Antisense inhibition of gene expression and growth in gram-negative bacteria by cell-penetrating peptide conjugates of peptide nucleic acids targeted to rpoD gene

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• 作者：Hui  ; Bai^a ;   ; ¹ ; Yu  ; You^b ;   ; ¹ ; Hua  ; Yan^a ;   ; ¹ ; Jingru  ; Meng^a ; Xiaoyan  ; Xue^a ; Zheng  ; Hou^a ; Ying  ; Zhou^a ; Xue  ; Ma^a ; Guojun  ; Sang^a ; Xiaoxing  ; Luo^a ;   ; ^{xxluo3@fmmu.edu.cn}
• 关键词：Cell-penentrating peptide conjugated peptide nucleic acid ; Antisense therapy ; Gram negative bacteria ; Antibiotic resistance ; RNA polymerase primary ¦Ò⁷⁰
• 刊名：Biomaterials
• 出版年：2012
• 出版时间：January 2012
• 年：2012
• 卷：33
• 期：2
• 页码：659-667
• 全文大小：937 K

文摘

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)₄XB- and (KFF)₃K-conjugated peptide nucleic acids (PPNAs) were developed to target rpoD, which encodes an RNA polymerase primary ¦Ò⁷⁰ 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)₄XB- and (KFF)₃K- 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)₄XB 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 ¦Ò⁷⁰ expression, as well as ¦Ò⁷⁰ downstream regulated genes including ftsZ, mazF, prfB, 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.