COMMENTARY COMMENTARY Systems approach to metal- based pharmacology Isolda Romero-Canelón and Peter J. Sadler1 There is an abundance of thiolate sulfur in Department of Chemistry, University of Warwick, Coventry CV4 7AL, United Kingdom cells. Most cells contain 2–14 mM glutathi- one, the tripeptide γ-L-glutamyl-L-cysteinyl- glycine (GSH), not only in the At least 20 elements of the periodic table are centuries. Preparations such as bismuth sub- but also in other organelles, e.g., mitochondria essential to humans, but understanding their salicylate (Pepto-Bismol), colloidal bismuth (5). Hong et al. have shown that uptake of Bi biochemistry presents many challenges. The subcitrate (CBS) (De-Nol), and ranitidine bis- into HK-2 human proximal tubular cells and genetic codes for metals such as iron, , muthcitrate(Pylorid)arewidelyusedtoday. bacteria (Escherichia coli) occurs by passive and are readily identified, but the same However, why is bismuth much more toxic to diffusion and is biphasic, the second phase cannot be said for vanadium, chromium, microbes such as Helicobacter pylori compared mirroring exponential growth of HK-2, but nickel, or tin, metals frequently in mineral with eukaryotic cells? In PNAS, Hong et al. (3) not E. coli. Moreover, Bi uptake greatly supplements. The genetic codes for metals show that a systems pharmacology approach increases GSH levels (approximately four are not just for the elements, they are for can provide new insights, an approach that, if times).Biuptakeis“self-propelled” by a feed- particular species of the elements. The metal applied more generally, will greatly advance back loop. Stimulation of GSH biosynthesis oxidation state, types and numbers of coor- the rational design of metallodrugs. by an external supply of cystine as a cysteine dinated ligands, as well as the coordination The speciation of bismuth itself is excep- precursor led to a remarkable accumulation of geometry are crucial for recognition by spe- tionally challenging: only one natural isotope 34 mM Bi in HK-2 cells. In contrast, on de- 209 cific uptake, transport, and delivery . ( Bi), a quadrupolar nucleus that is of little pletion of GSH using L-buthionine sulfoximine, Understanding and controlling the biochem- use for NMR, with no unpaired spins, no use- an inhibitor of γ-glutamyl-cysteine synthase istry of metal ions requires speciation on ful metal-centered electronic transitions, and (glutamate-cysteine ligase), the first step in timescales of nanoseconds to years (kinet- a highly variable and fluxional coordination GSH synthesis, CBS gave rise to HK-2 cell ics of ligand substitution), consideration of sphere with coordination numbers of 3–9. degradation and nucleus malformation. Co- + 3+ metal–ligand affinities (thermodynamics, equi- These features and the strong acidity of Bi3 - ordination of Bi to GSH in cells, forming ∼ librium constants), as well as redox potentials OH2 (pKa 1.5) lead to complicated stoichio- Bi(SG)3, triggers other events. This (con- + (1, 2). Medicines use not only essential, but metries for its compounds (“sub” salts). Bi3 is jugation) step may be aided by GSTs, which also nonessential elements. For example, bis- a borderline class “a/b,”“‘hard/soft’” metal ion, constitute up to 10% of cytosolic proteins. muth, the heaviest stable element, has been binding strongly not only to (hard) oxygen Thewidefamilyofmultidrugresistance- used for the treatment of infectious ligands, but also to (soft) sulfur, especially thi- associated proteins (MRPs) mediate ATP- and gastrointestinal conditions for over three olates (4). dependent transport of GSH conjugates, so decreasing cellular drug accumulation (con- ferring resistance). MRPs also pump conju- gates into perinuclear vesicles, where Bi is seen as dense black deposits, probably Bi sulfide (inclusion bodies). The nature of Bi (aswellasAgandPb)“intranuclear inclusion bodies” has long been a puzzle (6, 7). Impor- tantly, detoxification of Bi in human cells is not so effective in bacteria where GSH is less prevalent. Where does the sulfide come from? —via the action of cysteine desulfurase on cys- teine as it does for ferredoxins? Perhaps it arises from cleavage of the γ-glutamyl γ residue of Bi(SG)3 by -glutamyltransferase (γ-GT), which initiates mercapturic acid syn- thesis. In many anaerobic bacteria, GSH is absent and cysteine is the major thiol. Hence Bi drugs can be effective and selective against

Author contributions: I.R.-C. and P.J.S. wrote the paper. Fig. 1. Involvement of GSH, some GSH-dependent , and membrane pumps in cellular reactions of As(III), The authors declare no conflict of interest. Pt(II), Bi(III), and Au(I) drugs and essential Cu(I). Lysosomal deposits (gold aurosomes) or intranuclear inclusion bodies (Bi) are curious features also seen for some other metals. Pt(II) exchanges ligands more slowly and its reactions have See companion article on page 3211 in issue 11 of volume 112. some different features. These pathways can detoxify metals; their inhibition might be built into, e.g., an anticancer 1To whom correspondence should be addressed. Email: p.j.sadler@ strategy. Speciation of metal ions in intact cells is a major challenge. GSH also has redox functions in cells. warwick.ac.uk.

www.pnas.org/cgi/doi/10.1073/pnas.1503858112 PNAS | April 7, 2015 | vol. 112 | no. 14 | 4187–4188 Downloaded by guest on September 30, 2021 pathogens that cause syphilis and dysentery, inhibits GSH-peroxidase in vivo, which con- areas of research that use the rich structural for example. tributes to increased oxidative stress (13). diversity of metallic scaffolds for drug design. Many other soft metal ions have a high Auranofin exerts its antiinflammatory ac- Metal-based drugs can provide in-cell chem- affinity for thiolate sulfur and GSH seques- tivity by inhibiting mitochondrial thiore- ical reactions that are not accessible to purely tration pathways might be expected to be doxin reductase (TrxR) (14); nonetheless, organic drugs, accompanied by multitargeting involved in their too. These its anticancer activity can be enhanced by that can combat resistance (17). However, include anticancer drugs based on As(III) there is an urgent need for further devel- and Pt(II), antiarthritic Au(I) drugs, as well Hong et al. show that opment of methods to elucidate the spatial as the essential metal ion Cu(I) (Fig. 1). The a systems pharmacol- and temporal speciation of metallodrugs. As(III) drug Trisenox [As2O3,As(OH)3 in This requires a multidisciplinary effort with aqueous solution], the drug of choice for pro- ogy approach can pro- contributions like those of Hong et al. (3) myelocytic leukemia, forms GSH conjugates vide new insights, an to understand fully the implications of the that have been related to poor patient out- metal center in the metabolism and phar- come. These GSH-adducts are GST-substrates approach that, if macokinetic/dynamic properties of candi- easily detoxified by cancer cells (8). In con- applied more generally, date drugs. trast, the drug candidate Zinapar (darinapar- will greatly advance It is evident from this brief commentary sin, ZIO-101), dimethylarsenic conjugated to that progress in the design of metallodrugs glutathione, takes advantage of the excessive the rational design of requires a systems pharmacology approach. cystine/cysteine requirement of cancer cells. metallodrugs. Notonlydoweneedagoodunderstandingof This As(III) complex appears to be trans- the kinetics and thermodynamics of metal– formed by γ-GT at the cell surface with direct binding and subsequent inhibition of ligand exchange and redox reactions, but formation of the dimethylarsino-cysteine GST (15). Interestingly, a member of the also of the responses of cells in terms of adduct (DMAC), which is readily taken GST family, TDR1, activates antiparasitic attack on multiple targets, self-propelled up via cystine/cysteine importers. Hence in- antimonial prodrugs by deglutathionylation transport, and delivery processes related to creased cellular accumulation of As from of Sb(V)-glutathione adducts with reduction feedback loops (Fig. 1). For metallodrugs, it ZIO-101 occurs. This complex might be and release of Sb(III) (16). will often not be appropriate to describe their favored for multidrug-resistant tumors where There is a link to the metabolism of essential mechanisms of action in terms of a single chemoresistance is mediated by high GSH metal ions. GSH stimulates the uptake of target site. This “polypharmacology” offers levels (9). copper via the transporter Ctr1. Initial potential advantages because it is now be- I 2− In the case of Pt(II)-based anticancer drugs, formation of [Cu (SG)3] may be followed by coming apparent that resistance to single- GSH may be responsible for the inactivation delivery to P1B-type ATPases and superoxide target drugs can readily develop. The intro- ofcompoundssuchascisplatin,cis-diammi- dismutase mediated by the dynamic transfer of duction of systems pharmacology approaches nedichloroplatinum(II),oritsderivativescar- Cu(I) between cysteine thiolates of chaperones to unraveling the mechanism of action of boplatin and oxaliplatin. Indeed, at least 60% Atox1 and CCS in the cytoplasm (11). metallodrugs should lead to advances in their of cellular Pt is present as GSH-adducts and The field of and design and in their use so as to avoid un- increased levels of intracellular GSH are di- its applications in medicine are fast-growing wanted side effects. rectly linked to the development of resistance to Pt(II) drugs in lung and ovarian cancers (10). For ovarian cancer, proteomic studies 1 Chellan P, Sadler PJ (2015) The elements of life and medicines. 10 Sawers L, et al. (2014) Glutathione S-transferase P1 (GSTP1) have related differentially expressed GSTs Philos Trans A Math Phys Eng Sci 373(2037):20140182. directly influences platinum drug chemosensitivity in ovarian tumour 2 Barry NPE, Sadler PJ (2013) Exploration of the medical periodic cell lines. Br J Cancer 111(6):1150–1158. to Pt chemoresistance. Both and table: Towards new targets. Chem Commun (Camb) 49(45): 11 Dolgova NV, Nokhrin S, Yu CH, George GN, Dmitriev OY (2013) 5106–5131. carboplatin generate adducts in which Pt is Copper chaperone Atox1 interacts with the metal-binding domain of 3 Hong Y, Lai Y-T, Chan GC-F, Sun H (2015) Glutathione and Wilson’s in cisplatin detoxification. Biochem J 454(1): bound to the CXXC copper-binding motif multidrug resistance protein transporter mediate a self-propelled 147–156. disposal of bismuth in human cells. Proc Natl Acad Sci USA of methyl-CpG binding domain protein 2 – 112(11):3211–3216. 12 Ghadially FN (1979) The aurosome. J Rheumatol Suppl 5:45 50. (MBD2), and then excreted by ATP7B, 13 Reglinski J, et al. (1997) Myocrisin-mediated oxidative stress. Clin 4 Sun H (2011) Biological Chemistry of Arsenic, Antimony and – aCuexporter(11). Bismuth (Wiley, Chichester, UK). Chim Acta 268(1-2):85 99. Injectable Au(I) thiolate drugs such as 5 Marí M, et al. (2013) Mitochondrial glutathione: Features, 14 Casini A (2012) Exploring the mechanisms of metal-based regulation and role in disease. Biochim Biophys Acta 1830(5): pharmacological agents via an integrated approach. J Inorg Biochem Myochrysine (aurothiomalate) and the oral 3317–3328. 109:97–106. drug Ridaura [auranofin; (tetraacetylthioglu- 6 Wachstein M (1949) Studies on inclusion bodies; acid-fastness of 15 De Luca A, Hartinger CG, Dyson PJ, Lo Bello M, Casini A (2013) A nuclear inclusion bodies that are induced by ingestion of lead and new target for gold(I) compounds: Glutathione-S-transferase cose)Au(PEt3)] are used for treating rheuma- bismuth. Am J Clin Pathol 19(7):608–614. toid arthritis. It has long been known that inhibition by auranofin. J Inorg Biochem 119(I):38–42. 7 Fowler BA, Goyer RA (1975) Bismuth localization within nuclear 16 FyfePK,WestropGD,SilvaAM,CoombsGH,HunterWN inclusions by x-ray microanalysis. Effects of accelerating voltage. gold accumulates in lysosomes in cells as (2012) Leishmania TDR1 structure, a unique trimeric glutathione J Histochem Cytochem 23(10):722–726. dense deposits (aurosomes), widely believed transferase capable of deglutathionylation and antimonial 8 Sui M, Zhang Z, Zhou J (2014) Inhibition factors of arsenic trioxide prodrug activation. Proc Natl Acad Sci USA 109(29): to be sulfur-rich (12), but they are not well therapeutic effects in patients with acute promyelocytic leukemia. – characterized. In view of the observations on Chin Med J (Engl) 127(19):3503–3506. 11693 11698. 9 Garnier N, et al. (2014) The novel arsenical darinaparsin is 17 Romero-Canelón I, Sadler PJ (2013) Next-generation metal Bi by Hong et al. (3), it now seems to be the transported by cystine importing systems. Mol Pharmacol 85(4): anticancer complexes: Multitargeting via redox modulation. time to reexamine aurosomes. Myochrysine 576–585. Inorg Chem 52(21):12276–12291.

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