Kumar Prashant, Shenoi Rajesh A, Lai Benjamin F L, Nguyen Michael, Kizhakkedathu Jayachandran N, Straus Suzana K
Department of Chemistry, University of British Columbia , 2036 Main Mall, Vancouver, BC V6T 1Z1, Canada.
Biomacromolecules. 2015 Mar 9;16(3):913-23. doi: 10.1021/bm5018244. Epub 2015 Feb 20.
Aurein 2.2 is an antimicrobial peptide (AMP) whose mechanism of action is quite well-understood and that has good activity against Gram-positive bacteria. It is, however, highly cytotoxic. Poly(ethylene glycol) (PEG) conjugation (PEGylation) of protein and peptide drugs has been used for decades to improve their in vivo efficacy and blood circulation by enhancing the biocompatibility of the protein or peptide in question. However, the relatively large hydrodynamic size, high intrinsic viscosity, the limited number of functional groups available for conjugation, and immunogenicity of high molecular weight PEG limits its use in bioconjugation applications. Recently, hyperbranched polyglycerol (HPG) has been gaining attention as an alternative to PEG due to its excellent biocompatibility. Here, for the first time, we report the synthesis of HPG conjugates of antimicrobial peptides. Aurein 2.2 peptide was conjugated to high molecular weight HPG with a varying number of peptides per polymer, and the biocompatibility and antimicrobial activity of the conjugates were investigated. The antimicrobial activity of the peptide and its conjugates were determined by measuring the minimal inhibitory concentration (MIC) against Staphylococcus aureus and Staphylococcus epidermidis. The interaction of aurein 2.2 peptide and the conjugates with a model bacterial biomembrane was investigated using CD spectroscopy to understand the mode of action of the conjugates. The biocompatibility of the AMP-polymer conjugates was investigated by measuring red cell lysis, platelet activation and aggregation, complement activation, blood coagulation, and cell toxicity. Our results show that the size of the conjugates and the peptide density influence the biocompatibility of the antimicrobial conjugates. These results will help to further define the properties of HPG-AMP conjugates and set the stage for development of better therapeutic agents.
奥瑞因2.2是一种抗菌肽（AMP），其作用机制已为人所熟知，对革兰氏阳性菌具有良好的活性。然而，它具有高度的细胞毒性。蛋白质和肽类药物的聚乙二醇（PEG）缀合（PEG化）已被使用数十年，通过增强相关蛋白质或肽的生物相容性来提高其体内疗效和血液循环。然而，相对较大的流体力学尺寸、高特性粘度、可用于缀合的官能团数量有限以及高分子量PEG的免疫原性限制了其在生物缀合应用中的使用。最近，超支化聚甘油（HPG）因其优异的生物相容性作为PEG的替代品而受到关注。在此，我们首次报道了抗菌肽的HPG缀合物的合成。将奥瑞因2.2肽与高分子量HPG缀合，每个聚合物上肽的数量不同，并研究了缀合物的生物相容性和抗菌活性。通过测量对金黄色葡萄球菌和表皮葡萄球菌的最小抑菌浓度（MIC）来确定肽及其缀合物的抗菌活性。使用圆二色光谱法研究奥瑞因2.2肽及其缀合物与模型细菌生物膜的相互作用，以了解缀合物的作用方式。通过测量红细胞裂解、血小板活化和聚集、补体活化、血液凝固和细胞毒性来研究AMP-聚合物缀合物的生物相容性。我们的结果表明，缀合物的大小和肽密度会影响抗菌缀合物的生物相容性。这些结果将有助于进一步确定HPG-AMP缀合物的性质，并为开发更好的治疗剂奠定基础。
