Abstract
Solute carrier proteins (SLCs) control fluxes of ions and molecules across biological membranes and represent an emerging class of drug targets. SLC11A2 (hDMT1) mediates intestinal iron uptake and its inhibition might be used to treat iron overload diseases such as hereditary hemochromatosis. Here we report a micromolar (IC50 = 1.1 μM) pyrazolyl-pyrimidone inhibitor of radiolabeled iron uptake in hDMT1 overexpressing HEK293 cells acting by a non-competitive mechanism, which however does not affect the electrophysiological properties of the transporter. Isothermal titration calorimetry, competition with calcein, induced precipitation of radioactive iron and cross inhibition of the unrelated iron transporter SLC39A8 (hZIP8) indicate that inhibition is mediated by metal chelation. Mapping the chemical space of thousands of pyrazolo-pyrimidones and similar 2,2'-diazabiaryls in ChEMBL suggests that their reported activities might partly reflect metal chelation. Such metal chelating groups are not listed in pan-assay interference compounds (PAINS) but should be checked when addressing SLCs.
Highlights
Solute carrier proteins (SLCs) control fluxes of ions and molecules across biological membranes and represent an emerging class of drug targets
The synthesis of pyrazolones 21–25 only gave a single product whose structures were confirmed by an X-ray crystal structure of 24 as hydrochloride salt (Fig. 2)
Note that potentially metal chelating 2,2′-diazabiaryls such as pyrazolyl-pyrimidones are not listed in pan-assay interference compounds (PAINS).[39,40,41]
Summary
Pathologies such as hereditary hemochromatosis, β-thalassemia, Parkinson's disease and Alzheimer's disease, highlighting that its pharmacological inhibition may be beneficial to treat human diseases.[4,5,6,7,8,9] While metal chelators have been classically used to treat metal intoxication[10] and neurodegenerative diseases,[11,12] a recently reported highly potent and specific inhibitor of ferroportin (SLC40), a different iron transporter acting on the same pathway as hDMT1, has been shown to have clinical efficacy against β-thalassemia.[13]. Two families of small molecule hDMT1 inhibitors have been reported in the literature as the results of high-throughput screening campaigns, namely bis-cationic isothioureas such as dibenzofurans 1 and mesitylene 2,14 as well as pyrazolyl-pyridine 3 (Fig. 1).[15] In our own investigations on hDMT1, we used the inhibitors mentioned above as seeds for a ligand-based virtual screening campaign guided by 3D-shape and pharmacophore similarity[16] and discovered bis-isothiourea 4 and pyrazolylpyrimidone 5 as two additional hDMT1 inhibitors.[17] Kinetic studies, an X-ray structure of a related brominated bisisothiourea inhibitor in complex with a bacterial analog of the transporter and mutational studies recently showed that bisisothiourea-based compounds act as competitive inhibitors of hDMT1.18 On the other hand, pyrazolyl-pyrimidone 5, whose small size and better drug-like properties made it an attractive candidate compound, acted as a non-competitive inhibitor. We set out to investigate the pyrazolyl series (3 and 5) closer and understand its mechanism of action
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