本项目拟以细胞骨架成分-中间丝状蛋白α-螺旋保守基序肽（IFCM）为材料，N端融合具有细菌靶向及杀菌作用的生物活性肽S-thanatin （Ts），中间插入柔性连接片段（GGGS）及4个酸性氨基酸（EDED）组成的负电荷载药区，构成靶向融合肽Ts-linker-EDED-IFCM；用具有多苯环结构的阳离子抗生素米诺环素（Mc）激发，使之自组装形成一种具有细菌靶向、pH敏感释放、双重杀菌机制、耐药与非耐药菌同步杀灭的新型抗菌纳米；对其进行表征及急性/亚急性毒性试验研究；用标准菌株及临床分离多重耐药菌进行体外杀菌试验；建立革兰氏阳性和阴性标准菌株及耐药菌腹膜炎败血症模型，考察靶向载药纳米在体内对耐药和非耐药菌的作用效果，以获得结构-剂量-效应变化的多点数据，为靶向纳米抗菌药物的临床前研究打下基础；同时，也为除模式药物以外的其它种类抗生素或化合物自组装纳米的构建提供了理论和实验依据。

This study will firstly prepare a fusion peptide of Ts-linker-EDED-IFCM. Ts (S-thanatin) is an active peptide of specific bactericidal property, and IFCM is a short peptide derived from the α-helical consensus motif of the intermediate filament protein which constitutes cytoskeleton together with the microtubule and microfilament. In the recombinant Ts-linker-EDED-IFCM, IFCM is fused after the C-terminal of Ts with a flexible GGGS linker, and a negatively-charged region composed of 4 acidic amino-acid (EDED) is inserted between the linker and IFCM. After trigged with the positively-charged antibiotic of minocycline (Mc) with benzene rings structure, Ts-linker-EDED-IFCM will self-assemble into a spherical nanoparticle which is characterized of bacterial specific targeting potential , pH-sensitive release, double bactericidal mechanism, and synchronous killing both drug-susceptible and -resistant bacteria. The prepared nanoparticles will be characterized, and the acute- and subacute- toxicity has to be investigated. The in vitro antimicrobial potential could be evaluated by MICs and MBCs against standard strains and clinic isolates. The in vivo antimicrobial potential could be evaluated with a septic mouse model caused by different bacteria including drug-susceptible and -resistant bacteria. The in vivo structure-dosage-effect relationship and differences between drug-susceptible and -resistant bacteria or between Gram+ and Gram- bacteria will be studied as well. In conclusion, we will explore the feasibility to construct a novel self-assembly nanoparticle against bacteria based on the α-helical peptide. This research will provide theoretical basis and practical evidences for pre-clinic research and other nanoparticles preparation using chemicals or antibiotics beside the model drug minocycline used in this study.