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Targeting Viral RNA Using a Sequence Programmable Small Molecule-Oligonucleotide Conjugate

使用序列可编程小分子-寡核苷酸缀合物靶向病毒 RNA

基本信息

批准号：
10512627
负责人：
KEVAN M. SHOKAT
金额：
$ 545.04万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
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：
2022-05-16 至 2025-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10512627
关键词：
2019-nCoV 5&apos Untranslated Regions Address Affinity Antisense Oligonucleotides Base Pairing Binding Biochemical Biological Assay COVID-19 treatment Cells Chemicals Chikungunya virus Clinic Closure by clamp Collaborations Communicable Diseases Complex Coronavirus Crimean-Congo Hemorrhagic Fever Virus Cryoelectron Microscopy Crystallization Dose Dose-Limiting Elements Engineering Glues Head Human poliovirus In Vitro Laboratories Lassa virus Lead Length Letters Ligands Link Linker-Oligonucleotide Measures Mediating Messenger RNA Middle East Respiratory Syndrome Coronavirus Modality Modeling Modification Molecular Monitor Mutation Natural Products Obstruction Oligonucleotides Pharmaceutical Preparations Positioning Attribute Proteins Proteomics RNA RNA Sequences RNA Stability RNA Viruses Replicon Reporter Research Personnel Ribosomes Role SARS coronavirus SARS-CoV-2 variant Scanning Series Site Specificity Stretching Structure System Testing Therapeutic Toxic effect Traction Translational Repression Translations Variant Viral Viral Genome Viral Proteins Virus Virus Replication Zika Virus appendage arm base chemical genetics clinical candidate design experimental study helicase in vitro Assay in vivo inhibitor novel pandemic disease pre-clinical programs response rocaglamide small molecule structural biology translation factor viral RNA virology

项目摘要

PROJECT 1: TARGETING VIRAL RNA USING A SEQUENCE PROGRAMMABLE SMALL MOLECULE- OLIGONUCLEOTIDE CONJUGATE SUMMARY We propose to merge small molecule and antisense oligonucleotide (ASO) approaches to specifically inhibit viral RNA translation without inhibiting the translation of endogenous cellular mRNAs. Rocaglate natural products, such as rocaglamide A (Roc), demonstrate a unique RNA-targeting mechanism by which small molecule binding to a bimolecular cavity between eIF4A helicase and polypurine RNA (stretches of A and G) generates a steric clamp on the 5’ untranslated regions (UTRs) of target mRNAs. Due to obstruction of the scanning ribosome, this rocaglamide-mediated clamp results in translational inhibition of polypurine tract-containing mRNAs, including that of SARS-CoV-2 and other positive-sense RNA viruses. While pre-clinical viral replication assays indicate that zotatifin, a clinical candidate based on rocaglamide, may demonstrate a promising therapeutic window as an anti-viral, it is anticipated that the simultaneous inhibition of multiple polypurine tract-containing mRNAs within the cell could result in dose-limiting toxicities. We position rocaglamide as a unique protein-RNA molecular glue that can be directed to specific viral target sequences through the appendage of an additional RNA-targeting chemical element. We have designed rocaglamide-ASO (RocASO) molecules which link rocaglamide to an ASO whose binding is dependent upon complementary base-pairing with target RNA sequences. A model that juxtaposes the two therapeutic modalities reveals a permissible disposition of the two binding elements approximately 20 Å apart, which will be traversed by a variable chemical tether of sufficient length. RocASOs designed against the 5’ UTR of SARS-CoV-2 will be tested using biochemical ternary complex formation assays with eIF4A and viral RNA, as well as assayed using cellular reporters of SARS-CoV-2 5’ UTR translation and RNA stability. Finally, we will apply RocASOs in a SARS-CoV-2 replicon system to determine the relative tolerance of RocASOs to polypurine tract or ASO recognition sequence mutations. These experiments will also evaluate whether potentially emergent alterations to target sequences could be addressed by introducing concomitant modifications to RocASOs, assessing the platform as an adaptable RNA targeting modality for novel SARS-CoV-2 variants and other viral 5’ UTR sequences.

项目一：利用序列可编程小分子-靶向病毒RNA