Patent ID: 7750144

Claim:
A method of producing a dsRNAi agent having decreased off-target silencing activity, the method comprising: (a) identifying an off-target silencing activity mediated by a sense strand of a first dsRNAi agent, wherein the first dsRNAi agent directs cleavage by a RISC complex at a phosphodiester bond within a desired target mRNA; and (b) synthesizing a substituted dsRNAi agent comprising one or more substituted base pairs with respect to the first dsRNAi agent, wherein the substituted dsRNAi agent comprises a sense and an antisense strand, each strand having a 5′ end and a 3′ end, wherein the substituted dsRNAi agent directs cleavage by the RISC complex at the same phosphodiester bond within the desired target mRNA, wherein the one or more substituted base pairs are within about 5 base pairs from the 5′ end of the antisense strand (AS 5′) and the 3′ end of the sense strand (S 3′) and are selected from the group consisting of a mismatched base pair, a wobble base pair, a base pair comprising a rare nucleotide and a base pair comprising a base-modified nucleotide, such that the sense strand of the substituted dsRNAi agent is less effective than the sense strand of the first dsRNAi agent at entering the RISC complex; such that the substituted dsRNAi agent has decreased off-target silencing activity relative to the off-target silencing activity of the first dsRNAi agent.