| RNA ImmunoGenic Assay: A Method to Detect Immunogenicity of in vitro Transcribed mRNA in Human Whole Blood
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Author:
Date:
2020-12-20
[Abstract] The mRNA therapeutics is a new class of medicine to treat many various diseases. However, in vitro transcribed (IVT) mRNA triggers immune responses due to recognition by human endosomal and cytoplasmic RNA sensors, but incorporation of modified nucleosides have been shown to reduce such responses. Therefore, an assay signifying important aspects of the human immune system is still required. Here, we present a simple ex vivo method called ‘RNA ImmunoGenic Assay’ to measure immunogenicity of IVT-mRNAs in human whole blood. Chemically modified and unmodified mRNA ...
[摘要] [摘要] mRNA疗法是治疗多种疾病的新型药物。然而，我Ñ体外转录（IVT）的mRNA触发由于人类内体和细胞质RNA传感器，但是修饰的核苷的掺入识别的免疫应答已经示出吨ö减少此类反应。牛逼herefore ，测定标志着重要的环节人体免疫系统仍然需要。这里，我们提出一个简单的离体称为“RNA方法免疫ģ ENIC测定”测量的IVT-mRNA的免疫原性小号在人全血。将化学修饰和未修饰的mRNA与转染试剂（TransIT ）复合，并在人全血中共同孵育。特异性细胞因子测定（TNF- α ，INF- α ，INF- γ ，IL-6和IL-12p70的），使用的ELISA。进行qPCR分析以揭示特异性免疫途径的激活。所述RNA免疫ģ ENIC测定提供小号的简单且快速的方法来检测供体特异性-针对mRNA的治疗剂的免疫应答。
图形摘要：
RNA免疫基因测定的示意图
[背景] mRNA治疗是基因治疗的重要一类（Sahin等，2014 ；Antony等，2015 ...
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| Analysis of in vivo Interaction between RNA Binding Proteins and Their RNA Targets by UV Cross-linking and Immunoprecipitation (CLIP) Method
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Author:
Date:
2017-05-20
[Abstract] RNA metabolism is tightly controlled across different tissues and developmental stages, and its dysregulation is one of the molecular hallmarks of cancer. Through direct binding to specific sequence element(s), RNA binding proteins (RBPs) play a pivotal role in co- and post-transcriptional RNA regulatory events. We have recently demonstrated that, in pancreatic cancer cells, acquisition of a drug resistant (DR)-phenotype relied on upregulation of the polypyrimidine tract binding protein (PTBP1), which in turn is recruited to the pyruvate kinase pre-mRNA and favors splicing of the oncogenic ...
[摘要] RNA代谢在不同的组织和发育阶段被严格控制，其失调是癌症的分子特征之一。 通过直接结合特定的序列元件，RNA结合蛋白（RBP）在共转录和转录后调控事件中起关键作用。 我们最近证实，在胰腺癌细胞中，获得耐药（DR） - 表型取决于多聚嘧啶区结合蛋白（PTBP1）的上调，其又被引入丙酮酸激酶前mRNA并有利于剪接 致癌性PKM2变体。 在这里，我们描述了紫外（UV）光交联和免疫沉淀（CLIP）方法的逐步方案，以确定RBP与贴壁人细胞系中其目标RNA的特定区域的直接结合。
背景 在细胞核中转录时，新生的RNA立即与被称为RNA结合蛋白（RBP）的反式因子立即组装。这些因子直接与RNA分子中特定的顺式调控序列相互作用，从而形成核糖核蛋白（RNP）复合物（Dreyfuss et al。，2002; ...
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