First identification of small-molecule inhibitors of Pontin by combining virtual screening and enzymatic assay

2012 ◽  
Vol 443 (2) ◽  
pp. 549-559 ◽  
Author(s):  
Judith Elkaim ◽  
Michel Castroviejo ◽  
Driss Bennani ◽  
Said Taouji ◽  
Nathalie Allain ◽  
...  
Keyword(s):  
Virtual Screening ◽  
Small Molecule ◽  
Tumour Cell ◽  
Small Molecule Inhibitors ◽  
Colorimetric Assay ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Atp Binding Site ◽  
Set Up

The human protein Pontin, which belongs to the AAA+ (ATPases associated with various cellular activities) family, is overexpressed in several cancers and its silencing in vitro leads to tumour cell growth arrest and apoptosis, making it a good target for cancer therapy. In particular, high levels of expression were found in hepatic tumours for which the therapeutic arsenal is rather limited. The three-dimensional structure of Pontin has been resolved previously, revealing a hexameric assembly with one ADP molecule co-crystallized in each subunit. Using Vina, DrugScore and Xscore, structure-based virtual screening of 2200 commercial molecules was conducted into the ATP-binding site formed by a dimer of Pontin in order to prioritize the best candidates. Complementary to the in silico screening, a versatile and sensitive colorimetric assay was set up to measure the disruption of the ATPase activity of Pontin. This assay allowed the determination of inhibition curves for more than 20 top-scoring compounds, resulting in the identification of four ligands presenting an inhibition constant in the micromolar concentration range. Three of them inhibited tumour cell proliferation. The association of virtual screening and experimental assay thus proved successful for the discovery of the first small-molecule inhibitors of Pontin.

scholarly journals IN SILICO IDENTIFICATION OF APOBEC3B SMALL MOLECULE INHIBITORS FROM DTP-NCI LIBRARIES

2021 ◽  
pp. 165-170
Author(s):  
MARYAN MOHAMUD MOHAMED ◽  
NOR ATIQAH JUSRIL ◽  
MOHD ILHAM ADENAN ◽  
N. G. KWOK WEN
Keyword(s):  
Virtual Screening ◽  
Small Molecule ◽  
Small Molecule Inhibitors ◽  
3D Structure ◽  
In Vivo Studies ◽  
Amino Acid Residues ◽  
Autodock Vina ◽  
Raloxifene Hydrochloride

Objective: APOBEC3B (A3B) enzyme causes C-to-T or C-to-G somatic alteration in the cancer genome, leading to the evolution of a broad spectrum of human cancers. The present study aims to identify A3B small molecule inhibitors using a top-down approach via pharmacoinformatic virtual screening. Methods: Virtual screening of 2951 drug-alike molecules with diversified structures from the National Cancer Institute Development Therapeutics Program (DTP-NCI) compounds library was performed using GOLD and AutoDock Vina docking programs against the 3D structure of A3B (PDB ID: 5TD5). Results: Amongst the docked compounds, Nordracorubin, NSC641233 and Raloxifene hydrochloride showed the most potent binding affinities towards A3B on both Autodock/Vina and GOLD. Several significant similarities were observed between A3B and the three hits, including hydrogen bonds and pi-pi stacking. The three compounds also exhibited interaction with the centralized zinc cofactor and amino acid residues that directly contribute the deaminase activity of A3B enzyme. Conclusion: We hypothesize that the findings from this study could significantly shorten the quest for novel molecules against the A3B after confirmation with subsequent in vitro and in vivo studies in the near future.

scholarly journals A pharmacophore model for SARS-CoV-2 3CLpro small molecule inhibitors and in vitro experimental validation of computationally screened inhibitors

2021 ◽  
Author(s):  
Enrico Glaab ◽  
Ganesh Babu Manoharan ◽  
Daniel Abankwa
Keyword(s):  
Machine Learning ◽  
Molecular Dynamics ◽  
Virtual Screening ◽  
Small Molecule ◽  
Drug Target ◽  
Experimental Validation ◽  
Small Molecule Inhibitors ◽  
Pharmacophore Model ◽  
Computational Screening

AbstractAmong the biomedical efforts in response to the current coronavirus (COVID-19) pandemic, pharmacological strategies to reduce viral load in patients with severe forms of the disease are being studied intensively. One of the main drug target proteins proposed so far is the SARS-CoV-2 viral protease 3CLpro (also called Mpro), an essential component for viral replication. Ongoing ligand- and receptor-based computational screening efforts would be facilitated by an improved understanding of the electrostatic, hydrophobic and steric features that characterize small molecule inhibitors binding stably to 3CLpro, as well as by an extended collection of known binders.Here, we present combined virtual screening, molecular dynamics simulation, machine learning and in vitro experimental validation analyses which have led to the identification of small molecule inhibitors of 3CLpro with micromolar activity, and to a pharmacophore model that describes functional chemical groups associated with the molecular recognition of ligands by the 3CLpro binding pocket. Experimentally validated inhibitors using a ligand activity assay include natural compounds with available prior knowledge on safety and bioavailability properties, such as the natural compound rottlerin (IC50 = 37 μM), and synthetic compounds previously not characterized (e.g. compound CID 46897844, IC50 = 31 μM). In combination with the developed pharmacophore model, these and other confirmed 3CLpro inhibitors may provide a basis for further similarity-based screening in independent compound databases and structural design optimization efforts, to identify 3CLpro ligands with improved potency and selectivity.Overall, this study suggests that the integration of virtual screening, molecular dynamics simulations and machine learning can facilitate 3CLpro-targeted small molecule screening investigations. Different receptor-, ligand- and machine learning-based screening strategies provided complementary information, helping to increase the number and diversity of identified active compounds. Finally, the resulting pharmacophore model and experimentally validated small molecule inhibitors for 3CLpro provide resources to support follow-up computational screening efforts for this drug target.

scholarly journals Nanofiber-Mâché Hollow Ball Mimicking the Three-Dimensional Structure of a Cyst

Polymers ◽  
2021 ◽  
Vol 13 (14) ◽  
pp. 2273
Author(s):  
Wan-Ying Huang ◽  
Norichika Hashimoto ◽  
Ryuhei Kitai ◽  
Shin-ichiro Suye ◽  
Satoshi Fujita
Keyword(s):  
Three Dimensional ◽  
Hollow Structure ◽  
Dimensional Structure ◽  
The Novel ◽  
Fibrous Scaffold ◽  
Hydrogel Beads ◽  
Inner Surface ◽  
Intracranial Epidermoid ◽  
Hollow Ball

The occasional malignant transformation of intracranial epidermoid cysts into squamous cell carcinomas remains poorly understood; the development of an in vitro cyst model is urgently needed. For this purpose, we designed a hollow nanofiber sphere, the “nanofiber-mâché ball.” This hollow structure was fabricated by electrospinning nanofiber onto alginate hydrogel beads followed by dissolving the beads. A ball with approximately 230 mm3 inner volume provided a fibrous geometry mimicking the topography of the extracellular matrix. Two ducts located on opposite sides provided a route to exchange nutrients and waste. This resulted in a concentration gradient that induced oriented migration, in which seeded cells adhered randomly to the inner surface, formed a highly oriented structure, and then secreted a dense web of collagen fibrils. Circumferentially aligned fibers on the internal interface between the duct and hollow ball inhibited cells from migrating out of the interior, similar to a fish bottle trap. This structure helped to form an adepithelial layer on the inner surface. The novel nanofiber-mâché technique, using a millimeter-sized hollow fibrous scaffold, is excellently suited to investigating cyst physiology.

Discovery of Novel Small Molecule Inhibitors of Dengue Viral NS2B-NS3 Protease Using Virtual Screening and Scaffold Hopping

2012 ◽  
Vol 55 (14) ◽  
pp. 6278-6293 ◽  
Author(s):  
Jing Deng ◽  
Ning Li ◽  
Hongchuan Liu ◽  
Zhili Zuo ◽  
Oi Wah Liew ◽  
...  
Keyword(s):  
Virtual Screening ◽  
Small Molecule ◽  
Small Molecule Inhibitors ◽  
Scaffold Hopping ◽  
Ns3 Protease

EXTH-31. BRAIN DISTRIBUTION, CLEARANCE AND TOXICITY EVALUATION OF SMALL-MOLECULE INHIBITORS FOLLOWING CONVECTION-ENHANCED DELIVERY TO THE BRAINSTEM

2021 ◽  
Vol 23 (Supplement_6) ◽  
pp. vi170-vi170
Author(s):  
Erica Power ◽  
Juhee Oh ◽  
Jonghoon Choi ◽  
William Elmquist ◽  
David Daniels
Keyword(s):  
Small Molecule ◽  
Drug Concentration ◽  
Small Molecule Inhibitors ◽  
Efflux Pump ◽  
Sprague Dawley ◽  
Convection Enhanced Delivery ◽  
Efflux Pump Inhibitor ◽  
Delivery Technique ◽  
The Brain

Abstract BACKGROUND Diffuse midline gliomas (DMGs) harboring the H3K27M mutation are highly aggressive, fatal brainstem tumors that primarily occur in children. The blood-brain barrier (BBB) prevents numerous drugs from reaching CNS tumors, like DMG, at cytotoxic concentrations. Convection-enhanced delivery (CED) has emerged as a drug delivery technique that bypasses the BBB through a direct interstitial infusion under a pressure gradient. However, drug distribution and clearance from the brain following CED is poorly understood and has been cited as a potential reason for the lack of efficacy observed in prior clinical trials. OBJECTIVE The objective of this study was to understand how two small molecule inhibitors (alisertib, ponatinib) that inhibit cell growth and proliferation in DMG cells in vitro distribute and clear from the brain following CED to the brainstem. METHODS Sprague-dawley rats underwent a single 60mL CED infusion of drug to the brainstem (200mM alisertib, 10mM ponatinib) and were sacrificed 0.083, 1, 2, 4, 8 and 24 hours following the completion of the infusion. Brains were dissected and drug concentration was determined via HPLC analysis. RESULTS No rats showed any clinical or neurological signs of toxicity post-infusion. Both drugs showed significant differences in drug concentration based on anatomical brain region where higher concentrations were observed in the pons and cerebellum compared to the cortex. Drug half-life in the brain was ~0.5 hours for alisertib and ~1 hour for ponatinib, but this was not significantly increased following co-administration of elacridar, a BBB efflux pump inhibitor. CONCLUSIONS These results suggest that elimination of drugs from the brain in a complex, multifactorial mechanism that warrants further preclinical investigation prior to the initiation of a clinical trial.

scholarly journals Protein Folding: Search for Basic Physical Models

2003 ◽  
Vol 3 ◽  
pp. 623-635 ◽  
Author(s):  
Ivan Y. Torshin ◽  
Robert W. Harrison
Keyword(s):  
Protein Folding ◽  
Tertiary Structure ◽  
Three Dimensional ◽  
Physical Models ◽  
Dimensional Structure ◽  
Computational Techniques ◽  
Linear Sequence ◽  
Physical Forces

How a unique three-dimensional structure is rapidly formed from the linear sequence of a polypeptide is one of the important questions in contemporary science. Apart from biological context ofin vivoprotein folding (which has been studied only for a few proteins), the roles of the fundamental physical forces in thein vitrofolding remain largely unstudied. Despite a degree of success in using descriptions based on statistical and/or thermodynamic approaches, few of the current models explicitly include more basic physical forces (such as electrostatics and Van Der Waals forces). Moreover, the present-day models rarely take into account that the protein folding is, essentially, a rapid process that produces a highly specific architecture. This review considers several physical models that may provide more direct links between sequence and tertiary structure in terms of the physical forces. In particular, elaboration of such simple models is likely to produce extremely effective computational techniques with value for modern genomics.

scholarly journals Functional Analysis of the Mycobacterium tuberculosis FAD-Dependent Thymidylate Synthase, ThyX, Reveals New Amino Acid Residues Contributing to an Extended ThyX Motif

2008 ◽  
Vol 190 (6) ◽  
pp. 2056-2064 ◽  
Author(s):  
Jonathan E. Ulmer ◽  
Yap Boum ◽  
Christopher D. Thouvenel ◽  
Hannu Myllykallio ◽  
Carol Hopkins Sibley
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Thymidylate Synthase ◽  
Three Dimensional ◽  
Pyrimidine Ring ◽  
Dimensional Structure ◽  
Directed Mutagenesis ◽  
Catalytic Residue ◽  
Amino Acid Residues ◽  
Insight Into

ABSTRACT A novel FAD-dependent thymidylate synthase, ThyX, is present in a variety of eubacteria and archaea, including the mycobacteria. A short motif found in all thyX genes, RHRX7-8S, has been identified. The three-dimensional structure of the Mycobacterium tuberculosis ThyX enzyme has been solved. Building upon this information, we used directed mutagenesis to produce 67 mutants of the M. tuberculosis thyX gene. Each enzyme was assayed to determine its ability to complement the defect in thymidine biosynthesis in a ΔthyA strain of Escherichia coli. Enzymes from selected strains were then tested in vitro for their ability to catalyze the oxidation of NADPH and the release of a proton from position 5 of the pyrimidine ring of dUMP. The results defined an extended motif of amino acids essential to enzyme activity in M. tuberculosis (Y44X24 H69X25R95HRX7 S105XRYX90R199 [with the underlined histidine acting as the catalytic residue and the underlined serine as the nucleophile]) and provided insight into the ThyX reaction mechanism. ThyX is found in a variety of bacterial pathogens but is absent in humans, which depend upon an unrelated thymidylate synthase, ThyA. Therefore, ThyX is a potential target for development of antibacterial drugs.

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