CRISPR-based DNA methylation editing of NNT rescues the cisplatin resistance of lung cancer cells by reducing autophagy
CRISPR-based DNA methylation editing of NNT rescues the cisplatin resistance of lung cancer cells by reducing autophagy
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基于 CRISPR 的 NNT DNA 甲基化编辑通过减少自噬拯救肺癌细胞的顺铂耐药性
DOI：
10.1007/s00204-022-03404-0
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发表时间：
2022-11
影响因子：
6.1
通讯作者：
Daochuan Li
中科院分区：
文献类型：
--
作者：
Chi Xu;Shuyun Jiang;Xingyu Ma;Zeyong Jiang;Youguang Pan;Xiang Li;Liying Zhang;Hao Zhou;Shen Chen;Xiumei Xing;Liping Chen;Wenfan Fu;Qing Wang;Wen Chen;Daochuan Li
顺铂被推荐作为治疗晚期非小细胞肺癌（NSCLC）的一线化疗药物，但获得性耐药大大限制了其临床疗效。最近，DNA甲基化已被确定为化疗耐药性的重要因素。然而，顺铂耐药的确切DNA甲基化调控机制仍不清楚。在此，我们发现，在顺铂耐药的A549（A549/DDP）细胞中，烟酰胺核苷酸转氢酶（NNT）被DNA超甲基化所沉默。此外，NNT的DNA甲基化与NSCLC患者的不良预后呈正相关。NNT在A549/DDP细胞中的过表达可降低其顺铂耐药性，并抑制其细胞增殖和克隆形成等肿瘤恶性程度。NNT增强A549/DDP细胞对顺铂的敏感性与其调节NADPH和ROS水平的作用无关，而主要是由于NNT抑制了A549/DDP细胞的保护性自噬。进一步的研究表明，NNT可降低NAD+水平，从而抑制SIRT 1，阻断自噬通路，而补充NAD+前体可拮抗SIRT 1的再激活作用。此外，通过CRISPR/dCas 9-Tet 1系统靶向去甲基化NNT CpG岛显著降低了其DNA甲基化水平，抑制了A549/DDP细胞的自噬和顺铂耐药性。因此，本研究发现了一个新的耐药靶基因NNT，该基因在肺癌细胞顺铂耐药中起重要作用。我们的研究结果还表明，基于CRISPR的NNT DNA甲基化编辑可能是肺癌顺铂耐药的潜在治疗方法。
Cisplatin is recommended as a first-line chemotherapeutic agent against advanced non-small cell lung cancer (NSCLC), but acquired resistance substantially limits its clinical efficacy. Recently, DNA methylation has been identified as an essential contributor to chemoresistance. However, the precise DNA methylation regulatory mechanism of cisplatin resistance remains unclear. Here, we found that nicotinamide nucleotide transhydrogenase (NNT) was silenced by DNA hypermethylation in cisplatin resistance A549 (A549/DDP) cells. Also, the DNA hypermethylation of NNT was positively correlated to poor prognosis in NSCLC patients. Overexpression of NNT in A549/DDP cells could reduce their cisplatin resistance, and also suppressed their tumor malignancy such as cell proliferation and clone formation. However, NNT enhanced sensitivity of A549/DDP cells to cisplatin had little to do with its function in mediating NADPH and ROS level, but was mainly because NNT could inhibit protective autophagy in A549/DDP cells. Further investigation revealed that NNT could decrease NAD+level, thereby inactivate SIRT1 and block the autophagy pathway, while re-activation of SIRT1 through NAD+precursor supplementation could antagonize this effect. In addition, targeted demethylation of NNT CpG island via CRISPR/dCas9-Tet1 system significantly reduced its DNA methylation level and inhibited the autophagy and cisplatin resistance in A549/DDP cells. Thus, our study found a novel chemoresistance target gene NNT, which played important roles in cisplatin resistance of lung cancer cells. Our findings also suggested that CRISPR-based DNA methylation editing of NNT could be a potential therapeutics method in cisplatin resistance of lung cancer.
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影响因子：
8.1
作者：
Santos LRB;Muller C;de Souza AH;Takahashi HK;Spégel P;Sweet IR;Chae H;Mulder H;Jonas JC
通讯作者：
Jonas JC
影响因子：
82.9
作者：
Klughammer J;Kiesel B;Roetzer T;Fortelny N;Nemc A;Nenning KH;Furtner J;Sheffield NC;Datlinger P;Peter N;Nowosielski M;Augustin M;Mischkulnig M;Ströbel T;Alpar D;Ergüner B;Senekowitsch M;Moser P;Freyschlag CF;Kerschbaumer J;Thomé C;Grams AE;Stockhammer G;Kitzwoegerer M;Oberndorfer S;Marhold F;Weis S;Trenkler J;Buchroithner J;Pichler J;Haybaeck J;Krassnig S;Mahdy Ali K;von Campe G;Payer F;Sherif C;Preiser J;Hauser T;Winkler PA;Kleindienst W;Würtz F;Brandner-Kokalj T;Stultschnig M;Schweiger S;Dieckmann K;Preusser M;Langs G;Baumann B;Knosp E;Widhalm G;Marosi C;Hainfellner JA;Woehrer A;Bock C
通讯作者：
Bock C
重新利用用于靶向DNA甲基化的CRISPR-CAS9系统。
影响因子：
14.9
作者：
Vojta A;Dobrinić P;Tadić V;Bočkor L;Korać P;Julg B;Klasić M;Zoldoš V
通讯作者：
Zoldoš V
食管鳞状细胞癌的空间瘤内异质性和时间克隆进化
影响因子：
30.8
作者：
Hao, Jia-Jie;Lin, De-Chen;Dinh, Huy Q.;Mayakonda, Anand;Jiang, Yan-Yi;Chang, Chen;Jiang, Ye;Lu, Chen-Chen;Shi, Zhi-Zhou;Xu, Xin;Zhang, Yu;Cai, Yan;Wang, Jin-Wu;Zhan, Qi-Min;Wei, Wen-Qiang;Berrnan, Benjamin P.;Wang, Ming-Rong;Koeffler, H. Phillip
通讯作者：
Koeffler, H. Phillip
影响因子：
16.6
作者：
Xu X;Tan X;Tampe B;Wilhelmi T;Hulshoff MS;Saito S;Moser T;Kalluri R;Hasenfuss G;Zeisberg EM;Zeisberg M
通讯作者：
Zeisberg M