Exploiting the Hydrophobic Glycosyl Pocket of IgG1 for Imaging and Drug Delivery Applications

利用 IgG1 的疏水性糖基口袋进行成像和药物输送应用

• 批准号: 10619285
• 负责人: Lawrence Tumey
• 金额: $ 7.01万
• 依托单位: STATE UNIVERSITY OF NY,BINGHAMTON
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10619285
• 关键词: Achievement; Antibodies; B-Lymphocytes; Catabolism; Cells; Communities; Development; Drug Delivery Systems; Drug Kinetics; Environment; Enzymes; Esters; Evaluation; Exhibits; Goals; Hydrophobicity; IgG1; Image; Imaging Techniques; Immunoglobulin G; Immunology; Immunosuppressive Agents; Mediating; Methods; Molecular; Oncology; Plasma; Preparation; Property; Raman Spectrum Analysis; Reporting; Research; Site; Structure; Techniques; Technology; Therapeutic; Time; Xenograft Model; antibody conjugate; antibody engineering; chemical property; clinically relevant; functional group; imaging probe; interest; prevent; tool; trafficking

Project Summary: Due to the rapidly growing importance of antibody-mediated drug delivery, there is a critical need for simple and efficient site-specific conjugation techniques that do not require extensive antibody engineering efforts. Moreover, there is significant need to identify sites of conjugation that are protected from plasma enzymes and are useful for the attachment of hydrophobic payloads. The goal of this proposal is to optimize conjugation technology at the conserved Q295 residue in order to meet these challenges. In contrast to most sites of conjugation, the Q295 site is contained within a large hydrophobic cavity that is sterically shielded from plasma and is particularly amenable to the conjugation of large nonpolar payloads. Remarkably, the properties of this hydrophobic pocket are largely unexplored to-date and our preliminary research shows that there are significant untapped opportunities for exploiting the unique features of this site. The goal of this project is to demonstrate the broad utility of this site-specific conjugation technology through the preparation and evaluation of antibody conjugates for oncology, immunology, and imaging applications. We will accomplish this goal through the achievement of three aims. Aim#1 focuses on developing a thorough molecular understanding of the local environment around the Q295 residue and optimizing linkers that can place the payload within the associated hydrophobic pocket. The goal of this aim is to thoroughly understand the chemical properties of the hydrophobic pocket that surrounds the Q295 moiety. Aim#2 focuses on demonstrating the therapeutic utility of this technology through the preparation of ADCs that deliver a wide range of payloads – particularly focusing on payloads that have exhibited difficulties when attached through traditional (“stochastic”) conjugation approaches. Four particular payloads were selected: MMAE (due to its clinical relevance and known linker stability issues), Tubulysin (due to interest in payloads with low PGP efflux and also a labile ester functionality that has caused problems with traditional approaches), Thailanstatin A (due to its unique mechanism of action and to the labile functional groups in its structure), and Brequinar (due to its potency as an immunosuppressive agent and its very high hydrophobicity that has so-far prevented ADC delivery). The resulting B-cell targeting ADCs will be thoroughly evaluated for their pharmacokinetic profile and efficacy in a B-cell xenograft model. Aim#3 focuses on using the Q295 site for the development of Raman imaging probes that can be used for generating live-cell time lapse images. Importantly, there have been no reported attempts to use Raman imaging to study ADC trafficking. Traditional ADC conjugation methods cannot be employed for the attachment of the Raman tags due to their very high hydrophobicity. Successful achievement of these aims will provide the drug-delivery community with a valuable new tool for site-specific conjugation of problematic payloads and will establish new imaging techniques for the study of ADC trafficking and catabolism.

项目摘要： 由于抗体介导的药物递送的重要性迅速增长，因此对简单和 不需要广泛的抗体工程工作的有效特定地点集成技术。而且， 识别受保护不受血浆酶的共轭部位的重要需要，对 疏水有效载荷的附件。该提案的目的是优化在 保守的Q295居住，以应对这些挑战。与大多数共轭地点相反，Q295站点是 包含在一个大的疏水腔中，该腔体在空间上屏蔽了血浆，尤其适合于 大型非极有效载荷的结合。值得注意的是，这个疏水口袋的特性在很大程度上是出乎意料的 迄今为止和我们的初步研究表明，利用独特的机会有很大的尚未开发的机会 此网站的功能。该项目的目的是展示此特定地点共轭的广泛效用 通过制备和评估抗体结合物的肿瘤学，免疫学和成像的技术 申请。我们将通过实现三个目标来实现这一目标。目标＃1专注于开发 对Q295居住周围的当地环境的彻底理解，并优化可以 将有效载荷放在相关的疏水口袋中。这个目标的目的是彻底了解 周围Q295部分的疏水口袋的化学特性。目标＃2专注于展示 通过准备ADC提供广泛有效载荷的ADC，该技术的治疗实用性 - 特别是专注于通过传统附带的有效载荷（“随机”） 共轭方法。选择了四个特定的有效载荷：MMAE（由于其临床相关性和已知联系人 稳定性问题），Tubulysin（由于对PGP流出低的有效载荷的兴趣以及具有不稳定的酯功能 引起了传统方法的问题），泰兰施坦汀A（由于其独特的作用机理和不稳定的机理 其结构中的功能组）和brequinar（由于其作为免疫抑制剂的效力，并且非常高 疏水性具有SO-FAR阻止ADC的递送）。由此产生的B细胞靶向ADC将得到彻底评估 对于B细胞异种移植模型中的药代动力学概况和效率。 AIM＃3专注于使用Q295网站 拉曼成像问题的发展，可用于生成活细胞延时图像。重要的是， 没有据报道尝试使用拉曼成像研究ADC贩运的尝试。传统的ADC集成 由于其非常高的疏水性，无法使用拉曼标签的附着方法。成功的 这些目标的实现将为毒品交通社区提供一种有价值的特定网站的新工具 解决问题的有效载荷的结合，并将建立新的成像技术，以研究ADC贩运和 分解代谢。