Protein Engineering of a Biologic Drug Candidate

项目总结/摘要 吸烟每年造成近50万人死亡。在美国，直接的医疗成本 治疗吸烟和吸烟引起的疾病的费用超过1700亿美元/年。只有约20-25%的吸烟者能达到长期吸烟的目的。 使用标准药物治疗的长期戒烟，使大多数人仍然吸烟并寻求 替代品.本项目的重点是进一步评估一种新型尼古丁降解酶的蛋白质性质， 酶及其用作戒烟治疗的适用性。酶法降解尼古丁是一种新的 治疗戒烟的方法。已经建立了用于体内的临床“原理验证” 滥用药物的酶降解：两种可卡因降解酶已证明令人鼓舞 第1/2阶段的结果。 需要对这种酶的生物学特性进行工程改造，以降低潜在的免疫原性， 延长血清半衰期，以确保患者安全性、足够的稳定性和酶作用的持续时间。到 为此，我们将研究聚乙二醇化的效果，即大的无毒聚乙烯的化学缀合 二醇（PEG）部分结合到酶的表面，其提供潜在表位的空间掩蔽， 增强蛋白质的流体动力学半径，降低其肾清除率。聚乙二醇化先导化合物 将通过一系列体外酶和血清稳定性测定以及体内模型进行， 评估PEG化对免疫原性潜力和药代动力学性质的影响。 本项目的目标是降低免疫原性潜力，增加这种新型的血清稳定性。 酶通过优化各种聚乙二醇化策略，同时保持其有效的尼古丁降解 活动实现这些目标将为进一步优化电极导线提供支持性临床前证据 在成瘾的动物行为模型中进行的努力和多次剂量研究（第二阶段建议）; 将这种用于戒烟的新型尼古丁降解酶方法进展到临床前开发。

Project Summary/Abstract Cigarette smoking is responsible for nearly one-half million deaths/yr. in the U.S. where direct healthcare costs to treat smoking and smoking-attributable illness exceed $170 billion/year. Only ~20-25% smokers achieve long- term abstinence using standard of care pharmacotherapies leaving the majority still smoking and seeking alternatives. The focus of this project is to further assess the protein properties of a novel nicotine-degrading enzyme and its suitability for use as a smoking cessation treatment. Enzymatic degradation of nicotine is a new approach for treating smoking cessation. Clinical “proof-of-principle” has already been established for in vivo enzymatic degradation of drugs of abuse: two cocaine-degrading enzymes have demonstrated encouraging phase 1/2 results. Engineering of the biological properties of this enzyme is needed to reduce potential immunogenicity and to extend serum half-life, in order to ensure patient safety, sufficient stability and duration of enzymatic action. To this end we will investigate the effect of PEGylation, the chemical conjugation of large non-toxic polyethylene glycol (PEG) moieties to the surface of the enzyme, which provide steric masking of potential epitopes and enhanced hydrodynamic radius of the protein, decreasing its rate of renal clearance. PEGylated lead candidates will be progressed through a series of in vitro enzyme and serum stability assays, as well as in vivo models to assess PEGylation’s effects on immunogenic potential and pharmacokinetic properties. The goal of this project is to reduce the immunogenic potential and increase the serum stability of this novel enzyme though optimization of various PEGylation strategies, while maintaining its potent nicotine-degrading activity. Achieving these goals will provide supportive pre-clinical evidence to conduct further lead optimization efforts and multiple-dose studies in animal behavioral models of addiction (Phase II proposal); thereby progressing this novel nicotine degrading enzyme approach for smoking cessation into pre-clinical development.