Darvesh, S. | bioRxiv Science

bioRxiv2023

Dual inhibition of coronavirus Mpro and PLpro enzymes by phenothiazines and their antiviral activity

Coronavirus (CoV) replication requires efficient cleavage of viral polyproteins into an array of non-structural proteins involved in viral replication, organelle formation, viral RNA synthesis, and host shutoff. Human CoVs (HCoVs) encode two viral cysteine proteases, main protease (Mpro) and papain-like protease (PLpro), that mediate polyprotein cleavage. Using a structure-guided approach, a phenothiazine urea derivative that inhibits both SARS-CoV-2 Mpro and PLpro protease activity in vitro was identified. In silico docking studies also predicted binding of the phenothiazine to the active sites of Mpro and PLpro from distantly related alphacoronavirus, HCoV-229E (229E) and the betacoronavirus, HCoV-OC43 (OC43). The lead phenothiazine urea derivative displayed broad antiviral activity against all three HCoVs tested in cell culture infection models. It was further demonstrated that the compound inhibited 229E and OC43 at an early stage of viral replication, with diminished formation of viral replication organelles and the RNAs that are made within them, as expected following viral protease inhibition. These observations suggest that the phenothiazine urea derivative inhibits viral replication and may broadly inhibit proteases of diverse coronaviruses. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=100 SRC="FIGDIR/small/557219v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@fa2c92org.highwire.dtl.DTLVardef@844484org.highwire.dtl.DTLVardef@1ae5d5aorg.highwire.dtl.DTLVardef@730133_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LICoronavirus cysteine proteases Mpro and PLpro are targets for novel antiviral agents C_LIO_LIPhenothiazine ureas inhibit SARS-CoV-2 Mpro and PLpro protease activity C_LIO_LISome phenothiazine ureas inhibit replication of diverse coronaviruses with minimal cytotoxicity C_LIO_LIPhenothiazine ureas inhibit early stages of coronavirus replication consistent with failure of viral polyprotein cleavage C_LI

microbiology↗

bioRxiv2018

Tauopathy in the APPswe/PS1ΔE9 mouse model of familial Alzheimer’s disease

Despite compelling evidence that the accumulation of amyloid-beta (A{beta}) promotes cortical MAPT (tau) aggregation in familial and idiopathic Alzheimers disease (AD), murine models of cerebral amyloidosis are not considered to develop tau-associated pathology. The absence of neurofibrillary lesions in amyloidosis mice remains a challenge for the amyloidocentric paradigm of AD pathogenesis. It has resulted in the generation of transgenic mice harboring mutations in their tau gene, which may be inappropriate for studying a disease with no known TAU mutations, such as AD. Here, we have used APPswe/PS1{Delta}E9 mice to show that tau pathology can develop spontaneously in murine models of familial AD. Tauopathy was abundant around A{beta} deposits, with Gallyas- and thioflavin-S-positive perinuclear inclusions accumulating in the APPswe/PS1{Delta}E9 cortex by 18 months of age. Age-dependent increases in Gallyas signal correlated positively with binding levels of the paired helical filament (PHF) ligand [18F]Flortaucipir, in all brain areas examined. Sarkosyl-insoluble PHFs were visualized by electron microscopy. Tandem mass tag proteomics identified sequences of hyperphosphorylated tau in transgenic mice, along with signs of RNA missplicing, ribosomal dysregulation and disturbed energy metabolism. Human frontal gyrus tissue was used to validate these findings, revealing primarily quantitative differences between the tauopathy observed in AD patient vs. transgenic mouse tissue. Levels of tau mRNA were not different between APPswe/PS1{Delta}E9 and littermate control animals. As physiological levels of endogenous, wild-type tau aggregate secondarily to A{beta} in transgenic mice, this study demonstrates that amyloidosis is both necessary and sufficient to drive tauopathy in experimental models of familial AD.

neuroscience↗