英国剑桥大学MRC分子生物学实验室Philipp Holliger团队宣布他们开发出一种模块化的XNA核酶，可在生理条件下剪切长而有序的RNA，以用于等位基因特异性（allele-specific）基因沉默。2022年9月5日出版的《自然-化学》发表了这项成果。

核酸类催化剂（核酶、脱氧核酶和XNA核酶）切割靶标(m)RNA具有很高的特异性，但在临床应用中效果有限。在该研究中，团队通过体外进化和工程化改造，开发了一种高度特异性、模块化的具有RNA内切酶活性的XNA核酶——FR6_1，该XNA核酶由2’-脱氧-2’-氟代-β-D-阿拉伯糖核酸（FANA）组成。

FR6_1克服了之前的脱氧核酶和XNA核酶的活性限制，可以在生理Mg²⁺浓度下重新靶向切割高度结构化的全长（>5kb）的BRAF和KRAS的mRNA，且对肿瘤相关突变（BRAF V600E和KRAS G12D）具有等位基因特异性沉默。硫代磷酸酯（PS）-FANA修饰进一步增加了FR6_1的生物稳定性，使其在具有G12D等位基因的培养人腺癌细胞中，通过体内XNA核酶剪切活性实现对KRAS mRNA的快速降解。

研究人员表示，这些结果为开发基于FANA或其他XNA化学的改良版基因沉默剂提供了一个新起点。

附：英文原文

Title: A modular XNAzyme cleaves long, structured RNAs under physiological conditions and enables allele-specific gene silencing

Author: Taylor, Alexander I., Wan, Christopher J. K., Donde, Maria J., Peak-Chew, Sew-Yeu, Holliger, Philipp

Issue&Volume: 2022-09-05

Abstract: Nucleic-acid catalysts (ribozymes, DNA- and XNAzymes) cleave target (m)RNAs with high specificity but have shown limited efficacy in clinical applications. Here we report on the in vitro evolution and engineering of a highly specific modular RNA endonuclease XNAzyme, FR6_1, composed of 2′-deoxy-2′-fluoro-β-D-arabino nucleic acid (FANA). FR6_1 overcomes the activity limitations of previous DNA- and XNAzymes and can be retargeted to cleave highly structured full-length (>5kb) BRAF and KRAS mRNAs at physiological Mg2+ concentrations with allelic selectivity for tumour-associated (BRAF V600E and KRAS G12D) mutations. Phosphorothioate-FANA modification enhances FR6_1 biostability and enables rapid KRAS mRNA knockdown in cultured human adenocarcinoma cells with a G12D-allele-specific component provided by in vivo XNAzyme cleavage activity. These results provide a starting point for the development of improved gene-silencing agents based on FANA or other XNA chemistries.

DOI: 10.1038/s41557-022-01021-z

Source: https://www.nature.com/articles/s41557-022-01021-z