Biology subjects

Wilson, J. W.

Publications and source records attributed to Wilson, J. W..

6 recordsLinked to original sources

bioRxiv2025

Novel compounds derived from AR-12 that demonstrate host-directed clearance of intracellular Salmonella enterica Serovar Typhimurium

Salmonella infections, including typhoid and paratyphoid fevers, impose a substantial burden in low-income countries, contributing to significant morbidity and mortality associated with enteric and systemic infections. Developing effective treatments for Salmonella infections requires further consideration, especially due to widespread antibiotic resistance. Host-targeted therapies, such as AR-12, offer a promising solution to combat drug resistance. AR-12 has demonstrated broad-spectrum antimicrobial activity against various bacterial pathogens, including Salmonella enterica serovar Typhimurium, S. Typhi, Francisella tularensis, and F. novicida, as well as protozoan parasites and fungal pathogens. To expand its HDT potential against S. Typhimurium, we conducted a medicinal chemistry campaign using AR-12 as a scaffold with systematic optimization of various points of diversity and the pyrazole core to develop analogs informed by structure-activity relationship. This work led to the development of 81 AR-12 analogs. Primary screening identified 38 analogs that are both more potent and less cytotoxic than parent compound AR-12, while only three were less potent and more cytotoxic. Further, only seven compounds affected planktonic Salmonella growth below 20 {micro}M, suggesting host-directed activity in most of the compounds. Twelve analogs were chosen for secondary screening in MDR S. Typhimurium. Compounds 372, 373, and 378 demonstrated remarkable selectivity, with values exceeding 1500 for both susceptible and MDR S. Typhimurium, compared to AR-12s selectivity of around 20. This approximately 100-fold improvement, coupled with improved potency against intracellular Salmonella, suggests these analogs have significantly greater host-directed activity than direct antibacterial effects. Proteomic analysis for the two most potent compounds, 341 and 370 revealed enrichment of vesicle-mediated transport proteins, specifically with respect to retrograde transport at the trans-Golgi-network and intra-Golgi traffic. These results suggest that the analogs reduce intracellular S. Typhimurium replication by disrupting its exploitation of the host cells vesicle-mediated transport system.

microbiology↗

bioRxiv2025

Discovery of Host-Directed Small Molecules with Broad Anti-Leishmanial Efficacy

Leishmaniasis, a neglected tropical disease affecting nearly 10% of the global population, suffers from limited therapeutic options and rising drug resistance. To address this, we developed 343 analogs of AR-12, a compound that has previously illustrated host-directed anti-leishmanial effects. Primary screening using a luminescence-based assay revealed 66 analogs with greater selectivity than the parent compound, AR-12. Sixteen promising candidates, selected for high potency (IC < 1 {micro}M) or high selectivity (>15), underwent secondary screening via Giemsa staining. Four lead compounds (53, 134, 197, and 354) demonstrated therapeutic indices greater than 40. Tertiary assays confirmed their broad in vitro efficacy against both Leishmania donovani and L. mexicana. Notably, 197 exhibited potent host-directed activity and proteomic analysis identified lysozyme as a mechanistic target, implicating it in the host-mediated clearance of intracellular parasites. These findings highlight the dual host- and pathogen-directed mechanisms of these compounds and support their potential as the basis for new therapeutic strategies. Further optimization and clinical exploration of these leads are warranted to meet the urgent need for effective leishmaniasis treatments. AUTHOR SUMMARYLeishmaniasis is a parasitic infectious disease with limited therapeutic options and rising drug resistance. Drugs that act directly against Leishmania can further drive drug resistance. Host-directed therapies work to enable the host responses to enhance pathogen clearance and reduce disease progression. Host-directed therapies both limit the emergence of new drug resistance and combat drug resistant infections. In this work we screened a library of 343 AR-12 analogs for host-directed anti-leishmanial activity. This work identified 4 hit compounds. We then did proteomic analysis to understand the mechanism of action of the primary lead compound, identifying the protein lysozyme as having a role in host-directed activity against Leishmania infection.

biochemistry↗

bioRxiv2024

Epigenetic derepression of H3K9me3 mitigates Alzheimer-related pathology and improves cognition via immunomodulation and Vgf induction

We investigated the role of histone 3 lysine 9 trimethylation (H3K9me3), an epigenetic mechanism involved in the repression of synaptic plasticity and memory-related genes, within aging and Alzheimers disease (AD). Our study reveals that elevated cortical H3K9me3 strongly correlates with cognitive dysfunction in individuals with mild cognitive impairment (MCI) and AD. In old (18 months) and younger (14 months) APPSWE/PS1{Delta}E9 and 3xTg AD mouse models, inhibiting SUV39H1 methyltransferase with ETP69, substantially reduces cerebral H3K9me3 levels and attenuates amyloid-{beta} burden, tau pathology, and gliosis. Administration of ETP69 further promotes dendritic spine formation, leading to rapid and sustained improvements in cognitive function. Proteomics analysis indicates that a significant proportion of dysregulated proteins in the brains of AD-model mice are reversed by ETP69. These proteins are enriched for synaptic plasticity and learning-related pathways. ETP69 exerts its effects through multiple neuroprotective mechanisms, including regulation of neuroinflammation, induction of both blood and cerebral-infiltrating monocytes involved in cerebral A{beta} clearance. Moreover, ETP69 activates brain-derived neurotrophic factor (Bdnf) network, and particularly its downstream effector neurosecretory protein Vgf. These findings support the pharmacological inhibition of H3K9me3-mediated gene silencing to reverse AD-related pathology and cognitive decline.

neuroscience↗

bioRxiv2023

Systematic characterization of site-specific proline hydroxylation using hydrophilic interaction chromatography and mass spectrometry

We have developed a robust workflow for the identification of proline hydroxylation sites in proteins, using a combination of hydrophilic interaction chromatography (HILIC) enrichment and high-resolution nano-Liquid Chromatography-Mass Spectrometry (LC-MS) together with refining and filtering parameters during data analysis. Using this approach, we have combined data from cell lines treated with either the prolyl hydroxylase (PHD) inhibitor, Roxadustat (FG-4592), or with the proteasome inhibitor MG-132, or with a DMSO control, to identify a total of 4,993 and 3,247 proline hydroxylation sites, respectively, in HEK293 and RCC4 cells. A subset of 1,954 (HEK293) and 1,253 (RCC4) non-collagen proline hydroxylation sites with high confidence were inhibited by FG-4592 treatment. Features characteristic of proline hydroxylated peptides were identified, which were consistent between the HEK293 and RCC4 datasets. The more hydrophilic HILIC fractions were enriched in peptides containing hydroxylated proline residues, which showed characteristic differences in charge and mass distribution, as compared with either unmodified, or oxidised peptides. Furthermore, we discovered that the intensity of the diagnostic hydroxyproline immonium ion was dependent upon parameters including the MS collision energy setting, parent peptide concentration and the sequence of amino acids adjacent to the modified proline. We show using synthetic peptides that a combination of retention time in LC and optimised MS parameter settings allows reliable identification of proline hydroxylation sites in peptides, even when multiple prolines residues are present. Focussing on proteins in which newly identified proline hydroxylation sites were inhibited by the pan-PHD inhibitor, FG-4592, showed enrichment for proteins involved in metabolism of RNA, mRNA splicing and cell cycle regulation, including the protein phosphatase 1 regulatory subunit, Repo-Man (CDCA2). We show that Repo-Man is hydroxylated at P604 and in the accompanying study by Druker et al. 1, we present a combination of cellular and biochemical evidence that hydroxylation of Repo-Man at P604 is important for its function in controlling mitotic progression.

biochemistry↗

bioRxiv2021

High-throughput screening of the ReFRAME, Pandemic Box, and COVID Box drug repurposing libraries against SARS-CoV2 nsp15 endoribonuclease to identify small-molecule inhibitors of viral activity.

SARS-CoV-2 has caused a global pandemic, and has taken over 1.7 million lives as of mid-December, 2020. Although great progress has been made in the development of effective countermeasures, with several pharmaceutical companies approved or poised to deliver vaccines to market, there is still an unmet need of essential antiviral drugs with therapeutic impact for the treatment of moderate-to-severe COVID-19. Towards this goal, a high-throughput assay was used to screen SARS-CoV-2 nsp15 uracil-dependent endonuclease (endoU) function against 13 thousand compounds from drug and lead repurposing compound libraries. While over 80% of initial hit compounds were pan-assay inhibitory compounds, three hits were confirmed as nsp15 endoU inhibitors in the 1-20 M range in vitro. Furthermore, Exebryl-1, a {beta}-amyloid anti-aggregation molecule for Alzheimers therapy, was shown to have antiviral activity between 10 to 66 M, in VERO, Caco-2, and Calu-3 cells. Although the inhibitory concentrations determined for Exebryl-1 exceed those recommended for therapeutic intervention, our findings show great promise for further optimization of Exebryl-1 as an nsp15 endoU inhibitor and as a SARS-CoV-2 antiviral. Author summaryDrugs to treat COVID-19 are urgently needed. To address this, we searched libraries of drugs and drug-like molecules for inhibitors of an essential enzyme of the virus that causes COVID-19, SARS-CoV-2 nonstructural protein (nsp)15. We found several molecules that inhibited the nsp15 enzyme function and one was shown to be active in inhibiting the SARS-CoV-2 virus. This demonstrates that searching for SARS-CoV-2 nsp15 inhibitors can lead inhibitors of SARS-CoV-2, and thus therapeutics for COVID-19. We are currently working to see if these inhibitors could be turned into a drug to treat COVID-19.

microbiology↗

bioRxiv2019

Hypoxia induces rapid changes to histone methylation reprogramming chromatin for the cellular response

Molecular dioxygenases include JmjC-containing histone demethylases and PHD enzymes, but only PHDs are considered to be molecular oxygen sensors in cells. Although, it is known that hypoxia can alter chromatin, whether this is a direct effect on histone demethylases or due to hypoxia induced HIF-dependent transcriptional changes is not known. Here, we report that hypoxia induces a rapid and HIF-independent alteration to a variety of histone methylation marks. Genomic locations of H3K4me3 and H3K36me3 following short hypoxia predict the hypoxia gene signature observed several hours later in cells. We show that KDM5A inactivation mimics hypoxic changes to H3K4me3 in its targets and is required for the cellular response to hypoxia. Our results demonstrate a direct link between oxygen sensing and chromatin changes via KDM inhibition.\n\nOne Sentence SummaryRapid oxygen sensing by chromatin

biochemistry↗