Intronic Polyadenylation in Acquired Cancer Drug Resistance Circumvented by Utilizing CRISPR/Cas9 with Homology-Directed Repair: The Tale of Human DNA Topoisomerase IIα.
Intronic Polyadenylation in Acquired Cancer Drug Resistance Circumvented by Utilizing CRISPR/Cas9 with Homology-Directed Repair: The Tale of Human DNA Topoisomerase IIα.
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DNA拓扑异构酶IIα（170 kDa，TOP 2 α/170）通过产生短暂的双链DNA断裂来解决核酸拓扑缠结。TOP2α抑制剂/毒物稳定TOP2α-DNA共价复合物，导致持续的DNA损伤，并经常用于治疗各种癌症。对这些化疗药物的获得性耐药通常与TOP 2 α/170表达水平降低相关。研究表明，TOP2α/170的减少是由一种称为内含子多聚腺苷酸化（IPA）的替代多聚腺苷酸化引起的。作为IPA的结果，变体TOP2α mRNA转录物已被表征，其导致具有改变的生物活性的C-末端截短的TOP2α同种型的翻译。本文讨论了一个例子，其中通过利用外显子/内含子边界的CRISPR/Cas9特异性基因编辑，通过同源定向修复（HDR）减少TOP2α IPA，实现了对获得性TOP2α介导的耐药性的规避。这些结果说明了CRISPR/Cas9/HDR影响与异常IPA相关的耐药性的治疗潜力。内含子多聚腺苷酸化（IPA）通过促进转录组/蛋白质组改变在恶性转化、发展、进展和癌症化疗耐药性中起关键作用。DNA拓扑异构酶IIα（170 kDa，TOP 2 α/170）是抗癌药物的既定临床靶点，其疗效因耐药性而受损，通常与核TOP 2 α/170水平降低相关。在对TOP2α靶向药物具有获得性耐药性且TOP2α/170表达降低的白血病细胞系中，已报告了由于IPA导致C末端截短亚型翻译的变体TOP2α mRNA转录物，其核质分布发生改变或与野生型TOP2α/170发生异源二聚化。本文概述了调节前体mRNA加工和替代多聚腺苷酸化的各种机制，以及当剪接位点本质上较弱或可能突变时，通过同源定向修复（HDR）利用CRISPR/Cas9特异性基因编辑来减少IPA。在依托泊苷耐药的人白血病K562细胞中，讨论了TOP2α外显子19/内含子19剪接位点编辑的具体情况，作为规避获得性TOP2α介导的耐药的易处理策略。该实施例支持异常IPA在对TOP2α靶向药物的获得性耐药性中的重要性。此外，这些结果证明了CRISPR/Cas9/HDR影响与异常剪接/聚腺苷酸化相关的耐药性的治疗潜力。
DNA topoisomerase IIα (170 kDa, TOP2α/170) resolves nucleic acid topological entanglements by generating transient double-strand DNA breaks. TOP2α inhibitors/poisons stabilize TOP2α-DNA covalent complexes resulting in persistent DNA damage and are frequently utilized to treat a variety of cancers. Acquired resistance to these chemotherapeutic agents is often associated with decreased TOP2α/170 expression levels. Studies have demonstrated that a reduction in TOP2α/170 results from a type of alternative polyadenylation designated intronic polyadenylation (IPA). As a consequence of IPA, variant TOP2α mRNA transcripts have been characterized that have resulted in the translation of C-terminal truncated TOP2α isoforms with altered biological activities. In this paper, an example is discussed where circumvention of acquired TOP2α-mediated drug resistance was achieved by utilizing CRISPR/Cas9 specific gene editing of an exon/intron boundary through homology directed repair (HDR) to reduce TOP2α IPA. These results illustrate the therapeutic potential of CRISPR/Cas9/HDR to impact drug resistance associated with aberrant IPA. Intronic polyadenylation (IPA) plays a critical role in malignant transformation, development, progression, and cancer chemoresistance by contributing to transcriptome/proteome alterations. DNA topoisomerase IIα (170 kDa, TOP2α/170) is an established clinical target for anticancer agents whose efficacy is compromised by drug resistance often associated with a reduction of nuclear TOP2α/170 levels. In leukemia cell lines with acquired resistance to TOP2α-targeted drugs and reduced TOP2α/170 expression, variant TOP2α mRNA transcripts have been reported due to IPA that resulted in the translation of C-terminal truncated isoforms with altered nuclear-cytoplasmic distribution or heterodimerization with wild-type TOP2α/170. This review provides an overview of the various mechanisms regulating pre-mRNA processing and alternative polyadenylation, as well as the utilization of CRISPR/Cas9 specific gene editing through homology directed repair (HDR) to decrease IPA when splice sites are intrinsically weak or potentially mutated. The specific case of TOP2α exon 19/intron 19 splice site editing is discussed in etoposide-resistant human leukemia K562 cells as a tractable strategy to circumvent acquired TOP2α-mediated drug resistance. This example supports the importance of aberrant IPA in acquired drug resistance to TOP2α-targeted drugs. In addition, these results demonstrate the therapeutic potential of CRISPR/Cas9/HDR to impact drug resistance associated with aberrant splicing/polyadenylation.
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影响因子：
10.5
作者：
Chan SL;Huppertz I;Yao C;Weng L;Moresco JJ;Yates JR 3rd;Ule J;Manley JL;Shi Y
通讯作者：
Shi Y
影响因子：
10.5
作者：
Boutz PL;Bhutkar A;Sharp PA
通讯作者：
Sharp PA
DOI：
10.1038/nrm.2017.27
发表时间：
2017-07
期刊：
Nature reviews. Molecular cell biology
影响因子：
--
作者：
Baralle FE;Giudice J
通讯作者：
Giudice J
影响因子：
64.5
作者：
EARLY, P;ROGERS, J;HOOD, L
通讯作者：
HOOD, L
影响因子：
64.8
作者：
Dong, Ken C.;Berger, James M.
通讯作者：
Berger, James M.