5, and the antisense strand has at least 15 contiguous nucleotides of the antisense sequences of agent no. 3, 4 or 5, respectively.  
     
     
         7 . The iRNA agent of  claim 1 , wherein the iRNA agent significantly reduces the amount of BCR-ABL fusion protein levels present in cultured human cells after incubation with the agent compared to cells which have not been incubated with the agent, wherein the cells are preferably 32Dp210/e14a2, 32Dp210-T315I,  32 Dp210-H396P, 32Dp210/e13a2, 32Dp190/e1a2, M07p210/e14a2, K562, MEG-01, or SUP-B15, or have been isolated from a leukemic patient.  
     
     
         8 . The iRNA agent of  claim 1 , wherein the antisense RNA strand is 30 or fewer nucleotides in length, and the duplex region of the iRNA agent is 15-30 nucleotide pairs in length.  
     
     
         9 . The iRNA agent of  claim 1 , comprising a modification that causes the iRNA agent to have increased stability in a biological sample.  
     
     
         10 . The iRNA agent of  claim 9 , comprising a phosphorothioate or a 2′-modified nucleotide.  
     
     
         11 . The iRNA agent of  claim 10 , comprising at least one 5′-uridine-adenine-3′ (5′-ua-3′) dinucleotide wherein the uridine is a 2′-modified nucleotide; at least one 5′-uridine-guanine-3′ (5′-ug-3′) dinucleotide, wherein the 5′-uridine is a 2′-modified nucleotide; at least one 5′-cytidine-adenine-3′ (5′-ca-3′) dinucleotide, wherein the 5′-cytidine is a 2′-modified nucleotide; or at least one 5′-uridine-uridine-3′ (5′-uu-3′) dinucleotide, wherein the 5′-uridine is a 2′-modified nucleotide.  
     
     
         12 . The iRNA agent of  claim 10 , wherein the 2′-modified nucleotide comprises a modification selected from the group consisting of: 2′-deoxy, 2′-deoxy-2′-fluoro, 2′-O-methyl, 2′-O-methoxyethyl (2′-O-MOE), 2′-O-aminopropyl (2′-O-AP), 2′-O-dimethylaminoethyl (2′-O-DMAOE), 2′-O-dimethylaminopropyl (2′-O-DMAP), 2′-O-dimethylaminoethyloxyethyl (2′-O-DMAEOE), and 2′-O-N-methylacetamido (2′-O-NMA).  
     
     
         13 . The iRNA agent of any one of the preceding claims, comprising a nucleotide overhang having 1 to 4 unpaired nucleotides.  
     
     
         14 . The iRNA agent of  claim 13 , wherein the nucleotide overhang has 2 or 3 unpaired nucleotides.  
     
     
         15 . The iRNA agent of  claim 13 , wherein the nucleotide overhang is at the 3′-end of the antisense strand of the iRNA agent.  
     
     
         16 . The iRNA agent of any one of the preceding claims, comprising a ligand.  
     
     
         17 . The iRNA agent of  claim 16 , wherein the ligand is conjugated to the 3′-end of the sense strand of the iRNA agent.  
     
     
         18 . A pharmaceutical composition, comprising: 
 an iRNA agent of any one of the preceding claims, and    a pharmaceutically acceptable carrier.    
     
     
         19 . A method of reducing the amount of BCR-ABL RNA in a cell of a subject, comprising contacting the cell with an iRNA agent of any one of the preceding claims.  
     
     
         20 . A method of making an iRNA agent of  claim 1 , the method comprising the synthesis of the iRNA agent, wherein the sense and antisense strands comprise at least one modification that stabilizes the iRNA agent against nucleolytic degradation.  
     
     
         21 . A method of treating a human having or at risk for developing a proliferative disorder comprising administering an iRNA agent of  claim 1 , and preferably wherein the iRNA agent comprises a sense strand having at least 15 contiguous nucleotides of the sense strand sequences of the iRNA agents, agent numbers, 1, 2, 3, 4, 5, or 6, and an antisense strand having at least 15 contiguous nucleotides of the antisense sequences of the iRNA agents, agent numbers, 1, 2, 3, 4, 5, or 6.  
     
     
         22 . The method of  claim 21 , wherein the sense strand of the iRNA agent comprises at least 15 contiguous nucleotides of the sense sequence of agent no. 3, 4 or 5, and the antisense strand of the iRNA agent comprises at least 15 contiguous nucleotides of the antisense sequence of agent no. 3, 4 or 5, respectively.  
     
     
         23 . The method of  claim 22 , wherein the sense strand of the iRNA agent comprises at least 15 contiguous nucleotides of the sense sequence of agent no. 3, or 4, and the antisense strand of the iRNA agent comprises at least 15 contiguous nucleotides of the antisense sequence of agent no. 3, or 4, respectively, and the proliferative disorder is a Chronic Myeloid Leukemia.  
     
     
         24 . The method of  claim 22 , wherein the sense strand of the iRNA agent comprises at least 15 contiguous nucleotides of the sense sequence of agent no. 5, and the antisense strand of the iRNA agent comprises at least 15 contiguous nucleotides of the antisense sequence of agent no. 5, and the proliferative disorder is a Ph+acute lymphoblastic leukemia.  
     
     
         25 . The method of  claim 21 , wherein the iRNA agent is administered in an amount sufficient to reduce the level of BCR-ABL RNA in a cell or tissue of the human.