Off-target effect

Compound optimization, to screen a series of therapeutic diug candidates to find the compounds that are most specific for the target protein and those that cause unintended effects, i.e. improved understanding of the molecular mode of action including structure-activity relationships for on-target versus off-target effects... 

Eedorov Y, Anderson EM, Birmingham A, Reynolds A, KarpUow J, Robinson K, Leake D, Marshall WS, Khvorova A (2006) Off-target effects by siRNA can induce toxic phenotype. RNA 12 1188-1196... 

An analysis of more than 130 preclinical candidates that had attrited during further development showed the failure of the chemotype approach (i.e. that a compound of the same/similar chemotype will have similar risks of attrition and that a structurally diverse chemotype will offer the best approach to minimize attrition risk) and 2D structure-based methods to be able to effectively differentiate compounds [29]. Thus, the risk of failing or succeeding in development is not related to being of the same chemotype , and differentiation by this method may not be the most effective way dangers are both that a valuable series/chemotype could be discarded because of one bad result and that a structurally different compound may actually have similar off-target effects (e.g. due to the decoration versus the scaffold). 

Notes Analysis (Joost and Methner, Genome Biol., 3(11) research 0063.1-0063.16,2002. http //genomebiology.com/ 2002/3/11 /research/0063). Profiling is performed in two phases. The first phase consists of a panel of 25 GPCR representing the families most commonly associated with off-target effects. Once a compound shows activity in one of the assays from the first set, full dose response is run on receptors within the same target family to assess selectivity (Phase 2). 

Taken together, siRNAs can induce pronounced off-target effects and therefore it is important to design siRNAs and perform siRNA-based experiments in a way that reduces off- and increases on-target activities. 

A number of modified nucleotides have been tested and described in siRNA design. These are mostly modifications of the 2 OH group of the ribose. By the incorporation of chemically modified nucleotides into siRNAs, the on-target efficiency of the siRNAs can be increased (42 4). On the other hand, different types of siRNA off-target effects can be reduced by the use of chemically modified nucleotides immunostimulatory effects (reviewed in (20)) as well as sequence-dependent miRNA-like off-target effects (26, 44,45) and passenger strand incorporation (46, 47). In addition, chemical modifications can be used to improve the cellular delivery of siRNAs in living animals and are important tools to enhance the serum stability of siRNAs (48). 

Because of the intrinsic property of siRNAs to induce off-target effects in several ways as described in the introduction, we would like to underline the importance of minimizing siRNA off-target activity. We will focus mainly on siRNA activity towards modulation of off-target transcript levels. For an excellent review about siRNA-mediated off-target effects in general and for unspecific immune responses in particular, the reader is referred to reference (20) and Table 2. 

Sequence-dependent off-target effects Comparison of siRNA effects Comparison of knockdown efficiencies and resulting phenotypes of sequence-independent siRNAs against the same target... 

Since the described strategies to reduce off-target effects provide no guarantee that siRNA off-target activity can be neglected, it is essential to control all effects observed in RNAi experiments for specificity. The most simple way to do so is to use several siRNAs (at least... 

Jackson AL, Linsley PS (2010) Recognizing and avoiding siRNA off-target effects for target identification and therapeutic application. Nat Rev Drug Discov 9 57-67... 

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