Antisense inhibition of gene expression and growth in gram-negative bacteria by cell-penetrating peptide conjugates of peptide nucleic acids targeted to rpoD gene
Antisense inhibition of gene expression and growth in gram-negative bacteria by cell-penetrating peptide conjugates of peptide nucleic acids targeted to rpoD gene
复制标题
靶向 rpoD 基因的肽核酸的细胞穿透肽缀合物对革兰氏阴性细菌中基因表达和生长的反义抑制
DOI：
10.1016/j.biomaterials.2011.09.075
复制
发表时间：
2012-01-01
期刊：
影响因子：
14
通讯作者：
Luo, Xiaoxing
中科院分区：
文献类型：
--
作者：
Bai, Hui;You, Yu;Luo, Xiaoxing
革兰氏阴性菌（GNB）引起常见和严重的医院和社区获得性感染，具有较高的多药耐药（MDR）发生率和死亡率。MDR-GNB菌株的出现和传播限制了治疗选择，并突出了开发新治疗策略的必要性。在这项研究中，开发了肽（RXR）（4）XB-和（KFF）（3）K-缀合肽核酸（PPNA）以靶向rpoD，其编码被认为是细菌生长所必需的RNA聚合酶初级sigma（70）。在体外和感染模型中测试它们对MDR-GNB的不同临床分离株的抗微生物活性。（RXR）（4）XB-和（KFF）（3）K-缀合的PNA以浓度依赖性和序列选择性方式对MDR-GNB的不同菌株具有杀菌作用，而具有乱序碱基序列的PPNA对生长没有影响。在所测试的PPNAs中，（RXR）（4）XB缀合物PPNA 06对多重耐药大肠杆菌显示出更强的广谱抑制作用。肠道沙门氏菌、肺炎克雷伯氏菌和福氏志贺菌的体外和体内研究。结果与rpoD mRNA和sigma（70）表达的抑制以及sigma（70）下游调节基因包括ftsZ、迷宫prjB、rpoS、seqA、turfB和ygjD相关。PPNA 06对单个或多个MDR-GBN感染的人胃粘膜上皮细胞的处理证明了对细菌生长的完全抑制，并且对人细胞的形态和生长没有影响。与经典抗生素相比，PPNA 06在GNB中未显示抗生素抗性的诱导。这些发现首次证明rpoD是开发反义抗生素的潜在靶点，并表明抗rpoD PNA的肽缀合物在体外和体内对GNB具有活性。我们的研究结果为治疗MDR-GNB感染提供了一种可行的策略。(C)2011爱思唯尔有限公司保留所有权利。
Gram-negative bacteria (GNB) cause common and severe hospital- and community-acquired infections with a high incidence of multidrug resistance (MDR) and mortality. The emergence and spread of MDR-GNB strains limit therapeutic options and highlight the need to develop new therapeutic strategies. In this study, the peptide (RXR)(4)XB- and (KFF)(3)K-conjugated peptide nucleic acids (PPNAs) were developed to target rpoD, which encodes an RNA polymerase primary sigma(70) that is thought to be essential for bacterial growth. Their antimicrobial activities were tested against different clinical isolates of MDR-GNB in vitro and in infection models. The (RXR)(4)XB- and (KFF)(3)K- conjugated PNAs were bactericidal against different strains of MDR-GNB in concentration-dependent and sequence-selective manner, whereas a PPNA with a scrambled base sequence had no effect on growth. Among tested PPNAs, (RXR)(4)XB conjugate PPNA06 showed more potent and broad spectrum inhibition in multidrug-resistant Escherichia coli. Salmonella enterica, Klebsiella pneumoniae, and Shigella flexneri in vitro and in vivo. The results were associated with suppression of rpoD mRNA and sigma(70) expression, as well as sigma(70) downstream regulated genes including ftsZ, mazE prjB, rpoS, seqA, turfB and ygjD. The treatment of PPNA06 on mono- or multiple MDR-GBN infected human gastric mucosal epithelial cells demonstrated the complete inhibition on bacterial growth and no influence on morphology and growth of human cells. Also, PPNA06 did not show the induction of antibiotic resistance as compared with classical antibiotics in GNB. These findings firstly demonstrate that rpoD is potential target for developing antisense antibiotics, and indicate that peptide conjugates of anti-rpoD PNA are active against GNBs in vitro and in vivo. Our results offer a feasible strategy for treating MDR-GNB infections. (C) 2011 Elsevier Ltd. All rights reserved.