Technology to Create Spiegel ERAbodies on Demand: Biostable Universal Antibody Replacements

按需创建 Spiegel ERAbodies 的技术：生物稳定的通用抗体替代品

• 批准号: 10510985
• 负责人: Elisa Biondi
• 金额: $ 23.18万
• 依托单位: FOUNDATION FOR APPLIED MOLECULAR EVOLUTN
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-20 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10510985
• 关键词: Affinity; Amino Acids; Anions; Antibodies; Area; Attention; Behavior; Benchmarking; Binding; Binding Proteins; Biological; Biological Process; Biological Products; Biomedical Research; Cancer Center; Categories; Charge; Chemistry; Clinical; Consumption; DNA; Data; Diagnostic; Digestion; Discrimination; Disease; Evolution; Extracellular Protein; Foundations; Future; Genetic; Goals; Gold; Head; Hour; Image; Immune system; In Vitro; Label; Laboratories; Letters; Ligands; Medical; Methods; Modification; Molecular Evolution; Nobel Prize; Nucleotides; Organism; Palladium; Patients; Pattern; Peptides; Performance; Plague; Polymerase; Problem Solving; Procedures; Process; Proteins; RNA; RNA analysis; RNA-Directed DNA Polymerase; Reagent; Reproducibility; Research; Research Personnel; Ribonucleases; Role; Signal Transduction; Specificity; Structure; System; Techniques; Technology; Testing; Therapeutic; Therapeutic Agents; Time; Work; aptamer; base; biological systems; cancer cell; chemical synthesis; clinical practice; cost; density; design; functional group; high risk; innovation; interest; meetings; nanomolar; new technology; overexpression; prevent; programmed cell death ligand 1; receptor; scaffold; technology development; tool

Technology to Create Spiegel ERAbodies on Demand: Biostable Universal Antibody Replacements Foundation for Applied Molecular Evolution Elisa Biondi ABSTRACT Researchers in biomedical, diagnostic, and clinical areas want to create (or buy), on demand, reagents that bind to proteins and other targets that may be involved in a biological process that they are studying. Antibod- ies have long served this role. However, as biologics, antibodies are at the center of an "irreproducibility crisis" in biomedical research, and, even when suitable, take months and thousands of dollars to make. This has driven efforts to create antibody replacements, both protein (e.g. Darpins) and RNA (e.g. aptamers). The first are difficult to manipulate, while the second have low stability and disappointing affinity. We hypothesize that an unnatural platform with an "Expanded RNA Alphabet" (ERA) and extra functional groups with extra binding potential will meet this long-standing unmet need. While expanded DNA alphabets are now advanced, a first innovation is that ERAs have not yet been the target of any preliminary data. We hypothesize that nanomolar binding will be routinely achieved because ERAbodies will have access to (a) higher information density that will lead to (b) better defined folds, both by using an RNA scaffold and by having functionality that supports folding, (c) greater structural diversity that gives ERAbodies more modes for tight binding, and (d) more folding motifs that allow ERAbodies to have more compact structures. They are also hypothesized to have all of the advantages of classical aptamers, including value as the starting points for subsequent rounds of evolution, modifiability using signaling entities, low cost, fast turnaround, and direct chemical synthesis. This R21 project will prove the value of this new technology platform, which will allow researchers to order or directly create in weeks, binders for targets that they themselves select. We hypothesize a further innovation by merging yet unexplored ERA technology with the classical concept of mirror symmetry. To create ERAbodies that are stable in biological systems, we will make these in mirror image ("Spiegel") form. Aim 1. A single R21 demonstration project will show that ERAbodies can be made with building blocks created by palladium chemistry. Although this technology is agnostic with respect to applications (it can create binders for any target), this demonstration will have impact by targeting a protein with outsized medical significance, PD-L1 (Programmed Death Ligand 1). We will test the hypothesis that L-ERA reagents, much like standard L-RNA, are stable against RNases in living systems. This workflow step will consume 18 months. We will expand methods to sequence ERA and benchmark the fidelity of enzymatic synthesis. In the workflow, this will be completed in the first 6 months, as patterns of transliteration by various reverse transcriptases will be used to define a sequencing procedure applicable to various 6- and 8-letter ERA systems. The target metrics are binding affinity (nanomolar), binding specificity (100:1 discrimination), and in vitro sera stability (<0.1% RNase degradation over 2 hours).

按需创建Spiegel Erabodies的技术：可生物稳定的通用抗体 替换 应用分子进化基础 Elisa Biondi 抽象的 生物医学，诊断和临床领域的研究人员希望按需创建（或购买） 结合可能与他们正在研究的生物学过程有关的蛋白质和其他靶标。抗体 IE长期以来一直担任这个角色。但是，作为生物制剂，抗体是“不可夸大性危机”的中心 在生物医学研究中，即使在合适的情况下，也需要数月和数千美元才能赚钱。这就是 驱动抗体替代的驱动努力，包括蛋白质（例如DARPINS）和RNA（例如Aptamers）。第一个 很难操纵，而第二个则具有较低的稳定性和令人失望的亲和力。 我们假设一个不自然的平台，具有“扩展的RNA字母”（ERA）和额外功能 具有额外约束力的团体将满足这种长期的未满足需求。而扩展的DNA字母 现在是先进的，第一个创新是时代尚未成为任何初步的目标 数据。我们假设纳摩尔结合将通常实现，因为Erabodies可以访问 到（a）通过使用RNA脚手架和通过 具有支持折叠的功能，（c）更大的结构多样性，使Erabodies更多模式 紧密的结合，以及（d）更多的折叠基序，允许Erabodies具有更紧凑的结构。他们是 还假设具有经典适体的所有优势，包括价值作为起点 随后的进化回合，使用信号实体的可修改性，低成本，快速周转和直接 化学合成。 这个R21项目将证明这个新技术平台的价值，该平台将允许研究人员订购或 直接在几周内创建，他们自己选择的目标固定器。我们假设进一步的创新 通过将却没有开发的时代技术与经典的镜子对称性概念合并。创建 在生物系统中稳定的细胞代理，我们将以镜像形式（“ Spiegel”）形式制成这些。 AIM1。一个R21演示项目将表明可以用构建块制作Erabodies 由钯化学创建。尽管该技术在应用方面不可知（它可以创建 任何目标的粘合剂），该演示将通过靶向用大型医学来靶向蛋白质会产生影响 意义，PD-L1（编程死亡配体1）。我们将检验以下假设：L-ERA试剂类似 标准L-RNA对生物系统中的RNase是稳定的。这个工作流程将消耗18个月。 我们将将方法扩展到序列时代，并基于酶合成的保真度。在 工作流程，这将在最初的6个月内完成，作为通过各种反向音译的模式 转录酶将用于定义适用于各种6字母和8个字母时代系统的测序过程。 目标指标是结合亲和力（纳摩尔），结合特异性（100：1歧视）和体外 血清稳定性（2小时内<0.1％RNase降解）。