Mitigating ADA Through Site-specific Conjugation Technology

通过位点特异性缀合技术缓解 ADA

• 批准号: 10537800
• 负责人: Samantha Rene Benjamin
• 金额: $ 3.51万
• 依托单位: STATE UNIVERSITY OF NY,BINGHAMTON
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-11-21 至 2025-11-20
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10537800
• 关键词: Achievement; Affinity; Agonist; Allergic Reaction; Antibodies; Antibody Response; Antibody-drug conjugates; Antigen-Presenting Cells; Area; Attenuated; Binding; Biological; Biological Assay; CD80 gene; CD86 gene; Cells; Cysteine; Data; Development; Differentiation Antigens; Dose; Drug Delivery Systems; Engineering; Enzyme-Linked Immunosorbent Assay; Enzymes; Exhibits; FDA approved; Fc domain; Fluorescence; Fluorine; Funding; Goals; Hydrophobicity; IgG1; Immune; Immunologic Adjuvants; Immunooncology; Incidence; Location; Masks; Measurable; Mediating; Methods; Modeling; Molecular Conformation; Mus; Oregon; Oregon Green 488 carboxylic acid; Patients; Peptides; Pharmaceutical Preparations; Positioning Attribute; Proteins; Publishing; Reaction; Reporting; Risk; Sampling; Site; TLR7 gene; Technology; Therapeutic; Tumor Tissue; United States National Institutes of Health; antibody conjugate; base; biophysical tools; cancer therapy; clinical application; cytotoxic; design; drug clearance; experimental study; glycosylation; imaging system; immunogenicity; interest; live cell imaging; macrophage; negative affect; prevent; tool; tumor; uptake

Summary: Antibody drug conjugate (ADC) immunogenicity is a growing concern due to the increased interest in immune- stimulating drug delivery technology. A recent study by Novartis demonstrated that 14 out of 14 patients dosed with an experimental immune-stimulatory ADC developed a measurable anti-drug antibody (ADA) response. ADA negatively affects treatment by neutralizing drugs, increasing drug clearance, and causing severe allergic reactions. There is an urgent need for an effective method of conjugating payload to antibodies while minimizing the development and the effects of ADAs. We have developed an ADC bioconjugation technology that employs the conserved Q295 residue (continued funding provided by NIH 1RO1GM140026-O1A1) that we believe will mitigate many of these ADA concerns. The goal of this proposal is to demonstrate that this technology reduces Fc-gamma uptake in antigen presenting cells (APCs) and masks the drugs from binding to ADAs by hiding them in a sterically occluded hydrophobic pocket on the Fc domain. This goal will be achieved through the two aims. Aim #1 focuses on developing NMR and fluorescence-based tools to demonstrate that ADC payloads at the Q295 site are constrained within the sterically occluded IgG1 hydrophobic pocket. To this end we propose conjugating model payloads to the Q295 site using various linkers (such as short alkyl, peptide, and PEG) and determining their proximity to the hydrophobic pocket using two experimental methods: 1) a proximity induced fluorescent quenching assay using Oregon Green 488, due to its ability to self-quench and 2) F-19 protein NMR using a trifluoromethyl probe, due to its high sensitivity and low background level in biological samples. Aim #2 focuses on demonstrating that payloads attached to the sterically occluded Q295 position have reduced ADA binding and reduced potential for eliciting immunogenicity caused by Fc-gamma mediated uptake of ADCs. Achievement of the aims outlined herein will result in an antibody conjugation technology that exhibits a reduced risk of ADA and may be useful for the design of immune-stimulating antibody conjugates.

总结： 抗体药物偶联物（ADC）的免疫原性是一个日益关注的问题，由于增加的兴趣，免疫- 刺激药物输送技术。诺华公司最近的一项研究表明，14名患者中有14名服用了 用实验性免疫刺激性ADC产生了可测量的抗药抗体（ADA）应答。 ADA通过中和药物、增加药物清除率和引起严重过敏性反应而对治疗产生负面影响。 反应.迫切需要一种将有效载荷缀合至抗体的有效方法，同时 最大限度地减少ADA的发展和影响。我们开发了ADC生物偶联技术 使用保守的Q295残基（继续由NIH 1 RO 1GM 140026-O 1A 1提供资金）， 相信会减轻许多ADA的担忧。该提案的目的是证明， 该技术减少了抗原呈递细胞（APC）中Fc-γ的摄取，并掩盖了药物与 ADA通过将它们隐藏在Fc结构域上的空间封闭的疏水口袋中。这一目标将得以实现 通过这两个目标。目标#1专注于开发基于NMR和荧光的工具，以证明 在Q295位点的ADC有效载荷被限制在空间封闭的IgG 1疏水口袋内。本 最后，我们提出使用各种接头（例如短烷基， 肽和PEG），并使用两种实验方法测定它们与疏水口袋的接近度： 1)使用俄勒冈州绿色488的邻近诱导荧光猝灭测定，由于其自猝灭能力 和2）使用三氟甲基探针的F-19蛋白质NMR，由于其高灵敏度和低背景水平， 生物样本目标#2的重点是证明连接到空间闭塞Q295的有效载荷 位置具有降低的ADA结合和降低的引发由Fc-γ引起的免疫原性的可能性 介导的ADC摄取。本文概述的目的的实现将导致抗体缀合， 显示出ADA风险降低并可用于设计免疫刺激的技术。 抗体缀合物。