👤 Stojanović N

Inhibitors of poly(ADP-ribose) polymerase-1 (PARP-1) showed remarkable clinical efficacy in BRCA-mutated tumors. Based on the rational drug design, derivatives of PARP inhibitor 3-aminobenzamide (3-AB), 2-amino-4-methylbenzamide (L1) and 3-amino-N-methylbenzamide (L2), were coordinated to the ruthenium(II) ion, to form potential drugs affecting DNA and inhibiting PARP enzyme. The four conjugated complexes of formula: C1 [(ƞ⁶-toluene)Ru(L1)Cl]PF₆, C2 [(ƞ⁶-p-cymene)Ru(L1)Cl]PF₆, C3 [(ƞ⁶-toluene)Ru(L2)Cl₂] and C4 [(ƞ⁶-p-cymene)Ru(L2)Cl₂], have been synthesized and characterized. Colorimetric 3-(4.5-dimethylthiazol-2-yl)-2.5-diphenyltetrazolium bromide (MTT) assay showed the highest antiproliferative activity of C1 in HCC1937, MDA-MB-231, and MCF-7 breast cancer cells. Efficiency of inhibition of PARP-1 enzymatic activity in vitro decreased in order: C2 > C4 > 3-AB>C1 > C3. ICP-MS study of intracellular accumulation and distribution in BRCA1-mutated HCC1937 revealed that C1-C4 entered cells within 24 h. The complex C1 showed the highest intracellular accumulation, nuclear-targeting properties, and exhibited the highest DNA binding (39.2 ± 0.6 pg of Ru per μg of DNA) that resulted in the cell cycle arrest in the S phase. Show less

Novel phthiocol-based organometallics with in situ formed tridentate N,O,O-coordination motif were established via three-component microwave assisted one-pot reaction. These complexes exhibited enhanced stability in aqueous solution compared to the parental compound KP2048 and showed unexpected cytotoxic behaviour and selectivity in 2D and 3D cell cultures. Show less

A series of 15 piano-stool complexes featuring either a RuII, RhIII or IrIII metal center, a bidentate thiopyridone ligand, and different leaving groups was synthesized. The leaving groups were selected in order to cover a broad range of different donor atoms. Thus, 1-methylimidazole served as a N-donor, 1,3,5-triaza-7-phosphaadamantane (pta) as a P-donor, and thiourea as a S-donor. Additionally, three complexes featuring different halido leaving groups (Cl, Br, I) were added. Leaving group alterations were carried out with respect to a possible influence on pharmacokinetic and pharmacodynamic parameters, as well as the cytotoxicity of the respective compounds. The complexes were characterized via NMR spectroscopy, X-ray diffraction (where possible), mass spectrometry, and elemental analysis. Cytotoxicity was assessed in 2D cultures of human cancer cell lines by microculture and clonogenic assays as well as in multicellular tumor spheroids. Furthermore, cellular accumulation studies, flow-cytometric apoptosis and ROS assays, DNA plasmid assays, and laser ablation ICP-MS studies for analyzing the distribution in sections of multicellular tumor spheroids were conducted. This work demonstrates the importance of investigating each piano-stool complexes' properties, as the most promising candidates showed advantages over each other in certain tests/assays. Thus, it was not possible to single out one lead compound, but rather a group of complexes with enhanced cytotoxicity and activity. Show less

Due to the great potential expressed by an anticancer drug candidate previously reported by our group, namely, Ru-sq ([Ru(DIP)₂(sq)](PF₆) (DIP, 4,7-diphenyl-1,10-phenanthroline; sq, semiquinonate ligand), we describe in this work a structure-activity relationship (SAR) study that involves a broader range of derivatives resulting from the coordination of different catecholate-type dioxo ligands to the same Ru(DIP)₂ core. In more detail, we chose catechols carrying either an electron-donating group (EDG) or an electron-withdrawing group (EWG) and investigated the physicochemical and biological properties of their complexes. Several pieces of experimental evidences demonstrated that the coordination of catechols bearing EDGs led to deep-red positively charged complexes 1-4 in which the preferred oxidation state of the dioxo ligand is the uninegatively charged semiquinonate. Complexes 5 and 6, on the other hand, are blue/violet neutral complexes, which carry an EWG-substituted dinegatively charged catecholate ligand. The biological investigation of complexes 1-6 led to the conclusion that the difference in their physicochemical properties has a strong impact on their biological activity. Thus, complexes 1-4 expressed much higher cytotoxicities than complexes 5 and 6. Complex 1 constitutes the most promising compound in the series and was selected for a more in depth biological investigation. Apart from its remarkably high cytotoxicity (IC₅₀ = 0.07-0.7 μM in different cancerous cell lines), complex 1 was taken up by HeLa cells very efficiently by a passive transportation mechanism. Moreover, its moderate accumulation in several cellular compartments (i.e., nucleus, lysosomes, mitochondria, and cytoplasm) is extremely advantageous in the search for a potential drug with multiple modes of action. Further DNA metalation and metabolic studies pointed to the direct interaction of complex 1 with DNA and to the severe impairment of the mitochondrial function. Multiple targets, together with its outstanding cytotoxicity, make complex 1 a valuable candidate in the field of chemotherapy research. It is noteworthy that a preliminary biodistribution study on healthy mice demonstrated the suitability of complex 1 for further in vivo studies. Show less

The photoactivatable Ru (II) complex 1 [Ru(bipy)2(dpphen)]Cl2 (where bipy = 2,2'-bipyridine and dpphen = 2,9-diphenyl-1,10-phenanthroline) has been shown to possess promising anticancer activity against triple negative adenocarcinoma MDA-MB-231 cells. The present study aims to elucidate the plausible mechanism of action of the photoactivatable complex 1 against MDA-MB-231 cells. Upon photoactivation, complex 1 exhibited time-dependent cytotoxic activity with a phototoxicity index (P Index) of >100 after 72 h. A significant increase in cell rounding and detachment, loss of membrane integrity, ROS accumulation and DNA damage was observed. Flow cytometry and a fluorescent apoptosis/necrosis assay showed an induction of cell apoptosis. Western blot analysis revealed the induction of intrinsic and extrinsic pathways and inhibition of the MAPK and PI3K pathways. The photoproduct of complex 1 showed similar effects on key apoptotic protein expression confirming that it is behind the observed cell death. In conclusion, the present study revealed that complex 1 is a potent multi-mechanistic photoactivatable chemotherapeutic drug that may serve as a potential lead molecule for targeted cancer chemotherapy. Show less

Due to several negative issues, market available drugs have been gradually losing their importance in the treatment of cancer. With a view to discover suitable drugs capable of diagnosing as well as inhibiting the growth of cancer cells, we have aspired to develop a group of theranostic metal complexes which will be (i) target specific, (ii) cytoselective, thus rendering the normal cell unaffected, (iii) water-soluble, (iv) cancer cell permeable, and (v) luminescent, being beneficial for healing the cancer eternally. Therefore, to reach our goal, we have prepared novel Ru(II)- and Ir(III)-based bimetallic and hetero bimetallic scaffolds using click-derived pyridinyltriazolylmethylquinoxaline ligands followed by metal coordination. Most of the compounds have displayed significant cytoselectivity against colorectal adenocarcinoma (Caco-2) and epithiloid cervical carcinoma (HeLa) cells with respect to normal human embryonic kidney cells (HEK-293) compared to cisplatin [cis-diamminedichloroplatinum(II)] along with excellent binding efficacy with DNA as well as serum albumin. Complex [(η⁶-p-cymene)(η⁵-Cp*)Ru^IIIr^IIICl₂(K²-N,N-L)](PF₆)₂ [RuIrL] exhibited the best cytoselectivity against all the human cancer cells and was identified as the most significant cancer theranostic agent in terms of potency, selectivity, and fluorescence quantum yield. Investigation of the localization of complex [Ir₂L] and [RuIrL] in the more aggressive colorectal adenocarcinoma cell HT-29 indicates that mitochondria are the key cellular target for destroying cancer cells. Mitochondrial dysfunction and G2/M phase cell cycle arrest in HT-29 cell were found to be involved in the apoptotic cell death pathway induced by the test complexes [Ir₂L] and [RuIrL]. These results validate the concept that these types of complexes will be reasonably able to exert great potential for tumor diagnosis as well as therapy in the near future. Show less

Chemotherapy remains one of the dominant treatments to cure cancer. However, due to the many inherent drawbacks, there is a search for new chemotherapeutic drugs. Many classes of compounds have been investigated over the years to discover new targets and synergistic mechanisms of action including multicellular targets. In this work, we designed a new chemotherapeutic drug candidate against cancer, namely, [Ru(DIP)₂(sq)](PF₆) (Ru-sq) (DIP = 4,7-diphenyl-1,10-phenanthroline; sq = semiquinonate ligand). The aim was to combine the great potential expressed by Ru(II) polypyridyl complexes and the singular redox and biological properties associated with the catecholate moiety. Experimental evidence (e.g., X-ray crystallography, electron paramagnetic resonance, electrochemistry) demonstrates that the semiquinonate is the preferred oxidation state of the dioxo ligand in this complex. The biological activity of Ru-sq was then scrutinized in vitro and in vivo, and the results highlight the promising potential of this complex as a chemotherapeutic agent against cancer. Show less

Herein the design and synthesis of a new 8-hydroxyquinoline derivative, (S)-5-chloro-7-((proline-1-yl)methyl)8-hydroxyquinoline (HQCl-Pro), with good water solubility and multidrug resistance reversal activity are reported. In this work the proton dissociation processes of HQCl-Pro and its complex formation with [Rh(η⁵-C₅Me₅)(H₂O)₃]²⁺, [Ru(η⁶-p-cymene)(H₂O)₃]²⁺ and [Ru(η⁶-toluene)(H₂O)₃]²⁺ were investigated by the combined use of pH-potentiometry, UV-visible spectrometry and ¹H NMR spectroscopy. Our results revealed the prominent solution stability of the complexes in all cases. The lipophilicity of the complexes increased with the chloride ion concentration, and the complexes showed moderate log D values (-0.8 to +0.4) at pH 7.4 at all tested Cl^- concentrations. The formation of mixed hydroxido complexes from the aqua complexes was characterized by relatively high pK_a values (8.45-9.62 in chloride-free medium). Complexation processes are much slower with the Ru(η⁶-arene) triaqua cations than with [Rh(η⁵-C₅Me₅)(H₂O)₃]²⁺. Both the pK_a values and H₂O/Cl^- exchange constants of the Ru-complexes are lower by 0.5-1.0 orders of magnitude than those of the Rh analogue. Arene loss (p-cymene and toluene) and oxidation were found in the case of Ru-complexes when an excess of HQCl-Pro and aromatic (N,N) bidentate ligands was added. The cytotoxicity and antiproliferative effect of HQCl-Pro and its complexes were assayed in vitro. In contrast to the structurally familiar 8-hydroxyquinoline, HQCl-Pro and its Rh(η⁵-C₅Me₅) complex were somewhat more effective against drug resistant Colo 320 adenocarcinoma human cells compared to the drug sensitive Colo 205 cells. The Ru- and Rh-complexes showed a similar metal uptake level after 4 h, while a longer incubation time resulted in higher cellular Rh concentration. Show less

The reaction of 2-{2-(benzo[1,3]dioxol-5-yl)- diazo}-4-methylphenol (HL) with [Ru(PPh₃)₃Cl₂] in ethanol resulted in the carbonylated ruthenium complex [RuL(PPh₃)₂(CO)] (1), wherein metal-assisted decarbonylation via in situ ethanol dehydrogenation is observed. When the reaction was performed in acetonitrile, however, the complex [RuL(PPh₃)₂(CH₃CN)] (2) was obtained as the main product, probably by trapping of a common intermediate through coordination of CH₃CN to the Ru(II) center. The analogous reaction of HL with [Ir(PPh₃)₃Cl] in ethanol did not result in ethanol decarbonylation and instead gave the organoiridium hydride complex [IrL(PPh₃)₂(H)] (3). Unambiguous evidence for the generation of CO via ruthenium-assisted ethanol oxidation is provided by the synthesis of the ¹³C-labeled complex, [Ru(PPh₃)₂L(¹³CO)] (1A) using isotopically labeled ethanol, CH₃¹³CH₂OH. To summarize all the evidence, a ruthenium-assisted mechanistic pathway for the decarbonylation and generation of alkane via alcohol dehydrogenation is proposed. In addition, the in vitro antiproliferative activity of complexes 1-3 was tested against human cervical (HeLa) and human colorectal adenocarcinoma (HT-29) cell lines. Complexes 1-3 showed impressive cytotoxicity against both HeLa (half-maximal inhibitory concentration (IC₅₀) value of 3.84-4.22 μM) and HT-29 cancer cells (IC₅₀ values between 3.3 and 4.5 μM). Moreover, the complexes were comparatively less toxic to noncancerous NIH-3T3 cells. Show less

With the enormous progress in ruthenium complexes as promising anticancer agents after the entry of KP1019, KP1339, and NAMI-A in clinical trials, herein three arene ruthenium(II) NSAID (nonsteroidal anti-inflammatory drugs) complexes viz. [Ru(η⁶-p-cymene)(mef)Cl] (1), [Ru(η⁶-p-cymene)(flu)Cl] (2), and [Ru(η⁶-p-cymene)(dif)Cl] (3) are synthesized, characterized, and reported. Density functional theory (DFT) calculations were performed in support of the obtained experimental results by computing the equilibrium geometries, reactions pathways, relative Gibbs free energy, stability, and reactions barriers of the complexes. The present theoretical study shows that all the proposed structures of the complexes are energetically stable and favorable, and the results obtained are in close accordance with the experiment. Further, the in vitro cytotoxicity of the complexes was explored through MTT assay on MCF-7, Hela, A549, and HEK cell lines. It was found the complex 1 and 2 are significantly cytotoxic toward the MCF-7 cell line. These complexes have also shown a strong affinity toward CT-DNA and proteins (HSA and BSA) as analyzed through spectroscopic techniques. Further investigation of the mechanism of cell death of selected complexes was carried out by various staining, flow cytometry, and gene expression analysis obtained by RT-PCR. Show less

The synthesis and characterization of two half-sandwich complexes of Ru(II) and Ir(III) with thiabendazole as ancillary ligand and their DNA binding ability were investigated using experimental and computational methods. ¹H NMR and acid-base studies have shown that aquo-complexes are the reactive species. Kinetic studies show that both complexes bind covalently to DNA through the metal site and non covalently through the ancillary ligand. Thermal stability studies, viscosity, circular dichroism measurements and quantum chemical calculations have shown that the covalent binding causes breaking of the H-bonding between base pairs, bringing about DNA denaturation and compaction. Additionally, molecular dynamics (MD) simulations and quantum mechanics/molecular mechanics (QM/MM) calculations shed light into the binding features of the Ru(II) and Ir(III) complexes and their respective enantiomers toward double-helical DNA, highlighting the important role played by the NˆN ancillary ligand once the complexes are covalently linked to DNA. Moreover, metal quantification in the nucleus of SW480 colon adenocarcinoma cells were carried out by inductively coupled plasma-mass spectrometry (ICP-MS), both complexes are more internalized than cisplatin after 4 h of exposition. However, in spite of the dramatic changes in the helicity of the DNA secondary structure induced by these complexes and their nuclear localization, antiproliferative studies have revealed that both, Ru(II) and Ir(III) complexes, cannot be considered cytotoxic. This unexpected behavior can be justified by the fast formation of aquo-complexes, which may react with components of the cell culture medium or the cytoplasm compartment in such a way that they may become deactivated before reaching DNA. Show less

A two-photon excited "Ping-Pong" type energy transfer process is for the first time disclosed in a pyrene-modified Ir(iii) cyclometalated complex. The energy transfer, from the singlet excited state of the 4-(pyren-1-yl)-tpy (tpy-py) unit to the Ir(iii) moiety and then back again to the triplet excited state of the tpy-py unit, enhances both the two-photon absorption cross sections and singlet oxygen quantum yield of the complex, and dramatically boosts its two-photon photodynamic activity both in vitro and in 3D multicellular spheroids. Show less

Combination therapy targeting both tumor growth and vascularization is considered to be a cornerstone for colorectal carcinomas (CRC) treatment. However, the major obstacles of most clinical anticancer drugs are their weak selective activity towards cancer cells and inherent inner organs toxicity, accompanied with fast drug resistance development. In our effort to discover novel selective and non-toxic agents effective against CRC, we designed, synthesized and characterized a series of rhenium(I) tricarbonyl-based complexes with increased lipophilicity. Two of these novel compounds were discovered to possess remarkable anticancer, anti-angiogenic and antimetastatic activity in vivo (zebrafish-human HCT-116 xenograft model), being effective at very low doses (1-3 μM). At doses as high as 250 μM the complexes did not provoke toxicity issues encountered in clinical anticancer drugs (cardio-, hepato-, and myelotoxicity). In vivo assays showed that the two compounds exceed the anti-tumor and anti-angiogenic activity of clinical drugs cisplatin and sunitinib malate, and display a large therapeutic window. Show less

The potential chemotherapeutic properties coupled to photochemical transitions make the family of fac-[Re(CO)₃(N,N)X]^0/+ (N,N = a bidentate diimine such as 2,2'-bipyridine (bpy); X = halide, H₂O, pyridine derivatives, PR₃, etc.) complexes of special interest. We have investigated reactions of the aqua complex fac-[Re(CO)₃(bpy)(H₂O)](CF₃SO₃) (1) with potential anticancer activity with the amino acid L-cysteine (H₂Cys), and its derivative N-acetyl-L-cysteine (H₂NAC), as well as the tripeptide glutathione (H₃A), under physiological conditions (pH 7.4, 37 °C), to model the interaction of 1 with thiol-containing proteins and enzymes, and the impact of such coordination on its photophysical properties and cytotoxicity. We report the syntheses and characterization of fac-[Re(CO)₃(bpy)(HCys)]·0.5H₂O (2), Na(fac-[Re(CO)₃(bpy)(NAC)]) (3), and Na(fac-[Re(CO)₃(bpy)(HA)])·H₂O (4) using extended X-ray absorption spectroscopy, IR and NMR spectroscopy, electrospray ionization spectrometry, as well as the crystal structure of {fac-[Re(CO)₃(bpy)(HCys)]}₄·9H₂O (2 + 1.75 H₂O). The emission spectrum of 1 displays a variance in Stokes shift upon coordination of L-cysteine and N-acetyl-L-cysteine. Laser excitation at λ = 355 nm of methanol solutions of 1-3 was followed by measuring their ability to produce singlet oxygen (¹O₂) using direct detection methods. The cytotoxicity of 1 and its cysteine-bound complex 2 was assessed using the MDA-MB-231 breast cancer cell line, showing that the replacement of the aqua ligand on 1 with L-cysteine significantly reduced the cytotoxicity of the Re(I) tricarbonyl complex. Probing the cellular localization of 1 and 2 using X-ray fluorescence microscopy revealed an accumulation of 1 in the nuclear and/or perinuclear region, whereas the accumulation of 2 was considerably reduced, potentially explaining its reduced cytotoxicity. Replacing the aqua ligand with cysteine in the antitumor active fac-[Re(CO)₃(bpy)(H₂O)](CF₃SO₃) complex significantly reduced its cellular accumulation and cytotoxicity against the MDA-MB-213 breast cancer cell line, shifted its maximum emission to considerably higher energies, and decreased its fluorescence quantum yield. Show less

The synthesis and structural characterization of a newly synthesized mononuclear rhodium(iii) complex, Rhtrz, with a ligand (2,2,6-bis((4S,7R)-7,8,8-trimethyl-4,5,6,7-tetrahydro-1H-4,7-methanoindazol-3-yl)pyridine) and a ligand of 1,2,4-triazole, are presented in this paper. The kinetic interactions of the Rhtrz complex with essential biomolecules such as 5-GMP, L-Met, and GSH were examined. The study of the biological interactions was focused on the binding properties of the Rhtrz complex with CT-DNA and serum albumin. These interactions were investigated using UV-vis spectrophotometry, fluorescence spectroscopy, viscosity measurements, thermal denaturation studies, and electrophoresis. Fluorescence competition experiments with site-markers for BSA were used to locate the binding site of the Rhtrz-complex to the BSA. Molecular docking studies were carried out to obtain detailed binding information of the complex with CT-DNA, BSA, and HSA. Furthermore, the apparent distance between the donor (HSA) and acceptor (Rhtrz) was determined using fluorescence resonance energy transfer (FRET). The thermodynamic properties and relative stabilities of the Rhtrz complex were examined, constructing the two model equation by DFT calculations (B3LYP(CPCM)/LANL2DZp). In vitro cytotoxicity and redox status on the human epithelial colorectal cancer cell line (HCT-116) and healthy human fibroblast cell line (MRC-5) were also investigated. Show less

Iron is critical for virtually all organisms, yet major questions remain regarding the systems-level understanding of iron in whole cells. Here, we obtained Mössbauer and EPR spectra of Escherichia coli cells prepared under different nutrient iron concentrations, carbon sources, growth phases, and O2 concentrations to better understand their global iron content. We investigated WT cells and those lacking Fur, FtnA, Bfr, and Dps proteins. The coarse-grain iron content of exponentially growing cells consisted of iron-sulfur clusters, variable amounts of nonheme high-spin FeII species, and an unassigned residual quadrupole doublet. The iron in stationary-phase cells was dominated by magnetically ordered FeIII ions due to oxyhydroxide nanoparticles. Analysis of cytosolic extracts by size-exclusion chromatography detected by an online inductively coupled plasma mass spectrometer revealed a low-molecular-mass (LMM) FeII pool consisting of two iron complexes with masses of ∼500 (major) and ∼1300 (minor) Da. They appeared to be high-spin FeII species with mostly oxygen donor ligands, perhaps a few nitrogen donors, and probably no sulfur donors. Surprisingly, the iron content of E. coli and its reactivity with O2 were remarkably similar to those of mitochondria. In both cases, a "respiratory shield" composed of membrane-bound iron-rich respiratory complexes may protect the LMM FeII pool from reacting with O2 When exponentially growing cells transition to stationary phase, the shield deactivates as metabolic activity declines. Given the universality of oxidative phosphorylation in aerobic biology, the iron content and respiratory shield in other aerobic prokaryotes might be similar to those of E. coli and mitochondria. Show less

Third-generation aromatase inhibitors such as anastrozole (ATZ) and letrozole (LTZ) are widely used to treat estrogen receptor-positive ER+ breast cancers in postmenopausal women. Investigating their ability to coordinate metals could lead to the emergence of a new category of anticancer drug candidates with a broader spectrum of pharmacological activities. In this study, a series of ruthenium (II) arene complexes bearing the aromatase inhibitor anastrozole was synthesized and characterized. Among these complexes, [Ru(η ⁶ -C₆H₆)(PPh₃)(η ¹ -ATZ)Cl]BPh₄ (3) was found to be the most stable in cell culture media, to lead to the highest cellular uptake and in vitro cytotoxicity in two ER+ human breast cancer cell lines (MCF7 and T47D), and to induce a decrease in aromatase activity in H295R cells. Exposure of zebrafish embryos to complex 3 (12.5 μM) did not lead to noticeable signs of toxicity over 96 h, making it a suitable candidate for further in vivo investigations. Show less

Treatment of malignant and non-malignant cultured human cell lines with a cytotoxic IC₅₀ dose of ∼2 μM tris(4,7-diphenyl-1,10-phenanthroline)ruthenium(ii) chloride (RPC2) retards or arrests microtubule motion as tracked by visualizing fluorescently-tagged microtubule plus end-tracking proteins. Immunofluorescent microscopic images of the microtubules in fixed cells show substantial changes to cellular microtubule network and to overall cell morphology upon treatment with RPC2. Flow cytometry with MCF7 and H358 cells reveals only minor elevations of the number of cells in G₂/M phase, suggesting that the observed cytotoxicity is not tied to mitotic arrest. In vitro studies with purified tubulin reveal that RPC2 acts to promote tubulin polymerization and when imaged by electron microscopy, these microtubules look normal in appearance. Isothermal titration calorimetry measurements show an associative binding constant of 4.8 × 10⁶ M^-1 for RPC2 to preformed microtubules and support a 1 : 1 RPC2 to tubulin dimer stoichiometry. Competition experiments show RPC2 does not compete for the taxane binding site. Consistent with this tight binding, over 80% of the ruthenium in treated cells is co-localized with the cytoskeletal proteins. These data support RPC2 acting as an in vivo microtubule stabilizing agent and sharing many similarities with cells treated with paclitaxel. Show less

Of late, cancer has become a terrible disease affecting people throughout the world. Keeping this in mind, we tried to design drugs that are more lipophilic, target-specific, water-soluble, cytoselective and fluorescent. In this regard, we reported novel ruthenium(ii)-p-cymene imidazophenanthroline scaffolds as effective DNA targeting agents. The planarity of imidazophenanthroline ligands caused the Ru(ii) complex to be a good intercalator. An extended π-electronic conjugation was introduced in the imidazophenanthroline moieties through the Suzuki and Sonogashira coupling reactions. Here, we synthesized nine Ru(ii) complexes (16a-b, 17a-d, and 19a-c). Among these, [(η⁶-p-cymene)RuCl(K²-N,N-2-(4'-methyl-[1,1'-BIphenyl]-4-yl)-1H-imidazo[4,5-f][1,10]phenanthroline)]·PF₆ (16b) exhibited the best potency and selectivity with excellent cellular uptake; [(η⁶-p-cymene)RuCl(K²-N,N-2-(4-(phenylethynyl)phenyl)-1H-imidazo[4,5-f][1,10]phenanthroline)]·PF₆ (17a) acted as a cytoselective probe for live cell imaging. Show less

The monocationic chloro complexes containing chelating N∩N ligands: [(η⁶-p-cymene)Ru(L1-4)Cl]⁺ (1-4), where L1 = 4-methyl-1,10-phenantroline, L2 = dipyrido[3,2-a:2',3'-c]phenazine, L3 = 11-chloro-dipyrido[3,2-a:2',3'-c]phenazine, L4 = 11-nitro-dipyrido[3,2-a:2',3'-c]phenazine; p-cymene = 1-methyl-4-isopropylbenzene) have been prepared and characterized as the hexafluorophosphate salts. The biological activity of 1-4 has been investigated in selected 2D monolayer cell cultures (A549, PANC-1, MDA-MB-231, MRC-5). All investigated ruthenium complexes showed similar or even better cytotoxicity to cisplatin. However, there was no significant reduction in growth of PANC-1 cells in a 3D cell culture of multicellular tumor spheroids (MCTS) after treatment with 2-4, while the cisplatin treatment induced retardation in MCTS growth. Flow cytometry analysis of the cell cycle of PANC-1 cells shows that 3 caused changes of cell cycle phase distribution characterized by slight accumulation of cells in the G2-M phase. Absence of the Sub-G1 phase in the cell cycle of the treated cells indicated that there was no fragmentation of DNA for the analyzed time intervals (48 and 72 h treatment). Fluorescent microscopy, after acridine orange/ethidium bromide staining, revealed that the investigated ruthenium complexes induced some characteristics of apoptotic morphology (shrinking and condensation of chromatin) with notably preserved integrity of the plasma membrane. Investigation of cellular uptake and DNA - fraction accumulation performed by inductively coupled plasma mass spectrometry in PANC-1 cells with equimolar concentrations (5 μM) of 2-4 and cisplatin showed more efficient cellular uptake and DNA - fraction accumulation of complex 3 compared to complexes 2 and 4. Show less

Three new ruthenium(II)-arene complexes with pyrido[2',3':5,6]pyrazino[2,3-f][1, 10]phenanthroline (ppf) of general formula: C1 ([(ƞ⁶-benzene)Ru(ppf)Cl]PF₆, C2 ([(ƞ⁶-toluene)Ru(ppf)Cl]PF₆) and C3 ([(ƞ⁶-p-cymene)Ru(ppf)Cl]PF₆) have been synthesized. The structures of complexes were determined by elemental analysis, IR, ESI-MS, as well as with ¹H and ¹³C NMR spectroscopy. Cytotoxic activity has been evaluated in three different human neoplastic cell lines (A549, A375, LS 174T) and in one human non-tumor cell line (MRC-5), by the MTT assay. Complexes C1-C3 showed IC₅₀ values in the micromolar range below 100 µM. Complex C3, carrying ƞ⁶-p-cymene as the arene ligand, exhibited cytoselective activity toward human malignant melanoma A375 cells (IC₅₀ = 15.8 ± 2.7 µM), and has been selected for further analyses of its biological effects. Drug-accumulation study performed in the A375 cells disclosed that C3 possess lower ability of entering the cells compared to cisplatin and distributes approximately equally in the cytosol and membrane/organelle fraction of cells. Investigations in the 3D model of A375 cells, disclosed different effects of the complex C3 and cisplatin on growth of multicellular tumor spheroids (MCTSs). While the size of cisplatin-treated MCTSs decreased with time, MCTSs treated with C3 continued to growth. Differences in structural organization and biological activity of this type of ruthenium(II)-arene complexes versus cisplatin in A375 malignant melanoma cells pointed out their different modes of action, and necessity for further biological studies and optimizations for potential applications. Show less

Half-sandwich ruthenium(ii) complexes [(η⁶-p-cymene)Ru(C^N)-(X)]^0/+ (X = Cl, py or 4-NMe₂-py) containing a cyclometalated 2-ppy or 1-ppz with a non-coordinated CHO group as a handle for further functionalization have been synthesized to achieve selective cytotoxicity to cancer cells, the more potent compounds acting as proteosynthesis inhibitors; this is a new mode of action for half-sandwich metal complexes. Show less

A series of ruthenium(ii) complexes with N-heterocyclic carbene ligands were successfully synthesized by transmetalation reactions between silver(i) N-heterocyclic carbene complexes and [RuCl₂(p-cymene)]₂ in dichloromethane under Ar conditions. All new compounds were characterized by spectroscopic and analytical methods. These ruthenium(ii)-NHC complexes were found to be efficient precatalysts for the transfer hydrogenation of ketones by using 2-propanol as the hydrogen source in the presence of KOH as a co-catalyst. The antibacterial activity of ruthenium N-heterocyclic carbene complexes 3a-f was measured by disc diffusion method against Gram positive and Gram-negative bacteria. Compounds 3d exhibited potential antibacterial activity against five bacterial species among the six used as indicator cells. The product 3e inhibits the growth of all the six tested microorganisms. Moreover, the antioxidant activity determination of these complexes 3a-f, using 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azinobis-3-ethylbenzothiazoline-6-sulphonic acid (ABTS) as reagent, showed that compounds 3b and 3d possess DPPH and ABTS antiradical activities. From a concentration of 1 mg ml^-1, these two complexes presented a similar scavenging activity to that of the two used controls gallic acid (GA) and butylated hydroxytoluene (BHT). From a concentration of 10 mg ml^-1, the percentage inhibition of complexes 3b and 3d was respectively 70% and 90%. In addition, these two Ru-NHC complexes exhibited antifungal activity against Candida albicans. Investigation of the anti-acetylcholinesterase activity of the studied complexes showed that compounds 3a, 3b, 3d and 3e exhibited good activity at 100 μg ml^-1 and product 3d is the most active. In a cytotoxicity study the complexes 3 were evaluated against two human cancer cell lines MDA-MB-231 and MCF-7. Both 3d and 3e complexes were found to be active against the tested cell lines showing comparable activity with examples in the literature. Show less

Ru(ii) polypyridine complexes which can undergo photo-induced ligand dissociation and subsequent DNA covalent binding may potentially serve as photoactivated chemotherapeutic (PACT) agents. In this paper, three fluorinated dppz ligand coordinated Ru(ii) complexes (2-4) containing four monodentate pyridine ligands were studied. All complexes released one pyridine and covalently bound to DNA upon 470 nm irradiation. Compared with the parent complex [Ru(dppz)(py)₄]²⁺ (1), 2-4 displayed enhanced phototoxicity but diminished dark cytotoxicity, more favorable for PACT application. Complex 3 is the most efficient one with IC₅₀ values of about 8 μM toward HeLa and SKOV-3 cell lines, and also has a much higher IC₅₀ value toward normal L-02 cells. Our results indicate that fluorination on the retaining ligand may be an efficient way to improve the drug activity of Ru(ii) PACT agents. Show less

The use of ruthenium complexes as chemotherapeutic agents has been recently explored as one of the alternatives to conventional treatments. In the present study, two Ru(ii) polypyridyl complexes were synthesized and characterized: a strained [Ru(bipy)₂(BC)]Cl₂ (complex 1) where [bipy = 2,2'-bipyridine and BC = bathocuproine] along with the unstrained control [Ru(bipy)₂(phen)]Cl₂ (complex 2) where [phen = 1,10-phenanthroline]. The photophysical and photochemical analyses proved that unlike the photostable complex 2, complex 1 ejected both bipy and BC ligands at a ratio of 3 : 1 respectively. Results showed that the activity of complex 1 was significantly enhanced upon photoactivation. The response was however particularly significant in B16-F10 melanoma cells where phototoxicity index (PI = IC₅₀ dark/IC₅₀ light) was >900. When compared to cisplatin, the photoproducts were more potent against all tested cell lines, implying that the complex acquired significant chemotherapeutic potential upon irradiation. Cellular uptake of complex 1 and the free BC ligand were found to be significantly facilitated as evidenced by 400-600 fold increase in concentration of the compounds inside the cells relative to the extracellular culture medium. Complex 2 exhibited 35 times lower cellular concentration relative to complex 1. Flow cytometry and plasmid DNA gel electrophoresis measurements showed that complex 1 interacts with DNA inducing apoptosis in the dark and either late-apoptosis or necrosis upon irradiation. These findings corroborate the importance of lipophilic ligands such as BC to enhance uptake and subsequently improve the photochemotherapy potential of Ru(ii) polypyridyl complexes. Show less

Ru(ii) polypyridyl complexes which can undergo photo-induced ligand dissociation and DNA covalent binding are considered as potential photoactivated chemotherapeutic (PACT) agents. Herein four pyridine-2-sulfonate (py-SO3-) ligand based Ru(ii) complexes [Ru(N-N)2(py-SO3)]+ (1-4) were synthesized and studied. All the complexes can undergo fast py-SO3- ligand dissociation and DNA covalent binding upon visible light irradiation. However, only complex 4 exhibited high photo-induced anticancer activities towards a series of cancer cells, with half maximal inhibitory concentration (IC50) values in 100-300 nM regions and phototoxicity index (PI) values of about 100. In particular, complex 4 can also kill cisplatin resistant SKOV-3 and A549 cancer cells with IC50 values in 200-400 nM regions and PI values of about 50, which should be the first report of Ru(ii) based PACT agents that are also effective towards cisplatin resistant cancer cells. Complex 4 exhibited much higher cell uptake and nuclear accumulation levels, which may be the main reasons for its high anticancer activities. The in vivo anticancer experiments indicated that complex 4 can inhibit tumor growth significantly with fewer side effects. Our results may provide guidelines for developing novel photoactivatable Ru(ii) anticancer agents. Show less

Two new complexes of Ru(II) with mixed ligands were prepared: [Ru(bpy)₂smp](PF₆) (1) and [Ru(phen)₂smp](PF₆) (2), in which smp = sulfamethoxypyridazine; bpy = 2,2'-bipyridine; phen = 1,10-phenanthroline. The complexes have been characterized by elemental and conductivity analyses; infrared, NMR, and electrospray ionization mass spectroscopies; and X-ray diffraction of single crystal. Structural analyses reveal a distorted octahedral geometry around Ru(II) that is bound to two bpy (in 1) or two phen (in 2) via their two heterocyclic nitrogens and to two nitrogen atoms from sulfamethoxypyridazine-one of the methoxypyridazine ring and the sulfonamidic nitrogen, which is deprotonated. Both complexes inhibit the growth of chronic myelogenous leukemia cells. The interaction of the complexes with bovine serum albumin and DNA is described. DNA footprinting using an oligonucleotide as substrate showed the complexes' preference for thymine base rich sites. It is worth notifying that the complexes interact with the Src homology SH3 domain of the Abl tyrosine kinase protein. Abl protein is involved in signal transduction and implicated in the development of chronic myelogenous leukemia. Nuclear magnetic resonance (NMR) studies of the interaction of complex 2 with the Abl-SH3 domain showed that the most affected residues were T79, G97, W99, and Y115. Show less

Targeted delivery of clinically approved anticancer drug to tumor sites is an effective way to achieve enhanced drug efficacy as well as reduced side effects and toxicity. Here bicalutamide is caged by the Ru(II) center through the nitrile group, and three photoactive Ru(II) complexes were designed and synthesized. Docking study showed that the ruthenium(II) fragments can effectively block the binding of complexes 1-3 with AR (androgen receptor) owing to the large steric structures, thus bicalutamide in complexes 1-3 could not interact with AR-LBD (ligand binding domain). Once irradiation with blue light (465nm), complexes 1-3 can release bicalutamide and anticancer Ru(II) fragments, which possesses dual-action of AR binding and DNA interaction simultaneously. In vitro cytotoxicity study on these complexes further confirmed that complexes 1-3 exhibited considerable cytotoxicity upon irradiation with blue light. Significantly, complex 3 could be activated at 660nm, which greatly increases the scope of complex 3 to treat deeper within tissue. Theoretical calculations showed that the lowest singlet excitation energy of complex 3 is lower than those of complexes 1-2, which explains the experimental results well. Moreover, the ³MC (metal centered) states of these complexes are more stable than their ³MLCT (metal to ligand charge transfer) states, indicating that the photoactive processes of these complexes are likely to result in ligand dissociation. Show less

Eight new organometallic Ru(II)-arene complexes of the type [RuCl₂(η⁶-arene)(η¹-S-aroylthiourea)] (arene = p-cymene or benzene) were synthesized in order to evaluate the effect of the arene moiety and the substituent of the aroylthiourea ligand on the cytotoxicity of the complexes. The ligands (L1 and L2) and complexes (1-8) were characterized using analytical and spectroscopic (UV-visible, infrared, ¹H NMR, ¹³C NMR, and mass) methods. The structure of the ligands (L1 and L2) and complexes (1 and 3-6) was obtained from single-crystal X-ray diffraction studies. The cytotoxicity of the complexes was evaluated against four different cancer cell lines: MCF-7 (breast), COLO 205 (colon), A549 (lung), and IMR-32 (neuroblastoma). All the complexes showed good cytotoxicity and the highest was in the IMR-32 cell line, which articulates the specificity of these complexes toward the IMR-32 cancer cell line. The complexes 5, 7, and 8 exhibited remarkable cytotoxicity in the entire cancer cell lines tested, which was comparable with the standard drug, cisplatin. The anticancer mechanism of the complexes 3 and 7 in IMR-32 cells was evaluated by bright-field microscopy, intracellular reactive oxygen species (ROS), mitochondrial membrane potential (MMP), DNA damage, and caspase-3 analyses. The cells treated with the complexes showed upregulated caspase-3 compared to the control, and it was found that ROS and MMP were dose-dependent on analysis. Also, bright-field microscopy and 4',6-diamidino-2-phenylindole (DAPI) staining have correspondingly shown cellular membrane blebbing and DNA damage, which were morphological hallmarks of apoptosis. The study concluded that the complexes promoted the oxidative stress-mediated apoptotic death of the cancer cells through the generation of intracellular ROS, depletion of MMP, and damage of the nuclear material. Show less

Glioma stem cells (GSCs) are thought to be responsible for the recurrence and invasion of glioblastoma multiform (GBM), which have been evaluated and exploited as the therapeutic target for GBM. Cyclometalated iridium(III) complexes have been demonstrated as the potential anticancer agents, however, their antitumor efficacies against GSCs are still unknown. Herein, we investigated the antitumor activity of two cyclometalated iridium(III) complexes [Ir(ppy)₂L](PF₆) (Ir1) and [Ir(thpy)₂L](PF₆) (Ir2) (ppy = 2-phenylpyridine, thpy = 2-(2-thienyl)pyridine and L = 4,4'-Bis(hydroxymethyl)-2,2'-bipyridine) against GSCs. The results clearly indicate that Ir1 and Ir2 kill GSCs selectively with IC₅₀ values ranging from 5.26-9.05 μM. Further mechanism research display that Ir1 and Ir2 can suppress the proliferation of GSCs, penetrate into GSCs efficiently, localize to mitochondria, and induce mitochondria-mediated apoptosis, including the loss of mitochondrial membrane (MMP), elevation of intracellular reactive oxygen species (ROS) and caspases activation. Moreover, Ir1 and Ir2 can destroy the GSCs self-renewal and unlimited proliferation capacity by affecting the GSCs colony formation. According our knowledge, this is the first study to investigate the anti-GSCs properties of cyclometalated iridium(III) complexes. Show less