Recently, our group reported that a small interfering RNA (siRNA) targeting coagulation factor XII (siF12) leads to an unexpected prothrombotic response in a mouse model where venous thrombosis follows inhibition of endogenous anticoagulants. In this study, we aimed to clarify this unexpected response by evaluating the effects of this siF12 (here, siF12-A) on plasma coagulation through thrombin generation (TG). Besides a routine negative control siRNA (siNEG), we included extra siRNA controls: one siRNA similar to siF12-A except for positions 9-11 of the siRNA that are replaced with its complementary base pairs (siF12-AC9/11), and a second siRNA against F12 (siF12-B). Three days after injection, a significant increase in TG peak height was observed solely for animals injected with siF12-A and siF12-AC9/11, which is considered prothrombotic. As this change in coagulation was unrelated to FXII we conclude that it was off-target. For siRNA studies we now recommend to include mismatch siRNA controls, such as the C9/11 mismatch control used in this study, and to consider plasma coagulation in off-target analysis.

中文翻译：

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使用“ C9 / 11错配”对照siRNA可以揭示针对靶向小鼠凝血因子XII的siRNA的凝血相关序列的脱靶效应。

最近，我们的小组报告说，靶向凝血因子XII（siF12）的小干扰RNA（siRNA）会在小鼠血栓形成继而抑制内源性抗凝剂后导致意外的血栓形成反应。在这项研究中，我们旨在通过评估此siF12（此处为siF12-A）对凝血酶生成（TG）对血浆凝结的影响来澄清这种意外反应。除了常规的阴性对照siRNA（siNEG），我们还包括其他siRNA对照：一种与siF12-A相似的siRNA，但siRNA的9-11位被其互补碱基对（siF12-AC9 / 11）取代，针对F12的第二个siRNA（siF12-B）。注射后三天，仅观察到注射有siF12-A和siF12-AC9 / 11的动物的TG峰高显着增加，这被认为是血栓形成的。由于凝血的这种变化与FXII无关，因此我们得出结论，它没有达到目标。对于siRNA研究，我们现在建议包括错配siRNA对照，例如本研究中使用的C9 / 11错配对照，并在脱靶分析中考虑血浆凝结。