PRJNA706017: Exhaustive drug repurposing strategies support the pursuit of new chemical entities for the development of effective COVID-19 interventions
概要: The ongoing pandemic caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), necessitates strategies to identify prophylactic and therapeutic drug candidates for rapid clinical deployment. Here we describe a screening pipeline for the discovery of efficacious SARS-CoV-2 inhibitors. Two high-throughput, high-content imaging infection assays (one using HeLa cells expressing SARS-CoV-2 receptor ACE2 and the other using lung epithelial Calu-3 cells) were developed and used to screen ReFRAME, a best-in-class drug repurposing library. Among the promising hits, the antivirals nelfinavir and the parent of prodrug MK-4482 had most favorable in vitro activity, pharmacokinetic and human safety profiles, and both reduced SARS-CoV-2 replication in an orthogonal human differentiated primary cell model. However, only MK-4482 effectively blocked SARS-CoV-2 infection in a hamster model, likely due to inadequate plasma exposure of nelfinavir.
项目整体设计: Hamster lung from uninfected (U, n=2), vehicle treated (V, n=4) and MK-4482 (EIDD-2801) treated (T, n=4) samples were analyzed using RNASeq platform.
Hamster lung samples were collected after two different treatment regimens.
处理方案:
Hamster were infected with SARS-CoV-2 using a BSL-3 facility.
提取方案:
The lungs were harvested from the hamsters and homogenized in TRI-Reagent. RNA were extracted after infection using the Quick-RNA MicroPrep Kit (Zymo Research, USA) according to the manufacture’s instruction. RNA was converted into cDNA using the qScript™ cDNA SuperMix (Quantabio).
建库方案:
RNA sequencing libraries were generated using the Illumina TruSeq Stranded Total RNA Library Prep Gold with TruSeq Unique Dual Indexes (Illumina, San Diego, CA). Samples were processed following manufacturer’s instructions, except modifying RNA shear time to five minutes. Resulting libraries were multiplexed and sequenced with 100 basepair (bp) Paired End (PE100) to a depth of approximately 25-40 million reads per sample on an Illumina NovaSeq 6000. Samples were demuxltiplexed using bcl2fastq v2.20 Conversion Software (Illumina, San Diego, CA).
测序信息
分子类型:
polyA(+) RNA
库的片段类型:
PAIRED
库的链类型:
Forward
测序平台:
ILLUMINA
测序仪型号:
Illumina NovaSeq 6000
链特异性:
specific
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文章
Mucosal-associated invariant T cell responses differ by sex in COVID-19.
Med (New York, N.Y.) . 2021-04-13 [PMID:
33870241]
Drug repurposing screens identify chemical entities for the development of COVID-19 interventions.
