Department of Microbiology and Immunology, F. Edward Hébert School of Medicine, Uniformed Services University of the Health Sciences, Bethesda, Maryland, USA.
Antimicrob Agents Chemother. 2012 Jan;56(1):378-90. doi: 10.1128/AAC.00689-11. Epub 2011 Nov 7.
Helicobacter pylori has developed antimicrobial resistance to virtually all current antibiotics. Thus, there is a pressing need to develop new anti-H. pylori therapies. We recently described a novel oligo-acyl-lysyl (OAK) antimicrobial peptidomimetic, C(12)K-2β(12), that shows potent in vitro bactericidal activity against H. pylori. Herein, we define the mechanism of action and evaluate the in vivo efficacy of C(12)K-2β(12) against H. pylori after experimental infection of Mongolian gerbils. We demonstrate using a 1-N-phenylnaphthylamine (fluorescent probe) uptake assay and electron microscopy that C(12)K-2β(12) rapidly permeabilizes the bacterial membrane and creates pores that cause bacterial cell lysis. Furthermore, using nucleic acid binding assays, Western blots, and confocal microscopy, we show that C(12)K-2β(12) can cross the bacterial membranes into the cytoplasm and tightly bind to bacterial DNA, RNA, and proteins, a property that may result in inhibition of enzymatic activities and macromolecule synthesis. To define the in vivo efficacy of C(12)K-2β(12), H. pylori-infected gerbils were orogastrically treated with increasing doses and concentrations of C(12)K-2β(12) 1 day or 1 week postinfection. The efficacy of C(12)K-2β(12) was strongest in animals that received the largest number of doses at the highest concentration, indicating dose-dependent activity of the peptide (P < 0.001 by analysis of variance [ANOVA]) regardless of the timing of the treatment with C(12)K-2β(12). Overall, our results demonstrate a dual mode of action of C(12)K-2β(12) against the H. pylori membrane and cytoplasmic components. Moreover, and consistent with the previously reported in vitro efficacy, C(12)K-2β(12) shows significant in vivo efficacy against H. pylori when used as monotherapy. Therefore, OAK peptides may be a valuable resource for therapeutic treatment of H. pylori infection.
幽门螺杆菌对几乎所有当前的抗生素都产生了抗药性。因此，迫切需要开发新的抗幽门螺杆菌疗法。我们最近描述了一种新型的寡酰-赖氨酸（OAK）抗菌肽模拟物 C(12)K-2β(12)，它对幽门螺杆菌具有很强的体外杀菌活性。在此，我们定义了 C(12)K-2β(12) 的作用机制，并在蒙古沙土鼠感染实验后评估了其对幽门螺杆菌的体内疗效。我们通过 1-N-苯基萘胺（荧光探针）摄取测定和电子显微镜证明，C(12)K-2β(12)可快速穿透细菌膜并形成孔，导致细菌细胞裂解。此外，通过核酸结合测定、Western 印迹和共聚焦显微镜，我们表明 C(12)K-2β(12)可以穿过细菌膜进入细胞质，并与细菌 DNA、RNA 和蛋白质紧密结合，这种特性可能导致抑制酶活性和大分子合成。为了确定 C(12)K-2β(12)的体内疗效，在感染后 1 天或 1 周，用递增剂量和浓度的 C(12)K-2β(12)对感染了幽门螺杆菌的沙土鼠进行口腔灌胃治疗。在接受最高浓度和最大剂量的动物中，C(12)K-2β(12)的疗效最强，这表明该肽具有剂量依赖性活性（方差分析[ANOVA]的 P < 0.001），而与治疗 C(12)K-2β(12)的时间无关。总体而言，我们的结果表明 C(12)K-2β(12)对幽门螺杆菌膜和细胞质成分具有双重作用模式。此外，与之前报道的体外疗效一致，C(12)K-2β(12)作为单一疗法对幽门螺杆菌具有显著的体内疗效。因此，OAK 肽可能是治疗幽门螺杆菌感染的有价值资源。
