Small molecules that bind the SARS-CoV-2 nonstructural protein 3 Mac1 domain in place of ADP-ribose could be useful as molecular probes or scaffolds for COVID-19 antiviral drug discovery because Mac1 has been linked to the ability of coronaviruses to evade cellular detection. A high-throughput assay based on differential scanning fluorimetry (DSF) was therefore optimized and used to identify possible Mac1 ligands in small libraries of drugs and drug-like compounds. Numerous promising compounds included nucleotides, steroids, β-lactams, and benzimidazoles. The main drawback to this approach was that a high percentage of compounds in some libraries were found to influence the observed Mac1 melting temperature. To prioritize DSF screening hits, the shapes of the observed melting curves and initial assay fluorescence were examined, and the results were compared with virtual screens performed using AutoDock Vina. The molecular basis for alternate ligand binding was also examined by determining a structure of one of the hits, cyclic adenosine monophosphate, with atomic resolution.
The virus that causes COVID-19, SARS-CoV-2, has a large RNA genome that encodes numerous proteins that might be targets for antiviral drugs. Some of these proteins, such as the RNA-dependent RNA polymers, helicase and main protease, are well conserved between SARS-CoV-2 and the original SARS virus, but several others are not. This study examines one of the most novel proteins encoded by SARS-CoV-2, a macrodomain of nonstructural protein 3 (nsp3). Although 26% of the amino acids in this SARS-CoV-2 macrodomain differ from those seen in other coronaviruses, the protein retains the ability to bind ADP-ribose, which is an important characteristic of beta coronaviruses, and potential therapeutic target.
ABSTRACTThe Mac1 domain of the multifunctional SARS-CoV-2 non-structural protein 3 (nsp3) is a potential COVID-19 drug target because it is suspected to enhance the ability of the virus to evade the human immune system. The SARS-CoV-2 Mac1 domain binds ADP-ribose and proteins harboring this important post-translational modification. Small molecules that bind the Mac1 domain in place of ADP-ribose might therefore be useful as molecular probes or scaffolds for antiviral drug discovery. Two high throughput screens were used here to identify such ligands in small libraries of drugs and drug-like compounds. The first screen used differential scanning fluorimetry (DSF, aka the thermal shift or ThermoFluor assay) to examine the melting temperature of SARS-CoV-2 Mac1 domain in the presence of various compounds. In the second screen, various high-resolution SARS-CoV-2 Mac1 structures were used with Autodock VINA to identify potential ligands. Numerous hit compounds were either steroids (estradiol valerate & flunisolide), beta-lactams (cefaclor & cefatrizine), or benzimidazoles (telmisartan, rabeprazole, omeprazole, & esomeprazole). Isothermal titration calorimetry was used to confirm that rabeprazole, omeprazole, and compounds in other chemical classes, such as irinotecan, nifedipine, trifluoperazine, bind SARS-CoV-2 Mac1 with an affinity similar to ADP-ribose.
Histone
acetylation is a prominent epigenetic modification linked
to the memory loss symptoms associated with neurodegenerative disease.
The use of existing histone deacetylase inhibitor (HDACi) drugs for
treatment is precluded by their weak blood–brain barrier (BBB)
permeability and undesirable toxicity. Here, we address these shortcomings
by developing a new class of disulfide-based compounds, inspired by
the scaffold of the FDA-approved HDACi romidepsin (FK288). Our findings
indicate that our novel compound MJM-1 increases the overall level
of histone 3 (H3) acetylation in a prostate cancer cell line. In mice,
MJM-1 injected intraperitoneally (i.p.) crossed the BBB and could
be detected in the hippocampus, a brain region that mediates memory.
Consistent with this finding, we found that the post-training i.p.
administration of MJM-1 enhanced hippocampus-dependent spatial memory
consolidation in male mice. Therefore, MJM-1 represents a potential
lead for further optimization as a therapeutic strategy for ameliorating
cognitive deficits in aging and neurodegenerative diseases.
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