Liu Beijun, Huang Haifeng, Yang Zhibin, Liu Beiyin, Gou Sanhu, Zhong Chao, Han Xiufeng, Zhang Yun, Ni Jingman, Wang Rui
Key Laboratory of Preclinical Study for New Drugs of Gansu Province, School of Basic Medical Sciences, Lanzhou University, Lanzhou 730000, China; School of Pharmacy, Lanzhou University, Lanzhou 730000, China.
Shaanxi Provincial People's Hospital, Shanxi 710068, China.
Peptides. 2017 Feb;88:115-125. doi: 10.1016/j.peptides.2016.12.016. Epub 2016 Dec 28.
Currently, antimicrobial peptides have attracted considerable attention because of their broad-sprectum activity and low prognostic to induce antibiotic resistance. In our study, for the first time, a series of side-chain hybrid dimer peptides J-AA (Anoplin-Anoplin), J-RR (RW-RW), and J-AR (Anoplin-RW) based on the wasp peptide Anoplin and the arginine- and tryptophan-rich hexapeptide RW were designed and synthesized by click chemistry, with the intent to improve the antimicrobial efficacy of peptides against bacterial pathogens. The results showed that all dimer analogues exhibited up to a 4-16 fold increase in antimicrobial activity compared to the parental peptides against bacterial strains. Furthermore, the antimicrobial activity was confirmed by time-killing kinetics assay with two strains which showed that these dimer analogues at 1, 2×MIC were rapidly bactericidal and reduced the initial inoculum significantly during the first 2-6h. Notably, dimer peptides showed synergy and additivity effects when used in combination with conventional antibiotics rifampin or penicillin respectively against the multidrug-resistant strains. In the Escherichia coli-infected mouse model, all of hybrid dimer analogues had significantly lower degree of bacterial load than the untreated control group when injected once i.p. at 5mg/kg. In addition, the infected mice by methicillin-resistant (MRSA) strain could be effectively treated with J-RR. All of dimer analogues had membrane-active action mode. And the membrane-dependent mode of action signifies that peptides functions freely and without regard to conventional resistant mechanisms. Circular dichroism analyses of all dimer analogues showed a general predominance of α-helix conformation in 50% trifluoroethanol (TFE). Additionally, the acute toxicities study indicated that J-RR or J-AR did not show the signs of toxicity when adult mice exposed to concentration up to 120mg/kg. The 50% lethal dose (LD) of J-AA was 53.6mg/kg. In conclusion, to design and synthesize side chain-hybrid dimer analogues via click chemistry may offer a new strategy for antibacterial therapeutic option.
目前，抗菌肽因其广泛的活性和低诱导抗生素耐药性的可能性而备受关注。在我们的研究中，首次基于黄蜂肽Anoplin和富含精氨酸及色氨酸的六肽RW，通过点击化学设计并合成了一系列侧链杂合二聚体肽J-AA（Anoplin-Anoplin）、J-RR（RW-RW）和J-AR（Anoplin-RW），旨在提高肽对细菌病原体的抗菌效果。结果表明，与亲本肽相比，所有二聚体类似物对细菌菌株的抗菌活性提高了4至16倍。此外，通过对两种菌株的时间杀菌动力学分析证实了其抗菌活性，结果显示这些二聚体类似物在1、2×MIC时具有快速杀菌作用，并在最初的2至6小时内显著降低了初始接种量。值得注意的是，二聚体肽分别与传统抗生素利福平或青霉素联合使用时，对多重耐药菌株显示出协同和相加作用。在大肠杆菌感染的小鼠模型中，当以5mg/kg腹腔注射一次时，所有杂合二聚体类似物的细菌载量均显著低于未治疗的对照组。此外，J-RR可有效治疗耐甲氧西林（MRSA）菌株感染的小鼠。所有二聚体类似物均具有膜活性作用模式。这种依赖膜的作用模式表明肽可自由发挥作用，不受传统耐药机制的影响。所有二聚体类似物的圆二色性分析表明，在50%三氟乙醇（TFE）中，α-螺旋构象普遍占主导。此外，急性毒性研究表明，成年小鼠暴露于高达120mg/kg的浓度时，J-RR或J-AR未显示出毒性迹象。J-AA的50%致死剂量（LD）为53.6mg/kg。总之，通过点击化学设计和合成侧链杂合二聚体类似物可能为抗菌治疗提供一种新策略。
