Publications

@article{noKey,

title = {A novel screening assay for insulin receptor},

author = {Staňurová, Jana},

url = {https://www.biorxiv.org/content/10.1101/2025.03.20.644157v1.abstract},

doi = {https://doi.org/10.1101/2025.03.20.644157},

year  = {2025},

date = {2025-03-20},

abstract = {Insulin is a key hormone in glucose homeostasis. Its lack causes severe health complications and has to be compensated by regular administration of insulin. Despite intense long-lasting research, a more stable substitute has yet to be discovered to alleviate patients' issues. Here we report the development of a novel assay for screening potential insulin analogues based on the recently published method DIANA. Our assay meets the need for a fast, non-radioactive method as a sensitive alternative to the commonly used radioactive immunoassay.},

keywords = {MANTIS},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Disrupted α-ketoglutarate homeostasis trains monocyte-derived macrophages towards M2-like phenotype in long-term treated HIV-infection},

author = {Escós, Alejandra},

url = {https://www.biorxiv.org/content/10.1101/2025.01.15.633214v1.full},

doi = {https://doi.org/10.1101/2025.01.15.633214},

year  = {2025},

date = {2025-03-19},

abstract = {Cells of the myeloid lineage, particularly monocytes and macrophages, play a key role in HIV infection by contributing to viral replication, immune response, and maintaining immune balance during suppressive therapy. We hypothesized that metabolic reprogramming and altered chemokine signaling in people living with HIV (PWH) on long-term antiretroviral therapy (ART) affect monocyte transport and polarization due to ongoing inflammation. Therefore, the present study aimed to identify the mechanism of impaired monocyte/macrophage function in PWH on well-treated ART that can lead to clinical intervention strategies to improve health. Single-cell RNA sequencing, immune-phenotyping, and metabolic modeling identified altered expression of chemokine and metabolite receptors and altered metabolic flux in PWH monocytes that decreased monocyte migration. The plasma secretome revealed a nonclassical inflammatory microenvironment in PWH. Integrative multi-omics and single-cell proteomics of differentiated monocyte-derived macrophages (MDMs) detected metabolic reprogramming orchestrated by α-ketoglutarate (AKG) that affected macrophage function and HIV infection. Increased levels of AKG in plasma were shown to occur in PWH under ART. Therefore, when differentiating MDM with serum from PWH or AKG, macrophage function was found polarized towards an M2-like state. AKG alone was shown to increase CCR5 levels and increase HIV-1 infection in MDM. Here, we utilize systems biology-driven identification and ex vivo assays to show impaired macrophage polarization, due to metabolic training, can leads to a low-grade nonclassical inflammatory environment in well-treated PWH.},

keywords = {MANTIS},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {ActinoMation: a literate programming approach for medium-throughput robotic conjugation of Streptomyces spp.},

author = {Møller, Tenna A.},

url = {https://www.sciencedirect.com/science/article/pii/S2405805X25000389},

doi = {https://doi.org/10.1016/j.synbio.2025.03.005},

year  = {2025},

date = {2025-03-11},

abstract = {The genus Streptomyces are valuable producers of antibiotics and other pharmaceutically important bioactive compounds. Advances in molecular engineering tools, such as CRISPR, have provided some access to the metabolic potential of Streptomyces, but efficient genetic engineering of strains is hindered by laborious and slow manual transformation protocols. In this paper, we present a semi-automated medium-throughput workflow for the introduction of recombinant DNA into Streptomyces spp. using the affordable and open-sourced Opentrons (OT-2) robotics platform. To increase the accessibility of the workflow we provide an open-source protocol-creator, ActinoMation. ActinoMation is a literate programming environment using Python in Jupyter Notebook. We validated the method by transforming Streptomyces coelicolor (M1152 and M1146), S. albidoflavus (J1047), and S. venezuelae (DSM40230) with the plasmids pSETGUS and pIJ12551. We demonstrate conjugation efficiencies of 3.33*10-3/0.33% for M1152 with pSETGUS and pIJ12551; 2.96*10-3/0.29%for M1146 with pSETGUS and pIJ12551; 1.21*10-5/0.0012% for J1047 with pSETGUS and 4.70*10-4/0.047% with pIJ12551, and 4.97*10-2/4.97% for DSM40230 with pSETGUS and 6.13*10-2 /6.13% with pIJ12551 with a false positive rate between 8.33% and 54.54%. Automation of the conjugation workflow facilitates a streamlined workflow on a larger scale without any evident loss of conjugation efficiency.},

keywords = {MANTIS},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Discovery of inhibitors for bacterial Arr enzymes ADP-ribosylating and inactivating rifamycin antibiotics},

author = {Alaviuhkola, Juho},

url = {https://www.biorxiv.org/content/10.1101/2025.02.20.639278v1.full},

doi = {https://doi.org/10.1101/2025.02.20.639278},

year  = {2025},

date = {2025-02-22},

abstract = {ADP-ribosylation is an enzymatic process where an ADP-ribose moiety is transferred from NAD+ to an acceptor molecule. While ADP-ribosylation is well-established as a post-translational modification of proteins, rifamycin antibiotics are its only known small-molecule targets. ADP-ribosylation of rifampicin was first identified in Mycolicibacterium smegmatis, whose Arr enzyme transfers the ADP-ribose moiety to the 23-hydroxy group of rifampicin preventing its interaction with the bacterial RNA polymerase thereby inactivating the antibiotic. Arr homologues are widely spread among bacterial species and present in several pathogenic species often associated with mobile genetic elements. Inhibition of Arr enzymes offers a promising strategy to overcome ADP-ribosylation mediated rifamycin resistance. We developed a high-throughput activity assay, which was applied to screen an in-house library of human ADP-ribosyltransferase-targeted compounds. We identified 15 inhibitors with IC50 values below 5 µM against four Arr enzymes from M. smegmatis, Pseudomonas aeruginosa, Stenotrophomonas maltophilia and Mycobacteroides abscessus. The observed overall selectivity of the hit compounds over the other homologues indicated structural differences between the proteins. We crystallized M. smegmatis and P. aeruginosa Arr enzymes, the former in complex with its most potent hit compound with an IC50 value of 1.3 µM. We observed structural differences in the NAD+ binding pockets of the two Arr homologues explaining the selectivity. Although the Arr inhibitors did not sensitize M. smegmatis to rifampicin in a growth inhibition assay, the structural information and the collection of inhibitors provide a foundation for rational modifications and further development of the compounds.},

keywords = {MANTIS},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Amides of moronic acid and morolic acid with the tripeptides MAG and GAM targeting antimicrobial, antiviral and cytotoxic effects},

author = {Bildziukevich, Uladzimir},

url = {https://pubs.rsc.org/en/content/articlehtml/2024/md/d4md00742e},

doi = {https://doi.org/10.1039/d4md00742e},

year  = {2024},

date = {2024-10-29},

abstract = {A series of amides of selected plant triterpenoids, moronic acid and morolic acid, with the tripeptides MAG and GAM, was designed and synthesized. Two required tripeptides 5 and 10 were synthesized by a step-wise chain elongation of the ethyl esters of either glycine or L-methionine at their N-terminus using Boc-protected amino acids in each step. The tripeptides 5 and 10 were used for the synthesis of 13–23, the derivatives of moronic acid (11) and morolic acid (12), to get a series of amide derivatives of the less frequently studied triterpenoids 11 and 12. The target compounds, and their intermediates, were subjected to an investigation of their antimicrobial, antiviral and cytotoxic activity. Selectivity of the pharmacological effects was found. Generally, the target compounds inhibited only the G+ microorganisms. Compound 16 inhibited Staphylococcus aureus (I = 99.6%; c = 62.5 μM) and Enterococcus faecalis (I = 85%; c = 250 μM). Several compounds showed moderate antiviral effects, both anti-HIV-1, 19 (EC50 = 57.0 ± 4.1 μM, CC50 > 100 μM), 20 (EC50 = 17.8 ± 2.1 μM, CC50 = 41.0 ± 5.2 μM) and 23 (EC50 = 12.6 ± 0.82 μM, CC50 = 38.0 ± 4.2 μM), and anti-HSV-1, 22 (EC50 = 27.7 ± 3.5 μM, CC50 > 100 μM) and 23 (EC50 = 30.9 ± 3.3 μM, CC50 > 100 μM). The target compounds showed no cytotoxicity in cancer cells, however, several of their intermediates were cytotoxic. Compound 21 showed cytotoxicity in HeLa (IC50 = 7.9 ± 2.1 μM), G-361 (IC50 = 8.0 ± 0.6 μM) and MCF7 (IC50 = 8.6 ± 0.2 μM) cancer cell lines, while being non-toxic in normal fibroblasts (BJ; IC50 > 50 μM).},

keywords = {TEMPEST},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Development of a sensitive high-throughput enzymatic assay capable of measuring sub-nanomolar inhibitors of SARS-CoV2 Mpro},

author = {Kovar, Peter},

url = {https://www.sciencedirect.com/science/article/pii/S2472555224000418},

doi = {https://doi.org/10.1016/j.slasd.2024.100179},

year  = {2024},

date = {2024-09-01},

abstract = {The SARS-CoV-2 main protease (Mpro) is essential for viral replication because it is responsible for the processing of most of the non-structural proteins encoded by the virus. Inhibition of Mpro prevents viral replication and therefore constitutes an attractive antiviral strategy. We set out to develop a high-throughput Mpro enzymatic activity assay using fluorescently labeled peptide substrates. A library of fluorogenic substrates of various lengths, sequences and dye/quencher positions was prepared and tested against full length SARS-CoV-2 Mpro enzyme for optimal activity. The addition of buffers containing strongly hydrated kosmotropic anion salts, such as citrate, from the Hofmeister series significantly boosted the enzyme activity and enhanced the assay detection limit, enabling the ranking of sub-nanomolar inhibitors without relying on the low-throughput Morrison equation method. By comparing cooperativity in citrate or non-citrate buffer while titrating the Mpro enzyme concentration, we found full positive cooperativity of Mpro with citrate buffer at less than one nanomolar (nM), but at a much higher enzyme concentration (∼320 nM) with non-citrate buffer. In addition, using a tight binding Mpro inhibitor, we confirmed there was only one active catalytical site in each Mpro monomer. Since cooperativity requires at least two binding sites, we hypothesized that citrate facilitates dimerization of Mpro at sub-nanomolar concentration as one of the mechanisms enhances Mpro catalytic efficiency. This assay has been used in high-throughput screening and structure activity relationship (SAR) studies to support medicinal chemistry efforts. IC50 values determined in this assay correlates well with EC50 values generated by a SARS-CoV-2 antiviral assay after adjusted for cell penetration.},

keywords = {TEMPEST},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Discovery of an antivirulence compound that targets the Staphylococcus aureus SaeRS two-component system to inhibit toxic shock syndrome toxin-1 production},

author = {Dufresne, Karine},

url = {https://www.jbc.org/article/S0021-9258(24)01956-2/fulltext},

doi = {https://doi.org/10.1016/j.jbc.2024.107455},

year  = {2024},

date = {2024-07-01},

abstract = {Menstrual toxic shock syndrome (mTSS) is a rare but severe disorder associated with the use of menstrual products such as high-absorbency tampons and is caused by Staphylococcus aureus strains that produce the toxic shock syndrome toxin-1 (TSST-1) superantigen. Herein, we screened a library of 3920 small bioactive molecules for the ability to inhibit transcription of the TSST-1 gene without inhibiting the growth of S. aureus. The dominant positive regulator of TSST-1 is the SaeRS two-component system (TCS), and we identified phenazopyridine hydrochloride (PP-HCl) that repressed the production of TSST-1 by inhibiting the kinase function of SaeS. PP-HCl competed with ATP for binding of the kinase SaeS leading to decreased phosphorylation of SaeR and reduced expression of TSST-1 as well as several other secreted virulence factors known to be regulated by SaeRS. PP-HCl targets the virulence of S. aureus, and it also decreases the impact of TSST-1 on human lymphocytes without affecting the healthy vaginal microbiota. Our findings demonstrate the promising potential of PP-HCl as a therapeutic strategy against mTSS.},

keywords = {TEMPEST},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Discovery of JNJ-74856665: A Novel Isoquinolinone DHODH Inhibitor for the Treatment of AML},

author = {DeRatt, Lindsey G.},

url = {https://pubs.acs.org/doi/10.1021/acs.jmedchem.4c00809},

doi = {https://doi.org/10.1021/acs.jmedchem.4c00809},

year  = {2024},

date = {2024-06-18},

abstract = {Acute myelogenous leukemia (AML), a heterogeneous disease of the blood and bone marrow, is characterized by the inability of myeloblasts to differentiate into mature cell types. Dihydroorotate dehydrogenase (DHODH) is an enzyme well-known in the pyrimidine biosynthesis pathway and preclinical findings demonstrated that DHODH is a metabolic vulnerability in AML as inhibitors can induce differentiation across multiple AML subtypes. As a result of virtual screening and structure-based drug design approaches, a novel series of isoquinolinone DHODH inhibitors was identified. Further lead optimization afforded JNJ-74856665 as an orally bioavailable, potent, and selective DHODH inhibitor with favorable physicochemical properties selected for clinical development in patients with AML and myelodysplastic syndromes (MDS).},

keywords = {TEMPEST},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Machine learning accelerates pharmacophore-based virtual screening of MAO inhibitors},

author = {Cieślak, Marcin},

url = {https://www.nature.com/articles/s41598-024-58122-7},

doi = {https://doi.org/10.1038/s41598-024-58122-7},

year  = {2024},

date = {2024-04-08},

abstract = {Nowadays, an efficient and robust virtual screening procedure is crucial in the drug discovery process, especially when performed on large and chemically diverse databases. Virtual screening methods, like molecular docking and classic QSAR models, are limited in their ability to handle vast numbers of compounds and to learn from scarce data, respectively. In this study, we introduce a universal methodology that uses a machine learning-based approach to predict docking scores without the need for time-consuming molecular docking procedures. The developed protocol yielded 1000 times faster binding energy predictions than classical docking-based screening. The proposed predictive model learns from docking results, allowing users to choose their preferred docking software without relying on insufficient and incoherent experimental activity data. The methodology described employs multiple types of molecular fingerprints and descriptors to construct an ensemble model that further reduces prediction errors and is capable of delivering highly precise docking score values for monoamine oxidase ligands, enabling faster identification of promising compounds. An extensive pharmacophore-constrained screening of the ZINC database resulted in a selection of 24 compounds that were synthesized and evaluated for their biological activity. A preliminary screen discovered weak inhibitors of MAO-A with a percentage efficiency index close to a known drug at the lowest tested concentration. The approach presented here can be successfully applied to other biological targets as target-specific knowledge is not incorporated at the screening phase.},

keywords = {MANTIS},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {A novel micellular fluorogenic substrate for quantitating the activity of 1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase gamma (PLCγ) enzymes},

author = {Visvanathan, Ramya},

url = {https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0299541},

doi = {https://doi.org/10.1371/journal.pone.0299541},

year  = {2024},

date = {2024-03-29},

abstract = {The activities of the phospholipase C gamma (PLCγ) 1 and 2 enzymes are essential for numerous cellular processes. Unsurprisingly, dysregulation of PLCγ1 or PLCγ2 activity is associated with multiple maladies including immune disorders, cancers, and neurodegenerative diseases. Therefore, the modulation of either of these two enzymes has been suggested as a therapeutic strategy to combat these diseases. To aid in the discovery of PLCγ family enzyme modulators that could be developed into therapeutic agents, we have synthesized a high-throughput screening-amenable micellular fluorogenic substrate called C16CF3-coumarin. Herein, the ability of PLCγ1 and PLCγ2 to enzymatically process C16CF3-coumarin was confirmed, the micellular assay conditions were optimized, and the kinetics of the reaction were determined. A proof-of-principle pilot screen of the Library of Pharmacologically Active Compounds 1280 (LOPAC1280) was performed. This new substrate allows for an additional screening methodology to identify modulators of the PLCγ family of enzymes.},

keywords = {MANTIS},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {High-throughput screening assay for PARP-HPF1 interaction inhibitors to affect DNA damage repair},

author = {Dhakar, Saurabh S.},

url = {https://www.nature.com/articles/s41598-024-54123-8},

doi = {https://doi.org/10.1038/s41598-024-54123-8},

year  = {2024},

date = {2024-02-16},

abstract = {ADP-ribosyltransferases PARP1 and PARP2 play a major role in DNA repair mechanism by detecting the DNA damage and inducing poly-ADP-ribosylation dependent chromatin relaxation and recruitment of repair proteins. Catalytic PARP inhibitors are used as anticancer drugs especially in the case of tumors arising from sensitizing mutations. Recently, a study showed that Histone PARylation Factor (HPF1) forms a joint active site with PARP1/2. The interaction of HPF1 with PARP1/2 alters the modification site from Aspartate/Glutamate to Serine, which has been shown to be a key ADP-ribosylation event in the context of DNA damage. Therefore, disruption of PARP1/2-HPF1 interaction could be an alternative strategy for drug development to block the PARP1/2 activity. In this study, we describe a FRET based high-throughput screening assay to screen inhibitor libraries against PARP-HPF1 interaction. We optimized the conditions for FRET signal and verified the interaction by competing the FRET pair in multiple ways. The assay is robust and easy to automate. Validatory screening showed the robust performance of the assay, and we discovered two compounds Dimethylacrylshikonin and Alkannin, with µM inhibition potency against PARP1/2-HPF1 interaction. The assay will facilitate the discovery of inhibitors against HPF1-PARP1/2 complex and to develop potentially new effective anticancer agents.},

keywords = {MANTIS},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Discovery of Alternative Binding Poses through Fragment-Based Identification of DHODH Inhibitors},

author = {DeRatt, Lindsey G.},

url = {https://pubs.acs.org/doi/abs/10.1021/acsmedchemlett.3c00543},

doi = {https://doi.org/10.1021/acsmedchemlett.3c00543},

year  = {2024},

date = {2024-02-07},

abstract = {Dihydroorotate dehydrogenase (DHODH) is a mitochondrial enzyme that affects many aspects essential to cell proliferation and survival. Recently, DHODH has been identified as a potential target for acute myeloid leukemia therapy. Herein, we describe the identification of potent DHODH inhibitors through a scaffold hopping approach emanating from a fragment screen followed by structure-based drug design to further improve the overall profile and reveal an unexpected novel binding mode. Additionally, these compounds had low P-gp efflux ratios, allowing for applications where exposure to the brain would be required.},

keywords = {TEMPEST},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {High-Throughput GPCRome Screen of Pollutants Reveals the Activity of Polychlorinated Biphenyls at Melatonin and Sphingosine-1-phosphate Receptors},

author = {Wilkinson, Joshua C.},

url = {https://pubs.acs.org/doi/full/10.1021/acs.chemrestox.3c00388},

doi = {https://doi.org/10.1021/acs.chemrestox.3c00388},

year  = {2024},

date = {2024-01-31},

abstract = {Exposure to environmental pollutants is linked to numerous toxic outcomes, warranting concern about the effect of pollutants on human health. To assess the threat of pollutant exposure, it is essential to understand their biological activity. Unfortunately, gaps remain for many pollutants’ specific biological activity and molecular targets. A superfamily of signaling proteins, G-protein-coupled receptors (GPCRs), has been shown as potential targets for pollutant activity. However, research investigating the pollutant activity at the GPCRome is scarce. This work explores pollutant activity across a library of human GPCRs by leveraging modern high-throughput screening techniques devised for drug discovery and pharmacology. We designed and implemented a pilot screen of eight pollutants at 314 human GPCRs and discovered specific polychlorinated biphenyl (PCB) activity at sphingosine-1-phosphate and melatonin receptors. The method utilizes open-source resources available to academic and governmental institutions to enable future campaigns that screen large numbers of pollutants. Thus, we present a novel high-throughput approach to assess the biological activity and specific targets of pollutants.},

keywords = {MANTIS},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {MicroCycle: An Integrated and Automated Platform to Accelerate Drug Discovery},

author = {Brocklehurst, Cara E.},

url = {https://pubs.acs.org/doi/abs/10.1021/acs.jmedchem.3c02029},

doi = {https://doi.org/10.1021/acs.jmedchem.3c02029},

year  = {2024},

date = {2024-01-25},

abstract = {We herein describe the development and application of a modular technology platform which incorporates recent advances in plate-based microscale chemistry, automated purification, in situ quantification, and robotic liquid handling to enable rapid access to high-quality chemical matter already formatted for assays. In using microscale chemistry and thus consuming minimal chemical matter, the platform is not only efficient but also follows green chemistry principles. By reorienting existing high-throughput assay technology, the platform can generate a full package of relevant data on each set of compounds in every learning cycle. The multiparameter exploration of chemical and property space is hereby driven by active learning models. The enhanced compound optimization process is generating knowledge for drug discovery projects in a time frame never before possible.},

keywords = {MANTIS},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Unique and Common Agonists Activate the Insect Juvenile Hormone Receptor and the Human AHR},

author = {Sedlak, David},

url = {https://www.biorxiv.org/content/10.1101/2024.01.03.574093v1},

doi = {https://doi.org/10.1101/2024.01.03.574093},

year  = {2024},

date = {2024-01-04},

abstract = {Transcription factors of the bHLH-PAS family play vital roles in animal development, physiology, and disease. Two members of the family require binding of low-molecular weight ligands for their activity: the vertebrate aryl hydrocarbon receptor (AHR) and the insect juvenile hormone receptor (JHR). In the fly Drosophila melanogaster, the paralogous proteins GCE and MET constitute the ligand-binding component of JHR complexes. Whilst GCE/MET and AHR are phylogenetically heterologous, their mode of action is similar. JHR is targeted by several synthetic agonists that serve as insecticides disrupting the insect endocrine system. AHR is an important regulator of human endocrine homeostasis and it responds to environmental pollutants and endocrine disruptors. Whether AHR signaling is affected by compounds that can activate JHR has not been reported. To address this question, we screened a chemical library of 50,000 compounds to identify 93 novel JHR agonists in a reporter system based on Drosophila cells. Of these compounds, 26% modulated AHR signaling in an analogous reporter assay in a human cell line, indicating a significant overlap in the agonist repertoires of the two receptors. To explore the structural features of agonist-dependent activation of JHR and AHR, we compared the ligand-binding cavities and their interactions with selective and common ligands of AHR and GCE. Molecular dynamics modeling revealed ligand-specific as well as conserved side chains within the respective cavities. Significance of predicted interactions was supported through site-directed mutagenesis. The results have indicated that synthetic insect juvenile hormone agonists might interfere with AHR signaling in human cells.},

keywords = {TEMPEST},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Unified framework for patient-derived, tumor-organoid-based predictive testing of standard-of-care therapies in metastatic colorectal cancer},

author = {Tan, Tao, Mouradov, Dmitri, Sieber, Oliver},

url = {https://www.sciencedirect.com/science/article/pii/S2666379123005529},

doi = {https://doi.org/10.1016/j.xcrm.2023.101335},

year  = {2023},

date = {2023-12-19},

abstract = {Predictive drug testing of patient-derived tumor organoids (PDTOs) holds promise for personalizing treatment of metastatic colorectal cancer (mCRC), but prospective data are limited to chemotherapy regimens with conflicting results. We describe a unified framework for PDTO-based predictive testing across standard-of-care chemotherapy and biologic and targeted therapy options. In an Australian community cohort, PDTO predictions based on treatment-naive patients (n = 56) and response rates from first-line mCRC clinical trials achieve 83% accuracy for forecasting responses in patients receiving palliative treatments (18 patients, 29 treatments). Similar assay accuracy is achieved in a prospective study of third-line or later mCRC treatment, AGITG FORECAST-1 (n = 30 patients). “Resistant” predictions are associated with inferior progression-free survival; misclassification rates are similar by regimen. Liver metastases are the optimal site for sampling, with testing achievable within 7 weeks for 68.8% cases. Our findings indicate that PDTO drug panel testing can provide predictive information for multifarious standard-of-care therapies for mCRC.},

keywords = {MANTIS},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Lessons from assembling a microbial natural product and pre-fractionated extract library in an academic laboratory},

author = {Cook, Michael A},

url = {https://academic.oup.com/jimb/advance-article/doi/10.1093/jimb/kuad042/7459345},

doi = {https://doi.org/10.1093/jimb/kuad042},

year  = {2023},

date = {2023-12-05},

abstract = {Microbial natural products are specialized metabolites that are sources of many bioactive

compounds including antibiotics, antifungals, antiparasitics, anticancer agents, and probes of

biology. The assembly of libraries of producers of natural products has traditionally been the

province of the pharmaceutical industry. This sector has gathered significant historical

collections of bacteria and fungi to identify new drug leads with outstanding outcomes - upwards of 60% of drug scaffolds originate from such libraries. Despite this success, the repeated rediscovery of known compounds and the resultant diminishing chemical novelty contributed to a pivot from this source of bioactive compounds toward more tractable synthetic compounds in the drug industry. The advent of advanced mass spectrometry tools, along with rapid whole genome sequencing and in silico identification of biosynthetic gene clusters that encode the machinery necessary for the synthesis of specialized metabolites, offers the opportunity to revisit microbial natural product libraries with renewed vigor. Assembling a suitable library of microbes and extracts for screening requires the investment of resources and the development of methods that have customarily been the proprietary purview of large pharmaceutical companies. Here, we

report a perspective on our efforts to assemble a library of natural product-producing microbes

and the establishment of methods to extract and fractionate bioactive compounds using resources available to most academic labs. We validate the library and approach through a series of screens for antimicrobial and cytotoxic agents. This work serves as a blueprint for establishing libraries of microbial natural product producers and bioactive extract fractions suitable for screens of bioactive compounds.},

keywords = {TEMPEST},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Mechanistic plasticity in ApmA enables aminoglycoside promiscuity for resistance},

author = {Bordeleau, Emily},

url = {https://www.nature.com/articles/s41589-023-01483-3},

doi = {https://doi.org/10.1038/s41589-023-01483-3},

year  = {2023},

date = {2023-11-16},

abstract = {The efficacy of aminoglycoside antibiotics is waning due to the acquisition of diverse resistance mechanisms by bacteria. Among the most prevalent are aminoglycoside acetyltransferases (AACs) that inactivate the antibiotics through acetyl coenzyme A-mediated modification. Most AACs are members of the GCN5 superfamily of acyltransferases which lack conserved active site residues that participate in catalysis. ApmA is the first reported AAC belonging to the left-handed β-helix superfamily. These enzymes are characterized by an essential active site histidine that acts as an active site base. Here we show that ApmA confers broad-spectrum aminoglycoside resistance with a molecular mechanism that diverges from other detoxifying left-handed β-helix superfamily enzymes and canonical GCN5 AACs. We find that the active site histidine plays different functions depending on the acetyl-accepting aminoglycoside substrate. This flexibility in the mechanism of a single enzyme underscores the plasticity of antibiotic resistance elements to co-opt protein catalysts in the evolution of drug detoxification.},

keywords = {TEMPEST},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {High-Affinity Fluorogenic Substrate for Tissue Transglutaminase Reveals Enzymatic Hysteresis},

author = {Gates, Eric W.J.},

url = {https://pubs.acs.org/doi/abs/10.1021/acs.biochem.3c00337},

doi = {https://doi.org/10.1021/acs.biochem.3c00337},

year  = {2023},

date = {2023-10-19},

abstract = {Transglutaminases (TGases) are a family of calcium-dependent enzymes primarily known for their ability to cross-link proteins. Transglutaminase 2 (TG2) is one isozyme in this family whose role is multifaceted. TG2 can act not only as a typical transamidase through its catalytic core but also as a G-protein via its GTP binding site. These two discrete activities are tightly regulated by both environmental stimuli and redox reactions. Ubiquitously expressed in humans, TG2 has been implicated in numerous disease pathologies that require extensive investigation. The catalytic activity of TG2 can be monitored through various mechanisms, including hydrolysis, transamidation, or cleavage of isopeptide bonds. Activity assays are required to monitor the activity of this isozyme not only for studying its transamidation reaction but also for validation of therapeutics designed to abolish this activity. Herein, we present the design, synthesis, and evaluation of a new TG2 activity substrate based on a previously optimized inhibitor scaffold. The substrate APH7 exhibits excellent affinity, selectivity, and reactivity with TG2 (KM = 3.0 μM). Furthermore, its application also allowed the discovery of unique hysteresis at play within the catalytic activity and inhibition reactivity of TG2.},

keywords = {FAST},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Accelerated Hit Identification with Target Evaluation, Deep Learning and Automated Labs: Prospective Validation in IRAK1},

author = {Kamuntavicius, Gintautas},

url = {https://chemrxiv.org/engage/chemrxiv/article-details/6512960eade1178b2423365f},

doi = {10.26434/chemrxiv-2023-mh22x-v3},

year  = {2023},

date = {2023-09-26},

abstract = {In this study, we integrate Ro5’s target evaluation SpectraView and DL-driven virtual screening HydraScreen tools alongside Strateos' robotic cloud labs high-throughput screening platform to accelerate target and hit identification. Using SpectraView to select IRAK1 as the target, we prospectively validate HydraScreen, a structure-based deep learning model. We demonstrate that HydraScreen could identify up to 23.8% of all IRAK1 hits in the top 1% of the ranked compounds, simultaneously identifying the three most potent (nanomolar) scaffolds present in the library. The three nanomolar scaffolds identified in our project are novel for IRAK1 and lend themselves for future development. HydraScreen outperforms traditional virtual screening methods in an unbiased prospective evaluation and offers advanced features such as ligand pose confidence scoring. Thus, SpectraView and HydraScreen are innovative tools which can aid and expedite early stages of drug discovery.},

keywords = {TEMPEST},

pubstate = {published},

tppubtype = {article}

}