Protein Labeling and Crosslinking by Covalent Aptamers

共价适体的蛋白质标记和交联

基本信息

批准号：
10391640
负责人：
Alexander Deiters
金额：
$ 35.19万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-02-02 至 2026-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10391640
关键词：
Address Affinity Amino Acids Animal Model Antibodies Antibody-drug conjugates Antisense Oligonucleotides Area Binding Binding Proteins Biological Biological Models Biology Cell Surface Proteins Cell model Cell surface Cells Chemicals Chemistry Collection Data Detection Development Diagnostic Disease Drug Delivery Systems Embryo Engineering Evaluation Generations Goals Government Image Immobilization Immune Targeting Immune response In Vitro Kinetics Label Libraries Ligands Literature Mediating Methodology Modification Mus Nucleic Acids Nucleotides Oligonucleotides Persons Positioning Attribute Post-Translational Protein Processing Preparation Process Production Proteins RNA vaccine Reaction Reagent Reporting Research Research Personnel SARS-CoV-2 spike protein Site Specificity Structure-Activity Relationship Testing Therapeutic Thrombin Time Virus Zebrafish aptamer base circulating cancer cell cost covalent bond crosslink design experience improved in vivo novel nuclease nucleic acid-based therapeutics programs protein degradation residence small molecule inhibitor success tool transcription factor

项目摘要

PROJECT SUMMARY The overall goal of this proposal is to develop a fundamentally new class of oligonucleotide agents – covalent aptamers – that will have a major impact on nucleic acid-based reagents and therapeutics. Covalent aptamers will be an enabling methodology behind new aptamer applications and will provide new solutions to persistent limitations of this compound class. Aptamers are engineered oligonucleotides that bind protein targets and have found applications in a multitude of areas, including therapeutics, diagnostics, drug delivery, and imaging. They bind their targets with affinity and specificity that rivals those of antibodies, while displaying lower production cost, higher production consistency, and the ability to easily introduce chemical modifications. Capitalizing on the ability to amplify and sequence nucleic acids, aptamers are being generated through SELEX, a powerful in vitro selection process. Our central hypothesis is that the formation of a covalent bond between an aptamer – or a functional motif delivered by the aptamer – and its target protein will provide unprecedented residence time, increase nuclease stability (or obviate the need for it), and facilitate aptamer selection. Furthermore, covalent bond formation enables new applications based on the aptamer platform. The following aims will test this hy- pothesis. Aim 1: The chemistry of covalent aptamers. To gain a comprehensive understanding of the design principles behind covalent aptamers and to enable broad applicability, we will explore their chemistry based on our promising preliminary results. We will explore applications in antibody drug conjugate assembly, as well as catalytic covalent bond formation. Aim 2: Cell-based applications of covalent aptamers. We will develop several applications that are enabled by covalent aptamers and that showcase their utility as chemical biology probes. We are focusing on cell-surface proteins due to the rich collection of aptamers that have been reported. Aim 3: Selection of covalent aptamers. For the de novo generation of covalent aptamers capable of selectively cross- linking to any chosen protein target without prior sequence and/or structural information, we will develop a gen- eralized in vitro selection process that adds distinct advantages to traditional SELEX.

项目概要该提案的总体目标是开发一类全新的寡核苷酸试剂——共价键适配体——这将对基于核酸的试剂和治疗产生重大影响。将成为新适配体应用背后的一种可行方法，并将为持久性适配体提供新的解决方案该化合物类别的局限性是工程化的寡核苷酸，可结合蛋白质靶标并具有在许多领域都有应用，包括治疗、诊断、药物输送和成像。以与抗体相媲美的亲和力和特异性结合靶标，同时产量较低成本、更高的生产一致性以及轻松引入化学修饰的能力。核酸扩增和测序的能力，适体是通过 SELEX 产生的，SELEX 是一种强大的我们的中心假设是适体之间形成共价键 - 或由适体传递的功能基序及其目标蛋白将提供前所未有的停留时间，增加核酸酶稳定性（或消除对其的需要），并促进适体选择。此外，共价键的形成使得基于适体平台的新应用成为可能，以下目标将测试这种hy- 目标 1：共价适体的化学性质获得对设计的全面了解。共价适体背后的原理并实现广泛的适用性，我们将基于我们将探索在抗体药物偶联物组装方面的应用，以及我们有希望的初步结果。目标 2：共价适体的细胞应用。由共价适体实现的应用，并展示了它们作为化学生物学探针的实用性。由于已报道的适配体丰富，我们将重点放在细胞表面蛋白上，目标 3：共价适体的选择用于能够选择性交叉的共价适体的从头生成。在没有事先序列和/或结构信息的情况下连接到任何选定的蛋白质靶点，我们将开发一个基因- 实现了体外选择过程，为传统 SELEX 增添了明显的优势。