Versatile synthesis of cationic N-heterocyclic carbene–gold(i) complexes containing a second ancillary ligand. Design of heterobimetallic ruthenium–gold anticancer agents

ChemComm COMMUNICATION Versatile synthesis of cationic N-heterocyclic carbene–gold( ) complexes containing a second I Citethis:DOI:10.1039/c5cc09718e ancillary ligand. Design of heterobimetallic ruthenium–gold anticancer agents† Received24thNovember2015, Accepted14thJanuary2016 DOI:10.1039/c5cc09718e Jacob Ferna´ndez-Gallardo,a Benelita T. Elie,ab Mercedes Sanau´ c and Mar´ıa Contel*abd www.rsc.org/chemcomm Wedescribeaversatileandquickroutetocationicgold(I)complexes containing N-heterocyclic carbenes and a second ancillary ligand (suchasphosphanes,phosphites,arsinesandamines)ofinterestfor thesynthesisofcompoundswithpotentialcatalyticandmedicinal applications.Thegeneralsyntheticstrategyhasbeenappliedinthe preparationofnovelcationicheterobimetallicruthenium(II)–gold(I) complexes that are highly cytotoxic to renal cancer Caki-1 and coloncancerHCT116celllineswhileshowingasynergisticeffect andbeingmoreselectivethantheirmonometalliccounterparts. Scheme1 Preparationofgoldcomplexes[(IPr)Au(L)]A,L=PPh,A=ClO(cid:3) 3 4 (2a),A=CFSO(cid:3)(2b),A=ClO(cid:3),L=PEt (3),L=P(OPh) (4),L=AsPh (5), 3 3 4 3 3 3 The chemistry of gold(I) complexes containing N-heterocyclic L=bipy(6).(i)AgClO 4 orAgOSO 2 CF 3 (1eq.),CH 2 Cl 2 /diethylether1:1,from carbenes1 has developed rapidly in the past decade with the 01CtoRT,15min.(ii)L(1eq.),CH 2 Cl 2 /diethylether2:1,RT,15min. findingsoftheirrelevantapplicationsashomogeneouscatalysts1–7 and their potential as anticancer and antimicrobial agents7–14 by addition of the corresponding chlorido(phosphane) gold(I) (includingsomeheterometalliccomplexes15,16).Thebeneficial derivativeswithreactiontimesrangingfrom8to60hours,and effects of a second ligand in processes catalyzed by gold(I)–N- subsequentpurificationbyextractioninCH 2 Cl 2 /H 2 O.18,19This heterocycliccompoundshavebeendescribed.17Cationicgold(I)–N- method limits the nature of the anion to that present in the heterocyclic carbene complexes containing phosphanes of the imidazolium salt precursor. A second method developed by type [(NHC)AuPR ]I (NHC = 1,3-diethylbenzimidazol-2-ylidene) Nolan et al. for the preparation of [(IPr)Au(PR )]+ compounds 3 3 have been described recently by Ott et al. as very efficient involvesprotonolysisof[(IPr)Au(OH)]derivativeswithappropriate enzyme inhibitors with promising potential as anticancer PR (cid:2)HBF salts(RTandreactiontimesof14hours).20 3 4 chemotherapeutics.18,19 Wedescribehereasimpleandefficientsyntheticmethodto Thepreparationofthesecomplexesinvolvestheadditionof obtain gold(I)–N-heterocyclic carbenes with a second ancillary NaCO toN-heterocyclicimidazoliumiodideat501Cfollowed ligandbyabstractionofthechloridewithsilverperchloratein 3 compounds [(NHC)AuCl]21 and subsequent addition of the ancillary ligand (Scheme 1, total reaction time 30 min). The aDepartmentofChemistry,BrooklynCollegeandTheGraduateCenter, reactionschemeisquitegeneralanddifferentphosphanessuch TheCityUniversityofNewYork,Brooklyn,NY,11210,USA. E-mail:mariacontel@brooklyn.cuny.edu as PPh 3 and PEt 3 can be used. We also synthesized cationic bBiologyPhDProgram,TheGraduateCenter,TheCityUniversityofNewYork, complexescontainingNHCandaphosphiteP(OEt ) ,triphenyl- 33 365FifthAvenue,NewYork,NY,10016,USA arsineAsPh andbipyridine(bipy)assecondligands. cDepartamentodeQu´ımicaInorga´nica,UniversidaddeValencia,Burjassot, 3 Gold(I) compounds with hydrogen-bond-supported hetero- Valencia,46100,Spain dChemistryPhDProgram,TheGraduateCenter,TheCityUniversityofNewYork, cyclic carbene (HBHC) and nitrogen acyclic carbene (NAC) of 365FifthAvenue,NewYork,NY,10016,USA the type [(carbene)Au(AsPh 3 )][SbF 6 ] have been described by †Electronic supplementary information (ESI) available: Experimental section Espinetetal.byonepotreactionofgoldprecursors[(carbene)- (general information, instrumentation and synthetic procedures), NMR, IR, AuCl] and AgSBF 6 /AsPh 3 (2–5 h reactions).17 Cationic gold(I) UV-visandMS-ESI+HRspectra,crystallographicdata,interactionwithplasmid compoundsofthetype[(NHC)Au(a-diimine)]X(X=BF (cid:3),SbF (cid:3)) pBR322DNA,cellcultureandcellviabilityassaysandinhibitionofthioredoxin 4 6 havealsobeenreportedbyBraunsteinetal.byonepotreaction reductase.CCDC1436326and1436327.ForESIandcrystallographicdatainCIF orotherelectronicformatseeDOI:10.1039/c5cc09718e of[(NHC)AuCl]/a-diimineandTlPF 6 orAgX(24hreaction).21 Thisjournalis©TheRoyalSocietyofChemistry2016 Chem.Commun. .25:35:21 6102/10/52 no ytisrevinU notgnihsaW egroeG yb dedaolnwoD .6102 yraunaJ 52 no dehsilbuP View Article Online View Journal Communication ChemComm Fordifferentsynthetic/catalyticpurposesitmayberelevant to separate AgCl formed in the abstraction reaction before additionofthesecondancillaryligand.Wehavedemonstrated that the use of a silver salt containing an anion that can be covalently coordinated to gold allows for the separation of solutions (after filtration of AgCl) containing in situ generated [(NHC)AuOClO ]speciesthatarestableforatleast72hat51C, 3 andfor2hoursatRT.ThecovalentnatureoftheOClO (cid:3)group 3 in the intermediate has been confirmed by IR spectroscopy (spectraandexplanationsareprovidedintheESI†).Theaddition ofdifferentancillaryligandstothesesolutionsleadstoimmediate formation of stable cationic species (2–6) that precipitate in Scheme2 Preparation of heterometallic ruthenium–gold complexes the reaction media in high yields and that can be separated [Ru(p-cymene)Cl(m-dppm)Au(NHC)]ClO. (i) AgClO (1 eq.), CHCl/diethyl 2 4 4 2 2 by filtration without further purification (Scheme 1). The ether1:1,from01CtoRT,15min.(ii)[Ru(p-cymene)Cl(Z1-dppm)]9(1eq.), 2 reactioncanalsobeperformedusingsilvertriflate(AgOSO 2 CF 3 ) CH 2 Cl 2 /diethylether2:1,RT,15min. (2b).Itshouldbenotedthatcompound4isastableprecursor to the phosphane derivatives by displacement of the more labileAsPh . Following the general reaction scheme reported here we 3 Wehaveusedthisgeneralsyntheticmethodtoincorporate obtained cationic heterobimetallic Ru–Au complexes (13–16) a phosphane fragment containing a second metal center and with four different N-heterocyclic carbenes in high yields generatenewcationicheterobimetallicruthenium–goldcomplexes. (Scheme 2). These compounds could not have been prepared Ourgrouphasbeeninvolvedinthepreparationofheterometallic following any of the previously reported methods.17–20,22 The complexes containing gold(I) phosphane moieties as potential new heterometallic compounds (see the ESI† for a detailed cancer chemotherapeutics.23–26 The hypothesis is that the synthetic procedure and characterization) are soluble in DMSO/ incorporation of two active metals in the same molecule may H O,DMSO/PBSorDMSO/medium(1:99)mixturesatmicromolar 2 improve their activity as anti-tumor agents due to the inter- concentrations,whichisrelevantforsubsequentbiologicaltesting. action of the different metals with multiple biological targets Compounds 13–16 were characterized unambiguously by (cooperativeeffects)orbytheimprovedchemicophysicalproperties 31P{1H}, 1H and 13C{1H} NMR and IR spectroscopy, MS ESI oftheresultingheterometalliccompound(synergism).Wehave spectrometry and by elemental analysis (see the Experimental prepared a number of titanocene–gold derivatives with high sectionintheESI†).Inaddition,theionicnatureofthecompounds efficacy in ovarian and prostate cancer in vitro23 and in renal canbeascertainedbyconductivitymeasurementsthatconfirm cancer both in vitro24 and in vivo.25 We and others have also that they are 1:1 electrolytes. The anionic nature of ClO (cid:3) is 4 reported recently on the design of heterometallic ruthenium– also confirmed by IR spectroscopy (ESI†). The 31P{1H} NMR gold complexes with relevant in vitro properties against HCT spectra in CDCl of the heterometallic complexes show two 3 116coloncancercelllines(7and8inChart1).26Wefoundin distinct doublets at 26.7/20.0 ppm (13), 25.3/21.1 ppm (14), mostcasesasynergisticeffectoftheheterometalliccompound 28.7/21.5(15)and28.1/21.4ppm(16)duetocoordinationofthe whencomparedtoitsmonometalliccounterparts(eitheralone dppmligandtotheAu(I)andRu(II)centersrespectively.Moreover, orincombination).23–26 thesignalscorrespondingtothecarbonsoftheimidazoliumring We aimed to incorporate gold(I)–N-heterocyclic fragments show a doublet in all cases due to the coupling between the into [Ru(p-cymene)Cl (Z1-dppm)] 9 species to improve the carbenic (2J B 128 Hz) and the ethylenic (4J B 3 Hz) 2 PC PC pharmacological profile of previously reported heterometallic carbons and the phosphorous of the dppm ligand bound to complexes 7 and 8. Gold NHC complexes display strong anti- thegoldcenter. mitochondrial effects, which can be attributed to both their Single crystals of compounds 13 and 14 were isolated as cationicandlipophiliccharacteristics.27,28Combiningthepotential bright orange needles in dichloromethane/Et O mixtures. The 2 antimetastatic/cytotoxic properties of [Ru(p-cymene)Cl(Z1-dppm)] structureof13isdepictedbelowalongwithselectedanglesand 2 929andtheantimitochondrial/TrRxinhibitioneffectsof[(NHC)Au]+ distances.Thestructureof14iscollectedintheESI†alongwith and[(NHC)Au(PR )]+species10,18,19maygiverisetopotentcancer crystallographic data and tables of selected distances and 3 chemotherapeutics. anglesforboth13and14(quitesimilar)(Fig.1). Coordinationbondlengthsandanglesofthetwometalions in13and14(ESI†)areinagreementwiththosefoundforsimilar complexesretrievedinasearchintheCSD(v.1.16)26,30andinthe structureofpreviouslydescribed[Ru(p-cymene)Cl (m-dppm)AuCl] 2 (7).26ThebondlengthsAu(1)–P(2)andAu(1)–C(1)are,respectively, 2.228(7) and 2.052(12) Å while the angle P(2)–Au(1)–C(1) is Chart1 Previous heterometallic Ru–Au complexes synthesized in our 165.3(4)1showingastrongerdeviationofthelinearitypreferred group.26 by gold(I) and being considerably smaller than the angle of Chem.Commun. Thisjournalis©TheRoyalSocietyofChemistry2016 .25:35:21 6102/10/52 no ytisrevinU notgnihsaW egroeG yb dedaolnwoD .6102 yraunaJ 52 no dehsilbuP View Article Online ChemComm Communication Table1 IC values(mM)inhumancelllinesweredeterminedwithmono- 50 metallic[(NHC)AuCl]1,10–12monometallic[Ru(p-cymene)Cl(Z1-dppm)]9, 2 andnewheterometallicRu–Aucompounds(13–16)a Caki-1 HEK-293T HCT116 [(NHC)AuCl] 1 27.1(cid:4)2.0 61.5(cid:4)5.1 39.7(cid:4)4.9 10 21.2(cid:4)1.6 4100 31.2(cid:4)3.2 11 58.8(cid:4)3.9 4100 59.1(cid:4)5.8 12 17.5(cid:4)2.2 4100 27.7(cid:4)4.9 [Ru(p-cymene)Cl (Z1-dppm)]9 35.6(cid:4)3.7 81.6(cid:4)3.0 18.2(cid:4)2.2 2 NewcationicRu–Au 13 5.2(cid:4)0.9 73.2(cid:4)2.5 8.1(cid:4)1.8 14 14.1(cid:4)1.9 4100 5.22(cid:4)0.7 Fig.1 ORTEPviewofthemolecularstructuresof13showingthelabelling 15 3.8(cid:4)0.6 4100 6.4(cid:4)1.0 scheme.Labellingforhydrogenandcarbonatomsisomittedforclarity. 16 12.7(cid:4)2.7 4100 9.6(cid:4)3.1 A drawing of the molecular structure containing relevant carbon atoms Cisplatin 29(cid:4)4.11b 3.27(cid:4)0.13b — labelled is provided in the ESI.† Selected distances (Å) and angles (1): aAllcompoundsweredissolvedin1%ofDMSOanddilutedwithwater Au(1)–P(2) = 2.228(7), Au(1)–C(1) = 2.052(12), Ru(1)–P(1) = 2.352(3), before additionto cell culturemedium for a 72 h incubationperiod. Ru(1)–Cl(1) = 2.409(3), Ru(1)–Cl(2) = 2.422(3), Ru(1)–Z* = 1.719; CisplatinwasdissolvedinH O.bValuesfromref.24. 2 P(2)–Au(1)–C(1)=165.3(4),Cl(1)–Ru(1)–Cl(2)=87.65(11),Cl(1)–Ru(1)–P(1)= 85.39(10),Cl(2)–Ru(1)–P(1)=86.80(10),Ru(1)–P(1)–Z*=132.32.Z*=centroid also IC values above 100 micromolar for the HEK cell line but ofthep-cymenering. 50 is less toxic to the renal cancer cell line Caki-1. We studied the effect of the combination of monometallic gold 1, 10–12 and 177.88(9)1foundforheterobimetallic7.26Theruthenium(II)ion ruthenium9compounds(1:1equivalents)inCaki-1celllinesat exhibitstheexpectedpseudooctahedralthreeleggedpiano-stool 72 h which in all cases gave IC values larger than those of the 50 arrangement common for half-sandwich Ru(II) phosphine bimetallic compounds (ESI†). This fact supports the idea that complexesandasfoundin7. thereisindeedasynergisticeffectfortheheterometalliccomplexes The stability of the new heterometallic compounds 13–16 intheirinvitroactivityontherenalcancercellline. wasevaluatedby31P{1H}and1HNMRspectroscopyinDMSO-d We analyzed the interactions of 13–16 with representative 6 and by UV-vis spectroscopy in DMSO/PBS solution. NMR DNAmolecules.Specifically,wecarriedoutgelelectrophoresis experiments were performed in neat DMSO-d due to the low studies to disclose the effects of the new heterobimetallic 6 solubility of the compounds in DMSO-d /D O mixtures in compounds on plasmid (pBR322) DNA (ESI†). Compound 6 2 concentrations larger than 100 micromolar. The compounds [Ru(p-cymene)Cl (Z1-dppm)] 9 weakly reduced supercoiling in 2 arestableinDMSO-d withhalf-lifetimesrangingfrom2to15 plasmidDNAalthoughitwasabletobindDNAinanon-intercalative 6 days.TheUV-visspectraofselectedcompound15(micromolar fashion.31 Most gold(I)–N-heterocyclic carbene complexes (unless concentration) in 1:99 DMSO/PBS solution did not change in designedtotargetDNAquadruplexes)areknowntonotinteractor 24 h, with a half-life 472 h (see the ESI†) indicating the interactonlyweaklywithDNA.10Treatmentwithincreasingamounts stabilityofthesecomplexesinphysiologicalmedia. ofheterometallicRuAuderivatives13–16didnotaffectthemobility The cytotoxicity of the heterometallic complexes [Ru(p- ofthefaster-runningsupercoiledform(FormI)evenatthehighest cymene)Cl (m-dppm)Au(NHC)]ClO (13–16) and monometallic molarratios(ESI†).Thelackofinteractionbetweenthehetero- 2 4 gold(I) complexes [(NHC)AuCl] (1, 10–12 in Scheme 2) was bimetallic compounds and plasmid (pBR322) DNA (already assayedbymonitoringtheirabilitytoinhibitcellgrowthusing observed for previously described [Ru(p-cymene)Cl (m-dppm)AuX] 2 the BluePrestoTM assay (see details in the ESI†). In this assay, compounds, X = Cl 7, thiazoline 8) points out that other human renal Caki-1 and human colon HCT 116 and DLD1 biomolecular targets are probably implicated in the cell death cancercelllinesandnon-tumorigenichumanembryonickidney pathway. celllinesHEK-293Twereincubatedwiththeindicatedcompound Changes in cellanti-oxidant capacity are a characteristic of for 72 hours and compared for sensitivity to cisplatin, and manychemo-resistantcancers,andoverexpressionofthioredoxin [Ru(p-cymene)Cl (Z1-dppm)] 9. The results are summarized in reductase(TrRx)isamongthekeydefenseandsurvivalmechanisms 2 Table1(acompletetableincludingdataontheDLD1cellline of cisplatin-resistant cells. We have reported on the inhibition of and the combination of monometallic 9 and 1, 10–12 can be TrRx in Caki-1 cells by auranofin and heterometallic titanocene– found in the ESI†). The heterometallic compounds are more goldcomplexes.25SinceAu–NHCcompoundsareknowntoinhibit toxicto the renalcancer cell line (Caki-1cells)and colonHCT TrRx, we measured the activity of thioredoxin reductase in Caki-1 116 than cisplatin, monometallic [(NHC)AuCl] (1, 10–12) and cells, following incubation with compounds 13, 1 and 9. We [Ru(p-cymene)Cl (Z1-dppm)]9.29Importantly,allheterometallic foundthioredoxinreductaseactivitytobelowerincellstreated 2 RuAucomplexesareconsiderablylesstoxictothenon-tumorigenic with5mMof1,9and13,withanobservedinhibitionof17%, humanembryonickidneycellline(HEK-293T)thancisplatinand 44% and 63%, respectively, after a 24 hour incubation (Fig. 2 [Ru(p-cymene)Cl (Z1-dppm)] 9. Monometallic [(NHC)AuCl] has andseefiguresforallcompoundsintheESI†).Thisexperiment 2 Thisjournalis©TheRoyalSocietyofChemistry2016 Chem.Commun. .25:35:21 6102/10/52 no ytisrevinU notgnihsaW egroeG yb dedaolnwoD .6102 yraunaJ 52 no dehsilbuP View Article Online Communication ChemComm 2 Y.-M.Wang,A. D.LacknerandF. D.Toste, Acc.Chem.Res., 2014, 47,889. 3 D.Gatineau,J.P.Goddard,V.Mouries-MansuyandL.Fensterbank, Isr.J.Chem.,2013,53,892. 4 C.C.J.LohandD.Enders,Chem.–Eur.J.,2012,18,10212. 5 M.Nicolas,RSCCatal.Ser.,2011,6,317. 6 M. Alcazaro, T. Stork, A. Anoop, W. Thiel and A. Furstner, Angew. Chem.,Int.Ed.,2010,49,2542. 7 M.NicolasandS.P.Nolan,Chem.Soc.Rev.,2008,37,1776. 8 M.Tacke,J.Organomet.Chem.,2015,782,17. 9 F.LazregandC.S.J.Cazin,inN-HeterocyclicCarbenes,ed.S.P.Nolan, Wiley-VHC,2014,ch.7,p.73. 10 B.BertrandandA.Casini,DaltonTrans.,2014,43,4209. 11 L.OehningerandI.Ott,DaltonTrans.,2013,42,3269. 12 W.LiuandR.Gust,Chem.Soc.Rev.,2013,42,755. 13 K. M. Hindi, M. J. Panzer, C. A. Tessier, C. 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The biological Escajeda, A. Varela-Ramirez, R. J. Aguilera and M. Contel, Inorg. Chem.,2011,50,11099. activityofthesenewderivativesiscurrentlybeingfurtherexplored. 24 J.Fern´andez-Gallardo,B.T.Elie,J.Florian,J.Sulzmaier,M.Sanau´, We envision that the versatility of this method will enable the J.W.RamosandM.Contel,Organometallics,2014,33,6669. synthesisofabroadrangeofcompounds(e.g.multimetallicsystems, 25 J.Fern´andez-Gallardo,B.T.Elie,T.Sadhukha,S.Prabha,M.Sanau´, S. A. Rotenberg, J. W. Ramos and M. Contel, Chem. Sci., 2015, 6, polymers, nanocarriers) for which the incorporation of [Au(NHC)] 5269. speciesmayimprovetheirbiologicalorcatalyticapplications. 26 L.Massai,J.Fern´andez-Gallardo,A.Guerri,A.Arcangelic,S.Pillozzic, We thank the National Cancer Institute (NCI) for grant M.ContelandL.Messori,DaltonTrans.,2015,44,11067. 27 M.V.Baker,P.J.Barnard,S.J.Berners-Price,S.K.Brayshaw,J.L. 1SC1CA182844 (M. C.) M. 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