Copper Coordination Compounds As Biologically Active Agents

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International Journal of Molecular Sciences

Review Copper Coordination Compounds as Biologically Active Agents

Olga Krasnovskaya 1,2,*, Alexey Naumov 1, Dmitry Guk 1 , Peter Gorelkin 2 , Alexander Erofeev 1,2, Elena Beloglazkina 1 and Alexander Majouga 1,2,3

1 Chemistry Department, Lomonosov Moscow State University, Leninskie gory 1,3, 119991 Moscow, Russia; [email protected] (A.N.); [email protected] (D.G.); [email protected] (A.E.); [email protected] (E.B.); [email protected] (A.M.) 2 Department of Materials Science of Semiconductors and Dielectrics, National University of Science and Technology (MISIS), Leninskiy prospect 4, 101000 Moscow, Russia; [email protected] 3 Mendeleev University of Chemical Technology of Russia, Miusskaya Ploshchad’ 9, 125047 Moscow, Russia * Correspondence: [email protected]

Received: 4 May 2020; Accepted: 30 May 2020; Published: 31 May 2020

Abstract: Copper-containing coordination compounds attract wide attention due to the redox activity and biogenicity of copper ions, providing multiple pathways of biological activity. The pharmacological properties of metal complexes can be fine-tuned by varying the nature of the ligand and donor atoms. Copper-containing coordination compounds are effective antitumor agents, constituting a less expensive and safer alternative to classical platinum-containing chemotherapy, and are also effective as antimicrobial, antituberculosis, antimalarial, antifugal, and anti-inflammatory drugs. 64Cu-labeled coordination compounds are promising PET imaging agents for diagnosing malignant pathologies, including head and neck cancer, as well as the hallmark of Alzheimer’s disease amyloid-β (Aβ). In this review article, we summarize different strategies for possible use of coordination compounds in the treatment and diagnosis of various diseases, and also various studies of the mechanisms of antitumor and antimicrobial action.

Keywords: copper coordination compounds; antitumor drug; antibacterial agents; PET imagining agents; mycobacterium tuberculosis; Alzheimer’s disease

1. Introduction Metal-containing therapeutic agents comprise a fundamental class of drugs for treating tumors. Although many metal-containing drugs based on gold, ruthenium, gallium, titanium, and iron are in preclinical and clinical trials phases I and II [1], cisplatin and also second- and third-generation platinum coordination compounds (carboplatin, oxalyplatin, and picoplatin) are still the most eﬀective antitumor agents used in clinical practice [2]. The clinical use of platinum-based drugs entails many severe side eﬀects, such as nephrotoxicity [3], neurotoxicity [4], and also ototoxicity and myelosuppression [5]. It is assumed that antitumor drugs based on endogenous metals (Co, Cu, Zn, and Fe) are less toxic as compared with platinum analogues [6]. Copper-containing coordination compounds were found to be promising antitumor therapeutic agents that act by various mechanisms such as inhibition of proteasome activity [7,8], telomerase activity [9], reactive oxygen species (ROS) formation [10,11], DNA degradation [12], DNA intercalation [13], paraptosis [14], and others. Copper is an element of fundamental importance for the formation and functioning of several enzymes and proteins, such as cytochrome C oxidase and Cu/Zn superoxide dismutase, which are involved in the processes of respiration, energy metabolism, and DNA synthesis [15]. Most Cu(II) coordination compounds quickly form adducts with glutathione in the cell medium, which leads to

Int. J. Mol. Sci. 2020, 21, 3965; doi:10.3390/ijms21113965 www.mdpi.com/journal/ijms Int. J. Mol. Sci. 2020, 21, 3965 2 of 37 the formation of a coordination compound of monovalent Cu(I) capable of generating a superoxide anion, which can induce ROS formation in a fenton-like reaction [16]. Due to high redox activity, the therapeutic efficacy of copper coordination compounds is not limited to antiproliferative action. Copper coordination compounds can be highly effective in treating viral infections [17], inflammatory diseases [18], and microbial infections [19] by multiple mechanisms of action. A Cu(II) coordination compound based on indomethacin is currently used in veterinary practice as an anti-inflammatory drug [20]. Malignant and inflamed tissues metabolize an increased amount of copper as compared with healthy tissues [21], which gives copper-containing coordination compounds an additional advantage over other metal-containing drugs. Various delivery systems for copper-based therapeutic agents and also for copper and chelating ligand separate delivery have been developed to enhance their delivery into tumor tissues [22,23]. Development of novel copper coordination compounds with antitumor activity is a promising and relevant area of medical chemistry [24–27]. A number of copper/Disulfiram-based drug combinations are in recruiting clinical trials as inexpensive and highly effective antitumor agents for metastatic breast cancer therapy and as diagnostic tools [28]. Their clinical success has triggered the development of delivery and controlled release systems for copper coordination compounds, as well as a search for novel copper-containing anticancer agents [29,30]. A brief and clear summary of promising in vivo anticancer activity of this type of drugs, along with relevant and current clinical trials was reported by Tabti1 et al. [31], but rapid development of copper-based therapy has caused rapid changes in clinical data. In a search for methods to overcome the poor water solubility of copper complexes, Wehbe et al. briefly summarized the use of copper complexes as antineoplastic agents [32]. However, after the publication of a detailed high-quality review by Santini et al. [33], a small number of works examined, in detail, the latest biological aspects of the use of copper complexes as therapeutic agents. Recently, Ong et al. reported a metal application in tropical diseases treatment, expanding the understanding of the applications of copper-containing coordination compounds [34]. In this review we provide a summary of different publications of recent years, paying attention to the variety of biological studies on the therapeutic and diagnostic potential of copper coordination compounds. We have emphasized the most abundant methods used to assess the mechanism of antitumor action and other therapeutic effects. This review could be useful to researchers engaged in medicinal application of copper-containing agents, affecting various uses of copper coordination compounds such as anticancer, antituberculosis, antimicrobial, anti-inflammatory, antibacterial agents, as well as PET-imaging agents for the diagnosis of malignant neoplasms and Alzheimer’s disease.

2. Copper Coordination Compounds Based on Ligands with Various Donor Atoms

2.1. N- and O-Donor Ligands Casiopeínas comprise a family of copper coordination compounds with promising results for treating colorectal cancer and acute myeloid leukemia. Several Casiopeínas compounds have shown signiﬁcant therapeutic eﬃcacy, and two of them, Casiopeina III-ia 1 and Casiopeina II Gly 2 (Figure1), have underwent a number of clinical trials as drugs for the treatment of leukemia [35]. Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 3 of 38 Int. J. Mol. Sci. 2020, 21, 3965 3 of 37

Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 3 of 38

Figure 1. Chemical structures of Casiopeina II-gly 1 and Casiopeina-III-ia 2

Figure 1. Chemical structures of Casiopeina II-gly 1 and Casiopeina-III-ia 2. Several hypothesesFigure 1. Chemicalhave been structures developed of Casiopeina regarding II- glythe 1 mechanismand Casiopeina of- IIIaction-ia 2 of Casiopeinas, includingSeveral ROS hypotheses formation, have phosphate been developed hydrolysis, regarding DNA d theamage, mechanism and DNA of intercalation action of Casiopeinas, [36]. In includingaddition,Several ROS one hypotheses formation, of the latest phosphatehave studies been hydrolysis,developed [37] of coordination DNAregarding damage, compoundsthe mechanism and DNA of intercalation theof action Casiopeínas of [36Casiopeinas,]. class In addition, has oneincludingshown of the an latest ROSantiproliferative formation, studies [37 phosphateeffect] of coordination in Giardia hydrolysis, intestinalis compounds DNA trophozoite damage, of the andcultures, Casiope DNA íanas intercalationpathogen class has causing [3shown6]. Inan an antiproliferativeaddition,infectious one disease of e ff theect that latest in affectsGiardia studies residents intestinalis [37] of of coordination trophozoite developing compoundscultures, countries. a The pathogen of the antiproliferative Casiopeínas causing an class effect infectious has of diseaseshowncoordination thatan antiproliferative aff ects compounds residents iseffect of explained developing in Giardia by intestinalis their countries. ability trophozoite The to interact antiproliferative cultures, with the a pathogen celleffect membrane of causing coordination and an compoundsinfectiousincrease the disease is ROS explained concentration that affects by their residentsin abilitythe parasite to of interact developing from the with first co theuntries. hours cell of membrane The exposure antiproliferative (2 and–6 h increase). It was effect thefound of ROS coordination compounds is explained by their ability to interact with the cell membrane and concentrationthat these compounds in the parasite caused from the the death first hoursof trophozoite of exposure cells (2–6 as h).a result It was of found apoptosis. that these Guillermo compounds de increaseAnda-Jáuregui the ROS et concentration al. recently constructed in the parasite a network from the with first deregulated hours of exposure biological (2 pathways–6 h). It was featuring found caused the death of trophozoite cells as a result of apoptosis. Guillermo de Anda-Jáuregui et al. recently thatlinks these between compounds pathways caused that crosstalkthe death with of trophozoite each other. cellsThrough as a thisresult approach, of apoptosis. the following Guillermo three de constructed a network with deregulated biological pathways featuring links between pathways that Andafeatures-Jáuregui of Casiopeina et al. recently treatment constructed were identified: a network (a) with perturbation deregulated of biologicalsignaling pathways featuringrelated to crosstalk with each other. Through this approach, the following three features of Casiopeina treatment linksapoptosis between induction pathways, (b) perturbationthat crosstalk of with metabolic each other. pathways Through, and this(c) activation approach, of th immunee following responses three were identified: (a) perturbation of signaling pathways related to apoptosis induction, (b) perturbation features[38]. of Casiopeina treatment were identified: (a) perturbation of signaling pathways related to ofapoptosis metabolicCopper induction pathways, coordination, (b) and perturbation (c) compounds activation of metabolic of3– immune5 with pathways Schiffresponses , and bases [(c)38 ]. asactivation ligands of wereimmune obtained responses by [3condensation8].Copper coordination of 5-dimethylcyclohexane compounds 3–5-1,3with-dione Schi andff bases a hydrazine as ligands derivative were obtained by Shoair by condensation et al. [39] of(Figure 5-dimethylcyclohexane-1,3-dioneCopper 2). coordination compounds and a3 hydrazine–5 with Schiff derivative bases by as Shoair ligands et al. were [39] (Figure obtained2). by condensation of 5-dimethylcyclohexane-1,3-dione and a hydrazine derivative by Shoair et al. [39] (Figure 2).

Figure 2. Ligand synthesis scheme and chemical structures of coordination compounds 3–5. Figure 2. Ligand synthesis scheme and chemical structures of coordination compounds 3–5. Coordination compounds 3–5 showed the ability to intercalate calf thymus DNA and also showed cytotoxicCoordination activityFigure 2. on Ligand the compounds cell synthesis lines of3scheme–5 liver showed and cancer chemical the HepG-2 ability structures (human to intercalate of coordination liver cancer calf compounds thymus cell line DNA of 3 hepatocellular–5 . and also carcinoma)showed cytotoxic and breast activity cancer on the MCF-7 cell lines (breast of cancerliver cancer cell lineHepG of-2 invasive (human breastliver cancer ductal cell carcinoma) line of (Tablehepatocellular1Coordination). The toxicity carcinoma) compounds of the ligandsand breast3–5 and showed cancer their correspondingtheMCF ability-7 (breast to intercalatecancer coordination cell calfline compounds of thymus invasive DNA breast is comparable. and ductal also Complexshowedcarcinoma 4cytotoxicshowed) (Table activity the1). The greatest toxicityon the cytotoxic cellof the lines ligands activity of liver and on cancer their MCF-7 corresponding HepG cell lines.-2 (human coordination liver cancer compounds cell line of is hepatocellularcomparable.The antimicrobial Complex carcinoma) activity 4 showed and of breast ligandsthe greatest cancerL3– L5 MCFcytotoxicand-7 Cu(II)(breast activity complexescancer on MCF cell 3line-7– 5cell wereof lines.invasive tested breast against ductal bacteria andcarcinoma fungi. All) (Table ligands 1). andThe toxicity complexes of the were ligands found and to their have corresponding antibacterial activity coordination against compounds Gram-negative is Escherichiacomparable. coli Complex(except 3 4), showed Gram-positive the greatestStaphylococcus cytotoxic activity aureus ,on and MCFCandida-7 cell albicanslines. (Table2).

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Table 1. MTT data of coordination compounds 3–5 and ligands L3–L5 after 72 h of incubation [39]. Table 1. MTT data of coordination compounds 3–5 and ligands L3–L5 after 72 h of incubation [39]. IC50, µM ± S.D. Compound HepG-2 MCF-7IC 50, µM S.D.Compound HepG-2 MCF-7 ± L3 Compound6.88 ± HepG-20.5 27.19 MCF-7± 2.3 Compound3 HepG-241.77 ± 2.7 MCF-7 26.57 ± 1.9 L4 7.60 ± 0.9 14.65 ± 1.5 4 11.80 ± 1.3 9.38 ± 1.0 L3 6.88 0.5 27.19 2.3 3 41.77 2.7 26.57 1.9 L5 58.10 ± 3.4 ± 63.13 ± 3.6± 5 ± 67.66 ± 3.8 ± 46.75 ± 3.1 L4 7.60 0.9 14.65 1.5 4 11.80 1.3 9.38 1.0 The antimicrobial activity± of ligands ±L3–L5 and Cu(II) complexes± 3–5 were tested± against L5 58.10 3.4 63.13 3.6 5 67.66 3.8 46.75 3.1 bacteria and fungi. All ligands± and complexes± were found to have antibacterial± activity± against Gram-negative Escherichia coli (except 3), Gram-positive Staphylococcus aureus, and Candida albicans Table 2. Antibacterial and antifungal activities data of ligands L3–L5 and Cu(II) coordination (Table 2). compounds 3–5 [39].

Table 2. AntibacterialE. Coli and antifungal activities S. Aureus data of ligands L3–L5 and C. Cu(II) Albicans coordination compounds 3–5 [39]. Diameter of % Diameter of % Diameter of % Compound E.Inhibition Coli Activity SInhibition. Aureus Activity InhibitionC. Albicans Activity DiameterZone (Mm) of Index% DiameterZone (Mm) of Index% ZoneDiameter (Mm) of Index% Compound Inhibition Zone Activity Inhibition Zone Activity Inhibition Zone Activity L3 13 52.0 18 78.3 21 80.8 L4 (Mm)8 Index 32.0 (Mm) 11 Index 47.8 (Mm) 16 Index 61.5 L3L5 136 52.0 24.0 18 5 78.3 21.7 21 8 30.880.8 L43 8 332.0 12.0 11 8 47.8 34.8 1416 53.861.5 L54 6 924.0 36.0 5 16 21.7 69.6 198 73.130.8 35 3NA 12.0 —- 8 234.8 8.7 1014 38.553.8 Ampicillin4 925 36.0 100 16 23 69.6 100 NA19 73.1 —- Cloitrimazole5 NANA ---- —- NA2 8.7 —- 2610 10038.5 Ampicillin 25 100 23 100 NA ---- Cloitrimazole NA ---- NA ---- 26 100 Copper-containingCopper-containing antitumor antitumor agents, agents, with SchiffSchiff--basebase ligands ligands based based on on hydrazone hydrazone with with a a pyridinepyridine coligand, coligand were, were investigated investigatedby by QingYouQingYou MoMo et al. [40 [40].]. Introducing Introducing N N-containing-containing coligands coligands suchsuch as as imidazole, imidazole, pyridine, pyridine, quinoline, quinoline, phenanthroline, and and their their derivatives derivatives can can affect affect the the hydrophobicity,hydrophobicity, the the geometry geometry of the of coordination the coordination compound, compound, and consequently, and consequently the antitumor, the antitumor activity. Copperactivity. coordination Copper coordination compounds compounds6–8 with Schi 6ff–-base8 with ligands Schiff- andbase also ligands coordination and also compounds coordination9– 11 containingcompounds pyridine 9–11 ascontai a ligandning were pyridine obtained as (Figure a ligand3). Studies were obtained of antiproliferative (Figure 3). activity Studies showed of thatantiproliferative introducing a activity pyridine showed coligand that into introducing the structure a pyridine of the coligand coordination into the compound structure increases of the cytotoxiccoordination activity compound as expected. increases Coordination cytotoxic compounds activity as 9 expected.–11 containing Coordination a pyridine compounds coligand exhibit9–11 antiproliferativecontaining a pyridine activity coligandin vitro with exhibit an ICantiproliferative50 ranging from activity 1.12 to in 6.31 vitroµM with (MCF-7 an IC breast50 ranging cancer from cells), while1.12 non-coligandto 6.31 μM (MCF analogues-7 breast6 –cancer8 have cells), an IC while50 in non the- rangecoligand from analogues 3.66 to 18.616–8 haveµM an (MCF-7 IC50 in breastthe cancerrange cell from line 3.66 of invasiveto 18.61 µM breast (MCF ductal-7 breast carcinoma). cancer cell line of invasive breast ductal carcinoma).

Figure 3. Chemical structures of coordination compounds 6–8 with Schiﬀ-base ligands and 9–11 with a

Py coligand. Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 5 of 38 Int. J. Mol. Sci. 2020, 21, 3965 5 of 37 Figure 3. Chemical structures of coordination compounds 6–8 with Schiff-base ligands and 9–11 with a Py coligand. For coordination compounds 9–11, cytotoxicity studies were also conducted on cisplatin-resistant For coordination compounds 9–11, cytotoxicity studies were also conducted on lung cancer cell lines (A549cisR cisplatin-resistant lung cancer cell line of adenocarcinomic human cisplatin-resistant lung cancer cell lines (A549cisR cisplatin-resistant lung cancer cell line of alveolar basal epithelial cells). High toxicity was shown with an IC in the range of 3.77 to 6.03 µM adenocarcinomic human alveolar basal epithelial cells). High toxicity50 was shown with an IC50 in the µ (IC50 range> 50 ofM 3.77 for cisplatin).to 6.03 μM ( StudiesIC50 > 50 of μM the for mechanism cisplatin). Studies of the cytotoxicof the mechanism effect of of coordination the cytotoxic compound effect 11 showedof coordination that the drugcompound causes 11 DNA showed degradation, that the drug which causes triggers DNA degradation, the mechanism which of triggers ROS-mediated the apoptosismechanism of mitochondrial of ROS-mediated dysfunction. apoptosis of mitochondrial dysfunction. D. AnuD. Anu et al. et reportedal. reported tetra-nuclear tetra-nuclear mixed-valence mixed-valence copper copper (I/II) (I/II) coordination coordination compound compound 12 with12 with promisingpromising antitumor antitumor activity activity [41 [41] (Figure] (Figure4 ).4). The The structure structure of the the obtained obtained coordination coordination compound compound was confirmedwas confirmed by X-ray by X-ray diff diffraction.raction.

FigureFigure 4. Synthesis 4. Synthesis scheme scheme and and chemical chemical structure of of coordination coordination compound compound 12. 12. A spectrophotometric study of DNA intercalation by ligand L12 and coordination compound A spectrophotometric study of DNA intercalation by ligand L12 and coordination compound 12 was12 performedwas performed by titrationby titration of of a a calf calf thymus thymus DNA DNA solution with with the the solutions solutions of coordin of coordinationation compoundcompound12. 12. The The binding binding constants of of L12L12 andand coordination coordination compound compound 12 were12, respectivelywere, respectively,, (2.34 (2.34 ± 00.60).60) × 10105 M5 −M1 and1 and(3.50 (3.50± 0.73) ×0.73) 105 M−1,10 indicating5 M 1, indicating a weak interaction a weak with interaction the secondary with the structure secondary ± × − ± × − structureof DNA. of DNA. BSA protein BSA protein binding bindingwas confirmed was confirmed by fluorescence by fluorescence titration. The titration. antioxidant The activity antioxidant of activitycoordination of coordination compound compound 12 was prove12 wasn by proven the ability by the to abilitydecrease to the decrease reduction the of reduction Mo(VI) to of Mo(V) Mo(VI) to Mo(V)and and by by,, subsequent subsequent,, formation formation of a of complex a complex at acidic at acidic pH.pH. The The ability ability of L12 of L12andand coordination coordination compoundcompound12 to 12 cause to ca apoptosisuse apoptosis in MCF-7 in MCF cells-7 was cells proven was prove by then by acridine the acridine orange orange/ethidium/ethidium bromide bromide (AO/EtBr) staining method. (AO/EtBr) staining method. Studies of antiproliferative activity on breast cancer cells MCF-7 and lung cancer cells A-549 Studies of antiproliferative activity on breast cancer cells MCF-7 and lung cancer cells A-549 showed an IC50 of 32 ± 1.0 μM (MCF-7 breast cancer cell line of invasive breast ductal carcinoma), 15 showed an IC of 32 1.0 µM (MCF-7 breast cancer cell line of invasive breast ductal carcinoma), ± 1.5 μM (A50-549 lung± cancer cell line of adenocarcinomic human alveolar basal epithelial) for ligand 15 1.5 µM (A-549 lung cancer cell line of adenocarcinomic human alveolar basal epithelial) for ligand ± L12 and 25 ± 1.0 μM (MCF-7) and 12 ± 1.0 μM (A-549) for coordination compound 12. L12 and 25 1.0 µM (MCF-7) and 12 1.0 µM (A-549) for coordination compound 12. 2.2. N- ±and S-Donor Ligands ± 2.2. N- and S-Donor Ligands Elesclomol is an injectable chemotherapeutic agent ligand L13 with low molecular weight Elesclomol(Figure 5), which is an injectable demonstrated chemotherapeutic clinical efficacy agent in acute ligand myeloidL13 withleukemia low molecular[42]. This drug weight is also (Figure at 5), which demonstrated clinical efficacy in acute myeloid leukemia [42]. This drug is also at the first stage of clinical trials as a therapeutic agent for leukemia [43]. Elesclomol has been proven to exert an antitumor effect by forming a Cu(II) coordination compound in situ, and the corresponding coordination compound causes oxidative stress inside the malignant cell. The redox reaction Cu(II)/Cu(I) disrupts mitochondrial respiration and causes ROS formation. Ultimately, coordination of copper with the Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 6 of 38

the first stage of clinical trials as a therapeutic agent for leukemia [43]. Elesclomol has been proven to

Int.exert J. Mol. an antitumor Sci. 2020, 21, 3965effect by forming a Cu(II) coordination compound in situ, and the corresponding6 of 37 coordination compound causes oxidative stress inside the malignant cell. The redox reaction Cu(II)/Cu(I) disrupts mitochondrial respiration and causes ROS formation. Ultimately, coordination elesclomolof copper with ligand the disrupts elesclomol the ligand production disrupts and the metabolism production of cellularand metabolism energy and of cellular triggers energy the path and of mitochondrialtriggers the path apoptosis of mitochondrial in tumor cells,apoptosis leading in tumor to their cells death, leading [44]. to their death [44].

Figure 5. Chemical structure of e elesclomollesclomol ligand L13 and copper coordination compound 13..

An antitumorantitumor activityactivity of of the the redox-active redox-active copper copper coordination coordination compound compound was was also also confirmed confirmed in [45 in], where[45], where the potential the potential effectiveness effectiveness of elesclomol of elesclomol in treating in ovarian treating cancer ovarian was cancer shown. was The shown. elesclomol The ligandelesclomol showed ligand antiproliferative showed antiproliferative activity on six activity cell lines on of six gynecological cell lines of cancer gynecological with an IC cancer50 of 0.173 withµ anM andIC50 of an 0.173 IC90 ofµM 0.283 and µanM IC90 (tests of were 0.283 performed µM (tests withwere Cu-preincubatedperformed with Cu cell-preincubated lines). cell lines). Due to redox properties, copper coordination compounds not only are eeffectiveffective redox-activeredox-active antitumor agents but also are eeffectiveffective in treating bacterial and fungal infections. Tuberculosis (TB) caused by Mycobacterium tubtuberculosiserculosis (Mtb) is an infection causing more deaths than acquired acquired immunodeficiencyimmunodeficiency syndrome. First First-line-line drugs, such as rifampicin, successfully coped with bacterial pneumonia, but drug resistanceresistance requiresrequires seekingseeking newnew chemotherapeuticchemotherapeutic agents. A new triple-drugtriple-drug combination forfor treating treating TB TB is is a combinationa combination of oxidantof oxidant [46] [46] and redox-activeand redox-active drugs drugs [47] coupled [47] coupled with a thirdwith drug a third with drug a di ff witherent a mode different of action. mode Therefore, of action. the Therefore, redox activity the of redox copper activity ions coupled of copper with ions the factcoupled that thewith immune the fact system that the uses immune copper system to eliminate uses copper bacterial to infections eliminate makes bacterial copper infections coordination makes compoundscopper coordination promising compoundsantibacterial, promisingand in particular, antibacterial antituberculosis, and in chemotherapeutic particular, antituberculosis agents. chemotherapeuticRecent studies agents. by Ngwane et al. [48] demonstrated that elesclomol is relatively potent against Mtb H37RvRecent with studies a minimum by Ngwane inhibitory et al. concentration [48] demonstrated of 10 µ Mthat (4 elesclomol mg/L). In addition, is relatively against potent multidrug against resistantMtb H37Rv clinical with isolates a minimum of Mtb, it inhibitory displays additive concentration interactions of 10 with μM known (4 mg/L) tuberculosis. In addition, drugs against such as isoniazidmultidrug and resistant ethambutol, clinical and isolates a synergistic of Mtb, it interaction displays additive with rifampicin. interactions with known tuberculosis drugsControlled such as isoniazid supplementation and ethambutol, of elesclomol and a withsynergistic copper interaction leading to thewith formation rifampicin. of compound 13 in cultureControlled medium supplementation increased Mtb sensitivityof elesclomol by >with65-fold. copper (Table leading3) to the formation of compound 13 in culture medium increased Mtb sensitivity by >65-fold. (Table 3) Table 3. Effect of copper on antimycobacterial activity of ligand L13 against Mtb H37Rv [48]. Table 3. Effect of copper on antimycobacterial activity of ligand L13 against Mtb H37Rv [48]. Medium Used MIC (mg/L) MiddlebrookMedium 7H9Used * MIC (mg/L) 4 MiddlebrooksMiddlebrook 7H12 7H9 ** 4 4 HdBMiddlebrooks without CuSO 7H124 * >4 32 HdBHdB (CuSO without4 at 2 mgCuSO/L)4 >32 0.5 * amountHdB of copper (CuSO in4 mediumat 2 mg/L) was approximately0.5 1 mg/L. * amount of copper in medium was approximately 1 mg/L. Cu-ATSM 14 is a biologically active copper coordination compound based on thiosemicarbazides (FigureCu6-).ATSM This drug14 labeledis a with biologically the radioactive active isotopes copper 64 coordinationCu, 62Cu, and 60 compoundCu was used based as a PETon 64 62 60 hypoxiathiosemicarbazides imaging agent (Figure in head 6). This and drug neck cancerlabeled [ 49with]. It the demonstrated radioactive betterisotopes results Cu, in Cu, clinical and trialsCu thanwas usedthe 18-fluorodeoxyglucose as a PET hypoxia imaging used in agent clinical in practice head and [50 neck,51]. cancer [49]. It demonstrated better resultsDrug in clinical accumulation trials than in the hypoxic 18-fluorodeoxyglucose areas is associated used with in clinical redox practi transitionsce [50,51]. of Cu(II)/Cu(I). Labeled with a radionuclide tag, Cu-ATSM 14 penetrated into cells by passive diffusion and underwent glutathione reduction. Under normoxic conditions, the labile coordination compound Cu(I) is oxidized by intracellular oxygen to the coordination compound Cu(II) and leaves the cell. In contrast, under hypoxic conditions, the Cu(I) coordination compound dissociates into a ligand and a metal ion, which binds to intracellular chaperone proteins leading to the accumulation of a radionuclide in Int. J. Mol. Sci. 2020, 21, 3965 7 of 37 hypoxic regions of tumors. The oxidation process of Cu(I) is so fast that a noticeable intracellular reduction of Cu(II) ATSM occurs only in hypoxic (tumor) cells, while the drug leaves healthy cells without any changes [52] (Figure6). Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 7 of 38

FigureFigure 6. Chemical 6. Chemical structure structure and and cellular cellular accumulation scheme scheme of Cu of-ATSM Cu-ATSM 14. 14.

CoordinationDrug accumulation Cu-ATSM 14 inalso hypoxic proved areas to beis associated an eﬀective with drug redox capable transitions of slowing of Cu(II)/Cu(I). the progression of amyotrophicLabeled lateralwith a sclerosis radionuclide (ALS) tag, disease Cu-ATSM and 14 improving penetrated the into respiratory cells by passive and cognitive diffusion function and of patients.underwent Currently, glutathione the drug reduction. Cu(II)-ATSM Under is normoxic undergoing conditions, clinical the trials labile as coordination a drug for compound the treatment of Cu(I) is oxidized by intracellular oxygen to the coordination compound Cu(II) and leaves the cell. In ALS. Patient registration for phase III clinical trials of the treatment of this disease began in November contrast, under hypoxic conditions, the Cu(I) coordination compound dissociates into a ligand and a 2019 in Australiametal ion, [ which53]. binds to intracellular chaperone proteins leading to the accumulation of a Anjumradionuclide et al. in recently hypoxic reportedregions of tumors. eightthiosemicarbazido-based The oxidation process of Cu(I) Cu(II)is so fast complexes, that a noticeable Cu-ATSM analoguesInt. J. Mol.intr Sci.15acellular 20–2022, 2,1 with, reductionx FOR promising PEER of REVIEW Cu(II) antitumor ATSM occurs activity only [in54 hypoxic] (Figure (tumor)7). cells, while the drug leaves 8 of 38 healthy cells without any changes [52] (Figure 6). Coordination Cu-ATSM 14 also proved to be an effective drug capable of slowing the progression of amyotrophic lateral sclerosis (ALS) disease and improving the respiratory and cognitive function of patients. Currently, the drug Cu(II)-ATSM is undergoing clinical trials as a drug for the treatment of ALS. Patient registration for phase III clinical trials of the treatment of this disease began in November 2019 in Australia [53]. Anjum et al. recently reported eight thiosemicarbazido-based Cu(II) complexes, Cu-ATSM analogues 15–22, with promising antitumor activity [54] (Figure 7).

Figure 7. Chemical structures of bis(thiosemicarbazone)-based copper complexes 15–22. Figure 7. Chemical structures of bis(thiosemicarbazone)-based copper complexes 15–22. The toxicity of Cu(II) complexes 15–22 could be decreased by co-incubation with the nontoxic Cu The toxicity of Cu(II) complexes 15–22 could be decreased by co-incubation with the nontoxic chelator tetramolibdate (TM) or the antioxidant N-acetylcysteine (NAC), suggesting a mechanism of Cu chelator tetramolibdate (TM) or the antioxidant N-acetylcysteine (NAC), suggesting a Cu-induced oxidative stress. The redox behavior of Cu(II) complexes was also of interest. Depending on mechanism of Cu-induced oxidative stress. The redox behavior of Cu(II) complexes was also of interest. Depending on electron-donating effects of the di-substitutions on the diimine backbone, the Cu(II/I) redox potential itself was changed, and the cytotoxicity changed as a result. The Cu(II/I) redox potential was also proposed to govern the hypoxia selectivity of coordination compounds, but no selective toxicity under hypoxic conditions was shown. The ability of copper coordination compounds to successfully penetrate the blood-brain barrier has inspired some researchers to create copper-based preparations for visualizing pathological changes in Alzheimer’s disease. One of the major pathological hallmarks of the disease is the presence of extracellular senile plaques in the brain, consisting of an insoluble aggregated peptide called amyloid-β (Aβ), a 39−43 amino acid peptide [55]. Clinically used derivatives of benzothiazole, stilbene, and stripylpyridine labeled with 18-fluorine or 11-carbon are used for PET imaging of plaques by binding to a hydrophobic pocket of the peptide [56,57]. In addition, Zn2+ and Cu2+ cations have been proven to promote aggregation of amyloid plaques, which provides them with an advantage in binding to amyloid due to the increased consumption of these metals by amyloids [58,59]. A standard approach in developing Aβ PET imaging drugs is to modify a Cu-ATCM drug with a benzothiazole/stilbene moiety, which ensures the binding of the drug to the amyloid plaque. This developmental approach has been used by some Australian researchers (Figure 8).

Int. J. Mol. Sci. 2020, 21, 3965 8 of 37 electron-donating eﬀects of the di-substitutions on the diimine backbone, the Cu(II/I) redox potential itself was changed, and the cytotoxicity changed as a result. The Cu(II/I) redox potential was also proposed to govern the hypoxia selectivity of coordination compounds, but no selective toxicity under hypoxic conditions was shown. The ability of copper coordination compounds to successfully penetrate the blood-brain barrier has inspired some researchers to create copper-based preparations for visualizing pathological changes in Alzheimer’s disease. One of the major pathological hallmarks of the disease is the presence of extracellular senile plaques in the brain, consisting of an insoluble aggregated peptide called amyloid-β (Aβ), a 39 43 amino acid peptide [55]. Clinically used derivatives of benzothiazole, stilbene, − and stripylpyridine labeled with 18-ﬂuorine or 11-carbon are used for PET imaging of plaques by binding to a hydrophobic pocket of the peptide [56,57]. In addition, Zn2+ and Cu2+ cations have been proven to promote aggregation of amyloid plaques, which provides them with an advantage in binding to amyloid due to the increased consumption of these metals by amyloids [58,59]. A standard approach in developing Aβ PET imaging drugs is to modify a Cu-ATCM drug with a benzothiazole/stilbene moiety, which ensures the binding of the drug to the amyloid plaque. This developmental approach has been used by some Australian researchers (Figure8). Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 9 of 38

(a)

(b) (c)

Figure8. (a) Developed by Hickey et al. [60]. (b) Developed by SinChun Lim et al. [61]. (c) Developed

by McInnesFigure et 8. al. (a [)62 Developed]. Chemical by structures Hickey et of al. Cu-containing [60]. (b) Developed coordination by SinChun compounds Lim et al. for [61]. PET (c) imaging of Aβ plaques.Developed by McInnes et al. [62]. Chemical structures of Cu-containing coordination compounds for PET imaging of Aβ plaques.

Int. J. Mol. Sci. 2020, 21, 3965 9 of 37

Hickey et al. [60] succeeded in designing a copper radiopharmaceutical Cu(II)-ATSM with an appended stilbene functional group for Aβ plaque imaging. Binding of compounds 24 and 25 (coordination compound 23 was quite insoluble) to Aβ plaques was clearly evident as demonstrated by epi-fluorescent microscopy. Aβ-specific 1E8 antibody was used as a control. The biodistribution of coordination compounds 24 and 25 radiolabeled with 64Cu in wild-type mice after intravenous tail injection ( 13MBq) displayed good brain uptake of coordination compound 25 ∼ (1.11% ID/g) at 2 min after injection, dropping to 0.38% ID/g at 30 min. This indicates that coordination compound 25 can rapidly cross the blood-brain barrier of normal mice with a highly desirable fast washout from the brain as anticipated with no Aβ plaques to trap the imaging agent. Micro-PET images ofInt.pre-injected J. Mol. Sci. 2020 wild-type, 21, x FOR PEER mice REVIEW were also obtained. 10 of 38 The biodistribution of coordination compounds 27–30 radiolabeled with 64Cu in wild-type mice showedinjection the, dropping best brain to uptake 0.77% results ID/g at for 30 coordination min). TEM compound images of30 Aβ1−42(1.54% IDmodel/g at 2fibrils min after treate injection,d with droppingcompound to 28 0.77% or 30 ID demonstrated/g at 30 min). dramatic TEM images change of Asβ in1 the42 modelstructural fibrils morphology treated with. compound 28 or − 30 demonstratedAn alternative dramatic methodology changes is in based the structural on elemental morphology. mapping using laser ablation inductively coupledAn alternativeplasma mass methodology spectrometry is LA based-ICP on-MS elemental. A sample mapping of nonradioactive using laser isotopical ablation inductivelyly enriched 65 coupledCu-30 was plasma used. mass Coordination spectrometry compound LA-ICP-MS. 24 was A sample used as of a nonradioactive control. The benzofuran isotopically containing enriched 65 65complexCu-30 was Cu used.-30 appear Coordinationed to bind compound with improved24 was used differentiation as a control. whenThe benzofuran comparedcontaining with the 65 complexstyryl-pyridine65Cu- 30containingappeared complex to bind Cu with-24 and improved potentially diff offererentiationed better when sensitivity compared for amyloid. withthe styryl-pyridineOn the basis containing of these complex results, 65 radiolabeledCu-24 and potentially copper coordination offered better compound sensitivity could foramyloid. be used to assessOn amyloid the basis pathology of these results,in AD patients radiolabeled using copper PET. The coordination redox properties compound of copper could be ions, used the to ability assess amyloidto reduce pathology intracellularly, in AD patientsselective using accumulation PET. The redoxin hypoxic properties areas, of blood copper-brain ions, barr the abilityier penetration, to reduce intracellularly,and stability in selective a blood accumulationflow provides in copper hypoxic-containing areas, blood-brain therapeutic barrier agents penetration, features for and use stability as not inonly a blood therapeutic flow provides but also copper-containing diagnostic and theranostic therapeutic agents. agents features for use as not only therapeutic but also diagnostic and theranostic agents. 2.3. N/N-Donor Ligands 2.3. NBecause/N-Donor pathogens Ligands with multidrug resistance are emerging and new effective antibiotics againstBecause them pathogens are lacking, with metal multidrug-containing resistance coordination are emerging compounds and new have effective become antibiotics of interest against as themantibacterial are lacking, agents. metal-containing The effectiveness coordination of copper compounds coordination have becomecompounds of interest in the as treatment antibacterial of agents.bacterial The and eff ectiveness fungal infections of copper was coordination mentioned compounds above (coordination in the treatment compounds of bacterial 3 and and 5 fungal with infectionsantibacterial was and mentioned antifungal above activity (coordination [39] and compounds Escimolol-based3 and coordination5 with antibacterial compound and antifungal13 in Mtb activitytreatment [39 ] [48 and]). Escimolol-basedA copper-based coordination coordination compound compound13 in31Mtb with treatment antimalarial [48]). activity A copper-based against coordinationPlasmodium falciparum compound was31 with developed antimalarial by [63 activity] (Figure against 9). ThePlasmodium antimalarial falciparum activitieswas in developed vitro of bycompound [63] (Figure 31 9and). The its antimalarial ligand were activities respectivelyin vitro estimatedof compound as ED5031 and = 0.13its ligand and >30were mg respectively / ml for estimatedcoordination as ED50compound= 0.13 31 and and>30 liga mgnd/ L31ml for. coordination compound 31 and ligand L31.

Figure 9. Synthesis scheme and сhemical chemical structure of coordination compound 31..

Beeton et al. reported nine copper coordination compounds 32–39 based on 1,10-phenanthroline and also their platinum and palladium analogues compounds 40 and 41 with antimicrobial and antibiofilm activity [64] (Figure 10).

Int. J. Mol. Sci. 2020, 21, 3965 10 of 37

Figure 10. Chemical structure of coordination compounds 32–41. Figure 10. Chemical structure of coordination compounds 32–41. The resulting coordination compounds showed higher antimicrobial activity as compared with free The resulting coordination compounds showed higher antimicrobial activity as compared with ligands against Gram-positive and Gram-negative bacterial strains and also increased antibiotic activity free ligands against Gram-positive and Gram-negative bacterial strains and also increased antibiotic as compared with the standard preparation vancomycin against the clinical strain of methicillin-resistant activity as compared with the standard preparation vancomycin against the clinical strain of Staphylococcus aureus (MRSA) (Table4). methicillin-resistant Staphylococcus aureus (MRSA) (Table 4). Table4. Minimal inhibitory concentrations and hemolitic activity of coordination compounds 32–41 [64]. Table 4. Minimal inhibitory concentrations and hemolitic activity of coordination compounds 32–41 Compound[64]. S. Aureus S. Aureus E. Jaecalis E. Coli P. Aeruginosa % Lysis Rbcs MRSA252 MSSA209 NCTC775 NCTC86 ATCC27853 +/ (SD) Compound S. Aureus S. Aureus E. Jaecalis E. Coli P. Aeruginosa % Lysis −Rbcs 32 32MRSA252 MSSA209 32 NCTC775 32 NCTC86 64 ATCC27853>128 +/- SD 2.0) (0.4) 33 32 3232 3232 832 6464 >128>128 2.0 (0.4) 2.1 (0.1) 34 33 8832 1632 48 3264 > >128128 2.1 (0.1) 2.6 (0.3) 35 8 4 2 32 >128 2.2 (0.7) 34 88 16 4 32 > 128 2.6 (0.3) 36 4 4 4 16 >128 2.5 (0.3) 35 8 4 2 32 > 128 2.2 (0.7) 37 4 4 4 16 >128 2.0 (0.3) 38 36 24 24 24 1616 > >128128 2.5 (0.3) 3.1 (0.2) 39 37 24 24 24 1616 > >128128 2.0 (0.3) ND 40 38 1282 322 42 1616 > >128128 3.1 (0.2) ND 41 39 642 642 162 3216 > >128128 NDND Vancomycin40 0.25128 0.532 0.54 ND16 > 1 ND28 ND 2.6(0.2) Chloramphenicol41 IG64 1664 416 232 > 1 12828 ND ND CuClVancomycin2*2H2O >128 0.25 >1280.5 >1280.5 >ND128 ND>128 2.6(0.2) 2.0 (0.3) Chloramphenicol IG 16 4 2 128 ND TheCuCl authors2*2H associated2O >128 theaction mechanism>128 of>128 coordination >128 compounds> 12832 –41 with2.0 (0.3) interactions with the bacterial chromosome, which led to a decrease in bacterial reproduction. The redox activity of copperThe authorsions is dependent associated on the the action presence mechanism of reducing of agents. coordination This thiol compounds is glutathione 32–41 in mostwith Gram-negativeinteractions with bacteria the bacterial and bacillithiol chromosome, in several which Gram-positive led to a decrease bacteria in [65 bacterial]. Coordination reproduction. compounds The 40redoxand activity41 based of on copper Pt and Pd ions do is not dependent show significant on the antimicrobial presence of activity, reducing which agents. also indicates This thiol that is theglutathion antibacteriale in most activity Gram is- associatednegative bacteria with the and redox bacillithiol/nuclease in activity several of G copperram-positive ions. bacteria [65]. CoordinationCu(II) coordination compounds compounds 40 and 4132 –based41 are onless Pt active and than Pd dvancomycino not show on significant planktonic antimicrobial cells (Table4) andactivity, are relatively which also much indicates more activethat the on antibacterial biofilms. Copper-induced activity is associated DNA damage with the can redox/nuclease lead to death irrespectiveactivity of copper of the ions. physiological state or growth rate of the bacterial cells. Hence,Cu(II) coordination copper coordination compounds compounds 32–41can are not less only active be e ff thanective vancomycin synthetic antibacterial on planktonic agents cells but also(Table more 4) and effective are relatively as compared much with more classical active antibioticon biofilms. therapy Copper because-induced of theirDNA mechanism damage can of lead action. to death irrespective of the physiological state or growth rate of the bacterial cells. Hence, copper coordination compounds can not only be effective synthetic antibacterial agents but also more effective as compared with classical antibiotic therapy because of their mechanism of action. Brandão et al. reported Cu(II) coordination compounds based on thiochrome, the oxidized form of vitamin B1, with promising cytotoxic activity [66]. The ligand was obtained by oxidizing thiamine

Int. J. Mol. Sci. 2020, 21, 3965 11 of 37 Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 12 of 38 Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 12 of 38 with Brand copperão (II)et al. chloride. reported The Cu(II) resulting coordination coordination compounds compounds based on thiochrome, crystallize in the the oxidized form formof two of structuresvitaminwith copper B1,, compounds with (II) promising chloride. 42 and cytotoxic The 43 resulting (Figure activity coordination11 [66). ].Biological The ligand compounds studies was obtained on crystallizehuman by oxidizingcolon in theadenocarcinoma thiamine form of with two cellscopperstructures Caco (II)-,2 chloride.compounds showed Thethat 42 resultingboth and compounds 43 coordination(Figure 11reduce). Biological compounds the viability studies crystallize of onthese human in cells the more formcolon of thanadenocarcinoma two thiamine structures, or thiochrome.compoundscells Caco-2 To42showed andinvestigate43 that(Figure both the 11mechanismcompounds). Biological responsiblereduce studies the on viabilityfor human the cytotoxic of colon these adenocarcinoma effectcells more of compounds than cellsthiamine Caco-242 and or 43showedthiochrome., the authors that To both investigate tested compounds the the alleged reducemechanism involvement the viability responsible of of changes these for the cells in cytotoxic moreoxidat thanive effect stress thiamine of compounds levels. or thiochrome. By adding42 and ToN43-,acetylcysteine, investigate the authors the testedthey mechanism ruled the allegedout responsible ROS involvement formation, for the which cytotoxic of changes ultimately effect in of oxidatshowed compoundsive no stress change42 and levels. in43 the, the By cytotoxic authors adding effecttestedN-acetylcysteine, of the compound alleged involvementthey 42 ruled and onlyout of ROS changesa slight formation, in decrease oxidative which in stress theultimately cytotoxicity levels. showed By adding of compoundno change N-acetylcysteine, in43 the. Therefore, cytotoxic they oxidativeruledeffect out of compound ROSstress formation, does 42not and whichseem only t ultimatelyo aplay slight an decreaseimportant showed no in role change the in cytotoxicity the in themechanism cytotoxic of compound of eff ectbiological of 43 compound. Therefore,action 42of theseandoxidative only compounds, a stress slight decreasedoes which not indicates inseem the cytotoxicityto thatplay the an ability ofimportant compound to generate role43 .in Therefore, ROSthe mechanism is important oxidative of but stressbiological is not does always notaction seem the of maintothese play mechanismcompounds, an important of which the role cytotoxic inindicates the mechanism actionthat the of ability ofcopper biological to coordination generate action ROS of compounds. these is important compounds, A but comparison is which not always indicates of thethe cytotoxicthatmain the mechanism ability activity to generateofof thesethe cytotoxic compounds ROS is importantaction on ofcell copper but lines is not (coordinationIC50 always = 146 theμM compounds. mainfor compound mechanism A comparison 42 of and the IC cytotoxic50 of= 191the μMactioncytotoxic for of compound copper activity coordination of these43) with compounds compounds. cisplatin on (IC Acell50 comparison lines= 274 ( IC μM)50 of= the146 shows cytotoxicμM thatfor compound activitythese copper of these42 and coordination compounds IC50 = 191 μM for compound 43) with cisplatin (IC50 = 274 μM) shows that these copper coordination compoundson cell lines are (IC 50more= 146 effectiveµM for on compound human colon42 and adenocarcinoma IC50 = 191 µM cell for line compound of heterogeneous43) with cisplatin human compounds are more effective on human colon adenocarcinoma cell line of heterogeneous human (ICepithelial50 = 274 colorectalµM) shows adenocarcinoma that these copper Caco coordination-2. compounds are more effective on human colon adenocarcinomaepithelial colorectal cell adenocarcinoma line of heterogeneous Caco- human2. epithelial colorectal adenocarcinoma Caco-2.

Figure 11. ChemicalChemical structure structure of coordination compounds 42 and 43. Figure 11. Chemical structure of coordination compounds 42 and 43. Krasnovskaya etet al. [67 al.] reported[67] reported a Cu(II) coordination a Сu(II) compoundcoordination44 based compound on 2-aminoimidazolone 44 based on with2-aminoimidazoloneKrasnovskaya promising antitumor etwith al. activitypromising [67] (Figurereported antitumor 12 ). a Coordination С activityu(II) coordination(Figure compound 12). Coordination44 compoundshowed antiproliferative 44 compound based 44 on 2-aminoimidazolone with promising antitumor activity (Figure 12). Coordination compound 44 showedactivity higherantiproliferative than cisplatin activity (IC50 higherMCF-7 than 13.67 cisplatin0.81 µ(ICM).50 MCF-7 13.67 ± 0.81 μM). showed antiproliferative activity higher than cisplatin± (IC50 MCF-7 13.67 ± 0.81 μM).

Figure 12. ChemicalChemical structure structure of co coordinationordination compound 44. Figure 12. Chemical structure of coordination compound 44. Three copper copper complexes complexes with with potential potential anticancer anticancer and nonsteroidal and nonsteroidal anti-inflammatory anti-inﬂammatory activity wereactivity Three reported were copper reported by complexes Hussain by Hussain withet al.potentialet [68 al.] [68 (anticancerFigure] (Figure 13 ).and13 ). Coordination nonsteroidal Coordination anti compounds compounds-inflammatory 4545––4747 activity with benzimidazolebenzimidazole-derivedwere reported-derived by Hussain sca scaffoldsﬀolds et were were al. synthesized [ synthesized68] (Figure in accordance in13 accordance). Coordination with the with following the compounds following scheme. 45 scheme. In–47 addition with In additiontobenzimidazole antitumor to antitumor activity,-derived theactivity, scaffolds compounds the werecompounds were synthesized proposed were proposed as in potential accordance as candidatespotential with thecandidates for following NSAIDs. for scheme.NSAIDs .In addition to antitumor activity, the compounds were proposed as potential candidates for NSAIDs.

Int. J. Mol. Sci. 2020, 21, 3965 12 of 37 Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 13 of 38

Figure 13. Ligand synthesis scheme and chemical structure of coordination compoundscompounds 4545––4747..

HumanHuman serum serum albumin albumin (HAS) (HAS) binding binding of compounds of compounds45–47 was 45 evaluated–47 was using evaluated HAS fluorescence using HAS quenching in the presence of coordination compounds. The results showed that the K values (slope of fluorescence quenching in the presence of coordination compounds. The results showSV ed that the KSV 5 thevalues Stern–Volmer (slope of the plots) Stern were–Volmer of the plots) order were of 10 of, thusthe order indicating of 105, strong thus indicating quenching. strong quenching. AnAn Annexin Annexin/FITC/FITC assay assay showed showed that that the three the three complexes complexes45–47 exhibited45–47 exhibited an increase an in increase apoptotic in cellsapoptotic to a significant cells to a level significant followed level by followed necrosis. by Glutathione necrosis. Glutathionedepletion along depletion with ROS along formation with ROS in MCF-7formation cells in after MCF treatment-7 cells after with treatment coordination with compoundscoordination45 compounds–47 was also 45 shown.–47 was The also interaction shown. The of complexesinteraction withof complexes COX-2 inhibitor with COX was-2 alsoinhibitor confirmed, was also which confirmed, can be a which mechanism can be of a actionmechanism of these of potentialaction of these NSAIDs. potential Coordination NSAIDs. compoundsСoordination45 compounds–47 were tested 45–47in were vivo testedon albino in vivo rats on and albino mice rats for anti-inflammatory,and mice for anti-inflammatory, antipyretic, andantipyretic, analgesic and activities. analgesicThe activities. results The showed results that showed45 and that47 45have and significant47 have significant dose-dependent dose anti-inflammatory-dependent anti-inflammatory and analgesic activities and analgesic at a lower activities concentration. at a lower concentration.Sliwa et al. reported synthesis, characterization, and biological activity of three water-soluble copper(II)Sliwa et complexes al. reported [Cu(NOsynthesis,3)(PTA characterization,=O)(dmphen)][PF and biological6] 48, activity [Cu(Cl)(dmphen)2][PF of three water-6soluble] 49, and (Cu(NO ) (dmphen)) 50 [69] (Figure 14). copper(II) complexes3 2 [Cu(NO3)(PTA=O)(dmphen)][PF6] 48, [Cu(Cl)(dmphen)2][PF6] 49, and The cytotoxic activity of compounds 48–50 was evaluated on the normal human dermal fibroblast (Cu(NO3)2(dmphen)) 50 [69] (Figure 14). (NHDF), human lung carcinoma (A549), epithelioid cervix carcinoma (HeLa), colon cancer cell line of supraclavicular lymph node metastasis (LoVo), and breast cancer cell line of invasive breast ductal carcinoma (MCF-7) cell lines (Table5). All coordination compounds were more active than cisplatin but, expectedly, showed no significant selectivity to healthy cells. The interaction of compounds 48–50 with human apo-transferrin, causing a conformational change of the protein, was also proved using fluorescence and circular dichroism spectroscopy.

Table 5. MTT data of coordination compounds 48–50 after 72 h of incubation [69].

IC50, µM S.D. ± Cell Line/Compound 48 49 50 Cu(NO3)2 PTA = O Dmphen CDDP NHDF 0.57 0.08 0.23 0.03 1.72 0.25 310 47 Nd nd 16.6 2.1 ± ± ± ± ± A549 0.29 0.01 0.28 0.04 0.43 0.06 155 23 Nd nd 33.3 4.2 ± ± ± ± ± HeLa 1.12 0.16 1.13 0.17 0.43 0.06 19.1 2.9 Nd 720 108 16.6 3.1 ± ± ± ± ± ± MCF-7 0.57 0.08 0.57 0.08 3.45 0.51 155 23 Nd nd 33.3 4.2 ± ± ± ± ± LoVo 0.57 0.08 1.13 0.17 1.72 0.25 38.8 5.8 Nd 360 54 9.12 0.005 ± ± ± ± ± ±

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Figure 14. SynthesisSynthesis scheme and chemical structure of coordination compoundscompounds 4848––5050..

MilaniThe cytotoxic et al. reported activity seven of compounds novel Cu(I) complexes48–50 was51 – evaluated57 with potent on the antiproliferative normal human activity dermal for humanfibroblastInt. J. Mol. primary Sci. (NHDF), 2020, glioblastoma21, xhuman FOR PEER lung REVIEW cell carcinoma line U87cells (A549), [70] epithelioid (Figure 15 andcervix Table carcinoma6). (HeLa), colon cancer15 of 38 cell line of supraclavicular lymph node metastasis (LoVo), and breast cancer cell line of invasive breast ductal carcinoma (MCF-7) cell lines (Table 5). All coordination compounds were more active than cisplatin but, expectedly, showed no significant selectivity to healthy cells. The interaction of compounds 48–50 with human apo-transferrin, causing a conformational change of the protein, was also proved using fluorescence and circular dichroism spectroscopy.

Table 5. MTT data of coordination compounds 48–50 after 72 h of incubation [69].

IC50, µM ± S.D. Cell Line/Compound Figure48 15.15. Chemical49 structure of50 coordination Cu(NO compoundscompounds3)2 PTA =5151 O––57 57..Dmphen CDDP NHDF 0.57 ± 0.08 0.23 ± 0.03 1.72 ± 0.25 310 ± 47 Nd nd 16.6 ± 2.1 Table 6. MTT data of coordination compounds 51–57 on U87 cells after 72 h of incubation [70]. TableA549 6. MTT data0.2 of9 ± coordination 0.01 0.28 ± compounds 0.04 0.43 ± 51 0.06–57 on15 U875 ± cells23 afterNd 72 h of incubationnd [70]33.. 3 ± 4.2 HeLa 1.12 ± 0.16 1.13 ± 0.17 0.43 ± 0.06 19.1 ± 2.9 Nd 720 ± 108 16.6 ± 3.1 IC50, µM S.D. MCF -7 0.5 7 ± 0.08 0.57 ± 0.08 3.45 ± ±0.51 15IC5 50± ,2 µ3 M ± S.D.Nd nd 33.3 ± 4.2 CompoundCompoundLoVo 510.5 7 ± 0.08 52 521.1 3 ± 0.1753 53 1.72 ± 054.25 54 38.8 ± 5.8 5555 Nd 565636 0 ± 54 57579.1 2 ± 0.005 32.732.7 ± 00.6.6 31. 31.33 ± 11.7.7 2020 ± 1.51.5 46.46.77 ± 0.40.4 37.637.6 ± 1.11.1 34.534.50.9 ± 0.9 25.425.4 0.5± 0.5 ± ± ± ± ± ± ± Milani et al. reported seven novel Cu(I) complexes 51–57 with potent antiproliferative activity for humanFlow cytometryprimary glioblastoma was conducted cell line for U87cells accessing [70 the] (Figure apoptosis 15 and rate Table of 6 U87). cells treated with compounds 51–57 after 24 h. All the complexes, except compound 54, significantly induced apoptosis in U87 cells. The ability of compounds 51–57 to inhibit cancer cell growth by induction of cell cycle arrest was also estimated. Treatment of U87 cells with compounds 51–57 caused a marked arrest of G1, a growth phase that plays a key role in cell cycle progression and ensures that DNA is ready for synthesis. To validate the hypothesis that 51–57 triggered apoptosis in treated U87 cells by the effect on the expression level of apoptotic and anti-apoptotic genes, an RT-PCR assay was conducted. The results obtained showed an increased level of caspase-independent apoptosis genes including P53, P21, Bid, and Bax in U87 cells after treatment by all the complexes except compounds 54 and 55. The level of anti-apoptotic genes Bcl-2 and Bcl-xL was markedly inhibited in the presence of compounds 55 and 56. The investigation of the antitumor activity of Cu(I) coordination compounds 51–57, thus, demonstrated an inhibition of cell growth, cell cycle progression, migration ability, and expression level of anti-apoptotic genes and an induced apoptosis, necrosis, and expression level of apoptotic genes in a dose- and time-dependent manner in treated U87 cells. Kacar et al. reported coordination compound 58 based on a pyridyl-fluorobenzimidazole scaffold [71] (Figure 16).

Figure 16. Synthesis scheme, chemical structure of coordination compound 58.

Compound 58 showed antiproliferative and apoptotic effects on NIH/3T3 normal fibroblast cells and on SPC212 mesothelioma and DU145 prostate cancer cells. The most significant IC50 values were found against DU145, i.e., 37.0, 21.1, and 10.0 mM for the 24, 48, and 72 h treatments, respectively. A dose-dependent increase of pro-apoptotic Bax protein in DU145 preincubated with coordination compound 58 was observed.

Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 15 of 38

Figure 15. Chemical structure of coordination compounds 51–57.

Table 6. MTT data of coordination compounds 51–57 on U87 cells after 72 h of incubation [70].

IC50, µM ± S.D. Compound 51 52 53 54 55 56 57 Int. J. Mol. Sci. 2020, 21,32. 39657 ± 0.6 31.3 ± 1.7 20 ± 1.5 46.7 ± 0.4 37.6 ± 1.1 34.5 ± 0.9 25.4 ± 0.514 of 37

Flow cytometry was conducted for accessing the apoptosis rate of U87 cells treated with compoundsFlow cytometry 51–57 after was conducted 24 h. All for the accessing complexes the apoptosis, except ratecompound of U87 cells 54, treated significantly with compounds induced 51apoptosis–57 after in 24 U87 h. All cells. the The complexes, ability of except compounds compound 51–5754 to, significantly inhibit cancer induced cell growth apoptosis by induction in U87 cells. of Thecell cycle ability arrest of compounds was also estimated.51–57 to inhibit Treatment cancer of cell U87 growth cells with by induction compounds of cell 51– cycle57 caused arrest a was marked also estimated.arrest of G1, Treatment a growth of phase U87 cells that with plays compounds a key role 51in– cell57 caused cycle progression a marked arrest andof ensures G1, a growth that DNA phase is thatready plays for synthesis. a key role To in cellvalidate cycle the progression hypothesis and that ensures 51–57 that triggered DNA is apoptosis ready for in synthesis. treated U87 To validatecells by the hypothesis effect on the that expression51–57 triggered level apoptosis of apoptotic in treated and anti U87-apoptotic cells by the genes, effect an on RTthe-PCR expression assay level was ofconducted. apoptotic The and results anti-apoptotic obtained genes, showed an RT-PCRan increased assay level was conducted.of caspase-independent The results obtained apoptosis showed genes anincluding increased P53, level P21, of Bid, caspase-independent and Bax in U87 cells apoptosis after treatment genes includingby all the complexes P53, P21, Bid, except and compounds Bax in U87 cells54 and after 55. treatmentThe level of by anti all- theapoptotic complexes genes except Bcl-2 compoundsand Bcl-xL was54 andmarkedly55. The inhibited level of in anti-apoptotic the presence genesof compounds Bcl-2 and 55 Bcl-xL and 56 was. markedly inhibited in the presence of compounds 55 and 56. The investigation of the antitumor activity of Cu(I) coordination compounds 51–57, thus thus,, demonstrated an inhibition of cell growth, cell cycle progression, migration ability, and expressionexpression level ofof anti-apoptoticanti-apoptotic genes and an inducedinduced apoptosis,apoptosis, necrosis, and expression level of apoptoticapoptotic genes in a dose-dose- and time-dependenttime-dependent mannermanner inin treatedtreated U87U87 cells.cells. Kacar etet al. al. reported reported coordination coordination compound compound 58 based on a a pyridyl-fluorobenzimidazole pyridyl-fluorobenzimidazole scascaffoldffold [[7171]] (Figure(Figure 1616).).

Figure 16. SyntheSynthesissis scheme, chemical structure of coordination compound 5858..

Compound 5858showed showed antiproliferative antiproliferative and and apoptotic apoptotic eff ects effects on NIH on NIH/3T3/3T3 normal normal fibroblast fibroblast cells andcells onand SPC212 on SPC212 mesothelioma mesothelioma and DU145 and DU145 prostate prostate cancer cancer cells. Thecells. most The significantmost significant IC50 values IC50 values were foundwere found against against DU145, DU145 i.e., 37.0,, i.e., 21.1, 37.0, and 21.1, 10.0 and mM 10.0 for the mM 24, for 48, the and 24, 72 h48, treatments, and 72 h respectively. treatments, AInt.respectively dose-dependent J. Mol. Sci. 20. 20A , dose21, x increase FOR-dependent PEER of REVIEW pro-apoptotic increase of pro Bax-apoptotic protein in Bax DU145 protein preincubated in DU145 preincubated with coordination16 with of 38 compoundcoordination58 compoundwas observed. 58 was observed. Majouga etet al. reported al. reported mixed-valence mixed- Cu(IIvalence/I) copper Cu(II/I) compounds copper based compounds on 2-thioimidazolones based on with2 -thioimidazolones promising antitumor with promising activity [72 antitumor] (Figure activity17). [72] (Figure 17).

Figure 17. Chemical structure of coordination compounds 59–63. Figure 17. Chemical structure of coordination compounds 59–63.

An MTT test on MCF-7 (breast cancer cell line of invasive breast ductal carcinoma), SiHa (human cervical cancer cells with the modal chromosome number of 71), and HEK 293 (human embryonic kidney cell line) cell lines showed promising antitumor ability of compounds 59–62 (Table 7). The ability of compound 60 to damage DNA was confirmed by tunnel assay. Compound 60 also proved to be an effective telomerase inhibitor. Nuclear accumulation of labeled coordination compound 63 was proven using fluorescent microscopy.

Table 7. MTT data of coordination compounds 59–62 after 72 h of incubation [72].

IC50, µM ± S.D. Compound MCF-7 SiHa HEK293 59 3.7 ± 1.6 3.0 ± 0.2 2.5 ± 0.4 60 2.1 ± 0.8 2.2 ± 0.7 2.3 ± 0.9 61 7.4 ± 1.4 3.9 ± 2.3 25.3 ± 1.2 62 13.4 ± 3.8 8.5 ± 0.4 12.7 ± 3.7 Dox 2.1 ± 0.8 2.0 ± 0.8 1.1 ± 0.1 CDDP 64.1 ± 3.9 - 12.4 ± 3.9

An in vivo investigation of antitumor activity was conducted using breast adenocarcinoma Ca-755 inoculated into mice lines C57BL/6 (female). Treatment began 48 h after vaccination with compound 60 (24 and 12 mg/kg/d) injected intraperitoneally at 24-h intervals for five days. Indicators of tumor growth inhibition for mice with a course of the test substance at a dose of 12 mg/kg was 46.1% on day seven after the end of treatment and 36.1% on day 14 after the end of treatment. For a dose of 24 mg/kg, it was 73.5% on day seven after the end of treatment and 59.5% on day 14 after the end of treatment, animal’s body weight loss did not exceed 10%. Telomerase inhibitor compound 60, thus, proved to be an effective antitumor agent.

Int. J. Mol. Sci. 2020, 21, 3965 15 of 37

An MTT test on MCF-7 (breast cancer cell line of invasive breast ductal carcinoma), SiHa (human cervical cancer cells with the modal chromosome number of 71), and HEK 293 (human embryonic kidney cell line) cell lines showed promising antitumor ability of compounds 59–62 (Table7). The ability of compound 60 to damage DNA was confirmed by tunnel assay. Compound 60 also proved to be an effective telomerase inhibitor. Nuclear accumulation of labeled coordination compound 63 was proven using fluorescent microscopy.

Table 7. MTT data of coordination compounds 59–62 after 72 h of incubation [72].

IC50, µM S.D. ± Compound MCF-7 SiHa HEK293 59 3.7 1.6 3.0 0.2 2.5 0.4 ± ± ± 60 2.1 0.8 2.2 0.7 2.3 0.9 ± ± ± 61 7.4 1.4 3.9 2.3 25.3 1.2 ± ± ± 62 13.4 3.8 8.5 0.4 12.7 3.7 ± ± ± Dox 2.1 0.8 2.0 0.8 1.1 0.1 ± ± ± CDDP 64.1 3.9 - 12.4 3.9 ± ±

An in vivo investigation of antitumor activity was conducted using breast adenocarcinoma Ca-755 inoculated into mice lines C57BL/6 (female). Treatment began 48 h after vaccination with compound 60 (24 and 12 mg/kg/d) injected intraperitoneally at 24-h intervals for ﬁve days. Indicators of tumor growth inhibition for mice with a course of the test substance at a dose of 12 mg/kg was 46.1% on day seven after the end of treatment and 36.1% on day 14 after the end of treatment. For a dose of 24 mg/kg, it was 73.5% on day seven after the end of treatment and 59.5% on day 14 after the end of treatment, animal’s body weight loss did not exceed 10%. Telomerase inhibitor compound 60, thus, proved to be an eﬀective antitumor agent.

2.4. S/S-Donor Ligands The cytotoxic activity of copper coordination compounds can occur both when a coordination compound solution is administered in vivo/in cell and when a nontoxic ligand is administered with the cytotoxic coordination compound forming in situ (this approach has already been described for elesclomol in Section 2.2). Another example of the formation of a coordination compound during therapy is disulfiram (DSF), an FDA-approved drug for treating alcoholism. In recent years, the drug has attracted much attention as an antitumor inducer of ROS formation acting in combination with copper gluconate [73]. Disulfiram alone has a negligible effect on tumor cells, but in the presence of Cu(II) ions in a nanomolar range, it is effective against a wide range of tumor cell lines, as shown in [74]. In vivo administration of folate-targeted nanoparticles with encapsulated DSF to animals with subcutaneous models of breast cancer led to a decrease in tumor growth [75]. It was repeatedly shown [76] that the cytotoxic effect of the DSF emerges as a result of in situ formation of the coordination compound Cu-DSF. But a thorough study of DSF metabolism led to the conclusion that the coordination compound Cu-DSF does not in fact exist [77]. It was found in [78] that even in an aqueous solution, DSF does not form a coordination compound with copper and in fact decomposition into diethyldithiocarbamate (DDC) occurs. The resulting diethyldithiocarbamate (DDC) reacts with Cu(II) to form copper diethyldithiocarbamate 64, Cu(DDC)2 (Figure 18). Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 17 of 38

coordination compound Cu-DSF. But a thorough study of DSF metabolism led to the conclusion that the coordination compound Cu-DSF does not in fact exist [77]. It was found in [78] that even in an aqueous solution, DSF does not form a coordination compound with copper and in fact decomposition into diethyldithiocarbamate (DDC) occurs. The resulting diethyldithiocarbamate Int. J. Mol. Sci. 2020, 21, 3965 16 of 37 (DDC) reacts with Cu(II) to form copper diethyldithiocarbamate 64, Cu(DDC)2 (Figure 18).

Figure 18. Schemes of the interaction of copper (II) with disulfiram (DSF) and the formation of a coordination compound 64. Figure 18. Schemes of the interaction of copper (II) with disulfiram (DSF) and the formation of a Dithiocarbamatescoordination compound are a 64 known. class of copper chelating agents that exhibit significant antitumor activity in vitro against various tumor cell lines [79]. We note that the use of DSF in combination with copperDithiocarbamates gluconate is are in clinical a known trials class as a breast of copper cancer chelating treatment agents scheme that [80 ]. exhibit Liu et al. significant studied theantitumor mechanism activity of action in vitro in detail against and various reported tumor that the cell disulfiram lines [79]./ copper We note mixture that the inhibits use of Bcl2 DSF and in inducedcombination Bax proteinwith copper expression gluconate in all is GBMin clinical cell lines,trials inducesas a breast ROS cancer activity, treatment activates scheme the apoptosis [80]. Liu. JNKet al. pathway, studied causes the mechanism ROS-dependent of action activation in detail of the and JNK reported and p38 that pathways, the disulfiram/copper and inhibits NFkB mixture and ALDHinhibits activity Bcl2 and [81]. induced In a comment Bax protein to Liu et expression al., Cvek recalled in all GBM successful cell lines, trials for induces breast ROS cancer activity, using diethyldithiocarbamateactivates the apoptosis in JNK 1993 [pathway,82] and called causes for aROS return-dependent to undeservedly activation forgotten of the clinical JNK and trials p38 of thispathway inexpensives, and and inhibits effective NFkB drug and [83 ALDH]. Accordingly, activity [Cvek,81]. In disulfiram a comment could to be Liu used et al. for, Cvektreating recalled brain tumorssuccessful and trials even for other breast cancers. cancer using diethyldithiocarbamate in 1993 [82] and called for a return to undeservedlyAt present, forgotten DSF/Cu clinical antitumor trials activity of this inexpensive is still of interest. and effective Duan drug et al. [83 proposed]. According synergisticly, Cvek, breastdisulfiram tumor could therapy be used via for codelivery treating ofbrain doxorubicin tumors and and even disulfiram other cancers. cell killing using pH-sensitive core-shell-coronaAt present, DSF/Cu nanoparticles antitumor [84]. activity is still of interest. Duan et al. proposed synergistic breast tumorIn vivo therapyantitumor via codelivery efficacy was of proved doxorubici usingn and 4T1 tumor-bearing disulfiram cell mice. killing The using tumor pH inhibiting-sensitive ratecore- wasshell 34.81%-corona fornanoparticles the DSF-treated [84]. group, 68.27% for the DOX-treated group, 80.92% for the DSF +InDOX-treated vivo antitumor group, efficacy and 89.27%was proved for the using Co-NPs-treated 4T1 tumor-bearing group. mice. The tumor inhibiting rate was Wencheng 34.81% for Wu the published DSF-treated a delicate group, pH-sensitive 68.27% for Cu the/DSF DOX delivery-treated approach group, based80.92% on for constructing the DSF + mesoporousDOX-treated silica group nanoparticles, and 89.27% for enriched the Co with-NPs covalent-bonded-treated group. Cu2+ ions and physically bonded DSF [Wencheng85]. Under mild Wu acidicpublished conditions a delicate of the pH tumor-sensitive microenvironment, Cu/DSF delivery a rapid approach biodegradation based ofon nanoparticlesconstructing wasmesoporous assumed silicato occur nanoparticles with subsequent enriched Cu2 + withand DSF covalent release-bonded and an Cu instantaneous2+ ions and chelationphysicall reactiony bonded leading DSF [85 to] the. Under formation mild ofacidic a cytotoxic conditions agent. of Thisthe tumor approach microenvironment, was successfully a tested, rapid provingbiodegradation its efficacy of nanoparticlesin vitro and in was vivo assumed. Treating to 4T1occur tumor-bearing with subsequent female Cu2+ BALB and /DSFC nude release mice and with an 3.75instantaneous mg/kg of DSF chelation dose (as reaction part of leading the developed to the formation formulation) of a led cytotoxic to 71.4% agent tumor. This growth approach inhibition was aftersuccessfully two weeks tested, of treatment, proving and its efficacy no significant in vitro body-weight and in vivo changes. Treating was 4T1 observed. tumor-bearing female BALB/CYiqiu nude Li et mice al. with reported 3.75 mg/kg a DSF /ofCu DSF combination dose (as part to induceof the developed anti-NPC formulation) activity through led to a 71.4% joint actiontumor ofgrowth multiple inhibition apoptosis after pathways, two weeks such of treatment as an increasing, and no chloride significant channel-3 body-weight protein changes expression, was inducingobserved. ROS production, and decreasing NF-KB-p65 expression [86], and inhibiting the expression of α-SMAYiqiu in cancer-associated Li et al. reported fibroblastsa DSF/Cu combination (CAFs) [87]. toIn induce vivo, DSF anti/Cu-NPC combined activity with through cisplatin a joint (CDDP) action therapyof multiple was well apoptosis tolerated pathways, and could such significantly as an increasing suppress chloride the growth channel of nasopharyngeal-3 protein expression, (NPC) tissuesinducing [88 ROS]. McMahon production, et al. and recently decreasing summarized NF-KB all-p65 drug-delivery expression [86 approaches], and inhibiting and formulations the expression for DSFof α/Cu-SMA combinations in cancer-associated [89]. Several fibroblasts active clinical (CAFs) trials [87] testing. In vivo, the anticancerDSF/Cu combined efficacy of with DSF cisplatin against various(CDDP) cancers therapy are was underway, well tolerated such as and germ could cell tumorsignificantly treatment suppress [90] and the breast growth neoplasm of nasopharyngeal female and metastatic breast cancer [80,91]. Therefore, Disulfiram is a copper-based antitumor drug with great therapeutic potential for treating malignant tumors. Zinc pyrithione is an agent with antimicrobial activity [92]. The ligand pyrithione itself has no antiproliferative activity, but copper coordination compound 65 based on it demonstrated antitumor Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 18 of 38

(NPC)Int. J. Mol. tissues Sci. 2020 , [2881, ].x FOR McMahon PEER REVIEW et al. recently summarized all drug-delivery approaches18 andof 38 formulations for DSF/Сu combinations [89]. Several active clinical trials testing the anticancer efficacy(NPC) tissuesof DSF [against88]. McMahon various cancerset al. arerecently underway, summa suchrized as allgerm drug cell- deliverytumor treatment approaches [90] and bformulationsreast neoplasm for female DSF/С andu combinations metastatic breast [89]. cancer Several [80,91 active]. Therefore, clinical trialsDisulfiram testing is a the copper anticancer-based antitumorefficacy of drugDSF withagainst great various therapeutic cancers potential are underway, for treating such malignant as germ cell tumors. tumor treatment [90] and breastZinc neoplasm pyrithione female is an and agent metastatic with antimicrobial breast cancer activity [80,91]. [92Therefore,]. The ligand Disulfiram pyrithione is a copper itself has-based no aantitumorntiproliferative drug with activity, great but therapeutic copper coordination potential for compound treating malignant 65 based ontumors. it demonstrated antitumor activityZinc [93 pyrithione] (Figure is 19 an). Aagent study with of cytotoxicityantimicrobial of activity compound [92]. The65 on ligand breast pyrithione cancer cells itself (MCF has -no7) Int.antiproliferative J. Mol. Sci. 2020, 21 activity,, 3965 but copper coordination compound 65 based on it demonstrated antitumor17 of 37 showed significant activity of the coordination compound with IC50 = 0.375 μM. Activity was also activity [93] (Figure 19). A study of cytotoxicity of compound 65 on breast cancer cells (MCF-7) detected in U266 multiple myeloma cells with IC50 = 0.130 µM and in HepG2 liver cells with IC50 = activity0.495showed µM. [ 93significant ] (Figure 19activity). A study of the of cytotoxicitycoordination of compound 65withon breastIC50 = 0.375 cancer μM. cells Activity (MCF-7) was showed also detected in U266 multiple myeloma cells with IC50 = 0.130 µM and in HepG2 liver cells with IC50 = signiﬁcant activity of the coordination compound with IC50 = 0.375 µM. Activity was also detected in 0.495 µM. U266 multiple myeloma cells with IC50 = 0.130 µM and in HepG2 liver cells with IC50 = 0.495 µM.

Figure 19. Chemical structure of the coordination compound 65 (CuPT). Figure 19. Chemical structure of the coordination compound 65 (CuPT). The cytotoxicFigure effects 19. ofChemical compound structure 65 ofw theere coordinationevaluated excompound vivo on 65 bone (CuPT) marrow. cells from patientsThe with cytotoxic acute eff myeloidects of compound leukemia 65(AML)were evaluatedand on mononuclear ex vivo on bone cells marrowfrom peripheral cells from blood patients of The cytotoxic effects of compound 65 were evaluated ex vivo on bone marrow cells from withhealthy acute volunteers. myeloid leukemia In AML (AML) patients, and (CTR) on mononuclear CuPT and Bortezomib cells from peripheral, respectively blood, reduced of healthy the patients with acute myeloid leukemia (AML) and on mononuclear cells from peripheral blood of volunteers.viability of Inprimary AML patients, monocyte (CTR) cells CuPT with anand average Bortezomib, IC50 of respectively, 57.03 and reduced 20.50 nM, the while viability in an of healthy volunteers. In AML patients, (CTR) CuPT and Bortezomib, respectively, reduced the experiment with healthy cells (CTR), the average IC50 was respectively estimated at 101.08 and 74.23 primary monocyte cells with an average IC50 of 57.03 and 20.50 nM, while in an experiment with healthy nM.viability A 12 - ofh incubation primary monocyte of AML cells cells with with coordinati an averageon compoundIC50 of 57.03 65 in and doses 20.50 ranging nM, whilefrom 0.25 in anto cells (CTR), the average IC50 was respectively estimated at 101.08 and 74.23 nM. A 12-h incubation of experiment with healthy cells (CTR), the average IC50 was respectively estimated at 101.08 and 74.23 AML0.75 μM cells led with to coordination apoptosis, which compound was shown65 in doses by staining ranging with from annexin 0.25 to 0.75V/PIµ byM led flow to cytometry. apoptosis, nM. A 12-h incubation of AML cells with coordination compound 65 in doses ranging from 0.25 to whichCoordination was shown compound by staining 65, thus with, showed annexin efficacy V/PI by in flow AML cytometry. therapy Coordinationas compared with compound clinically65, used thus, 0.75 μM led to apoptosis, which was shown by staining with annexin V/PI by flow cytometry. showedBortezomib. efficacy in AML therapy as compared with clinically used Bortezomib. Coordination compound 65, thus, showed efficacy in AML therapy as compared with clinically used 2.5.Bortezomib. N N-,-, O-,O-, and S-DonorS-Donor Ligands

2.5. NThe-, O antitumorantitumor, and S-Donor activity activity Ligands of ofcopper copper coordination coordination compounds compounds with thiosemicarbazone with thiosemicarbazone has been has known been sinceknown the since 1960s. the Zhang 1960s. H etZhang al. reported H et Cu(II) al. reported coordination Cu(II) compound coordination66 with compound thiosemicarbazide 66 with 8-hydroxyquinoline-2-carboxaldehydethiosemicarbazideThe antitumor activity 8-hydroxyquinoline of copper coordination Cu(HQTS)-2-carboxaldehyde and compounds67 8-hydroxyquinoline-2-carboxaldehyde- with Cu(HQTS) thiosemicarbazone and has been67 known since the 1960s. Zhang H et al. reported Cu(II) coordination compound 66 with 4,4-dimethyl-3-thiosemicarbazide8-hydroxyquinoline-2-carboxaldehyde Cu(HQDMTS)-4,4-dimethyl (Figure-3-thiosemicarbazide 20)[94]. An IC50 of Cu(HQDMTS) 0.13 µM for compound (Figure 2066) thiosemicarbazide 8-hydroxyquinoline-2-carboxaldehyde Cu(HQTS) and 67 and[94]. 0.64An ICµ50M of for 0.13 compound μM for compound67 was obtained 66 and 0.64 as μM a result for compound of a study 67 on was SK-N-DZ obtained neuroblastoma as a result of a cellstudy8-hydroxyquinoline populations. on SK-N-DZ neuroblastoma-2-carboxaldehyde cell -populations.4,4-dimethyl -3-thiosemicarbazide Cu(HQDMTS) (Figure 20) [94]. An IC50 of 0.13 μM for compound 66 and 0.64 μM for compound 67 was obtained as a result of a study on SK-N-DZ neuroblastoma cell populations.

Figure 20. Chemical structures of the coordination compounds 66 and 67. Figure 20. Chemical structures of the coordination compounds 66 and 67.

Hancock et al.Figure reported 20. Chemical simultaneous structures administration of the coordination of a compounds thiosemicarbazone 66 and 67. copper-chelating agent in combination with a copper salt [95]. In situ formation of a cytotoxic coordination compound was assumed, as in the case of Disulfiram and elesclomol (Figure 21). A study of the cytotoxic effect of thiosemicarbazone ligand L68 in combination with an equimolar amount of copper chloride showed the induction of a cytotoxic effect by oxidative stress, glutathione depletion, and ROS formation. Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 19 of 38 Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 19 of 38 Hancock et al. reported simultaneous administration of a thiosemicarbazone copper-chelating Hancock et al. reported simultaneous administration of a thiosemicarbazone copper-chelating agent in combination with a copper salt [95]. In situ formation of a cytotoxic coordination compound agent in combination with a copper salt [95]. In situ formation of a cytotoxic coordination compound was assumed, as in the case of Disulfiram and elesclomol (Figure 21). A study of the cytotoxic effect was assumed, as in the case of Disulfiram and elesclomol (Figure 21). A study of the cytotoxic effect of thiosemicarbazone ligand L68 in combination with an equimolar amount of copper chloride of thiosemicarbazone ligand L68 in combination with an equimolar amount of copper chloride showed the induction of a cytotoxic effect by oxidative stress, glutathione depletion, and ROS showed the induction of a cytotoxic effect by oxidative stress, glutathione depletion, and ROS Int.formation. J. Mol. Sci. 2020, 21, 3965 18 of 37 formation.

Figure 21. Chemical structure of organic ligand L68. FigureFigure 21.21. Chemical structure of organic ligandligand L68L68.. InIn vivovivo toxicologicaltoxicological studiesstudies havehave shownshown thatthat thethe administrationadministration ofof 100100 mgmg/kg/kg ofof ligandligand in 100% In vivo toxicological studies have shown that the administration of 100 mg/kg of ligand in 100% DMSODMSO doesdoes notnot leadlead toto aa decreasedecrease inin tumortumor massmass inin mice.mice. AnAn equimolarequimolar mixturemixture ofof aa ligandligand L68L68 withwith DMSO does not lead to a decrease in tumor mass in mice. An equimolar mixture of a ligand L68 with coppercopper chloridechloride inin micemice showedshowed thethe maximummaximum toleratedtolerated dosedose ofof 1515 mgmg/kg,/kg, andand thethe administrationadministration ofof copper chloride in mice showed the maximum tolerated dose of 15 mg/kg, and the administration of 33 mgmg/kg/kg intravenouslyintravenously twotwo timestimes aa dayday duringduring ﬁvefive daysdays resultedresulted inin aa signiﬁcantsignificant (42%)(42%) decreasedecrease inin 3 mg/kg intravenously two times a day during five days resulted in a significant (42%) decrease in tumorstumors (human(human leukemialeukemia cellcell lineline HL60),HL60), withwith lessless thanthan 20%20% bodybody weightweight loss.loss. AnAn abilityability toto depletedeplete tumors (human leukemia cell line HL60), with less than 20% body weight loss. An ability to deplete glutathioneglutathione inin thethe cellcell byby thethe samesame mechanismmechanism as as arsenic arsenic oxide oxide (As (As2O2O33)) waswas alsoalso proven.proven. TheThe authorsauthors glutathione in the cell by the same mechanism as arsenic oxide (As2O3) was also proven. The authors suggestedsuggested that that a acombination combination of ligand of ligandL68 L68with with a copper a copper salt can salt be can used be in used conjunction in conjunction with classical with suggested that a combination of ligand L68 with a copper salt can be used in conjunction with chemotherapeuticclassical chemotherapeutic agents (cisplatin agents (cisplatin or Bortezomib). or Bortezomib). classical chemotherapeutic agents (cisplatin or Bortezomib). CarcelliCarcelli etet al. al. reported reported synthesis synthesis and and the the cytotoxic cytotoxic activity activity of Cu(II) of Cu(II) coordination coordination compounds compounds with Carcelli et al. reported synthesis and the cytotoxic activity of Cu(II) coordination compounds variouslywith variously substituted substituted salicylaldehyde salicylaldehyde thiosemicarbazone thiosemicarbazone ligands ligands [96] (Figure [96] 22(Figure). Inhibition 22). Inhibition doses in with variously substituted salicylaldehyde thiosemicarbazone ligands [96] (Figure 22). Inhibition thedoses low in nanomolar the low nanomolar range were range found were in somefound cases. in some cases. doses in the low nanomolar range were found in some cases.

FigureFigure 22.22. Synthesis scheme and chemical structurestructure ofof coordinationcoordination compoundscompounds 6969––7474.. Figure 22. Synthesis scheme and chemical structure of coordination compounds 69–74. TheThe inin vitrovitro activityactivity of of copper copper complexes complexes on on a a pair pair of of human human colon colon cancer cancer cell cell lines lines The in vitro activity of copper complexes on a pair of human colon cancer cell lines (LoVo(LoVo/LoVo/LoVo-OXP)-OXP) showed showed anticancer anticancer activity activity of of the the coordination coordination compounds compounds including including on on the the (LoVo/LoVo-OXP) showed anticancer activity of the coordination compounds including on the oxaliplatin-resistantoxaliplatin-resistant humanhuman coloncolon cancercancer cellcell lineline ofof supraclavicularsupraclavicular lymphlymph nodenode metastasismetastasis cellcell lineline oxaliplatin-resistant human colon cancer cell line of supraclavicular lymph node metastasis cell line LoVo-OXPLoVo-OXP(Table (Table8 ).8). LoVo-OXP (Table 8).

TableTable 8.8. MTTMTT datadata ofof coordinationcoordination compoundscompounds 6969––7474 afterafter 7272 hh ofof incubationincubation [[9696].]. Table 8. MTT data of coordination compounds 69–74 after 72 h of incubation [96].

IC50, µ M S.D.IC50, µM ± S.D. ± IC50, µM ± S.D. CompoundCompound LoVo LoVo LoVo LoVo-OXP-OXP Compound LoVo LoVo-OXP 69 0.031 ± 0.001 0.004 ± 0.001 6969 0.030.0311 ± 0.0010.001 0.00 0.0044 ± 0.0010.001 ± ± 70 70 0.020.0299 ± 0.0080.008 0.03 0.0300 ± 0.0100.010 70 0.029 ± ±0.008 0.030 ± 0.010± 71 71 0.030.0366 ± 0.0090.009 0.00 0.0088 ± 0.0020.002 71 0.036 ± ±0.009 0.008 ± 0.002± 72 72 0.020.0200 ± 0.0010.001 0.02 0.0200 ± 0.0010.001 72 0.020 ± ±0.001 0.020 ± 0.001± 73 73 0.20.211 ± 0.080.08 0.09 0.09 ± 0.010.01 73 0.21 ± ±0.08 0.09 ± 0.01± 74 74 0.030.0300 ± 0.0010.001 0.02 0.02 ± 0.010.01 74 0.030 ± ±0.001 0.02 ± 0.01± Oxaliplatin 2.17 1.37 13.92 1.68 ± ±

Three-dimensional MTT tests on colorectal adenocarcinoma cell line HCT-116, human pancreatic adenocarcinoma cell line PSN-1 spheroids also showed a high antiproliferative activity of coordination compounds 69–74 and a signiﬁcant eﬃcacy as compared with cisplatin (Table9). Int. J. Mol. Sci. 2020, 21, 3965 19 of 37

Table 9. MTT on three-dimensional (3D) spheroids of HCT-15 and PSN1 cancer cell spheroids of the coordination compounds 69–74 [96].

IC50, µM S.D. ± Compound HCT-116 PSN-1 69 1.08 0.38 0.90 0.02 ± ± 70 3.56 1.67 1.17 0.11 ± ± 71 1.25 0.98 0.90 0.30 ± ± 72 1.17 0.62 0.94 0.27 ± ± 73 1.69 0.45 1.18 0.23 ± ± 74 1.28 0.62 0.91 0.01 ± ± CDDP 68.20 4.57 52.60 3.78 ± ±

Cellular uptake and distribution were estimated using ICP-MS. A direct correlation between cellular accumulation and cytotoxic potency was not found by comparing uptake and cytotoxicity data in LoVo human colon cancer cells. Compounds 69–71 accumulated, substantially, in the mitochondria fraction and to a lesser extent in cytosolic fractions. To estimate if compounds 69–74 cause DNA damage, DNA fragmentation was evaluated using alkaline single-cell gel electrophoresis (comet assay), and no DNA fragmentation was observed. In addition, no glutathione depletion was observed, and an ROS evaluation confirmed that TSC complexes did not provoke a substantial increase of cellular ROS levels. Therefore, the promising antitumor activity of these coordination compounds is not associated with the redox activity of copper ions. In clarification of the mechanism of cytotoxic action, the complexes were found to inhibit the protein disulfide isomerase (PDI) enzyme. Therefore, it was hypothesized that the coordination compounds 69–74 interfere with PDI activity, possibly inhibiting its disulfide bond catalytic activity. The in vivo antitumor activity of compound 69 was evaluated in a solid tumor model, the highly aggressive syngeneic murine Lewis lung carcinoma (LLC). Tumor growth inhibition induced by compound 69 was compared with that of cisplatin (Table 10).

Table 10. In vivo antitumor activity of coordination compound 69 (cisplatin as a control) on Lewis lung carcinoma (LLC) tumor-bearing mice [96].

Daily Dose Average Tumor Weight Inhibition of Tumor (Mg/Kg) (Mean S.D., G) Growth (%) ± Control - 0.459 0.130 - ± 69 3 0.239 0.080 48.0 ± 69 6 0.118 0.090 74.3 ± CDDP 1.5 0.114 0.080 75.2 ±

A 6 mg/kg dose of coordination compound 69 induced tumor growth inhibition of about 74%, similar to cisplatin dosed at 1.5 mg/kg, but the time course of changes in body weight indicated that cisplatin induced elevated anorexia. In contrast, treatment with compound 69 did not induce a substantial body weight loss (<10%) throughout the therapeutic experiment. Once again, these results confirm the higher biocompatibility of copper-containing anticancer drugs as compared with classical platinum therapy. Kongot et al. developed Cu(II) coordination compound 75 based on a ligand obtained by the reaction between S-benzyldithiocarbamate and 2-hydroxy-5-(phenyldiazenyl)benzaldehyde [97] (Figure 23). The ability of compound 75 to bind to bovine serum albumin (BSA) was tested using its own fluorescence quenching method (titration of a protein solution with a solution of a coordination compound). The binding constant of coordination compound 75 calculated using the Benesi–Hildebrand equation was Ka = 0.94 104 M 1, which indicates a significant binding energy. × − Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 21 of 38 Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 21 of 38 own fluorescence quenching method (titration of a protein solution with a solution of a coordination compound). The binding constant of coordination compound 75 calculated using the Benesi– Int.own J. Mol. fluorescence Sci. 2020, 21, 3965quenching method (titration of a protein solution with a solution of a coordination20 of 37 compound).Hildebrand equation The binding was Kconstanta = 0.94 of× 10 coordination4 M−1, which compound indicates a75 significant calculated binding using theenergy. Benesi The– Hildebrandauthors believe equation that the was most Ka =probable 0.94 × 10 reason4 M−1, for which this indicatesaffinity is a the significant hydrogen binding bond between energy. Thethe Theauthorsamino authors groups believe believe of that the that aminothe the most most acid probable residues probable reason of reason the proteinfor for this this andaffinity affi thenity phenolicis isthe the hydrogen hydrogenoxygen ofbond the bond ligand.between between the the aminoamino groups groups of of the the amino amino acid acid residues residues of of the the proteinprotein andand the phenolic oxygen oxygen of of the the ligand. ligand.

Figure 23. Chemical structure of coordination compound 75. Figure 23. Chemical structure of coordination compound 75. Figure 23. Chemical structure of coordination compound 75. The cytotoxic activity of coordination compound 75 and the corresponding ligand was studied The cytotoxic activity of coordination compound 75 and the corresponding ligand was studied on humanThe cytotoxic cervical activity cancer cellsof coordination HeLa. Significant compound cell death 75 and was the achieved corresponding in the concentrationligand was studied range on human cervical cancer cells HeLa. Significant cell death was achieved in the concentration range onof 2human–10 μM. cervical The calculated cancer cells IC50 HeLa. values Significant were 4.46 μMcell fordeath coordination was achieved compound in the concentration 75 and 5.34 μM range for of 2–10 µM. The calculated IC50 values were 4.46 µM for coordination compound 75 and 5.34 µM ofthe 2 – corresponding10 μM. The calculated ligand. IC The50 values selectivity were of 4.46 the μM drugs for coordination obtained was compound evaluated 75 by and comparing 5.34 μM thefor for the corresponding ligand. The selectivity of the drugs obtained was evaluated by comparing the thecytotoxicity corresponding of the obtained ligand. The compounds selectivity on of healthy the drugs human obtained cells wasHEK evaluated-293, which by showed comparing a rather the cytotoxicity of the obtained compounds on healthy human cells HEK-293, which showed a rather high cytotoxicityhigh cell death of the at aobtained concentration compounds of 10 μM. on healthyOn the humanbasis of cellsthe obtainedHEK-293, data, which CC50 showed = 6.31 a μM rather for µ µ cellhighcoordination death cell at death a concentration compound at a concentration of75 10 andM. of 10.90 On 10 the μM. μM basis On for ofthe the the basis obtained ligand of the were data, obtained calculated. CC50 data,= 6.31 CC50 Thus,M for= coordination6.31 coordination μM for µ compoundcoordinationcompound75 75and compoundwas 10.90 foundM to75 forbe and selective the 10.90 ligand for μM werehealthy for calculated. the cells ligand at a concentration Thus, were calculated.coordination of IC 50 Thus, = compound4.46 coordinationμM obtained75 was µ foundcompoundon HeLa to be cell selective 75 lines, was found forwhich healthy to also be selectiveconfirm cells at aed for concentration the healthy selectivity cells of atof ICa the concentration50 =coordination4.46 M obtainedof compoundIC50 = 4.46 on HeLaμM with obtained respect cell lines, whichonto tumorHeLa also confirmed cellcells. lines, which the selectivity also confirm of theed coordinationthe selectivity compound of the coordination with respect compound to tumor with cells. respect to tumor cells. 2.6.2.6. Phosphine-Donor Phosphine-Donor Ligands Ligands 2.6.Cu(I) PhosphineCu(I) phosphine-based phosphine-Donor Ligands-based coordination coordination compounds compounds attract wide wide attention attention due due to to high high cytotoxicity, cytotoxicity, antibacterial,antibacterialCu(I) phosphine and, an anti-inﬂammatoryd anti-based-inflammatory coordination properties properties compounds [98].[98] The .attract The use of wide use a phosphine ofattention a phosphine due ligand to high prevents ligand cytotoxicity, prevents oxidation andantibacterialoxidation hydrolysis and, reactions anhydrolysisd anti due-inflammatory reactions to a strong due copper–phosphine propertiesto a strong[98] copper. The– interaction phosphine use of a [in33 phosphineteraction], which [33] allows ligand, which stabilization prevents allows of copperoxidationstabilization in and a monovalent of hydrolysis copper in state,areactions monovalent providing due tostate, a divers strong providing biological copper divers–phosphine activity. biological interaction activity. [33], which allows stabilizationKhanKhan et al.et of [99 al.copper] [99] reported inreported a monovalent nine ninecopper copper state, Cu(I) providing Cu(I) complexes complexes divers based biological based on thiphenilphosphine on activity. thiphe nilphosphine and thiourea and (Figurethiourea Khan24). (Figure et al. 24[99]). reported nine copper Cu(I) complexes based on thiphenilphosphine and thiourea (Figure 24).

Figure 24. Chemical structure of coordination compounds 76–84.

The synthesized compounds were utilized in diﬀerent biological assays, which showed antibacterial, antifungal, antilieshmanial, antioxidant, and cytotoxic properties against brine shrimps. Compounds 79 and 81 proved to be the most active molecules against bacteria, fungi, and the lieshmanial pathogen, in addition to having an excellent antioxidant activity. Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 22 of 38

Int. J. Mol. Sci. 2020, 21, x FOR PEERFigure REVIEW 24. Chemical structure of coordination compounds 76–84. 22 of 38

The synthesizedFigure 24. compounds Chemical structure were utilizedof coordination in different compounds biological 76–84. assays, which showed Int. J. Mol.antibacterial, Sci. 2020, 21, 3965 antifungal, antilieshmanial, antioxidant, and cytotoxic properties against brine 21 of 37 Theshrimps. synthesized Compounds compounds 79 and 81 were proved utilized to be the in most different active biological molecules assays, against which bacteria, showed fungi, and antibacterial,the lieshmanial antifungal, pathogen, antilieshmanial, in addition to antioxidant having an ,excellent and cytotoxic antioxidant properties activity. against brine shrimps. Compounds 79 and 81 proved to be the most active molecules against bacteria, fungi, and TapanelliTapanelli et al. [et100 al.] [100] reported reported two two water-soluble water-soluble Cu(I) Cu(I) phosphonate phosphonate complexes complexes compoun compoundsds 84 84 the lieshmanialand 85, which pathogen, showed in activity addition against to having Plasmodium an excellent early antioxidant sporogonic activity. (Figure 25). and 85,Tapanelli which showed et al. [100] activity reported against two water Plasmodium-soluble Cu(I) early phosphonate sporogonic complexes (Figure compoun 25). ds 84 and 85, which showed activity against Plasmodium early sporogonic (Figure 25).

Figure 25. Chemical structure of coordination compounds 84 and 85. Figure 25. Chemical structure of coordination compounds 84 and 85 .

CoordinationCoordination compoundFigure 25. compound Chemical84 with structure a more84 ofwith hydrophilic coordination a morecompounds and less hydrophilic bulky 84 and tris(hydroxymethyl)phosphane85. and less bulky showedtris(hydroxymethyl)phosphane inhibition of plasmodia growth showed at inhibition an early of stage plasmodia of the growth disease at an up early to 85 stage percent, of the while a stericallyCdiseaseoordination bulk up coordination to 85 percent, compound compound while 84a sterically 85withacted bulk a two coordination more to three hydrophilic compound times weaker 85 and acted in di two lessﬀerent to three bulky replicas. times At the tris(hydroxymethyl)phosphane showed inhibition of plasmodia growth at an early stage of the same time,weaker when in different the dose replicas. was reduced At the samefrom time,100 µ whenM, the the therapeutic dose was reduced eﬀect disappeared from 100 μM, completely. the disease up to 85 percent, while a sterically bulk coordination compound 85 acted two to three times For similartherapeutic gold and effect silver disappeared compounds, completely. inhibition For similar of parasite gold and growth silver in compounds, 80–85% occurred inhibition already of at weakerparasite in different growth replicas.in 80–85 % At occurred the same already time, at when concentrations the dose wasof 10 reducedμM. from 100 μM, the concentrations of 10 µM. therapeuticMashat effect et disappeared al. [101] reported completely. four Cu(I) For phenanthroline similar gold and-phosphine silver compounds, coordination inhibition compounds of 86– parasiteMashat89 (Figure growth et al. 26 in [101) .8 0–]85 reported% occurred four already Cu(I) at phenanthroline-phosphineconcentrations of 10 μM. coordination compounds 86–89 (Figure Mashat26). et al. [101] reported four Cu(I) phenanthroline-phosphine coordination compounds 86– 89 (Figure 26).

Figure 26. Chemical structure of coordination compounds 86–89.

Figure 26. Chemical structure of coordination compounds 86–89. CoordinationFigure compounds 26. Chemical 86–89 structureshowed DNAof coordination intercalation compounds ability. Strong86–89 DNA. binding is provided by triphenylphosphine ligands with a substituent at the 4-position of the phenyl ring Coordination compounds 86–89 showed DNA intercalation ability. Strong DNA binding is Coordinationcapable of forming compounds hydrogen86 bonds.–89 showed Coordination DNA compounds intercalation 86–89ability. showed IC Strong50 values DNA of 25 binding–91 is providedμM on by the triphenylphosphine MCF-7 cell line. The ligands highest with cytotoxicity a substituent was observed at the 4- positionfor compounds of the phenyl87 and 89 ring, which provided by triphenylphosphine ligands with a substituent at the 4-position of the phenyl ring capable capableshowed of forming the strongest hydrogen binding bonds. to DNA.Coordination compounds 86–89 showed IC50 values of 25–91 µ of formingμM on the hydrogenKomarnicka MCF-7 cell bonds. line et. The Coordination al. highest [102] cytotoxicity reported compounds was four observed 86 novel–89 forshowed compounds Cu(I) IC complexes50 87values and 89 of, basedwhich 25–91 onM on the MCF-7showed cellhydroxymethyldiphenylphosphine the line. strongest The highest binding cytotoxicityto DNA. 90 was–93, observedand four for compounds mixed sparfloxacin87 and 89 , which(HSf), showed i.e., the strongestKomarnickahydroxymethyldiphenylphosphine binding to etDNA. al. [102] reported coordination four compounds novel 94 Cu–(I)97 (Figure complexes 27). based on hydroxymethyldiphenylphosphine 90–93, and four mixed sparfloxacin (HSf), i.e., Komarnicka et al. [102] reported four novel Cu(I) complexes based on hydroxymethyldiphenylphosphine coordination compounds 94–97 (Figure 27). hydroxymethyldiphenylphosphine 90–93, and four mixed sparﬂoxacin (HSf), i.e., hydroxymethyldiphenylphosphine coordination compounds 94–97 (Figure 27). The cytotoxicity of the complexes synthesized and ligands (diimines, phosphines and phosphine oxides as potential decomposition products), starting compounds (CuNCS and CuI) was tested in vitro towards two cancer cell lines, i.e., mouse colon carcinoma (CT26) and human lung adenocarcinoma (A549). All tested complexes showed greater cytotoxicity than the corresponding ligands and copper iodide. In the case of the A549 line, dmp complexes compounds 90 and 91 ~25 mkM were twice as active as the bq complexes compounds 92 and 93 ~75 mkM. The activity against CT26 was slightly higher but was similar for both types of complexes. It was shown that the penetration of complexes into cells proceeds quickly and an increase in the incubation time of cells with drugs from 4 h to 24 h does not lead to an increase in cytotoxicity. Sparﬂoxacin moiety introduction into the structure of the phosphine ligand led to a two-fold increase in toxicity on both cell lines. Int. J. Mol. Sci. 2020, 21, 3965 22 of 37

FigureFigure 27. 27. ChemicalChemical structuresstructures ofof Cu(I)Cu(I) coordinationcoordination compounds compounds based based on on hydroxymethyldiphenylphosphinehydroxymethyldiphenylphosphine90–93, sparfloxacin90 (HSf)- hydroxymethyldiphenylphosphine93, sparfloxacin 94–97(HSf). 3. Drug-Basedhydroxymethyldiphenylphosphine Copper Coordination Compounds 94 – 97 The redox activity of copper cations along with their therapeutic efficacy, biogenicity, and ability to coordinate with various donor atoms opens up possibilities for synthesizing coordination compounds based on FDA-approved clinically used drugs with resultant target molecules having multiple biological effects. Copper coordination compounds based on ciprofloxacin [103], isoniazid [104], doxorubicin [105], indomethacin [106], and clioquinol [107] have been described. Coordination of copper cations with a drug molecule can change the pharmacodynamics of the drug and also enhance and complement therapeutic activity. A copper-containing gel based on indomethacin has shown increased activity as a local anti-inflammatory agent as compared with indomethacin [108]. A Cu(II) coordination compound based on indomethacin is capable of activating a copper-dependent opioid receptor and has a more effective analgesic effect than morphine with adjuvant arthritis after subcutaneous and oral administration [109]. Cu(II) coordination compound 98 with the anti-inflammatory drug Diclofenac was described by [110] (Figure 28). Diclofenac is one of the first anti-inflammatory NSAIDs used in medicine. Epidemiological studies have shown that chronic inflammation predisposes patients to the development of tumor diseases and that the long-term use of non-steroidal anti-inflammatory drugs reduced the riskInt. of J. developingMol. Sci. 2020, 2 malignant1, x FOR PEER neoplasms. REVIEW 24 of 38

FigureFigure 28. 28.Chemical Chemical structure structure of of coordination coordination compound compound 9898 [Cu(diclofenac)[Cu(diclofenac)22(H(H2O)2O)2] 2based] based on on the the NSAIDNSAID Diclofenac. Diclofenac.

Fenoprofen is a non-steroidal anti-inflammatory drug, a propionic acid derivative with anti-inflammatory, analgesic, and antipyretic effects. Gumilar et al. reported Cu(II) coordination compounds based on fenoprofen, caffeine, and DMF as coligands [111] (Figure 29). The coordination compounds Cu2(Fen)4(caf)2 (Fen-fenoprofenate anion; caf-caffeine) 95 and Cu2(Fen)2(DMF)2 96 have an analgesic effect, confirmed by studies in vitro and in vivo.

Figure 29. Chemical structure of coordination compounds 99 and 100 based on the NSAID drug fenoprofen.

The analgetic properties of coordination compounds 99 and 100 were also of interest. The visceral analgesic action on acetic acid-induced pain of both complexes was five to seven times more potent than fenoprofen salt at the same fenoprofen dose (20 mg/kg). Moreover, both complexes showed longer onset and shorter duration of writhing than fenoprofen salt (data not shown). This indicates that compounds 99 and 100 present a strong analgesic activity for visceral pain. Kovala-Demerzi et al. reported coordination compound 101 based on mefenamic acid ([Cu (Mef)2(H2O)]2) [112] (Figure 30). The cytotoxic activity of coordination compound 101 was tested in vitro on breast cancer cell line of invasive breast ductal carcinoma MCF-7, human bladder carcinoma cell line T24, lung cancer cell line of adenocarcinomic human alveolar basal epithelial cells A-549, and the mouse fibroblast cell line L-929. Coordination compound 101 exhibited greater cytotoxic activity as compared with the NSAID mefenamic acid (IC50 increased by two to six times). The IC50 values shown for 101 on the MCF-7 and L-929 tumor cell lines were compared with cisplatin (IC50 values were less than for cisplatin by 2.8 times for MCF-7 and 8.0 times for L-929). The coordination of mefenamic acid with copper (II) leads to the formation of an octahedral coordination compound, an increase in the cytotoxic activity of the initial ligand, and also new modes of cytotoxic action. Unfortunately, the low solubility of compound 101 prevents measuring anti-inflammatory effects in vivo.

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Int. J. Mol. Sci. 2020, 21, 3965 23 of 37

The cytotoxic effect of compound 98 was evaluated using the Uptiblue test on the following four human cell lines: human dermal fibroblast (HDF), human keratinocyte cell line HaCaT, and human colon adenocarcinoma cell lines SW620 and HT29. A comparison of the cytotoxic activity of the initial Cu(II) salt, Diclofenac, and coordination compound 98 ([Cu(diclofenac)2(H2O)2]) showed that the initial compounds have no cytotoxic activity (survival on tumor cell lines does not exceed 70% at a concentration 200 µM), while coordination compound 98 exhibits cytotoxicity on human colon adenocarcinoma cell lines SW620 and HT29 with the respective IC of 100 and 93 µM. Compound 98 Figure 28. Chemical structure of coordination compound 98 [Cu(diclofenac)50 2(H2O)2] based on the ([Cu(diclofenac)NSAID Diclofenac.2(H2O) 2]) is also the first Diclofenac-based coordination compound synthesized in a 100% aqueous medium. Fenoprofen is a non-steroidalnon-steroidal anti-inflammatory anti-inflammatory drug, a propionic acid derivative with anti-inflammatory,anti-inflammatory, analgesic, and and antipyretic antipyretic e effects.ffects. Gumilar et al. al. reported Cu(II) coordination coordination compounds based on fenoprofen,fenoprofen, cacaffeine,ffeine, andand DMFDMF asas coligandscoligands [[111111]] (Figure(Figure 2929).). The coordination compounds CuCu22(F(Fen)en)4(caf)(caf)22 (Fen(Fen-fenoprofenate-fenoprofenate anion; caf caf-ca-caffeine)ffeine) 9595 andand CuCu22(Fen)(Fen)22(DMF)2 96 have an analgesic eeffect,ffect, confirmedconfirmed byby studiesstudies inin vitrovitro andand inin vivo.vivo.

Figure 29. ChemicalChemical structure structure of of coordination coordination compounds compounds 99 and99 and100 based100 based on the on NSAID the NSAID drug drugfenoprofen. fenoprofen.

The analgetic analgetic properties properties of of coordination coordination compounds compounds99 and 99 100andwere 100 alsowere of also interest. of interest. The visceral The analgesicvisceral analgesic action on action acetic on acid-induced acetic acid-induced pain of bothpain complexesof both complexes was five was to sevenfive to times seven more times potent more thanpotent fenoprofen than fenoprofen salt at the salt same at the fenoprofen same fenoprofen dose (20 dose mg/ kg). (20 mg/kg). Moreover, Moreover, both complexes both complexes showed longershowed onset longer and onset shorter and duration shorter duration of writhing of writhing than fenoprofen than fenoprofen salt (data salt not (data shown). not This shown). indicates This thatindicates compounds that compounds99 and 100 99present and 100 a strongpresent analgesic a strong activityanalgesic for activity visceral for pain. visceral pain. Kovala-DemerziKovala-Demerzi et et al. al. reported reported coordination coordination compound compound 101 101basedbased on mefenamic on mefenamic acid ([Cuacid ([Cu (Mef) (H O)] ) [112] (Figure 30). The cytotoxic activity of coordination compound 101 was (Mef)2(H2O)]2 2)2 [1122] (Figure 30). The cytotoxic activity of coordination compound 101 was tested in testedvitro onin breast vitro on cancer breast cell cancer line of cell invasive line of breast invasive ductal breast carcinoma ductal carcinomaMCF-7, human MCF-7, bladder human carcinoma bladder carcinomacell line T24, cell lung line T24,cancer lung cell cancer line of cell adenocarcinomic line of adenocarcinomic human alveolar human alveolarbasal epithelial basal epithelial cells A- cells549, A-549,and the and mouse the mouse fibroblast fibroblast cell linecell line L-929. L-929. Coordination Coordination compound compound 101101 exhibitexhibiteded greater greater cytotoxic activity as compared with the NSAID mefenamic acid (IC increased by two to six times). The IC activity as compared with the NSAID mefenamic acid (IC5050 increased by two to six times). The IC5050 values shown for 101 on the MCF-7 and L-929 tumor cell lines were compared with cisplatin (IC values shown for 101 on the MCF-7 and L-929 tumor cell lines were compared with cisplatin (IC5050 values were less than for cisplatincisplatin by 2.8 timestimes forfor MCF-7MCF-7 andand 8.08.0 timestimes forfor L-929).L-929). The coordination of mefenamic acid with copper (II) leads to the formation of an octahedral coordination compound, an increase in the cytotoxic activity of the initial ligand, and also new modes of cyt cytotoxicotoxic action. Unfortunately, the low low solubility solubility of of compound compound 101101 preventsprevents measuring measuring anti anti-inflammatory-inflammatory effects effects in invivo vivo..

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Figure 30. Chemical structure of coordination compound 101 based on the NSAID mefenamic acid. FigureFigure 30. Chemical 30. Chemical structure structure of coordination of coordination compound compound 101 based 101 basedon the onNSAID the NSAID mefenamic mefenamic acid. acid. Xiangchao Shi et al. developed copper (II) coordination compounds 102 and 103 based XiangchaoXiangchao Shi et Shial. etdeveloped al. developed copper copper (II) coordination (II) coordination compounds compounds 102 and 102 103 and based 103 based on a on a on a phenanthroline derivative and aspirin [113] (Figure 31). Compound 102 effectively induces phenanthrolinephenanthroline derivative derivative and aspirinand aspirin [113] [113 (Figure] (Figure 31). Compound31). Compound 102 effectively102 effectively induces induces mitochondrial dysfunction and promotes early apoptosis in ovarian cancer cells. It also inhibits mitochondrialmitochondrial dysfunction dysfunction and promotes and promotes early apoptosisearly apoptosis in ovarian in ovarian cancer cancer cells. Itcells. also It inhibits also inhibits the the the expression of cyclooxygenase-2 (COX-2), a key enzyme involved in the inflammatory response. expressionexpression of cyclooxygenase of cyclooxygenase-2 (COX-2 -(COX2), a -key2), aenzyme key enzyme involved involved in the in inflammatory the inflammatory response. response. A A A similar coordination compound 103 CuL without an aspirin ligand has a similar effect on cell redox similarsimilar coordination coordination compound compound 103 CuL 103 wi CuLthout wi thoutan aspirin an aspirin ligand ligand has a hassimilar a similar effect effecton cell on redox cell redox homeostasis and cell cycle progression, but its cytotoxic activity is relatively low because its effect on homeostasishomeostasis and cell and cycle cell progression,cycle progression, but its but cytotoxic its cytotoxic activity activity is relatively is relatively low because low because its effect its effecton on mitochondrial function is mild and it cannot inhibit COX-2. mitochondrialmitochondrial function function is mild is andmild it and cannot it cannot inhibit inhibit COX- 2.COX -2.

Figure 31. Synthesis scheme of copper coordination compounds 102 and 103 with mixed FigureFigure 31. Synthesis 31. Synthesis scheme of copperscheme coordination of copper compounds coordination102 compoundsand 103 with mixed102 and phenanthroline 103 with mixed phenanthroline and the NSAID aspirin. and thephenanthroline NSAID aspirin. and the NSAID aspirin.

The IC50 value50 on ovarian cancer cell line of ovarian serous cystadenocarcinoma SKOV-3, The ICThe50 value IC onvalue ovarian on ovariancancer cell cancer line of cell ovarian line ofserous ovarian cystadenocarcinoma serous cystadenocarcinoma SKOV-3, cervical SKOV-3, cancercervicalcervical cell cancer line cancer cell HeLa, line cell and HeLa, line human HeLa, and human normal and human normal kidney normal kidney cell line kidney cell HK-2 line cell for HK line coordination-2 HKfor -coordination2 for coordination compound compound102 compoundis 10–30%102 is 10210 less– 30is %1 than0 –less30% thatthan less of that than compound of that compound of compound103, which103, which 103 does, which does not contain notdoes contain not an contain aspirin an aspirin an fragment, aspirin fragment, fragment, and and three three and to three fiveto five timesto times five less timesless than than less that thatthan of theof that the ligand of ligand the (Table ligand (Table 11 (Table). 11 Coordination). Coordination 11). Coordination compound compound compound102 102was was shown102 shown was to shown haveto have an to have antiproliferativean antiproliferativean antiproliferative effect effect through effect through DNA through degradation DNA DNA degradation and degradation mitochondrial and mitochondrialand dysfunction. mitochondrial dysfunction. The introduction dysfunction. The The ofintroduction theintroduction aspirin of moiety the of aspirin the not aspirin only moiety increases moiety not only not the increases only antitumor increases the effi antitumor cacy the antitumor of the efficacy drug efficacy but of the also of drug the reduces drugbut also the but also inflammatoryreducesreduces the inflammatory the threat. inflammatory threat. threat.

Table Table11. MTT 11. data MTT of data coordination of coordination compounds compounds 102 and 102 103 and, and 103 the, and phenanthroline the phenanthroline ligand ligand after 72 after 72 h of incubation.h of incubation. [113]. [113].

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Table 11. MTT data of coordination compounds 102 and 103, and the phenanthroline ligand after 72 h of incubation [113]. Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 26 of 38

IC50, µM S.D. ± IC50, µM ± S.D. Compound SKOV-3 HeLa HK-2 Compound SKOV-3 HeLa HK-2 102 (with aspirin) 1.1 0.6 1.5 0.5 4.4 0.5 102 (with aspirin) 1.1 ± ±0.6 1.5 ± 0.5 4.4± ± 0.5 103 (without aspirin) 1.5 0.4 1.8 0.5 4.6 0.8 103 (without aspirin) 1.5 ± ±0.4 1.8 ± 0.5 4.6± ± 0.8 L102 5.4 1.2 6.8 1.2 12.3 1.6 L102 5.4 ± ±1.2 6.8 ± 1.2 12.3± ± 1.6

TheThe COX-2 COX-2 levellevel inin lipopolysaccharide-stimulated lipopolysaccharide-stimulated RAW macrophages macrophages was investigated on a flowflow cytometer cytometer after after treatment treatment with coordination with coordination compounds compounds102 and 103 ,102 and theand anti-inflammatory 103, and the potentialanti-inflammatory of coordination potential compound of coordination102 was compound confirmed. 102 was confirmed. NitroimidazoleNitroimidazole derivativesderivatives areare widely widely used used drugs drugs with with multiple multiple pharmacological pharmacological eff ectseffects such such as antifungalas antifungal [114 [114],], antibacterial antibacterial [115 [115],], and and cytotoxic cytotoxic [ 116[116].]. They They also also represent represent aa classclass ofof hypoxiahypoxia indicatorsindicators that that have have been been investigated investigated for for hypoxia-selective hypoxia-selective cytotoxicity cytotoxicity andand radiosensitizationradiosensitization of of hypoxichypoxic cellscells [[117117].]. The eeffectivenessffectiveness of these molecules depends on the generationgeneration of aa nitroradicalnitroradical anionanion by intracellular reduction, which makes 5-nitroimidazoles5-nitroimidazoles suitablesuitable forfor penetration into cells byby passivepassive didiffusion,ffusion, creatingcreating aa favorablefavorable concentrationconcentration gradient.gradient. Once inside the cell,cell, thethe nitroradicalnitroradical anionanion interactsinteracts withwith DNADNA andand destroysdestroys thethe doubledouble helix.helix. Cu(II) coordination compounds 104 andand 105105 andand Cu(I)Cu(I) coordinationcoordination compoundscompounds 106106 andand 107107 withwith ligandsligands basedbased onon 5-nitroimidazole5-nitroimidazole werewere synthesizedsynthesized inin [[118118]] (Figure(Figure 3232).).

Figure 32. L104 and L105 ligand synthesis scheme and the chemical structures of coordination compoundsFigure 32. L104104– 107andbased L105 on ligand the antifungal synthesis drug scheme Metronidazole. and the chemical structures of coordination compounds 104–107 based on the antifungal drug Metronidazole. MTT tests on oxaliplatin-resistant and non-resistant human colon cancer cell line of supraclavicular lymphМТТ node tests metastasis on oxaliplatin LoVo-OXP-resistant and LoVo and of compoundsnon-resistant104 human–107 showed colon a cancer signiﬁcant cell increase line of ofsupraclavicular antiproliferative lymph activity node of metastasisCu(II) compounds LoVo-OXP104 and 105LoVoand of Cu(I) compounds compounds 104–106107 andshowed107 as a comparedsignificant with increase ligands of in antiproliferativ monolayer culturese activity of various of Cu(II) lines compounds of human tumor 104 and cells. 105 Water-soluble and Cu(I) Cu(I)compounds coordination 106 and compound 107 as compared106 showed with higher ligands cytotoxicity in monolayer as compared cultures of with various Cu(II) lines coordination of human compoundtumor cells.104 Water(Table- 12soluble). The data Cu(I) obtained coordination indicates compound that water-soluble 106 showed Cu(I) coordination higher cytotoxicity compounds as havecompared a better with cellular Cu(II accumulation) coordination than compound the Cu(II) 104 analogues. (Table 12 This). The hypothesis data obtained was tested indicate usingsAAS, that andwater intracellular-soluble Cu(I) accumulation coordination of coordination compounds compound have a better106 (R cellular= H) was accumulation shown to be than better the than Cu(II) the Cu(II)analogues. analogue. This hypothesis was tested using AAS, and intracellular accumulation of coordination compound 106 (R = H) was shown to be better than the Cu(II) analogue.

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Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 27 of 38 Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 27 of 38 Int. J.Int. Mol. J. Mol. Sci.Table 20Sci.20 12.2020, 21,MTT ,x 21 FOR, x dataFOR PEER PEER of REVIEW coordination REVIEW compounds 104–107 after 72 h of incubation [118]. 27 of 27 38 of 38 Table 12. MTT data of coordination compounds 104–107 after 72 h of incubation. [118]. Table 12. MTT data of coordination compounds 104–107 after 72 h of incubation. [118]. TableTable 12. MTT12. MTT data data of coordination of coordinationIC compounds50, compoundsµM S.D. 104 –104–107107 after after 72 h 72 of h incubation. of incubation. [118] [118].. ± IC50, µM ± S.D. IC50, µM ± S.D. Compound CompoundLoVo LoVo LoVo-OXP IC50 LoVo-OXP, µM ± S.D. Compound LoVo IC50, µ LoVo-OXPM ± S.D. CompoundCompound104 Cu(II) 104 Cu(II)LoVo5.9LoVo 5.9 ± 0.6 0.6 5.1 0.5 LoVo LoVo-OXP- 5.1OXP ± 0.5 104 Cu(II) 5.9 ± 0.6± ± 5.1 ± 0.5 104 104Cu(II)105 Cu(II)Cu(II) 105 Cu(II)5.9 ±5.94.3 0 4.3.6 ±± 0.60.50.5 4.6 1.0 5.1 ± 5.1 4.60.5 ±± 0.51.0 105 Cu(II) 4.3 ± 0.5± ± 4.6 ± 1.0 105 105Cu(II)106 Cu(II) Cu(I) 106 Cu(I)4.3 ±4.32.1 0 2.1.5 ±± 0.51.11.1 1.9 0.9 4.6 ± 4.6 1.91.0 ±± 1.00.9 106 Cu(I) 2.1 ± 1.1± ± 1.9 ± 0.9 106 106Cu(I)107 Cu(I)Cu(I) 107 Cu(I)2.1 ±2.14.9 1 4.9.1 ±± 1.11.01.0 4.6 0.8 1.9 ± 1.9 4.60.9 ±± 0.90.8 107 Cu(I) 4.9 ± 1.0± ± 4.6 ± 0.8 107 107Cu(I) Cu(I) 4.9 ±4.9 1.0 ± 1.0 4.6 ± 4.6 0.8 ± 0.8 Nitroimidazole derivatives are a promising platform for developing biologically active NitroimidazoleNitroimidazole derivatives derivatives are are a promisinga promising platform platform for for developing developing biologically biologically activeactive coordinationNitroimidazoleNitroimidazole compounds, derivatives derivatives and arewater-soluble are a promisinga promising Cu(I) platform co platformordination for for developingcompounds developing capable biologically biologically of high active cellularactive coordinationcoordination compounds, compounds, and water-solubleand water-soluble Cu(I) Cu(I) coordination coordination compounds compounds capable capable of of high high cellular cellular coordinationcoordinationaccumulation compounds, compounds, are a promising and and water water-solublealternative-soluble Cu(I)to theCu(I) coordination classical coordination Cu(II) compounds coordinaticompounds capableon capablecompounds, of high of high cellular showing cellular a accumulationaccumulation are a promisingare a promising alternative alternative to the to classicalthe classical Cu(II) Cu(II) coordination coordination compounds, compounds, showing showing a a accumulationsignificant improvement are a promising of the alternative cytotoxic potency.to the classical Cu(II) coordination compounds, showing a accumulationsignificant areimprovement a promising of alternative the cytotoxic to thepotency. classical Cu(II) coordination compounds, showing a signiﬁcantsignificant improvementThe 8-hydroxyquinoline improvement of the of cytotoxic the (8-HQ)cytotoxic potency. derivatives potency. comprise a class of antifungal or antimicrobial significantThe improvement 8-hydroxyquinoline of the cytotoxic (8-HQ) potency. derivatives comprise a class of antifungal or antimicrobial Theagents. 8-hydroxyquinolineThe Developing 8-hydroxyquinoline Cu(II) (8-HQ) coordination (8-HQ) derivatives derivatives compound comprise comprises with aligands classa class based of of antifungal antifungalon oxyquinolones or or antimicrobial antimicrobial opens up agents.The 8 -Developinghydroxyquinoline Cu(II) coordination (8-HQ) derivatives compound comprises with aligands class ofbased antifungal on oxyquinolones or antimicrobial opens up agents.agents.opportunities Developing Developing Cu(II)for designingCu(II) coordination coordination agents compounds with compound multiple withs with biological ligands ligands based activity. based on on oxyquinolonesTardito oxyquinolones et al. opensreported opens up up a agents.opportunities Developing for Cu(II) designing coordination agents compoundswith multiple with biological ligands based activity. on oxyq Tarditouinolones et al. opens reported up a opportunitiesopportunitieshalogenated for designing clioquinol for designing agents (CQ), with agentswhich multiple iswith an biologicalanalogmultipleue ofbiological activity. 8-HQ, Tarditoand activity. copper et al.Tardito coordination reported et aal. halogenated compoundsreported a opportunitieshalogenated for clioquinol designing (CQ), agents which with is an multiple analog ue biological of 8-HQ, activity. and copper Tardito coordination et al. reported compounds a clioquinolhalogenated108– (CQ),115 based which clioquinol on is it an [119] analogue (CQ), (Figure which of 33). 8-HQ, is anMTT andanalog data copperue for of coordination 8-HQ, and copper compounds coordination 108108––115115 compounds onbased cervical on halogenated108–115 based clioquinol on it (CQ),[119] which(Figure is 33). an ManalogueTT data of for 8 -coordinationHQ, and copper compounds coordination 108– 115compounds on cervical it [119108]cancer (Figure–115 cell based 33 line). MTT onHeLa itdata [119] and for human(Figure coordination prostate 33). MTT cancer compounds data cellfor linecoordination108 PC3–115 areon given compounds cervical in Table cancer 108 13.– cell115 lineon cervical HeLa 108–cancer115 based cell line on HeLait [119 and] (Figure human 33 prostate). MTT datacancer for cell coordination line PC3 are compounds given in Table 108 –13.115 on cervical andcancer humancancer cell prostate cellline line HeLa HeLa cancer and and human cell human line prostate PC3 prostate are cancer given cancer cell in cellTableline line PC3 13 PC3. are aregiven given in Table in Table 13. 13.

Figure 33. Estimated сhemical structures of coordination compounds 108–115. Figure 33. Estimated сhemical structures of coordination compounds 108–115. FigureFigureFigure 33. 33.Estimated Estimated33. Estimated chemical сhemical сhemical structuresstructures structures of coordination of coordination compounds compounds compounds 108108 –108115–115–.115 . . Table 13. The structures of coordination compounds 108–115 based on 8-HQ and their cytotoxic Table 13. The structures of coordination compounds 108–115 based on 8-HQ and their cytotoxic Table 13.TableactivityThe 13. structures data The [119]. structures of coordination of coordination compounds compounds108–115 108based–115 on based 8-HQ on and 8-HQ their and cytotoxic their activitycytotoxic Tableactivity 13. The data structures [119]. of coordination compounds 108–115 based on 8-HQ and their cytotoxic dataactivity [119activity]. data data [119] [119].. IC50, µM ± S.D. IC50, µM ± S.D. Compound Chemical Structure LogP (Ligand) IC 50, µICM50 ,± ICµM HeLaS.D.50 ,± µ S.D.M S.D. PC 3 Compound Chemical Structure LogP (Ligand) HeLa ± PC3 CompoundCompoundCompound Chemical Chemical Structure Structure LogP LogP (Ligand) LogP (Ligand)(Ligand) HeLa HeLa HeLa PC PC3 3 PC3

Cu(5-SO3-8-HQ) Cu(5Cu(5-SO-SO3-83--8-HQ)HQ) −0.21 n.d. n.d. Cu(5-SOCu(5-SO-8-HQ)1083-8-HQ) −0.21− 0.21 n.d. n.d. n.d. n.d. 3101088 −0.21 0.21n.d. n.d. n.d.n.d. 108 108 −

Cu(5-SO3-7-I-8-HQ) Cu(5-SO3-7-I-8-HQ) 0.70 n.d. n.d. Cu(5-SOCu(5-SO3-7-I-8-HQ)1093-7-I-8-HQ) 0.70 n.d. n.d. Cu(5-SO3109-7-I-8-HQ) 0.70 0.70n.d. n.d. n.d.n.d. 109 109 0.70 n.d. n.d. 109

Cu(8-HQ) 1.84 1.9 1.3 Cu(8-HQ)Cu(8-HQ)110 Cu(8-HQ) 1.841.84 1.9 1.3 1.3 110Cu(8-HQ)110 1.84 1.9 1.3 110 1.84 1.9 1.3 110

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Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW Table 13. Cont. 28 of 38 Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 28 of 38 Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 28 of 38 Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW IC50, µM S.D. 28 of 38 Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW ± 28 of 38 Compound Chemical Structure LogP (Ligand) HeLa PC3

Cu(5-Cl-8-HQ) Cu(5-Cl-8-HQ) 2.58 3.1 2.3 Cu(5-Cl-8-HQ)111 2.58 3.1 2.3 Cu(5-Cl-8-HQ)111 2.58 3.1 2.3 Cu(5-Cl-8-HQ)Cu(5-Cl-8-HQ)111 2.58 3.1 2.3 111 2.58 2.583.1 3.1 2.3 2.3 111 111

Cu(5,7-Me-8-HQ) Cu(5,7-Me-8-HQ) 2.66 2.7 1.9 Cu(5,7-Me-8-HQ)112 2.66 2.7 1.9 Cu(5,7-Me-8-HQ)Cu(5,7-Me-8-HQ)112 2.66 2.7 1.9 Cu(5,7-Me-8-HQ)112 2.66 2.662.7 1.9 1.9 112 112 2.66 2.7 1.9 112

Cu(5,7-Cl-8-HQ) Cu(5,7-Cl-8-HQ) 3.22 5.3 4.7 Cu(5,7-Cl-8-HQ)Cu(5,7-Cl-8-HQ)113 3.22 5.3 4.7 Cu(5,7-Cl-8-HQ)113 3.223.22 5.3 4.7 4.7 Cu(5,7-Cl-8-HQ)113 113 3.22 5.3 4.7 113 3.22 5.3 4.7 113

Cu(CQ) Cu(CQ)Cu(CQ) 3.50 8.9 9.0 Cu(CQ)114 3.50 3.508.9 8.9 9.0 9.0 114Cu(CQ)114 3.50 8.9 9.0 Cu(CQ)114 3.50 8.9 9.0 114 3.50 8.9 9.0 114

Cu(5,7-I-8-HQ)Cu(5,7-I-8-HQ) Cu(5,7-I-8-HQ) 3.75 3.7512.8 12.8 16.216.2 Cu(5,7-I-8-HQ)115 115 3.75 12.8 16.2 Cu(5,7-I-8-HQ)115 3.75 12.8 16.2 Cu(5,7-I-8-HQ)115 3.75 12.8 16.2 115 3.75 12.8 16.2 115

A structure–activityA structure–activity relationship relationship (SAR) (SAR) study study was conducted. was conducted. Cellular Cellular accumulation accumulation of the drugof the A structure–activity relationship (SAR) study was conducted. Cellular accumulation of the drug Acan structure–activity occur via active transport,relationship as suggested (SAR) study by the was hCTR1 conducted. copper Cellulartransporter accumulation for cisplatin of [120], the can occurdrug viaAcan structure–activity activeoccur via transport, active transport,relationship as suggested as suggested(SAR) by thestudy by hCTR1 thewas hCTR1 conducted. copper copper transporter Cellulartransporter accumulation for for cisplatin cisplatin [of120 [120], the], ordrug by canpassive occur diffusion via active through transport, the asplasma suggested membrane, by the hCTR1 in which copper case transporterthe drug should for cisplatin be endowed [120], or by passivedrugor by can passive di occurﬀusion diffusion via throughactive through transport, the plasma the as plasma suggested membrane, membrane, by the in whichhCTR1 in which casecopper case the transporterthe drug drug should should for cisplatin be be endowed endowed [120], withor by appropriate passive diffusion lipophilicity through to thepass plasma through membrane, the cell membrane in which andcase reachthe drug a sufficient should beintracellular endowed with appropriateorwith by appropriate passive lipophilicity diffusion lipophilicity through to pass to thepass through plasma through the membrane, cellthe cell membrane membrane in which and caseand reach reachthe adrug sua ﬃsufficient shouldcient intracellular beintracellular endowed concentration.with appropriate An lipophilicity excessively tolipophilic pass through compound the cell accumulates membrane and in thereach membrane, a sufficient while intracellular greater concentration.withconcentration. appropriate An excessivelyAn lipophilicity excessively lipophilic to lipophilic pass compoundthrough compound the accumulatescell accumulates membrane in and in the thereach membrane, membrane, a sufficient while while intracellular greater greater hydrophilicityconcentration. preventsAn excessively interaction lipophilic with thecompound lipid bilayer. accumulates Of the studiedin the membrane,derivatives, whilecoordination greater hydrophilicityconcentration.hydrophilicity prevents Anprevents excessively interaction interaction lipophilic with with the lipidthecompound lipid bilayer. bi layer.accumulates Of theOf the studied studiedin the derivatives, membrane,derivatives, coordination while coordination greater compoundshydrophilicity based prevents on interactionthe most with hydrophilic the lipid biligandslayer. Of L108 the studiedand L109derivatives, (5-SO3-8-HQ coordination and compoundshydrophilicitycompounds based onbased prevents the moston interaction hydrophilicthe most with hydrophilic ligands the lipidL108 biligandsandlayer.L109 Of L108 the(5-SO3-8-HQ studied and L109derivatives, and (5-SO3-8-HQ 5-SO3-7-I-8-HQ) coordination and 5-SO3-7-I-8-HQ)compounds based do noton exhibitthe most cytotoxic hydrophilic activity, whileligands ligands L108 ofand the mostL109 active(5-SO3-8-HQ coordination and do notcompounds5-SO3-7-I-8-HQ) exhibit cytotoxic based do activity, noton exhibitthe while most cytotoxic ligands hydrophilic of activity, the most ligandswhile active ligands coordinationL108 ofand the L109most compounds active(5-SO3-8-HQ coordination are ligands and compounds5-SO3-7-I-8-HQ) are ligandsdo not exhibitwith intermediate cytotoxic activity, lipophilicity, while ligandsnamely, of ligands the most L110 active to L112coordination (8-HQ, with intermediate5-SO3-7-I-8-HQ)compounds lipophilicity,are doligands not exhibitwith namely, intermediate cytotoxic ligands activity,L110 lipophilicity,to L112while(8-HQ, ligandsnamely, 5,7-Me-8-HQ, of ligandsthe most L110 andactive to 5-Cl-8-HQ). coordinationL112 (8-HQ, 5,7-Me-8-HQ,compounds are and ligands 5-Cl-8-HQ). with intermediate lipophilicity, namely, ligands L110 to L112 (8-HQ, Coordinationcompounds5,7-Me-8-HQ, compoundsare and ligands 5-Cl-8-HQ). 110withand intermediate114 (Cu-CQ andlipophilicity, Cu-8-HQ) namely, were shown ligands to inhibit L110 theto proteasomeL112 (8-HQ, 5,7-Me-8-HQ,Coordination and 5-Cl-8-HQ).compounds 110 and 114 (Cu-CQ and Cu-8-HQ) were shown to inhibit the activity.5,7-Me-8-HQ, MTTCoordination tests showed and 5-Cl-8-HQ).compounds that coordination 110 and compound 114 (Cu-CQ114 (Cu-CQ)and Cu-8-HQ) is at least were 10 timesshown more to inhibit cytotoxic the proteasomeCoordination activity. compounds МТТ tests showed 110 and that 114 coordination (Cu-CQ and compound Cu-8-HQ) 114 were (Cu-CQ) shown is at to least inhibit 10 times the than ligandproteasomeCoordinationL114 activity.administered compounds МТТ tests separately showed110 and whenthat 114 coordination tested(Cu-CQ for and 48compound hCu-8-HQ) on HeLa 114 were cell(Cu-CQ) linesshown is (IC at to least =inhibit8.9 10 timesµ Mthe moreproteasome cytotoxic activity. than МТТligand tests L114 showed administered that coordination separately compound when tested 114 for(Cu-CQ) 48 h on is atHeLa50 least cell 10 timeslines for coordination(ICproteasomemore50 = cytotoxic8.9 μM compoundactivity. for than coordination МТТligand114 tests L114and compoundshowed 93administeredµM that for114 coordinationthe and separately ligand) 93 μM forcompound and when the PC3 ligand) tested cell114 and for lines(Cu-CQ) 48PC3 (ICh cellon is at =HeLalines least9.0 (ICµ cell10M50 times lines= for 9.0 (ICmore50 = cytotoxic 8.9 μM for than coordination ligand L114 compound administered 114 and separately 93 μM for when the ligand) tested andfor 48PC3 h cellon50 HeLalines (ICcell50 lines= 9.0 μmore(ICM50 for = cytotoxic 8.9 coordination μM for than coordination compoundligand L114 compound 114 µadministered and >100 114 μ andM separately for 93 theμM ligand).for when the ligand) tested andfor 48 PC3 h oncell HeLa lines (ICcell50 lines= 9.0 coordinationμ(ICM50 for = 8.9 compoundcoordination μM for coordination114 compoundand >100 compound 114M and for >100 the114 ligand).μandM for 93 theμM ligand).for the ligand) and PC3 cell lines (IC50 = 9.0 (ICμM50 for = 8.9 coordination μM for coordination compound compound 114 and >100 114 μandM for93 μtheM ligand).for the ligand) and PC3 cell lines (IC50 = 9.0 μM forShah coordination et al. demonstrated compound significant 114 and >100 antibacterial μM for the activity ligand). of compound 114 (Cu-CQ) and its μM forShah coordination et al. demonstrated compound significant 114 and >100 antibacterial μM for the activity ligand). of compound 114 (Cu-CQ) and its strongShah gain et by al. copper demonstrated ions [121]. significant The antibacterial antibacterial activity activity of compound of compound 110 was114 confirmed(Cu-CQ) and using its Mtb-infectedstrongShah gain et by al.macrophages copper demonstrated ions in [121]. the significant presence The antibacterial antibacterialor absence activity of activity7.5 μofM compound CuSOof compound4. 110 was114 (Cu-CQ)confirmed and using its Mtb-infectedstrong gain by macrophages copper ions in [121]. the presenceThe antibacterial or absence activity of 7.5 μofM compound CuSO4. 110 was confirmed using strongMtb-infected gain by macrophages copper ions in[121]. the presenceThe antibacterial or absence activity of 7.5 μofM compound CuSO4. 110 was confirmed using Mtb-infected macrophages in the presence or absence of 7.5 μM CuSO4. Mtb-infected macrophages in the presence or absence of 7.5 μM CuSO4.

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Shah et al. demonstrated significant antibacterial activity of compound 114 (Cu-CQ) and its strong gain by copper ions [121]. The antibacterial activity of compound 110 was confirmed using Mtb-infected macrophages in the presence or absence of 7.5 µM CuSO4. Copper coordination compounds based on 8-hydroxyquinolines exhibit both cytotoxic and antibacterial properties. The molecular design allows varying their lipophilicity and cellular accumulation. The results obtained together with the promising data obtained for elesclomol L13 [38] Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 29 of 38 Int.confirm J. Mol. Sci. the 2020 promising, 21, x FOR PEER use REVIEW of copper coordination compounds in treating bacterial pneumonia29 of 38 caused by Mtb. The therapy certainly owes its success to the redox activity of copper cations. Both therapeutic causedcaused by Mtb. by Mtb.The therapyThe therapy certainly certainly owes owes its success its success to the to redox the redox activity activity of copper of copper cations. cations. Both Both regimens involving separate uses of copper and a ligand and in situ drug formation open up great therapeutictherapeutic regimens regimens involving involving separate separate uses uses of copper of copper and aand ligand a ligand and inand situ in drugsitu drug formation formation opportunities for developing formulations, including those that are selective for healthy tissues. openopen up great up great opportunities opportunities for developing for developing formulations, formulations, including including those those that thatare selective are selective for for Silva et al. reported a nanostructured lipid system for low-soluble isoniazid-based healthyhealthy tissues tissues. . copperSilva complexes et al. reported compounds a nanostructured116 ([CuCl (INH) lipid sy]stemH O, for117 low[Cu(NCS)-soluble isoniazid(INH) ]-5HbasedO, copper and 118 Silva et al. reported a nanostructured lipid2 system2 · for2 low-soluble isoniazid2 -based2 · copper2 [Cu(NCO)complexes(INH) compounds] 4H O with116 antimycobacterial ([CuCl2(INH)2]*H activity2O, 117 (Figure [Cu(NCS) 34). The2(INH) nano-sized2]*5H2O, drug and delivery118 complexes compounds2 2 · 2 116 ([CuCl2(INH)2]*H2O, 117 [Cu(NCS)2(INH)2]*5H2O, and 118 [Cu(NCO)2(INH)2]*4H2O with antimycobacterial activity (Figure 34). The nano-sized drug delivery [Cu(NCO)systems2(INH) increased2]*4H their2O with antimycobacterial antimycobacterial activity, activity decreased (Figure 34 cytotoxicity). The nano-sized against drug the delivery Vero cell line, systemsandsystems consequently increased increased their improved theirantimycobacterial antimycobacterial the selectivity activity, indexactivity, decreased [122 decreased]. cytotoxicity cytotoxicity against against the Vero the Vero cell line,cell line, and consequentlyand consequently improved improved the selectivity the selectivity index index [122]. [122 ].

116 116 117 117 118 118

FigureFigureFigure 34. 34.Chemical 34.Chemical Chemical structures structures structures of coordination ofof coordinationcoordination compounds compounds 116– 118116116 –based–118118 based basedon the on onantituberculosis the the antituberculosis antituberculosis drugdrug drugisoniazid. isoniazid. isoniazid.

A recentAA recent recent study study study on onthe on the cyto the cyto-genotoxicity -cytogenotoxicity-genotoxicity of Cu(II) of of Cu(II) Cu(II) compounds compounds compounds 116–116 118116– –118with118with withINH INH INHwas wascwasonducted conductedconducted by byFregonezi Fregoneziby Fregonezi et et al. al. andet and al. also alsoand concluding also concluding concluding that that the thatthe compounds compounds the compounds show show no show cytotoxicity no cytotoxicity no cytotoxicity in therapeutic in therapeutic in therapeutic doses [123 ]. dosesdoses [123]. [ 123]. 4. Natural Product-Based Copper Coordination Compounds 4. Natural4. Natural Product Product-Based-Based Copper Copper Coordination Coordination Compounds Compounds Natural products (NPs) have attracted lots of attention as biological active ligands for copper coordinationNaturalNatural products compounds, products (NPs) (NPs) hav duee hav toattracted thee attracted fact lots that oflots nearly attention of attention 60% as of biological clinicallyas biological active approved active ligands ligands anticancer for copper for drugscopper are coordination compounds, due to the fact that nearly 60% of clinically approved anticancer drugs are coordinationassociated compounds with NPs. Advances, due to the of fact metal that complexesnearly 60% withof clinically natural approved product-like anticancer compounds drugs haveare been associated with NPs. Advances of metal complexes with natural product-like compounds have been associatedrecently with summarized NPs. Advances by Heras of metal et al. complexes [124], herein with a few natural examples product of-like those compounds design. have been recently summarized by Heras et al.[124], herein a few examples of those design. recentlyFei summarized et al. [125 by] Heras reported et al. two[124], copper herein a (II) few complexes, examples of compoundsthose design. 119 and 120, based on Fei et al. [125] reported two copper (II) complexes, compounds 119 and 120, based on dehydroabieticFei et al. [125] acid reported (DHA), thetwo main copper component (II) complexes of traditional, compounds Chinese 119 medicine and 120 rosin, based (Figure on 35). dehydroabieticdehydroabietic acid acid(DHA), (DHA), the main the main component component of tra ofditional traditional Chinese Chinese medicine medicine rosin rosin (Figure (Figure 35). 35).

FigureFigure 35. Synthesis 35. Synthesis scheme scheme and chemical and chemical structure structure of coordination of coordination compounds compounds 119 and 119 120and based 120 based Figure 35. Synthesis scheme and chemical structure of coordination compounds 119 and 120 based on on theon dehydroabietic the dehydroabietic acid. acid. the dehydroabietic acid. The Theability ability of compounds of compounds 119 and119 and120 to120 interact to interact with with calf thymuscalf thymus DNA DNA (CT (CT DNA) DNA) via via intercalation,intercalation, as well as well as albumin as albumin binding binding ability ability has been has been proven proven by various by various physicochemical physicochemical methods.methods. MTT MTT assay assay illustrates illustrates that thethat selective the selective cytotoxic cytotoxic activity activity of compound of compound 119 was 119 betterwas better than than that that of ligand of ligand L119 L119, compound, compound 120, 120 cisplatin, cisplatin, and, oxaliplatin.and oxaliplatin. The exposureThe exposure of compound of compound 119 to119 to MCFMCF-7 cells-7 cells resulted resulted in cell in cycle cell cycle arrest arrest in G1 in phase, G1 phase, apoptosis, apoptosis, mitochondrial mitochondrial dysfunction dysfunction, and, and elevatedelevated ROS ROSlevel, level, also compoundalso compound 119 proved119 proved to induce to induce apoptosis apoptosis through through intrinsic intrinsic and extrinsicand extrinsic pathways,pathways, autophagy autophagy, and, DNAand DNA damage damage in MCF in MCF-7 cells.-7 cells. Compound Compound 119 is119 assumed is assumed to have to havethe the

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The ability of compounds 119 and 120 to interact with calf thymus DNA (CT DNA) via intercalation, as well as albumin binding ability has been proven by various physicochemical methods. MTT assay illustrates that the selective cytotoxic activity of compound 119 was better than that of ligand L119, compound 120, cisplatin, and oxaliplatin. The exposure of compound 119 to MCF-7 cells resulted in cell cycle arrest in G1 phase, apoptosis, mitochondrial dysfunction, and elevated ROS level, also compound Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 30 of 38 119 proved to induce apoptosis through intrinsic and extrinsic pathways, autophagy, and DNA damage inability MCF-7 to resist cells. metastasis Compound and119 angiogenesisis assumed due to have to downregulation the ability to resist of VEGFR2, metastasis MMP and-2 angiogenesis, and MMP-9 dueexpression to downregulation levels in tumor of VEGFR2, cells. MMP-2, and MMP-9 expression levels in tumor cells. Chen et et al. al. [7] [ 7reported] reported a coordination a coordination compound compound of copper of copper 121 based121 onbased Hinokitiol on Hinokitiol,, a natural amonoterpenoid natural monoterpenoid (Figure 36 (Figure). A coordination 36). A coordination compound compound was formed was in formedsitu while in situusing while equimolar using equimolarmixtures of mixtures L121 and of L121 CuSOand4,CuSO as was4, as waspreviously previously described described for for Disulfiram, Disulﬁram, e elesclomollesclomol and and thiosemicarbazonethiosemicarbazone ligandligand L68L68..

Figure 36. Chemical structure of Hinokitiol L121..

Ligand L121L121 inin thethe presencepresence of of CuSO CuSO4 4induces induces striking striking accumulation accumulation of of ubiquitinated ubiquitinated proteins proteins in A549in A549 and and K562 K562 cells, cells, which which means means that it is that capable it is of capable inhibiting of inhibit the activitying the of the activity 19S proteasomal of the 19S DUBsproteasomal much DUBsmore e muchﬀectively more than effectively it does thanthe chymotrypsin-like it does the chymotrypsin activity- oflike the activity 20S proteasome. of the 20S Coordinationproteasome. compound Coordination121 eﬀ compoundectively induces 121 caspase-independenteffectively induces and caspase paraptosis-like-independent cell death and in A549paraptosis and K562-like cellcells, death and thein A549resulted and cell K56 death2 cells, has and been the proven resulted to dependcell death on ATF4-assosiatedhas been provenER to stressdepend but on not ATF4 ROS-assosiated generation. ER stress but not ROS generation.

5. Conclusions 5. Conclusions Copper-containing coordination compounds are a promising class of drugs with multiple Copper-containing coordination compounds are a promising class of drugs with multiple therapeutic effects from antitumor to anti-inflammatory activity. This review summarizes the successful therapeutic effects from antitumor to anti-inflammatory activity. This review summarizes the use of copper coordination compounds as antitumor, antimalarial, antituberculosis, antifungal, successful use of copper coordination compounds as antitumor, antimalarial, antituberculosis, and anti-inflammatory drugs. antifungal, and anti-inflammatory drugs. An antitumor activity of copper coordination compounds was repeatedly proven in vivo. An antitumor activity of copper coordination compounds was repeatedly proven in vivo. 60 Imidazolin-4-oneImidazolin-4-one based coordination compound 60 showed 73.5%73.5% of breast adenocarcinoma Ca-755Ca-755 growth inhibitioninhibition inin sevenseven days days of of treatment treatment with with a a 24 24 mg mg/kg/kg dose; dose animal’s; animal’s body body weight weight loss loss did did not exceed 10%. Thiosemicarbazone-based ligand L68 + CuCl2 showed 42% of monocytic leukemia HL60 not exceed 10%. Thiosemicarbazone-based ligand L68 + CuCl2 showed 42% of monocytic leukemia growthHL60 growth inhibition inhibition in five daysin five of treatmentdays of treatment with a 3 mgwith/kg a dose,3 mg/kg with dose, less than with 20% less body than weight 20% body loss. Theweight DSF loss/Cu. nanoparticleThe DSF/Cu deliverynanoparticle system delivery showed system 71.4% showed of breast 71 cancer.4% of 4T1 breast growth cancer inhibition 4T1 growth after twoinhibition weeks after of treatment two weeks with of treatment a 3.75 mg /withkg of a DSF3.75 dose,mg/kg and of DSF there dose, were and no there significant were no body-weight significant changesbody-weight observed. changes Thiosemicarbazone-based observed. Thiosemicarbazone coordination-based compoundcoordination69 compoundshowed 74% 69 of showed Lewis lung74% carcinomaof Lewis lung (LLC) carcinoma tumor growth (LLC)inhibition tumor growth in seven inhibition days of in treatment seven day withs of a trea 6 mgtment/kg dose, with weight a 6 mg/kg loss bodydose, weightweight lossloss didbody not weight exceed loss 10%. did not exceed 10%. Regarding thethe mechanismmechanism of of antitumor antitumor activity, activity, the the vast vast majority majority of coordinationof coordination compounds compounds act throughact through the ROS the formation ROS formation (1, 2, L13 (1, + 2,Cu L13, 42 +, 43Cu,, 45 – 42,47 ,43, Disulfiram-based 45–47, Disulfiram coordination-based coordination compound 64compound, L68 + Cu 64,, 119 L68), glutation+ Cu, 119), depletion glutation ( 45depletion–47, L68 (+45Cu–47, ,69 L68–74 +), Cu, proteasome 69–74), proteasome inhibition ( 7inhibition, 8, 110, 114 (7,, L1218, 110,+ 114,Cu), L121 DNA + degradationCu), DNA degradation (1, 2, 11, 60 ,(1,102 2,, 11,119 60,), DNA 102, 119 intercalation), DNA intercalation (1, 2, 3–5, L13 (1, 2,+ Cu,3–5, 86L13–89 +, 119Cu,, 86120–),89, apoptosis 119, 120), induction apoptosis ( 1inducti, 2, 65,on102 (,1,119 2, ),65, and 102, cell 119 cycle), and arrest cell (cycle51–57 arrest, 103, 119(51–).57 Despite, 103, 119 the). factDespite that the most fact coordination that most coordination compounds ofcompounds copper exhibit of copper ROS-mediated exhibit ROS cytotoxicity,-mediated examplescytotoxicity, of examples of coordination compounds acting by other mechanisms are also described. Thus, the cytotoxic activity of coordination compounds 42, 43, 69–74, L121 + Cu is not associated with ROS formation and does not decrease under the influence of ROS inhibitors. Redox-active drugs proved to be an effective supplement in addition to antituberculous drugs, or even being an independent therapy. Thus, elesclomol-based copper coordination compound 13, Cu-CQ–based coordination compound 114, and isoniazid-based coordination compounds 116–118 showed promising activity against Mycobacterium tuberculosis. Phosphine-based coordination compounds 79 and 81 showed a promising activity against bacteria, fungi, and the lieshmanial

Int. J. Mol. Sci. 2020, 21, 3965 30 of 37 coordination compounds acting by other mechanisms are also described. Thus, the cytotoxic activity of coordination compounds 42, 43, 69–74, L121 + Cu is not associated with ROS formation and does not decrease under the influence of ROS inhibitors. Redox-active drugs proved to be an effective supplement in addition to antituberculous drugs, or even being an independent therapy. Thus, elesclomol-based copper coordination compound 13, Cu-CQ–based coordination compound 114, and isoniazid-based coordination compounds 116–118 showed promising activity against Mycobacterium tuberculosis. Phosphine-based coordination compounds 79 and 81 showed a promising activity against bacteria, fungi, and the lieshmanial pathogen. In addition, 1,10-phenanthroline–based coordination compounds 32–39 showed higher antibiofilm activity than the clinically used Vancomycin, which is also associated with redox activity of copper cations. The ability of copper coordination compounds to penetrate through the blood-brain barrier along with their stability in the bloodstream allows development of 64Cu-marked PET-imagine agents. Thiosemicarbazone-based coordination compounds 23–30 are Aβ-targeted PET-visualizers of Alzheimer’s disease showed the ability to rapidly crossing the blood-brain barrier, as well as good brain uptake and Aβ plaques affinity. In addition, CuATSM coordination compounds are hypoxia-sensitive PET-visualizers of malignant neoplasms, including head and neck cancer. Cu(II/I) redox potential was repeatedly proven to govern the hypoxia selectivity of CuATSM coordination compounds. Anti-inflammatory properties of coper-containing coordination compounds are interesting due to the possibility of twin antitumor/anti-inflammatory drug development. Thus, aspirin-based coordination compound 102 showed COX-2 inhibition due to aspirin moiety, whereas coordination compounds 45–47 showed analgesic properties themselves. It is also worth noting that using both a ligand and a copper salt is as effective as using a coordination compound. Copper-containing coordination compounds of disulfiram metabolite are always formed in situ, and the same approach has been successfully applied in vitro and in vivo to a number of compounds, such as L13 + Cu, L68 + Cu, L108–L113 + Cu, and L121 + Cu. The redox activity of copper ions along with the their biogenicity, the stability of copper coordination compounds in the bloodstream, and the highly promising therapeutic results in vitro and in vivo prove the potential of copper coordination compounds to become widely used in clinical practice.

Funding: This research was funded by Ministry of Education and Science of the Russian Federation implemented by a governmental decree dated 16 March 2013, no. 211, The Russian Science Foundation, grant number 19-74-10059” and Russian Foundation for Basic Research, grant number 19-29-08007. Acknowledgments: We acknowledge the financial support from the Ministry of Education and Science of the Russian Federation in the framework of the Increase Competitiveness Program of NUST “MISIS” implemented by a governmental decree dated 16 March 2013, no. 211. (Introduction and Conclusions). The Russian Science Foundation supported the section “Drug-based copper coordination compounds” (grant no. 19-74-10059), and the Russian Foundation for Basic Research supported the section “Copper coordination compounds based on ligands with various donor atoms” (grant no. 19-29-08007). Conflicts of Interest: The authors declare no conflict of interest.

References

1. Dömötör, O.; de Almeida, R.F.M.; Côrte-Real, L.; Matos, C.P.; Marques, F.; Matos, A.; Real, C.; Kiss, T.; Enyedy, É.A.; Garcia, M.H.; et al. Studies on the mechanism of action of antitumor bis(aminophenolate) ruthenium (III) complexes. J. Inorg. Biochem. 2017, 168, 27–37. [CrossRef][PubMed] 2. Dabrowiak, J.C. Metals in Medicine; Wiley: Hoboken, NJ, USA, 2009; pp. 49–249. 3. Volarevic, V.; Djokovic, B.; Jankovic, M.G.; Harrell, C.R.; Fellabaum, C.; Djonov, V.; Arsenijevic, N. Molecular mechanisms of cisplatin-induced nephrotoxicity: A balance on the knife edge between renoprotection and tumor toxicity. J. Biomed. Sci. 2019, 26, 1–14. [CrossRef][PubMed] 4. McWhinney,S.R.; Goldberg, R.M.; McLeod, H.L. Platinum Neurotoxicity Pharmacogenetics. Mol. Cancer Ther. 2009, 8, 10–16. [CrossRef][PubMed] Int. J. Mol. Sci. 2020, 21, 3965 31 of 37

5. Wheate, N.J.; Walker, S.; Craig, G.E.; Oun, R. The Status of Platinum Anticancer Drugs in the Clinic and in Clinical Trials. Dalton Trans. 2010, 39, 8113–8127. [CrossRef] 6. Gaál, A.; Orgován, G.; Mihucz, V.G.; Pape, I.; Ingerle, D.; Streli, C.; Szoboszlai, N.J. Metal Transport Capabilities of Anticancer Copper Chelators. Trace Elem. Med. Biol. 2018, 47, 79–88. 7. Xin, C.; Xiao, Z.; Jinghong, C.; Qianqi, Y.; Li, Y. Hinokitiol copper complex inhibits proteasomal deubiquitination and induces paraptosis-like cell death in human cancer cells. Eur. J. Pharmacol. 2017, 815, 147–155. 8. Zeeshan, M.; Murugadas, A.; Ghaskadbi, S.; Rajendran, R.B.; Akbarsha, M.A. ROS dependent copper toxicity in Hydra-biochemical and molecular study. Comp. Biochem. Physiol. C Toxicol. Pharmacol. 2016, 185, 1–12. [CrossRef] 9. Qin, Q.-P.; Meng, T.; Tan, M.-X.; Liu, Y.-C.; Luo, X.-J.; Zou, B.-Q.; Liang, H. Synthesis, crystal structure and biological evaluation of a new dasatinib copper(II) complex as telomerase inhibitor. Eur. J. Med. Chem. 2018, 143, 1597–1603. [CrossRef][PubMed] 10. Hernandes, M.S.; Britto, L.R. NADPH oxidase and neurodegeneration. Curr. Neuropharmacol. 2012, 10, 321–327. [CrossRef] 11. Fatfat, M.; Merhi, R.A.; Rahal, O.; Stoyanovsky, D.A.; Zaki, A.; Haidar, H.; Kagan, V.E.; Gali-Muhtasib, H.; Machaca, K. Copper Chelation Selectively Kills Colon Cancer Cells through Redox Cycling and Generation of Reactive Oxygen Species. BMC Cancer. 2014, 14, 527. [CrossRef] 12. Kremer, M.L. Mechanism of the Fenton reaction. Evidence for a new intermediate. Phys. Chem. Chem. Phys. 1999, 1, 3595–3605. [CrossRef] 13. Sangeetha, S.; Murali, M. Non-covalent DNA binding, protein interaction, DNA cleavage and cytotoxicity of [Cu(quamol)Cl] H O. Int. J. Biol. Macromol. 2018, 107, 2501–2511. [CrossRef][PubMed] · 2 14. Martinez-Bulit, P.; Garza-Ortíz, A.; Mijangos, E.; Barrón-Sosa, L.; Sánchez-Bartéz, F.; Gracia-Mora, I.; Flores-Parra, A.; Contreras, R.; Reedijk, J.; Barba-Behrens, N. 2,6-Bis(2,6-diethylphenyliminomethyl)pyridine coordination compounds with cobalt(II), nickel(II), copper(II), and zinc(II): Synthesis, spectroscopic characterization, X-ray study and in vitro cytotoxicity. J. Inorg. Biochem. 2015, 142, 1–7. [CrossRef] 15. Fengyi, Z.; Weifan, W.; Wen, L.; Li, X.; Shilong, Y.; Xu-Min, C.; Mengyi, Z.; Meng, L.; Mengtao, M.; Hai-Jun, X.; et al. High anticancer potency on tumor cells of dehydroabietylamine Schiff-base derivatives and a copper(II) complex. Eur. J. Med. Chem. 2018, 146, 451–459. 16. Chudal, L.; Pandey, N.K.; Phan, J.; Johnson, O.; Lin, L.; Yu, H.; Shu, Y.; Huang, Z.; Xing, M.; Liu, J.P.; et al. Copper-Cysteamine Nanoparticles as a Heterogeneous Fenton-Like Catalyst for Highly Selective Cancer Treatment. ACS Appl. Bio Mater. 2020, 3, 1804–1814. [CrossRef] 17. Lebon, F.; Boggetto, N.; Ledecq, M.; Durant, F.; Benatallah, Z.; Sicsic, S.; Lapouyade, R.; Kahn, O.; Mouithys-Mickalad, A.; Deby-Dupont, G.; et al. Metal-Organic Compounds: A New Approach for Drug Discovery: N1-(4-Methyl-2-Pyridyl)-2,3,6-Trimethoxybenzamide Copper(II) Complex as an Inhibitor of Human Immunodeficiency Virus 1 Protease. Biochem. Pharmacol. 2002, 63, 1863–1873. [CrossRef] 18. Roch-Arveiller, M.; Huy, D.P.; Maman, L.; Giroud, J.-P.; Sorenson, J.R.J. Non-Steroidal Anti-Inflammatory Drug-Copper Complex Modulation of Polymorphonuclear Leukocyte Migration. Biochem. Pharmacol. 1990, 39, 569–574. [CrossRef] 19. Stanila, A.; Braicu, C.; Stanila, S.; Pop, R.M. Antibacterial Activity of Copper and Cobalt Amino Acids Complexes. Not. Bot. Horti Agrobot. Cluj-Na. 2011, 39, 124–129. [CrossRef] 20. Weder, J.E.; Hambley, T.W.; Kennedy, B.J.; Lay, P.A.; MacLachlan, D.; Bramley, R.; Delfs, C.D.; Murray, K.S.; Moubaraki, B.; Warwick, B.; et al. Anti-Inflammatory Dinuclear Copper(II) Complexes with Indomethacin. Synthesis, Magnetism and EPR Spectroscopy. Crystal Structure of the N,N-Dimethylformamide Adduct. Inorg. Chem. 1999, 38, 1736–1744. [CrossRef] 21. Liao, Y.; Zhao, J.; Bulek, K.; Tang, F.; Chen, X.; Cai, G.; Jia, S.; Fox, P.; Huang, E.; Pizarro, T.; et al. Inflammation mobilizes copper metabolism to promote colon tumorigenesis via an IL-17-STEAP4-XIAP axis. Nat. Commun. 2020, 11, 900. [CrossRef] 22. Chen, K.T.J.; Anantha, M.; Leung, A.W.Y.; Kulkarni, J.A.; Militao, G.G.C.; Wehbe, M.; Sutherland, B.; Cullis, P.R.; Bally, M.B. Characterization of a liposom al copper(II)-quercetin formulation suitable for parenteral use. Drug Delivery Transl. Res. 2020, 10, 202–215. [CrossRef][PubMed] Int. J. Mol. Sci. 2020, 21, 3965 32 of 37

23. Liu, X.; Chu, H.; Cui, N.; Wang, T.; Dong, S.; Cui, S.; Dai, Y.; Wang, D. In vitro and in vivo evaluation of biotin-mediated PEGylated nanostructured lipid as carrier of disulfiram coupled with copper ion. J. Drug Delivery Sci. Technol. 2019, 51, 651–661. [CrossRef] 24. Pellei, M.; Bagnarelli, L.; Luciani, L.; Del Bello, F.; Giorgioni, G.; Piergentili, A.; Quaglia, W.; De Franco, M.; Gandin, V.; Marzano, C.; et al. Synthesis and Cytotoxic Activity Evaluation of New Cu(I) Complexes of Bis(pyrazol-1-yl) Acetate Ligands Functionalized with an NMDA Receptor Antagonist. Int. J. Mol. Sci. 2020, 21, 2616. [CrossRef][PubMed] 25. Takjoo, R.; Centore, R.; Hayatolgheibi, S. Mixed ligand complexes of cadmium(II) and copper(II) dithiocarbazate: Synthesis, spectral characterization, X-ray crystal structure. Inorg. Chim. Acta. 2018, 471, 587–594. [CrossRef] 26. Khan, M.H.; Cai, M.; Deng, J.; Yu, P.; Liang, H.; Yang, F. Anticancer Function and ROS-Mediated Multi-Targeting Anticancer Mechanisms of Copper (II) 2-hydroxy-1-naphthaldehyde Complexes. Molecules 2019, 24, 2544. [CrossRef] 27. Sabithakala, T.; Chittireddy, V.R.R. DNA Binding and in vitro anticancer activity of 2-((1H-benzimidazol- 2-yl)methylamino)acetic acid and its copper(II) mixed-polypyridyl complexes: Synthesis and crystal structure. Appl. Organomet. Chem. 2018, 32, 4550. [CrossRef] 28. Denoyer, D.; Clatworthy, S.A.S.; Cater, M.A. Copper Complexes in Cancer Therapy. Metallo-Drugs Dev. Action Anticancer Agents 2018, 16, 469–506. 29. Wang, Z.; Tan, J.; McConville, C.; Kannappan, V.; Tawari, P.; Brown, J.; Ding, J.; Armesilla, A.; Irache, J.; Mei, Q.; et al. Poly lactic-co-glycolic acid controlled delivery of disulfiram to target liver cancer stem-like cells. Nanomed. Nanotechnol. Biol. Med. 2017, 13, 641–657. 30. Banerjee, P.; Geng, T.; Mahanty, A.; Li, T.; Zong, L.; Wang, B. Integrating the drug, disulfiram into the vitamin E-TPGS-modified PEGylated nanostructured lipid carriers to synergize its repurposing for anti-cancer therapy of solid tumors. Int J. Pharm. 2019, 557, 374–389. [CrossRef][PubMed] 31. Tabti, R.; Tounsi, N.; Gaiddon, C.; Bentouhami, E.; Désaubry, L. Progress in Copper Complexes as Anticancer Agents. Med. Chem. 2017, 7, 1–55. [CrossRef] 32. Wehbe, M.; Leung, A.W.Y.; Abrams, M.J.; Orvig, C.; Bally, M.B. A Perspective – can copper complexes be developed as a novel class of therapeutics? Dalton Trans. 2017, 46, 10758–10773. [CrossRef][PubMed] 33. Santini, C.; Pellei, M.; Gandin, V.; Porchia, M.; Tisato, F.; Marzano, C. Advances in Copper Complexes as Anticancer Agents. Chem. Rev. 2014, 114, 815–862. [CrossRef][PubMed] 34. Ching Ong, Y.; Roy, S.; Andrews, P.; Gasser, G. Metal Compounds against Neglected Tropical Diseases. Chem. Rev. 2019, 119, 730–796. [CrossRef][PubMed] 35. Ruiz-Azuara, L.; Bastian, G.; Bravo-Gómez, M.E.; Cañas, R.C.; Flores-Alamo, M.; Fuentes, I.; Mejia, C.; García-Ramos, J.; Serrano, A. Abstract CT408: Phase I study of one mixed chelates copper(II) compound, Casiopeína CasIIIia with antitumor activity and its mechanism of action. AACR. Cancer Res. 2014, 74 (Suppl. 19). [CrossRef] 36. Galindo-Murillo, R.; Garcia-Ramos, J.C.; Ruiz-Azuara, L.; Cheatham, T.E.; Cortes-Guzman, F. Intercalation processes of copper complexes in DNA. Nucleic Acids Res. 2015, 43, 5364–5376. [CrossRef] 37. Rufino-González, Y.; Ponce-Macotela, M.; García-Ramos, J.C.; Martínez-Gordillo, M.N.; Galindo-Murillo, R.; González-Maciel, A.; Reynoso-Robles, R.; Tovar-Tovar, A.; Flores-Alamo, M.; Toledano-Magaña, Y.; et al. Antigiardiasic activity of Cu(II) coordination compounds: Redox imbalance and membrane damage after a short exposure time. J. Inorg. Biochem. 2019, 195, 83–90. [CrossRef] 38. Anda-Jáuregui, G.; Espinal-Enríquez, J.; Hur, J.; Alcalá-Corona, S.A.; Ruiz-Azuara, L.; Hernández-Lemus, E. Identification of Casiopeina II-gly secondary targets through a systems pharmacology approach. Comput. Biol. Chem. 2019, 78, 127–132. [CrossRef] 39. Shoair, A.F.; El-Bindary, A.A.; El-Ghamaz, N.A.; Rezk, G.N. Synthesis, characterization, DNA binding and antitumor activities of Cu(II) complexes. J. Mol. Liq. 2018, 269, 619–638. [CrossRef] 40. Mo, Q.; Deng, J.; Liu, Y.; Huang, G.; Li, Z.; Yu, P.; Gou, Y.; Yang, F. Mixed-ligand Cu(II) hydrazone complexes designed to enhance anticancer activity. Eur. J. Med. Chem. 2018, 156, 368–380. [CrossRef] 41. Anu, D.; Naveen, P.; Vijaya, P.B.; Frampton, C.S.; Kaveri, M.V. An unexpected mixed valence tetranuclear Copper (I/II) Complex: Synthesis, Structural Characterization, DNA/Protein Binding, Antioxidant and Anticancer Properties. Polyhedron. 2019, 167, 137–150. [CrossRef] Int. J. Mol. Sci. 2020, 21, 3965 33 of 37

42. Synta and GlaxoSmithKline Announce Elesclomol Granted Orphan Drug Designation by the FDA. Available online: https://ir.madrigalpharma.com/news-releases/news-release-details/synta-and-glaxosmithkline- announce-elesclomol-granted-orphan (accessed on 28 January 2008). 43. Hedley, D.; Shamas-Din, A.; Chow, S.; Sanfelice, D.; Schuh, A.C.; Brandwein, J.M.; Seftel, M.D.; Gupta, V.; Yee, K.W.L.; Schimmer, A.D. A phase I study of elesclomol sodium in patients with acute myeloid leukemia. Leuk. Leuk. Lymphoma. 2016, 57, 2437–2440. [CrossRef][PubMed] 44. Vincent, G.; Fayewicz, S.L.; Bateman, N.W.; Hood, B.L.; Sun, M.; Suhan, J.; Duensing, S.; Yin, Y.; Sander, C.; Kirkwood, J.M.; et al. Mitochondrial respiration-an important therapeutic target in melanoma. PLoS ONE. 2012, 7, 40690. 45. Gorshkov, K.; Sima, N.; Sun, W.; Lu, B.; Huang, W.; Travers, J.; Klumpp-Thomas, C.; Michael, S.G.; Xu, T.; Huang, R. Quantitative Chemotherapeutic Profiling of Gynecologic Cancer Cell Lines Using Approved Drugs and Bioactive Compounds. Transl. Oncol. 2019, 12, 441–452. [CrossRef] 46. Haynes, R.K.; Cheu, K.-W.; Chan, H.-W.; Wong, H.-N.; Li, K.-Y.; Tang, M.M.-K.; Chen, M.-J.; Guo, Z.-F.; Guo, Z.-H.; Sinniah, K. Interactions between Artemisinins and other Antimalarial Drugs in Relation to the Cofactor Model—A Unifying Proposal for Drug Action. Chem. Med. Chem. 2012, 7, 2204–2226. [CrossRef] 47. Rathore, S.; Datta, G.; Kaur, I.; Malhotra, P.; Mohmmed, A. Disruption of cellular homeostasis induces organelle stress and triggers apoptosis like cell-death pathways in malaria parasite. Cell Death Dis. 2015, 6, 1803. [CrossRef] 48. Ngwane, A.H.; Petersen, R.D.; Baker, B.; Wiid, I.; Wong, H.N.; Haynes, R.K. The evaluation of the anti-cancer drug elesclomol that forms a redox-active copper chelate as a potential anti-tubercular drug. IUBMB Life. 2019, 71, 532–538. [CrossRef] 49. Paul, D.B.; Simon, R.B.; Mark, B.M.; Helen, M.B.; Jason, S.L.; Jonathan, R.D. Nitroimidazole conjugates of bis (thiosemicarbazonato) 64Cu (II) – Potential combination agents for the PET imaging of hypoxia. J. Inorg. Biochem. 2010, 104, 126–135. 50. Maurer, R.I.; Blower, P.J.; Dilworth, J.R.; Reynolds, C.A.; Zheng, Y.; Mullen, G.E.D. Studies on the Mechanism of Hypoxic Selectivity in Copper Bis(Thiosemicarbazone) Radiopharmaceuticals. J. Med. Chem. 2002, 45, 1420–1431. [CrossRef] 51. Basu, S.; Zhuang, H.; Torigian, D.A.; Rosenbaum, J.; Chen, W.; Alavi, A. Functional imaging of inflammatory diseases using nuclear medicine techniques. Semin. Nucl. Med. 2009, 39, 124–145. [CrossRef] 52. Colombié, M.; Gouard, S.; Frindel, M.; Vidal, A.; Chérel, M.; Kraeber-Bodéré, F.; Rousseau, C.; Bourgeois, M. Focus on the Controversial Aspects of (64) Cu-ATSM in Tumoral Hypoxia Mapping by PET Imaging. Front. Med. 2015, 2, 58. [CrossRef] 53. Clinicaltrials.gov. Phase 1 Dose Escalation and PK Study of Cu(II)ATSM in ALS/MND. NCT02870634. 54. Anjum, R.; Palanimuthu, D.; Kalinowski, D.S.; Lewis, W.; Park, K.C.; Kovacevic, Z.; Khan, I.U.; Richardson, D.R. Synthesis, Characterization, and in Vitro Anticancer Activity of Copper and Zinc Bis(Thiosemicarbazone) complexes. Inorg. Chem. 2019, 58, 13709–13723. [CrossRef][PubMed] 55. Hardy, J.A.; Higgins, G.A. Alzheimer’s disease: The amyloid cascade hypothesis. Science. 1992, 256, 184–185. [CrossRef][PubMed] 56. Kung, H.F.; Choi, S.R.; Qu, W.; Zhang, W.; Skovronsky, D. 18F Stilbenes and Styrylpyridines for PET Imaging of Aβ Plaques in Alzheimer’s Disease: A Miniperspective. J. Med. Chem. 2010, 53, 933–941. [CrossRef] 57. Clark, C.M.; Pontecorvo, M.J.; Beach, T.G.; Bedell, B.J.; Coleman, R.E.; Doraiswamy, P.M.; Fleisher, A.S.; Reiman, E.M.; Sabbagh, M.N.; Sadowsky, C.H.; et al. Cerebral PET with florbetapir compared with neuropathology at autopsy for detection of neuritic amyloid-β plaques: A prospective cohort study. Lancet Neurol. 2012, 11, 669–678. [CrossRef] 58. Bagheri, S.; Squitti, R.; Haertlé, T.; Siotto, M.; Saboury, A.A. Role of Copper in the Onset of Alzheimer’s Disease Compared to Other Metals. Front. Aging. Neurosci. 2018, 9, 446. [CrossRef][PubMed] 59. Kim, A.C.; Lim, S.; Kim, Y.K. Metal Ion Effects on Aβ and Tau Aggregation. Int. J. Mol. Sci. 2018, 19, 128. [CrossRef] 60. Lim, S.C.; Paterson, B.M.; Fodero-Tavoletti, M.T.; O’Keefe, G.J.; Cappai, R.; Barnham, K.J.; Villemagne, V.L.; Donnelly, P.S. A copperradiopharmaceutical for diagnostic imaging of Alzheimer’s disease: A bis(thiosemicarbazonato)copper(ii) complex that binds to amyloid-β plaques. Chem. Commun. 2010, 46, 5437–5439. [CrossRef] Int. J. Mol. Sci. 2020, 21, 3965 34 of 37

61. Hickey, J.L.; Lim, S.C.; Hayne, D.J.; Paterson, B.M.; White, J.M.; Villemagne, V.L.; Roselt, P.; Binns, D.; Cullinane, C.; Jeffery, C.M.; et al. Diagnostic Imaging Agents for Alzheimer’s Disease: Copper Radiopharmaceuticals that Target Aβ Plaques. J. Am. Chem. Soc. 2013, 135, 16120–16132. [CrossRef] 62. McInnes, L.E.; Noor, A.; Kysenius, K.; Cullinane, C.; Roselt, P.; McLean, C.A.; Chiu, F.C.K.; Powell, A.K.; Crouch, P.J.; White, J.M.; et al. Potential Diagnostic Imaging of Alzheimer’s Disease with Copper-64 Complexes That Bind to Amyloid-β Plaques. Inorg. Chem. 2019, 5, 3382–3395. [CrossRef] 63. Gokhale, N.H.; Padhye, S.B.; Billington, D.C.; Rathbone, D.L.; Croft, S.L.; Kendrick, H.D.; Anson, C.E.; Powell, A.K. Synthesis and characterization of copper(II) complexes of pyridine-2-carboxamidrazones as potent antimalarial agents. Inorg. Chim. Acta. 2003, 349, 23–29. [CrossRef] 64. Beeton, M.L.; Aldrich-Wright, J.R.; Bolhuis, A. The antimicrobial and antibiofilm activities of copper (II) complexes. J. Inorg. Biochem. 2014, 140, 167–172. [CrossRef][PubMed] 65. Newton, G.L.; Rawat, M.; La Clair, J.J.; Jothivasan, V.K.; Budiarto, T.; Hamilton, C.J.; Claiborne, A.; Helmann, J.D.; Fahey, R.C. Bacillithiol is an antioxidant thiol produced in Bacilli. Nat. Chem. Biol. 2009, 5, 625–627. [CrossRef][PubMed] 66. Brandão, P.; Guieu, S.; Correia-Branco, A.; Silva, C.; Martel, F. Development of novel Cu(I) compounds with vitamin B1 derivative and their potential application as anticancer drugs. Inorg. Chim. Acta. 2019, 487, 287–294. [CrossRef] 67. Krasnovskaya, O.O.; Fedorov, Y.V.; Gerasimov, V.M.; Skvortsov, D.A.; Moiseeva, A.A.; Mironov, A.V.; Beloglazkina, E.K.; Zyk, N.V.; Majouga, A.G. Novel 2-aminoimidazole-4-one complexes of copper(ii) and cobalt(ii): Synthesis, structural characterization and cytotoxicity. Arab. J. Chem. 2019, 12, 835–846. [CrossRef] 68. Hussain, A.; AlAjmi, M.F.; Rehman, M.T.; Amir, S.; Husain, F.M.; Alsalme, A.; Siddiqui, M.A.; AlKhedhairy, A.A.; Khan, R.A. Copper(II) complexes as potential anticancer and Nonsteroidal anti-inflammatory agents: In vitro and in vivo studies. Sci. Rep. 2019, 9, 5237. [CrossRef] 69. Sliwa,´ E.I.; Sliwińska-Hill,U.;´ Ba˙zanów, B.; Siczek, M.; Kłak, J.; Smoleński,P. Synthesis, Structural, and Cytotoxic Properties of New Water-Soluble Copper(II) Complexes Based on 2,9-Dimethyl-1,10- Phenanthroline and Their One Derivative Containing 1,3,5-Triaza-7-Phosphaadamantane-7-Oxide. Molecules 2020, 25, 741. 70. Milani, N.C.; Maghsoud, Y.; Hosseini, M.; Babaei, A.; Rahmani, H.; Roe, S.M.; Gholivand, K. A new class of copper(I) complexes with imine-containing chelators which show potent anticancer activity. Appl. Organometal. Chem. 2020, 34, 5526. 71. Kacar, S.; Unver, H.; Sahinturk, V. A mononuclear copper(II) complex containing benzimidazole and pyridyl ligands: Synthesis, characterization, and antiproliferative activity against human cancer cells. Arabian J. Chem. 2020, 13, 4310–4323. [CrossRef] 72. Majouga, A.G.; Zvereva, M.I.; Rubtsova, M.P.; Skvortsov, D.A.; Mironov, A.V.; Azhibek, D.M.; Krasnovskaya, O.O. Mixed Valence Copper(I,II) Binuclear Complexes with Unexpected Structure: Synthesis, Biological Properties and Anticancer Activity. J. Med. Chem. 2014, 14, 6252–6258. [CrossRef] 73. Hald, J.; Jacobsen, E. A Drug Sensitizing the Organism to Ethyl Alcohol. Lancet. 1948, 252, 1001–1004. [CrossRef] 74. Tawari, P.E.; Wang, Z.; Najlah, M.; Tsang, C.W.; Kannappan, V.; Liu, P.; McConville, C.; He, B.; Armesilla, A.L.; Wang, W. The Cytotoxic Mechanisms of Disulfiram and Copper(II) in Cancer Cells. Toxicol. Res. 2015, 4, 1439–1442. [CrossRef] 75. Fasehee, H.; Dinarvand, R.; Ghavamzadeh, A.; Esfandyari-Manesh, M.; Moradian, H.; Faghihi, S.; Ghaffari, S.H. Delivery of Disulfiram into Breast Cancer Cells Using Folate-Receptor-Targeted Plga-Peg Nanoparticles: In Vitro and in Vivo Investigations. J. Nanobiotechnol. 2016, 14, 32. [CrossRef][PubMed] 76. Sedlacek, J.; Martins, L.M.D.R.S.; Danek, P.; Pombeiro, A.J.L.; Cvek, B. Diethyldithiocarbamate Complexes with Metals Used as Food Supplements Show Different Effects in Cancer Cells. J. Appl. Biomed. 2014, 12, 301–308. [CrossRef] 77. Wu, X.; Xue, X.; Wang, L.; Wang, W.; Han, J.; Sun, X.; Zhang, H.; Liu, Y.; Che, X.; Yang, J.; et al. Suppressing autophagy enhances disulfiram/copper-induced apoptosis in non-small cell. Eur. J. Pharmacol. 2018, 827, 1–12. [CrossRef][PubMed] 78. Lewis, D.J.; Deshmukh, P.; Tedstone, A.A.; Tuna, F.; O’Brien, P. On the Interaction of Copper(II) with Disulfiram. Chem. Commun. 2014, 50, 13334–13337. [CrossRef][PubMed] Int. J. Mol. Sci. 2020, 21, 3965 35 of 37

79. Wang, F.; Jiao, P.; Qi, M.; Frezza, M.; Dou, Q.P.; Yan, B. Turning Tumor-Promoting Copper into an Anti-Cancer Weapon Via High-Throughput Chemistry. Curr. Med. Chem. 2010, 17, 2685–2698. [CrossRef] 80. Phase II Trial of Disulfiram With Copper in Metastatic Breast Cancer (DISC). The Institute of Molecular and Translational Medicine, Czech Republic. Clinicaltrials.gov Identifier: NCT03323346. 81. Liu, P.; Brown, S.; Goktug, T.; Channathodiyil, P.; Kannappan, V.; Hugnot, J.-P.; Guichet, P.-O.; Bian, X.; Armesilla, A.L.; Darling, J.L.; et al. Cytotoxic effect of disulfiram/copper on human glioblastoma cell lines and ALDH-positive cancer-stem-like cells. Br. J. Cancer 2012, 107, 1488–1497. [CrossRef] 82. Cvek, B. Comment on ‘Cytotoxic effect of disulfiram/copper on human glioblastoma cell lines and ALDH-positive cancer-stem-like cells’. Br. J. Cancer. 2013, 108, 993. [CrossRef] 83. Dufour, P.; Lang, J.M.; Giron, C.; Duclos, B.; Haehnel, P.; Jaeck, D.; Jung, J.M.; Oberling, F. Sodium ditiocarb as adjuvant immunotherapy for high risk breast cancer: A randomized study. Biotherapy 1993, 6, 9–12. [CrossRef] 84. Tao, X.; Gou, J.; Zhang, Q.; Tan, X.; Ren, T.; Yao, Q.; Tian, B.; Kou, L.; Zhang, L.; Tang, X. Synergistic breast tumor cell killing achieved by intracellular co-delivery of doxorubicin and disulfiram via core-shell-corona nanoparticles. Biomater. Sci. 2018, 6, 1869–1881. [CrossRef] 85. Wu, W.; Yu, L.; Jiang, Q.; Huo, M.; Lin, H.; Wang, L.; Chen, Y.; Shi, J. Enhanced Tumor-Specific Disulfiram Chemotherapy by In Situ Cu2+ Chelation-Initiated Nontoxicity-to-Toxicity Transition. J. Am. Chem. Soc. 2019, 29, 11531–11539. [CrossRef][PubMed] 86. Xu, X.; Xu, J.; Zhao, C.; Hou, X.; Li, M.; Wang, L.; Chen, L.; Chen, Y.; Zhu, L.; Yang, H. Antitumor effects of disulfiram/copper complex in the poorly-differentiated nasopharyngeal carcinoma cells via activating ClC-3 chloride channel. Biomed. Pharmacother. 2019, 120, 109529. [CrossRef][PubMed] 87. Yaping, Y.; Mengjia, L.; Xiaoxue, S.; Congran, Z.; Yawei, W.; Liwei, W.; Lixin, C.; Zhihong, L.; Linyan, Z.; Haifeng, Y. The selective cytotoxicity of DSF-Cu attributes to the biomechanical properties and cytoskeleton rearrangements in the normal and cancerous nasopharyngeal epithelial cells. Int. J. Biochem. Cell Biol. 2017, 84, 96–108. 88. Li, Y.; Chen, F.; Chen, J.; Chan, S.; He, Y.; Liu, W.; Zhang, G. Disulfiram/Copper Induces Antitumor Activity against Both Nasopharyngeal Cancer Cells and Cancer-Associated Fibroblasts through ROS/MAPK and Ferroptosis Pathways. Cancers 2020, 12, 138. [CrossRef] 89. McMahon, A.; Chen, W.; Li, F. Old wine in new bottles: Advanced drug delivery systems for disulfiram-based cancer therapy. J. Control Release 2020, 319, 352–359. [CrossRef] 90. Phase II Study of Disulfiram and Cisplatin in Refractory TGCTs. National Cancer Institute, Slovakia. ClinicalTrials.gov Identifier: NCT03950830. 91. Phase II Study of Vinorelbine, Cisplatin, Disulfiram and Copper in CTC_EMT Positive Refractory Metastatic Breast Cancer. National Cancer Institute, Slovakia. ClinicalTrials.gov Identifier: NCT04265274. 92. Blanchard, C.; Brooks, L.; Ebsworth-Mojica, K.; Didione, L.; Wucher, B.; Dewhurst, S.; Krysan, D.; Dunman, P.M.; Wozniak, R.A.F. Zinc Pyrithione Improves the Antibacterial Activity of Silver Sulfadiazine Ointment. mSphere 2016, 5, 00194-16. [CrossRef] 93. Liu, N.N.; Liu, C.J.; Li, X.F.; Liao, S.Y.; Song, W.B.; Yang, C.S.; Zhao, C.; Huang, H.B.; Guan, L.X.; Zhang, P.Q.; et al. A Novel Proteasome Inhibitor Suppresses Tumor Growth Via Targeting Both 19s Proteasome Deubiquitinases and 20s Proteolytic Peptidases. Sci. Rep. 2014, 4, 5240. [CrossRef][PubMed] 94. Zhang, H.; Thomas, R.; Oupicky, D.; Peng, F. Synthesis and characterization of new copper thiosemicarbazone complexes with an ONNS quadridentate system: Cell growth inhibition, S-phase cell cycle arrest and proapoptotic activities on cisplatin-resistant neuroblastoma cells. J. Biol. Inorg. Chem. 2008, 13, 47–55. [CrossRef] 95. Hancock, C.N.; Stockwin, L.H.; Han, B.; Divelbiss, R.D. A copper chelate of thiosemicarbazone NSC 689534 induces oxidative/ER stress and inhibits tumor growth in vitro and in vivo. Free Radical Biol. Med. 2011, 50, 110–121. [CrossRef] 96. Carcelli, M.; Tegoni, M.; Bartoli, J.; Marzano, C.; Pelosi, G.; Salvalaio, M.; Rogolino, D.; Gandin, V. In vitro and in vivo anticancer activity of tridentate thiosemicarbazone copper complexes: Unravelling an unexplored pharmacological target. Eur. J. Med. Chem. 2020, 194, 112266. [CrossRef] 97. Kongot, M.; Reddy, D.; Singh, V.; Patel, R.; Singhal, N.K.; Kumar, A. Potent drug candidature of an ONS donor tethered copper (II) complex: Anticancer activity, cytotoxicity and spectroscopically approached BSA binding studies. Spectrochim. Acta Part A 2019, 212, 330–342. [CrossRef][PubMed] Int. J. Mol. Sci. 2020, 21, 3965 36 of 37

98. Balakrishna, M.S. Copper(I) Chemistry of Phosph ines, Functionalized Phosphines and Phosphorus Heterocycles; Elsevier: Amsterdam, The Netherlands, 2019; pp. 1–442. 99. Khan, S.I.; Ahmad, S.; Altaf, A.A.; Rauf, M.K.; Badshah, A.; Azam, S.S.; Tahir, M.N. Heteroleptic copper(I) halides with triphenylphosphine and acetylthiourea: Synthesis, characterization and biological studies (experimental and molecular docking). New J. Chem. 2019, 43, 19318–19330. [CrossRef] 100. Tapanelli, S.; Habluetzel, A.; Pellei, M.; Marchiò, L.; Tombesi, A.; Capparè, A.; Santini, C. Novel metalloantimalarials: Transmission blocking effects of water soluble Cu(I), Ag(I) and Au(I) phosphane complexes on the murine malaria parasite Plasmodium berghei. J. Inorg. Biochem. 2017, 166, 1–4. [CrossRef] [PubMed] 101. Komarnicka, U.K.; Starosta, R.; Kyziołb, A.; Je˙zowska-Bojczuk,M. Copper(I) complexes with phosphine derived from sparfloxacin. Part I – structures, spectroscopic properties and cytotoxicity. Dalton Trans. 2015, 44, 12688–12699. [CrossRef][PubMed] 102. Mashat, K.H.; Babgi, B.A.; Hussien, M.A.; Arshad, M.N.; Abdellattif, M.H. Synthesis, structures, DNA-binding and anticancer activities of some copper(I)-phosphine complexes. Polyhedron 2019, 158, 164–172. [CrossRef] 103. Wallis, S.C.; Gahan, L.R.; Charles, B.G.; Hambley, T.W.; Duckworth, P.A. Copper(II) Complexes of the Fluoroquinolone Antimicrobial Ciprofloxacin. Synthesis, X-Ray Structural Characterization, and Potentiometric Study. J. Inorg. Biochem. 1996, 62, 1–16. [CrossRef] 104. Hanson, J.C.; Camerman, N.; Camerman, A. Structure of a Copper-Isoniazid Complex. J. Med. Chem. 1981, 24, 1369–1371. [CrossRef] 105. Bottari, B.; Maccari, R.; Monforte, F.; Ottana, R.; Rotondo, E.; Vigorita, M.G. Isoniazid Related Copper(II) and Nickel(II) Complexes with Antimycobacterial in Vitro Activity. Bioorg. Med. Chem. Lett. 2000, 10, 657–660. [CrossRef] 106. Sukul, A.; Poddar, S.K.; Haque, S.; Kumar, S.; Das, S.; Mahmud, Z.; Rahman, A. Synthesis, Characterization and Comparison of Local Analgesic, Anti-Inflammatory, Anti-Ulcerogenic Activity of Copper and Zinc Complexes of Indomethacin. Anti-Inflamm. Anti-Allergy Agents Med. Chem. 2017, 15, 221–233. [CrossRef] 107. Di Vaira, M.; Bazzicalupi, C.; Orioli, P.; Messori, L.; Bruni, B.; Zatta, P. Clioquinol, a Drug for Alzheimer’s Disease Specifically Interfering with Brain Metal Metabolism: Structural Characterization of Its Zinc(II) and Copper(II) Complexes. Inorg. Chem. 2004, 43, 3795–3797. [CrossRef] 108. Yassin, N.Z.; El-Shenawy, S.M.; Abdel-Rahman, R.F.; Yakoot, M.; Hassan, M.; Helmy, S. Effect of a Topical Copper Indomethacin Gel on Inflammatory Parameters in a Rat Model of Osteoarthritis. Drug Des. Devel. Ther. 2015, 9, 1491–1498. [PubMed] 109. Okuyama, S.; Hashimoto, S.; Aihara, H.; Willingham, W.M.; Sorenson, J.R. Copper Complexes of Non-Steroidal Antiinflammatory Agents: Analgesic Activity and Possible Opioid Receptor Activation. Agents Actions. 1987, 21, 130–144. [CrossRef][PubMed] 110. Sayen, S.; Carlier, A.; Tarpin, M.; Guillon, E. A Novel Copper(II) Mononuclear Complex with the Non-Steroidal Anti-Inflammatory Drug Diclofenac: Structural Characterization and Biological Activity. J. Inorg. Biochem. 2013, 120, 39–43. [CrossRef][PubMed] 111. Gumilar, F.; Agotegaray, M.; Bras, C.; Gandini, N.A.; Minetti, A.; Quinzani, O. Anti Nociceptive Activity and Toxicity Evaluation of Cu(II)-Fenoprofenate Complexes in Mice. Eur. J. Pharmacol. 2012, 675, 32–39. [CrossRef] 112. Kovala-Demertzi, D.; Hadjipavlou-Litina, D.; Staninska, M.; Primikiri, A.; Kotoglou, C.; Demertzis, M.A. Anti-oxidant, in vitro, in vivo anti-inflammatory activity and antiproliferative activity of mefenamic acid and its metal complexes with manganese(II), cobalt(II), nickel(II), copper(II) and zinc(II). J. Enzyme Inhib. Med. Chem. 2009, 24, 742–752. [CrossRef] 113. Shi, X.; Fang, H.; Guo, Y.; Yuan, H.; Guo, Z.; Wang, X. Anticancer copper complex with nucleus, mitochondrion and cyclooxygenase-2 as multiple targets. J. Inorg. Biochem. 2019, 190, 38–44. [CrossRef] 114. Olender, D.; Zwawiak,˙ J.; Lukianchuk, V.; Lesyk, R.; Kropacz, A.; Fojutowski, A.; Zaprutko, L. Synthesis of some N-substituted nitroimidazole derivatives as potential antioxidant and antifungal agents. Eur. J. Med. Chem. 2009, 44, 645–652. [CrossRef] 115. Varshney, V.; Mishra, N.N.; Shukla, P.K.; Sahu, D.P. Synthesis of nitroimidazole derived oxazolidinones as antibacterial agents. Eur. J. Med. Chem. 2010, 45, 661–666. [CrossRef] Int. J. Mol. Sci. 2020, 21, 3965 37 of 37

116. Mushtaque, M.; Avecilla, F.; Haque, A.; Yab, Z.; Rizvi, M.M.A.; Khan, M.S. Synthesis, structural and biological activity of N-substituted 2-methyl-4-/5-nitroimidazole derivatives. J. Mol. Struct. 2019, 1185, 440–449. [CrossRef] 117. Ding, W.Q.; Liu, B.L.; Vaught, J.L.; Yamauchi, H.; Lind, S.E. Anticancer Activity of the Antibiotic Clioquinol. Cancer Res. 2005, 65, 3389–3395. [CrossRef] 118. Pellei, M.; Gandin, V.; Cimarelli, C.; Quaglia, W.; Mosca, N.; Bagnarelli, L.; Marzano, C.; Santini, C. Syntheses and biological studies of nitroimidazole conjugated heteroscorpionate ligands and related Cu(I) and Cu(II) complexes. J. Inorg. Biochem. 2018, 187, 33–40. [CrossRef][PubMed] 119. Tardito, S.; Barilli, A.; Bassanetti, I.; Tegoni, M.; Bussolati, O.; Franchi-Gazzola, R.; Mucchino, C.; Marchiò, L. Copper-Dependent Cytotoxicity of 8-Hydroxyquinoline Derivatives Correlates with Their Hydrophobicity and Does Not Require Caspase Activation. J. Med. Chem. 2012, 55, 10448–10459. [CrossRef][PubMed] 120. Harrison, M.D.; Jones, C.E.; Solioz, M.; Dameron, C.T. Intracellular Copper Routing: The Role of Copper Chaperones. Trends Biochem. Sci. 2000, 25, 29–32. [CrossRef] 121. Shah, S.; Dalecki, A.G.; Malalasekera, A.P.; Crawford, C.L.; Michalek, S.M.; Kutsch, O.; Sun, J.; Bossmann, S.H.; Wolschendorf, F. 8-Hydroxyquinolines Are Boosting Agents of Copper-Related Toxicity in Mycobacterium Tuberculosis. Antimicrob. Agents Chemother. 2016, 60, 5765–5776. [CrossRef][PubMed] 122. Silva, P.B.; Souza, P.C.; Calixto, G.M.F.; Lopes, E.D.O.; Frem, R.C.G.; Netto, A.V.G.; Mauro, A.E.; Pavan, F.R.; Chorilli, M. In Vitro Activity of Copper(II) Complexes, Loaded or Unloaded into a Nanostructured Lipid System, against Mycobacterium tuberculosis. Int. J. Mol. Sci. 2016, 17, 745. [CrossRef] 123. Fregonezi, N.F.; de Souza, F.A.; Aleixo, N.A.; da Silva Gomes, P.S.; Silvestre, R.B.; De Grandis, A.; da Silva, P.B.; Pavan, F.R.; Chorilli, M.; Resende, F.A. Cyto-genotoxic evaluation of novel anti-tubercular copper (II) complexes containing isoniazid-based ligands. Regul. Toxicol. Pharmacol. 2020, 113, 104653. [CrossRef] 124. Heras, B.L.; Amesty, Á.; Estévez-Braun, A.; Hortelano, S. Metal Complexes of Natural Product Like-compounds with Antitumor Activity. Anti-Cancer Agents Med. Chem. 2019, 19, 48–65. [CrossRef] 125. Fei, B.-L.; Tu, S.; Wei, Z.; Wang, P.; Long, J.-Y.; Qiao, C.; Chen, Z.-F. Biological evaluation of optically pure chiral binuclear copper(II) complexes based on a rosin derivative as highly potential anticancer agents. Dalton Trans. 2019, 48, 15646–15656. [CrossRef]

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