Manipulating Aromaticity: characterization of an ultra-rapid insulin analog

控制芳香度：超快速胰岛素类似物的表征

• 批准号: 8395099
• 负责人: Bruce Hill Frank
• 金额: $ 30.17万
• 依托单位: THERMALIN DIABETES, LLC
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-16 至 2014-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8395099
• 关键词: Algorithms; Amino Acids; Artificial Pancreas; Blood; Blood Glucose; Boxing; Budgets; Buffers; Clinical; Clinical Research; Clinical Trials; Color; Cystine; Development; Diabetes Mellitus; Digestion; Dose; Drug Formulations; Drug Kinetics; Enzyme-Linked Immunosorbent Assay; Event; Family suidae; Feedback; Generations; Glycosylated hemoglobin A; Gold; Gray unit of radiation dose; Hyperglycemia; Hypoglycemia; Infusion procedures; Injection of therapeutic agent; Insulin; Insulin Infusion Systems; Insulin, Aspart, Human; Insulin, Lispro, Human; Insulin-Dependent Diabetes Mellitus; Ions; Kinetics; Lead; Left; Letters; Licensing; Life; Light; Mediating; Modeling; Mutagenesis; Mutation; NPM1 gene; National Institute of Diabetes and Digestive and Kidney Diseases; NovoLog; Octreotide; Parents; Partner in relationship; Patients; Performance; Pharmacodynamics; Phase; Physical condensation; Positioning Attribute; Principal Investigator; Production; Proteins; Protomer; Pump; Rattus; Regimen; Relative (related person); Risk; Safety; Sampling; Series; Serum; Side; Site-Directed Mutagenesis; Small Business Innovation Research Grant; Solubility; Stream; Streptozocin; Structure; Study Section; Subcutaneous Injections; Suggestion; Sulfur; System; Technology; Testing; Thermodynamics; Time; TimeLine; Trypsin; Zinc; absorption; analog; animal data; base; career; chemical stability; chemical synthesis; design; dimer; glucose monitor; glycemic control; improved; in vivo; innovation; insulin dimers; non-diabetic; pre-clinical; programs; response; subcutaneous

DESCRIPTION (provided by applicant): We seek to develop an ultra-fast insulin analog formulation for the treatment of diabetes mellitus. Ultra-fast pharmacokinetic/dynamic (PK/PD) promises to enable superior performance of pump therapy (continuous subcutaneous insulin infusion; CSII) with enhanced safety and more robust integration with continuous glucose monitors (CGM). Our product promises to improve the safety and efficacy of algorithm-controlled closed-loop systems (the "artificial pancreas"); ultra-fast PK/PD would also facilitate post-prandial glycemic control in conventional multi-injection regimens. An innovative structural approach is proposed based on non-standard mutagenesis of the insulin molecule. This application has been revised based on the suggestions of the Study Section, most saliently to enhance its focus on stability and to provide supporting animal data. The current barrier to more rapid subcutaneous absorption is the rate of disassembly of the zinc insulin hexamer. This rate is controlled by the stability of an anti-parallel ¿-sheet at the dimer interface of the hexamr. Partial progress toward accelerated disassembly following subcutaneous injection was achieved in the 1990s by conventional site-directed mutagenesis at the periphery of this ¿-sheet. Insulin aspart (the active component of Novolog(R); Novo-Nordisk) and insulin lispro (the active component of Humalog(R); Eli Lilly) contain such partially destabilizing mutations (ProB28->Asp in Novolog; ProB28->Lys and LysB29->Pro in Humalog ("KP")). Because their designs were limited by conventional mutagenesis, however, neither of these products could fully exploit structural strategies to promote disassembly of the core ¿-sheet in the insulin hexamer. The core ¿ -sheet of the insulin hexamer is remarkable for the confluence of 8 conserved aromatic rings (TyrB16, PheB24, PheB25, TyrB26, and their dimer-related mates). The thermodynamic and kinetic stability of this interface is regulated by an intricate series of aromatic-aromatic interactions. We have discovered that the substitution of PheB24 by its non-aromatic analog cyclohexanylalanine (Cha) is compatible with native-like structure and function, but markedly enhances the rate of disassembly of KP-insulin. Since submission of the original application, we have also invented an ultra-stable version of this analog containing additional A-chain substitution ThrA8->Glu. We therefore propose to synthesize and characterize ChaB24-KP-insulin and ChaB24- GluA8-insulin as candidate ultra-rapid formulations. Dr. B. Frank (principal investigator) was co-inventor of Humalog(R) during his prior career at Eli Lilly. Thermalin Diabetes, LLC has an exclusive license to ChaB24- related IP, which is owned by CWRU. We thank the Study Section for its guidance. PUBLIC HEALTH RELEVANCE: By manipulating the aromaticity of a key side chain within the insulin molecule, we have conferred accelerated disassembly of the insulin hexamer. In a pig model this invention appears to provide ultra-rapid absorption of the insulin formulation into the blood stream and so promises to enable patients with diabetes mellitus to achieve enhanced glycemic control with reduced risk of meal-related hyper- or hypoglycemia. Having reduced this idea to practice, we have developed Cyclolog-1 and Cyclolog-1E (extended shelf life); these promise to provide significantly improved clinical benefits relative to other fast-acting analogs i present use or (to our knowledge) in development with application to the safety and efficacy of insulin pumps and closed-loop systems. This project will complete feasibility testing on Cyclolog-1 and Cyclolog-1E.

描述（由申请人提供）：我们寻求开发用于治疗糖尿病的超快速胰岛素类似物制剂。超快速药代动力学/动力学（PK/PD）有望实现泵治疗（连续皮下胰岛素输注; CSII）的上级性能，同时增强安全性，并与动态血糖监测仪（CGM）更稳健地集成。我们的产品有望提高算法控制闭环系统（“人工胰腺”）的安全性和有效性;超快PK/PD也将促进传统多次注射方案中的餐后血糖控制。基于胰岛素分子的非标准突变，提出了一种创新的结构方法。根据研究部分的建议对本申请进行了修订，最显著的是加强了对稳定性的关注，并提供了支持性动物数据。 目前阻碍更快皮下吸收的是锌胰岛素六聚体的分解速率。该速率由六聚体的二聚体界面处的反平行半折叠的稳定性控制。在20世纪90年代，通过在该折叠的外围进行常规定点诱变，实现了皮下注射后加速分解的部分进展。门冬胰岛素（Novolog（R）的活性组分; Novo-Nordisk）和赖脯胰岛素（Humidine（R）的活性组分; Eli Lilly）含有这种部分不稳定突变（Novolog中的ProB28-> Asp; Humidine（"KP"）中的ProB28-> Lys和LysB29-> Pro）。然而，由于他们的设计受到常规诱变的限制，这些产品都不能充分利用结构策略来促进胰岛素六聚体中核心折叠的分解。 核心？胰岛素六聚体的β-折叠对于8个保守芳环（TyrB16、PheB24、PheB25、TyrB26和它们的二聚体相关配偶体）的汇合是显著的。该界面的热力学和动力学稳定性由一系列复杂的芳香族-芳香族相互作用调节。我们已经发现，PheB24的非芳香族类似物环己基丙氨酸（Cha）的取代是兼容的天然样的结构和功能，但显着提高分解的速度KP-胰岛素。自从提交原始申请以来，我们还发明了这种类似物的超稳定形式，其含有额外的A链取代ThrA8-> Glu。因此，我们建议合成和表征ChaB24-KP-胰岛素和ChaB24-GluA8-胰岛素作为候选超快速制剂。B医生。Frank（首席研究员）在Eli Lilly的前职业生涯中是Humanoid（R）的共同发明者。Thermalin Diabetes，LLC拥有ChaB24相关知识产权的独家许可，该知识产权归CWRU所有。我们感谢研究科的指导。 公共卫生相关性：通过操纵胰岛素分子内关键侧链的芳香性，我们已经赋予了胰岛素六聚体的加速分解。在猪模型中，本发明似乎提供了胰岛素制剂超快速吸收到血流中，因此有望使糖尿病患者实现增强的血糖控制，同时降低与膳食相关的高血糖或低血糖的风险。在将这一想法付诸实践后，我们开发了Cyclolog-1和Cyclolog-1E（延长保质期）;这些药物有望提供相对于目前使用或（据我们所知）正在开发的其他速效类似物显著改善的临床获益，用于胰岛素泵和闭环系统的安全性和有效性。该项目将完成Cyclolog-1和Cyclolog-1E的可行性测试。