Monday, April 30, 2018

Dimethyl fumarate targets GAPDH and aerobic glycolysis to modulate immunity

Michael D. Kornberg, Pavan Bhargava, Paul M. Kim, Vasanta Putluri, Adele M. Snowman, Nagireddy Putluri, Peter A. Calabresi Solomon H. Snyder

Science, 2018: eaan4665
DOI: 10.1126/science.aan4665

Highlight in "In the Pipeline"

Activated immune cells undergo a metabolic switch to aerobic glycolysis akin to the Warburg effect, thereby presenting a potential therapeutic target in autoimmune disease. Dimethyl fumarate (DMF), a derivative of the Krebs cycle intermediate fumarate, is an immunomodulatory drug used to treat multiple sclerosis and psoriasis. Although its therapeutic mechanism remains uncertain, DMF covalently modifies cysteine residues in a process termed succination. We found that DMF succinates and inactivates the catalytic cysteine of the glycolytic enzyme glyceraldehyde 3-phosphate dehydrogenase (GAPDH) in mice and humans, both in vitro and in vivo. It thereby down-regulates aerobic glycolysis in activated myeloid and lymphoid cells, which mediates its anti-inflammatory effects. Our results provide mechanistic insight into immune modulation by DMF and represent a proof of concept that aerobic glycolysis is a therapeutic target in autoimmunity.

Saturday, April 28, 2018

Affinity‐driven Covalent Modulator of Glyceraldehyde‐3‐phosphate Dehydrogenase (GAPDH) Cascade


Jeffy Chern  Chun-Ping Lu  Zhanxiong Fang  Ching-Ming Chang  Kuo-Feng Hua Yi-Ting Chen  Cheng Yang Ng  Yi-Lin Sophia Chen  Yulin Lam  Shih-Hsiung Wu

Angewandte Chemie, 2018

doi: 10.1002/ange.201801618

Traditional medicines provide a fertile ground to explore potent lead compounds; yet, their transformation into modern drugs is fraught with challenges in deciphering the target that is mechanistically valid for its biological activity. Herein we revealed Z‐(+)‐isochaihulactone 1 exhibited significant inhibition against MDR cancer cell lines and mice xenografts. By NMR spectroscopy, 1 was shown to resist to an off‐targeting thiolate, thus giving 1 a target covalent inhibitor‐like (TCI) property. By identifying the pharmacophore of 1 (α,β‐unsaturated moiety), a 1‐derived probe was designed and synthesized for TCI‐oriented activity‐based proteome profiling (TCI‐ABPP). With MS/MS and computer‐guided molecular biology approaches, we elucidated that an affinity‐driven Michael addition of the noncatalytic C247 of GAPDH controlled the "ON/OFF" switch of apoptosis through non‐canonically nuclear GAPDH translocation, which bypasses the common apoptosis‐resistant route of MDR cancers.

Thursday, April 26, 2018

The Discovery of Osimertinib (TAGRISSO™): An Irreversible Inhibitor of Activating and T790M Resistant Forms of the Epidermal Growth Factor Receptor Tyrosine Kinase for the Treatment of Non‐Small Cell Lung Cancer

Michael J. Waring

Successful Drug Discovery, 2018
doi: 10.1002/9783527808694.ch12

Non‐small cell lung cancer (NSCLC) consists of a group of diseases that account for 80‐85% of lung cancer cases. Signaling through epidermal growth factor receptor (EGFR) is important for the maintenance of healthy tissue in particular epithelial cells, but it is also highly expressed in a number of cancers and in particular NSCLC. Despite initial encouraging responses to treatment in EGFR activating mutant tumors, it has been observed that resistance to tyrosine kinase inhibitor (TKI) therapy develops fairly rapidly. Pioneering work at Zeneca Pharmaceuticals in the 1990s led to the discovery of the anilinoquinazoline class of EGFR inhibitors. An analysis of responses to therapy led to the discovery of activating mutations in the EGFR kinase domain. This mutation is a point mutation at threonine 790, which becomes methionine (T790M). This chapter shows tumor responses in T790M‐positive patients in response to osimertinib treatment.

Saturday, April 21, 2018

Identification of Noncatalytic Lysine Residues from Allosteric Circuits via Covalent Probes

Jens Bongard, Marian Lorenz, Ingrid R. Vetter, Patricia Stege, Arthur T. Porfetye, Anna Laura Schmitz, Farnusch Kaschani, Alex Wolf, Uwe Koch, Peter Nussbaumer, Bert Klebl, Markus Kaiser, and Michael Ehrmann

ACS Chem. Biol., Article ASAP
DOI: 10.1021/acschembio.8b00101

Covalent modifications of nonactive site lysine residues by small molecule probes has recently evolved into an important strategy for interrogating biological systems. Here, we report the discovery of a class of bioreactive compounds that covalently modify lysine residues in DegS, the rate limiting protease of the essential bacterial outer membrane stress response pathway. These modifications lead to an allosteric activation and allow the identification of novel residues involved in the allosteric activation circuit. These findings were validated by structural analyses via X-ray crystallography and cell-based reporter systems. We anticipate that our findings are not only relevant for a deeper understanding of the structural basis of allosteric activation in DegS and other HtrA serine proteases but also pinpoint an alternative use of covalent small molecules for probing essential biochemical mechanisms.
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Reactive Chemical Probes: Beyond the Kinase Cysteinome

Lyn Jones

Angewandte Chemie, 2018

doi: 10.1002/ange.201802693

The reaction of small molecule chemical probes with proteins has been harnessed to develop covalent inhibitor drugs and protein profiling technologies. This Essay discusses some of the recent enhancements to the chemical biology toolkit that are enabling the study of previously unchartered areas of chemoproteomic space. An analysis of the kinome is used to illustrate the potential for these approaches to pursue new targets using reactive chemical probes.

Tuesday, April 17, 2018

Cysteine-reactive probes and their use in chemical proteomics

Dominic G. Hoch, Daniel Abegg, and Alexander Adibekian [web]

Chem. Commun., 2018, doi: 10.1039/C8CC01485J 

Proteomic profiling using bioorthogonal chemical probes that selectively react with certain amino acids is now a widely used method in life sciences to investigate enzymatic activities, study posttranslational modifications and discover novel covalent inhibitors. Over the past two decades, researchers have developed selective probes for several different amino acids, including lysine, serine, cysteine, threonine, tyrosine, aspartate and glutamate. Among these amino acids, cysteines are particularly interesting due to their highly diverse and complex biochemical role in our cells. In this feature article, we focus on the chemical probes and methods used to study cysteines in complex proteomes.

Tuesday, April 10, 2018

Structure-based design of targeted covalent inhibitors

Richard Lonsdale  and  Richard A. Ward

Chem. Soc. Rev., 2018
doi: 10.1039/C7CS00220C

Covalent inhibition is a rapidly growing discipline within drug discovery. Many historical covalent inhibitors were discovered by serendipity, with such a mechanism of action often regarded as undesirable due to potential toxicity issues. Recent progress has seen a major shift in this outlook, as covalent inhibition shows promise for targets where previous efforts to identify non-covalent small molecule inhibitors have failed. Targeted covalent inhibitors (TCIs) can offer drug discovery scientists the ability to increase the potency and/or selectivity of small molecule inhibitors, by attachment of reactive functional groups designed to covalently bind to specific sites in a target. In this tutorial review we introduce the broader concept of covalent inhibition, discuss the potential benefits and challenges of such an approach, and provide an overview of the current status of the field. We also describe some strategies and computational tools to enable successful covalent drug discovery.

Thursday, April 5, 2018

Reversible Covalent Reaction of Levosimendan with Cardiac Troponin C in Vitro and in Situ

Brittney A. Klein, Béla Reiz, Ian M. Robertson, Malcolm Irving, Liang Li , Yin-Biao Sun, and Brian D. Sykes

Biochemistry, 2018
doi: 10.1021/acs.biochem.8b00109

The development of calcium sensitizers for the treatment of systolic heart failure presents difficulties, including judging the optimal efficacy and the specificity to target cardiac muscle. The thin filament is an attractive target because cardiac troponin C (cTnC) is the site of calcium binding and the trigger for subsequent contraction. One widely studied calcium sensitizer is levosimendan. We have recently shown that when a covalent cTnC–levosimendan analogue is exchanged into cardiac muscle cells, they become constitutively active, demonstrating the potency of a covalent complex. We have also demonstrated that levosimendan reacts in vitro to form a reversible covalent thioimidate bond specifically with cysteine 84, unique to cTnC. In this study, we use mass spectrometry to show that the in vitro mechanism of action of levosimendan is consistent with an allosteric, reversible covalent inhibitor; to determine whether the presence of the cTnI switch peptide or changes in either Ca2+ concentration or pH modify the reaction kinetics; and to determine whether the reaction can occur with cTnC in situ in cardiac myofibrils. Using the derived kinetic rate constants, we predict the degree of covalently modified cTnC in vivo under the conditions studied. We observe that covalent bond formation would be highest under the acidotic conditions resulting from ischemia and discuss whether the predicted level could be sufficient to have therapeutic value. Irrespective of the in vivo mechanism of action for levosimendan, our results provide a rationale and basis for the development of reversible covalent drugs to target the failing heart.

Tuesday, April 3, 2018

Itaconate is an anti-inflammatory metabolite that activates Nrf2 via alkylation of KEAP1

Evanna L. Mills, Dylan G. Ryan, Hiran A. Prag, Dina Dikovskaya, Deepthi Menon, Zbigniew Zaslona, Mark P. Jedrychowski, Ana S. H. Costa, Maureen Higgins, Emily Hams, John Szpyt, Marah C. Runtsch, Martin S. King, Joanna F. McGouran, Roman Fischer, Benedikt M. Kessler, Anne F. McGettrick, Mark M. Hughes, Richard G. Carroll, Lee M. Booty, Elena V. Knatko, Paul J. Meakin, Michael L. J. Ashford, Louise K. Modis, Gino Brunori, Daniel C. Sévin, Padraic G. Fallon, Stuart T. Caldwell, Edmund R. S. Kunji, Edward T. Chouchani, Christian Frezza, Albena T. Dinkova-Kostova, Richard C. Hartley, Michael P. Murphy & Luke A. O’Neill

Nature, 2018
doi:10.1038/nature25986

The endogenous metabolite itaconate has recently emerged as a regulator of macrophage function, but its precise mechanism of action remains poorly understood1,2,3. Here we show that itaconate is required for the activation of the anti-inflammatory transcription factor Nrf2 (also known as NFE2L2) by lipopolysaccharide in mouse and human macrophages. We find that itaconate directly modifies proteins via alkylation of cysteine residues. Itaconate alkylates cysteine residues 151, 257, 288, 273 and 297 on the protein KEAP1, enabling Nrf2 to increase the expression of downstream genes with anti-oxidant and anti-inflammatory capacities. The activation of Nrf2 is required for the anti-inflammatory action of itaconate. We describe the use of a new cell-permeable itaconate derivative, 4-octyl itaconate, which is protective against lipopolysaccharide-induced lethality in vivo and decreases cytokine production. We show that type I interferons boost the expression of Irg1 (also known as Acod1) and itaconate production. Furthermore, we find that itaconate production limits the type I interferon response, indicating a negative feedback loop that involves interferons and itaconate. Our findings demonstrate that itaconate is a crucial anti-inflammatory metabolite that acts via Nrf2 to limit inflammation and modulate type I interferons.

Thiol Reactivity of N-Aryl α-Methylene-γ-lactams: Influence of the Guaianolide Structure [@KayBrummond]

 Daniel P. Dempe, Chong-Lei Ji, Peng Liu, and Kay M. Brummond The Journal of Organic Chemistry, 2020 DOI: 10.1021/acs.joc.2c01530 The α-meth...