The Identification, Characterization and Optimization of Small Molecule Probes of Cysteine Proteases: Experiences of the Penn Center for Molecular Discovery with Cathepsin B and Cathepsin L

Huryn, Donna M.; Smith, Amos B.

doi:10.2174/156802609789753653

Cited by 7 publications

(5 citation statements)

References 40 publications

(40 reference statements)

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“…25−28 Both in vivo and in vitro studies have demonstrated that certain cathepsin B inhibitors reduce tumor cell motility and invasiveness. 29,30 Because of these links, considerable effort has been made to identify cathepsin B inhibitors.…”

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confidence: 99%

“…The system is applied to cathepsin B, a cysteine protease implicated in tumorigenesis, arthritis, and parasite infection. − Both in vivo and in vitro studies have demonstrated that certain cathepsin B inhibitors reduce tumor cell motility and invasiveness. , Because of these links, considerable effort has been made to identify cathepsin B inhibitors. Successful inhibitors include epoxysuccinyl, aziridinyl, biguanide, and β-lactam derivatives. , The MS screen used here reveals several potential new inhibitors.…”

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confidence: 99%

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Droplet Electrospray Ionization Mass Spectrometry for High Throughput Screening for Enzyme Inhibitors

Sun

Kennedy

2014

Anal. Chem.

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High throughput screening (HTS) is important for identifying molecules with desired properties. Mass spectrometry (MS) is potentially powerful for label-free HTS due to its high sensitivity, speed, and resolution. Segmented flow, where samples are manipulated as droplets separated by an immiscible fluid, is an intriguing format for high throughput MS because it can be used to reliably and precisely manipulate nanoliter volumes and can be directly coupled to electrospray ionization (ESI) MS for rapid analysis. In this study, we describe a “MS Plate Reader” that couples standard multiwell plate HTS workflow to droplet ESI-MS. The MS plate reader can reformat 3072 samples from eight 384-well plates into nanoliter droplets segmented by an immiscible oil at 4.5 samples/s and sequentially analyze them by MS at 2 samples/s. Using the system, a label-free screen for cathepsin B modulators against 1280 chemicals was completed in 45 min with a high Z-factor (>0.72) and no false positives (24 of 24 hits confirmed). The assay revealed 11 structures not previously linked to cathepsin inhibition. For even larger scale screening, reformatting and analysis could be conducted simultaneously, which would enable more than 145 000 samples to be analyzed in 1 day.

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confidence: 99%

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confidence: 99%

Droplet Electrospray Ionization Mass Spectrometry for High Throughput Screening for Enzyme Inhibitors

Sun

Kennedy

2014

Anal. Chem.

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“…However, no orally bioavailable, brain effecting, cathepsin B-specific inhibitor is currently available and developing such will be difficult and will take considerable resources and time. For example, a high throughput screen of the National Institutes of Health (NIH) small molecule library for a cathepsin B-specific inhibitor did not find a druggable hit ( 309 ). The known cathepsin B-specific inhibitor, CA-074, is not orally druggable and when made more so with the addition of a methyl group, it looses specificity.…”

Section: E64d Is a Promising Drug Inhibitor Of Cathepsin B For Preclimentioning

confidence: 99%

Cathepsin B is a New Drug Target for Traumatic Brain Injury Therapeutics: Evidence for E64d as a Promising Lead Drug Candidate

et al. 2015

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There is currently no therapeutic drug treatment for traumatic brain injury (TBI) despite decades of experimental clinical trials. This may be because the mechanistic pathways for improving TBI outcomes have yet to be identified and exploited. As such, there remains a need to seek out new molecular targets and their drug candidates to find new treatments for TBI. This review presents supporting evidence for cathepsin B, a cysteine protease, as a potentially important drug target for TBI. Cathepsin B expression is greatly up-regulated in TBI animal models, as well as in trauma patients. Importantly, knockout of the cathepsin B gene in TBI mice results in substantial improvements of TBI-caused deficits in behavior, pathology, and biomarkers, as well as improvements in related injury models. During the process of TBI-induced injury, cathepsin B likely escapes the lysosome, its normal subcellular location, into the cytoplasm or extracellular matrix (ECM) where the unleashed proteolytic power causes destruction via necrotic, apoptotic, autophagic, and activated glia-induced cell death, together with ECM breakdown and inflammation. Significantly, chemical inhibitors of cathepsin B are effective for improving deficits in TBI and related injuries including ischemia, cerebral bleeding, cerebral aneurysm, edema, pain, infection, rheumatoid arthritis, epilepsy, Huntington’s disease, multiple sclerosis, and Alzheimer’s disease. The inhibitor E64d is unique among cathepsin B inhibitors in being the only compound to have demonstrated oral efficacy in a TBI model and prior safe use in man and as such it is an excellent tool compound for preclinical testing and clinical compound development. These data support the conclusion that drug development of cathepsin B inhibitors for TBI treatment should be accelerated.

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“…They are viewed as potentially useful drug targets, as their catalytic activity can be inhibited by small molecules that have become an attractive field for the pharmaceutical industry [24][25][26][27]. Cathepsin L is a lysosomal protease that is the main catalyst in intracellular protein catabolism, is associated with atherosclerosis, metabolic syndrome and cancer, and has also been related to bone resorption, sperm cell maturation, macrophage malfunction and Alzheimer's disease [28][29][30][31].…”

Section: Introductionmentioning

confidence: 99%

Molecular dynamics simulation of halogen bonding mimics experimental data for cathepsin L inhibition

Celis-Barros

Saavedra-Rivas

Salgado

et al. 2014

J Comput Aided Mol Des

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A MD simulation protocol was developed to model halogen bonding in protein-ligand complexes by inclusion of a charged extra point to represent the anisotropic distribution of charge on the halogen atom. This protocol was then used to simulate the interactions of cathepsin L with a series of halogenated and non-halogenated inhibitors. Our results show that chloro, bromo and iodo derivatives have progressively narrower distributions of calculated geometries, which reflects the order of affinity I > Br > Cl, in agreement with the IC50 values. Graphs for the Cl, Br and I analogs show stable interactions between the halogen atom and the Gly61 carbonyl oxygen of the enzyme. The halogen-oxygen distance is close to or less than the sum of the van der Waals radii; the C-X···O angle is about 170°; and the X···O=C angle approaches 120°, as expected for halogen bond formation. In the case of the iodo-substituted analogs, these effects are enhanced by introduction of a fluorine atom on the inhibitors' halogen-bonding phenyl ring, indicating that the electron withdrawing group enlarges the σ-hole, resulting in improved halogen bonding properties.

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The Identification, Characterization and Optimization of Small Molecule Probes of Cysteine Proteases: Experiences of the Penn Center for Molecular Discovery with Cathepsin B and Cathepsin L

Cited by 7 publications

References 40 publications

Droplet Electrospray Ionization Mass Spectrometry for High Throughput Screening for Enzyme Inhibitors

Droplet Electrospray Ionization Mass Spectrometry for High Throughput Screening for Enzyme Inhibitors

Cathepsin B is a New Drug Target for Traumatic Brain Injury Therapeutics: Evidence for E64d as a Promising Lead Drug Candidate

Molecular dynamics simulation of halogen bonding mimics experimental data for cathepsin L inhibition

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