Cytokine screening identifies TNF to potentially enhance immunogenicity of pediatric sarcomas.

Introduction: Pediatric sarcomas, including osteosarcoma (OS), Ewing sarcoma (EwS) and rhabdomyosarcoma (RMS) carry low somatic mutational burden and low MHC-I expression, posing a challenge for T cell therapies. Our previous study showed that mediators of monocyte maturation sensitized the EwS cell line A673 to lysis by HLA-A*02:01/CHM1-specific allorestricted T cell receptor (TCR) transgenic CD8 T cells (CHM1 CD8 T cells).

Methods: In this study, we tested a panel of monocyte maturation cytokines for their ability to upregulate immunogenic cell surface markers on OS, EwS and RMS cell lines, using flow cytometry.

50 Years of Organoselenium Chemistry, Biochemistry and Reactivity: Mechanistic Understanding, Successful and Controversial Stories.

In 1973, two major discoveries changed the face of selenium chemistry: the identification of the first mammal selenoenzyme, glutathione peroxidase 1, and the discovery of the synthetic utility of the so-called selenoxide elimination. While the chemical mechanism behind the catalytic activity of glutathione peroxidases appears to be mostly unveiled, little is known about the mechanisms of other selenoproteins and, for some of them, even the function lies in the dark. In chemistry, the capacity of organoselenides of catalyzing hydrogen peroxide activation for the practical manipulation of organic functional groups has been largely explored, and some mechanistic details have been clearly elucidated.

The glutathione peroxidase family: Discoveries and mechanism.

The discoveries leading to our present understanding of the glutathione peroxidases (GPxs) are recalled. The cytosolic GPx, now GPx1, was first described by Mills in 1957 and claimed to depend on selenium by Rotruck et al., in 1972.

Selenium-Catalyzed Reduction of Hydroperoxides in Chemistry and Biology.

Among the chalcogens, selenium is the key element for catalyzed HO reduction. In organic synthesis, catalytic amounts of organo mono- and di-selenides are largely used in different classes of oxidations, in which HO alone is poorly efficient. Biological hydroperoxide metabolism is dominated by peroxidases and thioredoxin reductases, which balance hydroperoxide challenge and contribute to redox regulation.

Proton Transfer and S 2 Reactions as Steps of Fast Selenol and Thiol Oxidation in Proteins: A Model Molecular Study Based on GPx.

Invited for this month's cover are collaborating groups from Università degli Studi di Padova, Vrije Universiteit Amsterdam, and Universidad de la República Uruguay. The cover picture shows two lorries along the road directed to the destination 'H O reduction', and the selenol (SeH) lorry is faster than the thiol (SH) lorry. This cartoon represents the situation of glutathione peroxidase (GPx), in which the presence of selenium rather than sulfur warrants a significantly faster hydroperoxide reduction along the same mechanistic path.

Looking Back at the Early Stages of Redox Biology.

The beginnings of redox biology are recalled with special emphasis on formation, metabolism and function of reactive oxygen and nitrogen species in mammalian systems. The review covers the early history of heme peroxidases and the metabolism of hydrogen peroxide, the discovery of selenium as integral part of glutathione peroxidases, which expanded the scope of the field to other hydroperoxides including lipid hydroperoxides, the discovery of superoxide dismutases and superoxide radicals in biological systems and their role in host defense, tissue damage, metabolic regulation and signaling, the identification of the endothelial-derived relaxing factor as the nitrogen monoxide radical (more commonly named nitric oxide) and its physiological and pathological implications. The article highlights the perception of hydrogen peroxide and other hydroperoxides as signaling molecules, which marks the beginning of the flourishing fields of redox regulation and redox signaling.

Proton Transfer and S 2 Reactions as Steps of Fast Selenol and Thiol Oxidation in Proteins: A Model Molecular Study Based on GPx.

The so-called peroxidatic cysteines and selenocysteines in proteins reduce hydroperoxides through a dual attack to the peroxide bond in a two-step mechanism. First, a proton dislocation from the thiol/selenol to a close residue of the enzymatic pocket occurs. Then, a nucleophilic attack of the anionic cysteine/selenocysteine to one O atom takes place, while the proton is shuttled back to the second O atom, promoting the formation of a water molecule.

A dual attack on the peroxide bond. The common principle of peroxidatic cysteine or selenocysteine residues.

The (seleno)cysteine residues in some protein families react with hydroperoxides with rate constants far beyond those of fully dissociated low molecular weight thiol or selenol compounds. In case of the glutathione peroxidases, we could demonstrate that high rate constants are achieved by a proton transfer from the chalcogenol to a residue of the active site [Orian et al. Free Radic.

Regulatory Phenomena in the Glutathione Peroxidase Superfamily.

The selenium-containing Glutathione peroxidases (GPxs)1-4 protect against oxidative challenge, inhibit inflammation and oxidant-induced regulated cell death. GPx1 and GPx4 dampen phosphorylation cascades predominantly prevention of inactivation of phosphatases by HO or lipid hydroperoxides. GPx2 regulates the balance between regeneration and apoptotic cell shedding in the intestine.

Selenoprotein Gene Nomenclature.

The human genome contains 25 genes coding for selenocysteine-containing proteins (selenoproteins). These proteins are involved in a variety of functions, most notably redox homeostasis. Selenoprotein enzymes with known functions are designated according to these functions: TXNRD1, TXNRD2, and TXNRD3 (thioredoxin reductases), GPX1, GPX2, GPX3, GPX4, and GPX6 (glutathione peroxidases), DIO1, DIO2, and DIO3 (iodothyronine deiodinases), MSRB1 (methionine sulfoxide reductase B1), and SEPHS2 (selenophosphate synthetase 2).

Selenium and redox signaling.

Selenium compounds that contain selenol functions or can be metabolized to selenols are toxic via superoxide and HO generation, when ingested at dosages beyond requirement. At supra-nutritional dosages various forms of programmed cell death are observed. At physiological intakes, selenium exerts its function as constituent of selenoproteins, which overwhelmingly are oxidoreductases.

Helmut Sies and the compartmentation of hydroperoxide metabolism.

The early work of Helmut Sies on mammalian hydroperoxide metabolism is reviewed with particular emphasis on the in situ function of catalase and glutathione peroxidase1. Starting out from a catalase-dominated thinking in the middle of the last century, Sies first demonstrated, by whole organ spectroscopy, that H2O2 is generated in rat liver and metabolized by catalase. In a joined effort with the author's group, he then worked out that glutathione peroxidase can kinetically compete with catalase in hydroperoxide metabolism in situ.

Identification of Novel Chemical Scaffolds Inhibiting Trypanothione Synthetase from Pathogenic Trypanosomatids.

Background: The search for novel chemical entities targeting essential and parasite-specific pathways is considered a priority for neglected diseases such as trypanosomiasis and leishmaniasis. The thiol-dependent redox metabolism of trypanosomatids relies on bis-glutathionylspermidine [trypanothione, T(SH)2], a low molecular mass cosubstrate absent in the host. In pathogenic trypanosomatids, a single enzyme, trypanothione synthetase (TryS), catalyzes trypanothione biosynthesis, which is indispensable for parasite survival.

Selenocysteine oxidation in glutathione peroxidase catalysis: an MS-supported quantum mechanics study.

Glutathione peroxidases (GPxs) are enzymes working with either selenium or sulfur catalysis. They adopted diverse functions ranging from detoxification of H(2)O(2) to redox signaling and differentiation. The relative stability of the selenoenzymes, however, remained enigmatic in view of the postulated involvement of a highly unstable selenenic acid form during catalysis.

Genetic and chemical analyses reveal that trypanothione synthetase but not glutathionylspermidine synthetase is essential for Leishmania infantum.

Trypanothione is a unique and essential redox metabolite of trypanosomatid parasites, the biosynthetic pathway of which is regarded as a promising target for antiparasitic drugs. Synthesis of trypanothione occurs by the consecutive conjugation of two glutathione molecules to spermidine. Both reaction steps are catalyzed by trypanothione synthetase (TRYS), a molecule known to be essential in Trypanosoma brucei.

Identification of M. tuberculosis thioredoxin reductase inhibitors based on high-throughput docking using constraints.

A virtual screening campaign is presented that led to small molecule inhibitors of thioredoxin reductase of Mycobacterium tuberculosis (MtTrxR) that target the protein-protein interaction site for the substrate thioredoxin (Trx). MtTrxR is a promising drug target because it dominates the Trx-dependent hydroperoxide metabolism and the reduction of ribonucleotides, thus facilitating survival and proliferation of M. tuberculosis.

Molecular dynamics reveal binding mode of glutathionylspermidine by trypanothione synthetase.

The trypanothione synthetase (TryS) catalyses the two-step biosynthesis of trypanothione from spermidine and glutathione and is an attractive new drug target for the development of trypanocidal and antileishmanial drugs, especially since the structural information of TryS from Leishmania major has become available. Unfortunately, the TryS structure was solved without any of the substrates and lacks loop regions that are mechanistically important. This contribution describes docking and molecular dynamics simulations that led to further insights into trypanothione biosynthesis and, in particular, explains the binding modes of substrates for the second catalytic step.

The fairytale of the GSSG/GSH redox potential.

Background: The term GSSG/GSH redox potential is frequently used to explain redox regulation and other biological processes.

Scope Of Review: The relevance of the GSSG/GSH redox potential as driving force of biological processes is critically discussed. It is recalled that the concentration ratio of GSSG and GSH reflects little else than a steady state, which overwhelmingly results from fast enzymatic processes utilizing, degrading or regenerating GSH.

The trypanothione system and its implications in the therapy of trypanosomatid diseases.

Biosynthesis and the use of trypanothione, a redox metabolite of parasitic trypanosomatids, are reviewed here with special emphasis on the development of trypanocidal drugs. This metabolic system is unique to and essential for the protozoal parasites. Selective inhibition of key elements of trypanothione metabolism, therefore, promises eradication of the parasites without affecting the host.

A fast virtual screening approach to identify structurally diverse inhibitors of trypanothione reductase.

Trypanothione reductase (TryR) is one of the favorite targets for those designing drugs for the treatment of Chagas disease. We present the application of a fast virtual screening approach for designing hit compounds active against TryR. Our protocol combines information derived from structurally known inhibitors and from the TryR receptor structure.

The trypanothione system and the opportunities it offers to create drugs for the neglected kinetoplast diseases.

Parasitic trypanosomatids (Kinetoplastida) are the causative agents of devastating and hard-to-treat diseases such as African sleeping sickness, Chagas disease and various forms of Leishmaniasis. Altogether they affect > 30 Million patients, account for half a million fatalities p.a.