Strategies for development of antimicrobial peptides and proteins

Reihaneh Manteghi, K. Kristó, G. Szakonyi, I. Csóka
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Abstract

Many researches have been done with the aim of overcoming AMPs challenges. Different modification strategies as well as different delivery development ideas were offered by these researches. However, peptide modifications and formulation of peptide delivery systems are challenging tasks and hide several risks. Understanding and evaluating the cause - effect relations within the initial Risk Assessment (RA) step in case of all attributes is novelty since it gives the basis for the experimental design as the next step, and aids the formulation development in order to get the final product in the targeted quality range. It also helps to focus on the resources (human, financial, time) related to the final product quality aimed at. By means of RA method within QbD approach of early pharmaceutical development we monitored the factors with highly risk potential in the PEGylation process and risks such as loosing antimicrobial activity of peptide are prevented. The selection of CQAs, CQAs, QTPPs, CQAs and CPPs/CMAs of a PEGylated PGLa formula was performed and interdependence rating among the QTPP elements and CQAs, as well as among the CPPs and CQAs was performed. This careful theoretical study led to the selection of the right methodologies and materials in the synthesis of PEGylated AMPs and their formulation and consequently resulted in obtaining optimized formulation. In our second work, LYZ encapsulated in a novel polyelectrolyte core-shell nanoparticles through the LBL technique utilized as a carrier system to control the release of protein. The preparation of LYZ NPs was made according to 23 full factorial design with QbD approach. Our aim was to understand the effect of process parameters through the determination of mathematical equations, based on which the optimization parameters can be predicted under different process parameters. The optimization parameters were encapsulation efficiency, particle size, enzyme activity, and the amount of α-helix structure. The nanoparticles were analyzed with transmission electron microscopy (TEM), Fourier-transform infrared spectroscopy (FTIR), and circular dichroism (CD) spectroscopy. Based on our results, we found that pH was the most important factor and pH 10 was recommended during the formulation. Enzyme activity and α-helix content correlated with each other very well, and particle size and encapsulation efficiency also showed a very good correlation with each other. The results of the α-helix content of FTIR and CD measurements were very similar for the precipitated lysozyme due to the solid-state of lysozyme.
抗菌肽和蛋白质的开发策略
为了克服amp的挑战,已经进行了许多研究。这些研究提供了不同的修改策略和不同的交付发展思路。然而,肽修饰和肽传递系统的制定是具有挑战性的任务,隐藏着一些风险。在所有属性的情况下,了解和评价初始风险评估(RA)步骤中的因果关系是新颖的,因为它为下一步的实验设计提供了基础,并有助于配方开发,以使最终产品达到目标质量范围。它还有助于关注与最终产品质量相关的资源(人力,财务,时间)。通过早期药物开发QbD方法中的RA方法,我们监测了聚乙二醇化过程中具有高风险潜力的因素,并预防了肽失去抗菌活性等风险。对聚乙二醇化PGLa分子式的CQAs、CQAs、QTPPs、CQAs和CPPs/CMAs进行选择,并对QTPP元件与CQAs、CPPs与CQAs之间的相互依赖进行评级。这一仔细的理论研究导致正确的方法和材料的选择在聚乙二醇化amp的合成和他们的配方,从而导致获得优化的配方。在我们的第二项工作中,通过LBL技术将LYZ封装在一种新型的多电解质核壳纳米颗粒中,作为一种载体系统来控制蛋白质的释放。采用QbD方法,采用23全因子设计制备LYZ NPs。我们的目的是通过确定数学方程来了解工艺参数的影响,并在此基础上预测不同工艺参数下的优化参数。优化参数为包封率、粒径、酶活性、α-螺旋结构量。采用透射电子显微镜(TEM)、傅里叶变换红外光谱(FTIR)和圆二色光谱(CD)对纳米颗粒进行了分析。根据我们的研究结果,我们发现pH是最重要的因素,建议在配制过程中pH为10。酶活性与α-螺旋含量的相关性很好,粒径与包封效率的相关性也很好。由于溶菌酶呈固态,溶菌酶α-螺旋含量的FTIR和CD测量结果非常接近。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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