Biology subjects

Murakami, E.

Publications and source records attributed to Murakami, E..

4 recordsLinked to original sources

bioRxiv2024

Discovery and Synthesis of GS-7682, a Novel Prodrug of a 4'-CN-4-Aza-7,9-Dideazaadenosine C-Nucleoside with Broad-Spectrum Potency Against Pneumo- and Picornaviruses and Efficacy in RSV-Infected African Green Monkeys.

Acute respiratory viral infections (ARVI), such as pneumovirus and respiratory picornavirus infections, exacerbate disease in COPD and asthma patients. A research program targeting respiratory syncytial virus (RSV) led to the discovery of GS-7682 (1) a novel phosphoramidate prodrug of a 4'-CN-4-aza-7,9-dideazaadenosine C-nucleoside GS-646089 (2) with broad antiviral activity against RSV EC50 = 3-46 nM, human metapneumovirus (hMPV) EC50 = 210 {+/-} 50 nM, human rhinovirus (RV) EC50 = 54-61 nM, and enterovirus (EV) EC50 = 83-90 nM. Prodrug optimization for cellular potency and lung cell metabolism identified the 5-methyl((S)-hydroxy(phenoxy)phosphoryl)-L-alaninate in combination with 2,3-diisobutyrate promoieties as optimal for high intracellular triphosphate formation in vitro and in vivo. 1 demonstrated significant reductions of viral loads in the lower respiratory tract of RSV-infected African green monkeys when administered once daily via intratracheal nebulized aerosol. Together these finding support additional evaluation of 1 and its analogs as a potential therapeutic for pneumo- and picornaviruses.

biochemistry↗

bioRxiv2021

Therapeutic efficacy of an oral nucleoside analog of remdesivir against SARS-CoV-2 pathogenesis in mice.

The COVID-19 pandemic remains uncontrolled despite the rapid rollout of safe and effective SARS-CoV-2 vaccines, underscoring the need to develop highly effective antivirals. In the setting of waning immunity from infection and vaccination, breakthrough infections are becoming increasingly common and treatment options remain limited. Additionally, the emergence of SARS-CoV-2 variants of concern with their potential to escape therapeutic monoclonal antibodies emphasizes the need to develop second-generation oral antivirals targeting highly conserved viral proteins that can be rapidly deployed to outpatients. Here, we demonstrate the in vitro antiviral activity and in vivo therapeutic efficacy of GS-621763, an orally bioavailable prodrug of GS-441524, the parental nucleoside of remdesivir, which targets the highly conserved RNA-dependent RNA polymerase. GS-621763 exhibited significant antiviral activity in lung cell lines and two different human primary lung cell culture systems. The dose-proportional pharmacokinetic profile observed after oral administration of GS-621763 translated to dose-dependent antiviral activity in mice infected with SARS-CoV-2. Therapeutic GS-621763 significantly reduced viral load, lung pathology, and improved pulmonary function in COVID-19 mouse model. A direct comparison of GS-621763 with molnupiravir, an oral nucleoside analog antiviral currently in human clinical trial, proved both drugs to be similarly efficacious. These data demonstrate that therapy with oral prodrugs of remdesivir can significantly improve outcomes in SARS-CoV-2 infected mice. Thus, GS-621763 supports the exploration of GS-441524 oral prodrugs for the treatment of COVID-19 in humans.

microbiology↗

bioRxiv2020

Remdesivir potently inhibits SARS-CoV-2 in human lung cells and chimeric SARS-CoV expressing the SARS-CoV-2 RNA polymerase in mice.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged in 2019 as the causative agent of the novel pandemic viral disease COVID-19. With no approved therapies, this pandemic illustrates the urgent need for safe, broad-spectrum antiviral countermeasures against SARS-CoV-2 and future emerging CoVs. We report that remdesivir (RDV), a monophosphoramidate prodrug of an adenosine analog, potently inhibits SARS-CoV-2 replication in human lung cells and primary human airway epithelial cultures (EC50 = 0.01 M). Weaker activity was observed in Vero E6 cells (EC50 = 1.65 M) due to their low capacity to metabolize RDV. To rapidly evaluate in vivo efficacy, we engineered a chimeric SARS-CoV encoding the viral target of RDV, the RNA-dependent RNA polymerase, of SARS-CoV-2. In mice infected with chimeric virus, therapeutic RDV administration diminished lung viral load and improved pulmonary function as compared to vehicle treated animals. These data provide evidence that RDV is potently active against SARS-CoV-2 in vitro and in vivo, supporting its further clinical testing for treatment of COVID-19.

microbiology↗

bioRxiv2020

Genome editing in mammals and plants using CRISPR type I-D nuclease

Adoption of the CRISPR-Cas system has revolutionized genome engineering in recent years; however, application of genome editing with CRISPR type I--the most abundant CRISPR system in bacteria--has been less developed. Type I systems in which Cas3 nuclease degrades the target DNA are known; in contrast, for the sub-type CRISPR type I-D (TiD), which lacks a typical Cas3 nuclease in its cascade, the mechanism of target DNA degradation remains unknown. Here, we found that Cas10d--a nuclease in TiD--is multi-functional in PAM recognition, stabilization and target DNA degradation. TiD can be used for targeted mutagenesis of genomic DNA in human cells, directing both bi-directional long-range deletions and short insertions/deletions. TiD off-target effects, which were dependent on the mismatch position in the protospacer of TiD, were also identified. Our findings suggest TiD as a unique effector pathway in CRISPR that can be repurposed for genome engineering in eukaryotic cells.

molecular biology↗