In mature erythrocytes, glucose-6-phosphate dehydrogenase (G6PDH) and 6-phosphogluconate dehydrogenase (6PGDH) yield NADPH, a crucial cofactor of the enzyme glutathione reductase (GR) converting glutathione disulfide (GSSG) into its reduced state (GSH). GSH is essential for detoxification processes in and survival of erythrocytes. We explored whether the anti-inflammatory compounds Bay 11-7082, parthenolide and dimethyl fumarate (DMF) were able to completely deplete a common target (GSH), and to impair the function of upstream enzymes of GSH recycling and replenishment. Treatment of erythrocytes with Bay 11-7082, parthenolide or DMF led to concentration-dependent eryptosis resulting from complete depletion of GSH. GSH depletion was due to strong inhibition of G6PDH activity. Bay 11-7082 and DMF, but not parthenolide, were able to inhibit the GR activity. This approach "Inhibitors, Detection of their common target that is completely depleted or inactivated when pharmacologically relevant concentrations of each single inhibitor are applied, Subsequent functional analysis of upstream enzymes for this target" (IDS), can be applied to a broad range of inhibitors and cell types according to the selected target. The specific G6PDH inhibitory effect of these compounds may be exploited for the treatment of human diseases with high NADPH and GSH consumption rates, including malaria, trypanosomiasis, cancer or obesity.

Centre for Ophthalmology Institute for Ophthalmic Research Eberhard Karls University Tübingen Frondsbergstr 23 72076 Tübingen Germany

Department of Biology Faculty of Medicine and Dentistry Palacky University Hnevotinska 3 77515 Olomouc Czech Republic

Department of Cardiology Vascular Medicine and Physiology University of Tübingen Gmelinstr 5 72076 Tübingen Germany

Department of Dermatology and Allergy Biedersteinerstr 29 Technische Universität München 80802 München Germany

Department of Dermatology; Eberhard Karls University Tübingen Germany

Department of Life Sciences Laboratory of Pharmacology and Toxicology University of Siena Via A Moro 2 53100 Siena Italy

Division of Radiobiology and Molecular Environmental Research Department of Radiation Oncology Roentgenweg 11 Eberhard Karls University Tübingen 72076 Tübingen Germany

Institute of Physiology 2 Keplerstr 15 Eberhard Karls University of Tübingen 72074 Tübingen Germany

Laboratory of Biophysics Saarland University Campus A2 4 66123 Saarbrücken Germany

Psoriasis Center Department of Dermatology University Medical Center Schleswig Holstein Campus Kiel Schittenhelmstr 7 Kiel 24105 Germany

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