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Two new Ru(II)-based complexes containing 2-thiouracil derivatives, known as 2-thiouracil (2TU) and 6-methyl-2-thiouracil (6m2TU), were synthesized using cis,trans-[RuCl₂(PPh₃)₂(bipy)] as a precursor. The obtained compounds with a general formula trans-[Ru(2TU)(PPh₃)₂(bipy)]PF₆ (1) and trans-[Ru(6m2TU)(PPh₃)₂(bipy)]PF₆ (2) were characterized by analytical techniques such as NMR, UV-vis, and IR spectroscopies, elementary analysis, mass spectrometry, and single-crystal X-ray diffraction. Moreover, the investigation of the complexes-DNA interaction were carried out using spectrophotometric titrations and showed that the complexes present a weak interaction with this biomolecule. The compounds were evaluated against HL-60, K-562, HepG2, and B16-F10 cancer cells and against noncancer cells (PBMCs). The results of the biological assay revealed that complex 2 is more promising than complex 1. Finally, the present study suggests that complexes 1 and 2 causes cell death by apoptosis, significantly increasing the percentage of apoptotic HL-60 cells, in which the compounds altered the cell cycle, reducing the cells in G₁/G₀, G₂/M, and S phases. Show less

Title: Green Light-Triggered Photocatalytic Anticancer Activity of Terpyridine-Based Ru(II) Photocatalysts. Abstract: The relentless increase in drug resistance of platinum-based chemotherapeutics has opened the scope for other new cancer therapies with novel mechanisms of action (MoA). Recently, photocatalytic cancer therapy, an intrusive catalytic treatment, is receiving significant interest due to its multitargeting cell death mechanism with high selectivity. Here, we report the synthesis and characterization of three photoresponsive Ru(II) complexes, viz., [Ru(ph-tpy)(bpy)Cl]PF6 (Ru1), [Ru(ph-tpy)(phen)Cl]PF6 (Ru2), and [Ru(ph-tpy)(aip)Cl]PF6 (Ru3), where, ph-tpy = 4'-phenyl-2,2':6',2″-terpyridine, bpy = 2,2'-bipyridine, phen = 1,10-phenanthroline, and aip = 2-(anthracen-9-yl)-1H-imidazo[4,5-f][1,10] phenanthroline, showing photocatalytic anticancer activity. The X-ray crystal structures of Ru1 and Ru2 revealed a distorted octahedral geometry with a RuN5Cl core. The complexes showed an intense absorption band in the 440-600 nm range corresponding to the metal-to-ligand charge transfer (MLCT) that was further used to achieve the green light-induced photocatalytic anticancer effect. The mitochondria-targeting photostable complex Ru3 induced phototoxicity with IC50 and PI values of ca. 0.7 μM and 88, respectively, under white light irradiation and ca. 1.9 μM and 35 under green light irradiation against HeLa cells. The complexes (Ru1-Ru3) showed negligible dark cytotoxicity toward normal splenocytes (IC50s > 50 μM). The cell death mechanistic study revealed that Ru3 induced ROS-mediated apoptosis in HeLa cells via mitochondrial depolarization under white or green light exposure. Interestingly, Ru3 also acted as a highly potent catalyst for NADH photo-oxidation under green light. This NADH photo-oxidation process also contributed to the photocytotoxicity of the complexes. Overall, Ru3 presented multitargeting synergistic type I and type II photochemotherapeutic effects. Show less

A series of arene Ru(II) complexes, [(η⁶-MeC₆H₅)Ru(L)Cl]Cl, (L=o-ClPIP, 1; m-ClPIP, 2 and p-ClPIP, 3) (o-ClPIP=2-(2-chlorophenyl)imidazo[4,5-f][1,10]phenanthroline; m-ClPIP=2-(3-chlorophenyl)imidazo[4,5-f][1,10]phenanthroline; p-ClPIP=2-(4-chlorophenyl)imidazo[4,5-f][1,10]phenanthroline) was synthesized and investigated as a potential apoptosis inducer in chemotherapy. Spectroscopy and molecular docking simulations show that 1 exhibits moderated binding affinity to KRAS G-quadruplex DNA by groove mode. Further, in vitro studies reveal that 1 displays inhibitory activity against MCF-7 growth with IC₅₀ = 3.7 ± 0.2 μM. Flow cytometric analysis, comet assay, and immunofluorescence confirm that 1 can induce the apoptosis of MCF-7 cells and G0/G1 phase arrest through DNA damage. In summary, the prepared arene Ru(II) complexes can be developed as a promising candidate for targeting G-quadruplex structure to induce the apoptosis of breast cancer cells via binding and stabilizing KRAS G-quadruplex conformation on oncogene promoter. Show less

Cisplatin-based chemotherapy is a common regimen for bladder cancer, a life-threatening cancer with more than 500,000 new cases worldwide annually. Like many other metallodrugs, cisplatin causes severe side effects for its general toxicity. Organoruthenium is known for its structural stability, good anticancer activity, and possible low general toxicity. Here, we have prepared and characterized a series of water-soluble ruthenium-arene complexes with N,N'-chelating ligands: [Ru(II)-η⁶-arene-(4,4'-(X)₂-2,2'-bipyridine)Cl]Cl (arene = p-cymene, X = C₄H₉ (1), COOH (2), COOCH₃ (3), COOC₂H₅ (4); arene = benzene, X = C₄H₉ (5), COOCH₃ (6), COOC₂H₅ (7)). These complexes are carefully characterized using single-crystal X-ray diffraction, UV-vis, IR, ¹H NMR, and MALDI-TOF MS spectroscopy. Their DFT-calculated structural and thermodynamic properties are consistent with the experimental observations. Biophysicochemical studies of complex interaction with CTDNA and BSA supported by molecular docking simulations reveal suitable properties of 1-7 as anticancer agents. Cytotoxicities of 1-7 are evaluated on healthy human MCF-10a-breast epithelial and African green monkey Vero cells, and carcinoma human HepG-2-hepatic, T24-bladder, and EAhy-926-endothelial cells. All complexes exhibit much higher cytotoxicity for T24 than cisplatin. Particularly, 1 and 2 are also highly selective toward T24. Fluorescence imaging and flow cytometry demonstrate that 1 and 2 penetrate T24 cell membrane and induce early apoptosis at their respective IC₅₀ concentrations, which ultimately lead to cell death. Statistical analysis suggests that the order of importance for T24 cell antiproliferation is protein binding, Log p, Ru-Cl bond length, while DNA binding is the least important. This study is the first to report the anti-bladder cancer efficacy of Ru-arene-2,2'-bipyridine complexes, and may provide insights for rational design of organoruthenium drugs in the enduring search for new chemotherapeutic agents. Show less

In this study, half-sandwich Ru(II) complexes containing acylthiourea ligands of the general type [Ru(η⁶-p-cymene)(PPh₃)(S)Cl]PF₆ (1m-6m) and [Ru(η⁶-p-cymene)(PPh₃)(S-O)]PF₆ (1b-6b) where S/S-O = N',N'-disubstituted acylthiourea were synthesized and characterized (via elemental analyses, IR spectroscopy, ¹H NMR spectroscopy, ¹³C{¹H} NMR spectroscopy, and X-ray diffractometry), and their cytotoxic activity was evaluated. The different coordination modes of the acylthiourea ligands, monodentately via S (1m-6m) and bidentately via S,O (1b-6b), to ruthenium were modulated from different synthetic routes. The cytotoxicity of the complexes was evaluated in five human cell lines (DU-145, A549, MDA-MB-231, MRC-5, and MCF-10A) by MTT assay. The IC₅₀ values for prostate cancer cells (2.89-7.47 μM) indicated that the complexes inhibited cell growth, but that they were less cytotoxic than cisplatin (2.00 μM). Unlike for breast cancer cells (IC₅₀ = 0.28-0.74 μM) and lung cancer cells (IC₅₀ = 0.51-1.83 μM), the complexes were notably more active than the reference drug, and a remarkable selectivity index (SI 4.66-19.34) was observed for breast cancer cells. Based on both the activity and selectivity, complexes 5b and 6b, as well as their respective analogous complexes in the monodentate coordination 5m and 6m, were chosen for further investigation in the MDA-MB-231 cell line. These complexes not only induced morphology changes but also were able to inhibit colony formation and migration. In addition, the complexes promoted cell cycle arrest at the sub-G₁ phase inducing apoptosis. Interaction studies by viscosity measurements, gel electrophoresis, and fluorescence spectroscopy indicated that the complexes interact with the DNA minor groove and exhibit an HSA binding affinity. Show less

A novel series of ionic Ru(II) arene Cp* sandwich complexes has been synthesized and characterized. Screening results for cytotoxicity against a range of human tumor cell lines and normal human cells indicate that the complexes show promising anticancer activity, which varies with changes in the arene ligand and the anionic counterion. Show less

Background/objectives

Breast cancer (BC) remains one of the most prevalent and deadly cancers worldwide, with limited access to advanced treatments in developing regions. There is a critical need for novel therapies with unique mechanisms of action, especially to overcome resistance to conventional platinum-based drugs. This study investigates the anticancer potential of the ruthenium complex Bis(quinolin-8-olato)bis(triphenylphosphine)ruthenium(II) (Ru(quin)₂) in ER-positive (T47D) and triple-negative (MDA-MB-231) BC cell lines.

Results

Ru(quin)₂ demonstrated dose-dependent cytotoxicity, with IC50 values of 48.3 μM in T47D cells and 45.5 μM in MDA-MB-231 cells. Its cytotoxic effects are primarily driven by apoptosis, as shown by increased BAX expression, enhanced caspase-3 activity, reduced Aurora B kinase levels, and elevated histone release. Ru(quin)₂ also induced autophagy, evidenced by LC3-I to LC3-II conversion and reduced SQSTM1, partially mediated through MAPK signaling. Furthermore, Ru(quin)₂ induced G0/G1 cell cycle arrest by downregulating cyclin D1, CDK4, and CDK6, alongside upregulation of the CDK inhibitor p21.

Conclusions

Ru(quin)₂ emerges as a potent candidate for BC treatment, with multiple mechanisms of action involving apoptosis, autophagy, and cell cycle arrest. Further studies are warranted to elucidate its detailed molecular mechanisms and evaluate its therapeutic potential in vivo, moving toward clinical applications for both ER-positive and triple-negative BC management. Show less

In recent years, photodynamic therapy (PDT) has emerged as a promising alternative to classical chemotherapy for treating cancer. PDT is based on a nontoxic prodrug called photosensitizer (PS) activated by light at the desired location. Upon irradiation, the PS reacts with the oxygen present in the tumor, producing cytotoxic reactive oxygen species (ROS). Compounds with highly conjugated π-bond systems, such as porphyrins and chlorins, have proven to be excellent light scavengers, and introducing a metal atom in their structure improved the generation of ROS. In this work, a series of tetrapyrrole-ruthenium(II) complexes derived from protoporphyrin IX and the commercial drug verteporfin were designed as photosensitizers for PDT. The complexes were almost nontoxic on human gastric cancer cells under dark conditions, revealing remarkable cytotoxicity upon irradiation with light. The ruthenium atom in the central cavity of the chlorin ligand allowed combined mechanisms in photodynamic therapy, as both singlet oxygen and superoxide radicals were detected. Additionally, one complex produced large amounts of singlet oxygen under hypoxic conditions. Biological assays demonstrated that the ruthenium derivatives caused cell death through a caspase 3 mediated apoptotic pathway and via CHOP, an endoplasmic reticulum stress-inducible transcription factor involved in apoptosis and growth arrest. Show less

Novel metal complexes have received great attention in the last decades due to their potential anticancer activity. Notably, ruthenium-based complexes have emerged as good alternative to the currently used platinum-based drugs for cancer therapy, providing less toxicity and side effects to patients. Glioblastoma is an aggressive and invasive type of brain tumor and despite of advances is the field of neurooncology there is no effective treatment until now. Therefore, we sought to investigate the potential antiproliferative activity of phosphine-ruthenium-based complexes on human glioblastoma cell lines. Due to its octahedral structure as opposed to the square-planar geometry of platinum(II) compounds, ruthenium(II) complexes exhibit different structure-function relationship probably acting through a different mechanism from that of cisplatin beyond their ability to bind DNA. To better improve the pharmacological activity of metal complexes we hypothesized that neutron activation of ruthenium in the complexes would allow to decrease the effective concentration of the compound needed to kill tumor cells. Herein we report on the effect of unmodified and neutron activated phosphine ruthenium II complexes on glioblastoma cell lines carrying wild-type and mutated p53 tumor suppressor gene. Induction of apoptosis/authophagy as well as generation of reactive oxygen species were determined. The phosphine ruthenium II complexes tested were highly active against glioblastoma cell lines inducing cell death both through apoptosis and autophagy in a p53 independent fashion. Neutron activation of ruthenium compounds rendered them more active than their original counterparts suggesting a new strategy to improve the antitumor activity of these compounds. Show less

Title: Comparative studies on Abstract: On the basis of our previous comparative studies on the DNA binding of a pair of ruthenium(II) complex enantiomers, Δ-[Ru(bpy)2PBIP]2+ and Λ-[Ru(bpy)2PBIP]2+ {bpy = 2,2'-bipyridine, PBIP = 2-(4-bromophenyl)imidazo[4,5-f]1,10-phenanthroline}, in this study, their antitumor activities and mechanisms were further investigated comparatively. The cytotoxicity assay demonstrated that both the enantiomers exerted selective antiproliferative effects on cancer cell lines A2780 and PC3. Fluorescence localization experiments suggested that both the enantiomers effectively permeated the nucleus of HeLa cells and co-localized with DNA, resulting in their DNA damage and apoptosis. Flow cytometry experiments showed that the apoptosis was enhanced by increasing the concentration of each enantiomer. Western blotting analyses indicated that both extrinsic and intrinsic apoptosis pathways were activated by the two enantiomers. miRNA microarray analyses displayed that both the enantiomers up- and downregulated multiple miRNAs, some of which were predicted to be associated with carcinogenesis. The above experimental results also showed that the Δ-enantiomer exerted a more potent antitumor activity, a higher efficiency of entering cancer cells and a stronger apoptosis-inducing effect compared with the Λ-enantiomer. Combined with the previously published research results, experimental results from this study implied that the antitumor activity of a metal complex might have originated from the conformation change of DNA in tumor cells caused by the intercalation of the complex, that the antitumor mechanism of a metal complex could be related to its DNA-binding mode, and that the antitumor efficiency of a metal complex could result from its DNA-binding strength. Show less

Title: Analysis of antiproliferative activity of new half-sandwich arene Ru(II) thiophene based aroylhydrazone complexes. Abstract: Efforts in researching the efficient anti-tumor properties of three novel arene ruthenium(II) complexes incorporating thiophene-based aroylhydrazone ligands have been undertaken. The complexes' elemental composition was [(η6-p-cymene)Ru(L)Cl]. They were comprehensively characterized through elemental and spectroscopic analyses (FT-IR, UV-vis, NMR, and HR-MS). Single crystal X-ray diffraction studies revealed a pseudo-octahedral geometry with bidentate coordination of the ligands in a representative complex. The in vitro assessment of the complexes' cancer cell growth inhibition was conducted using the MTT assay against A549 (human lung carcinoma), HeLa (human cervical carcinoma), HuH-7 (hepatocellular carcinoma), and NIH-3T3 (mouse fibroblast non-cancerous cell line). Results indicated significant cytotoxicity across all cancer cell lines, with IC50 concentrations of complex 2 being 6.8 μM for A549, 11.6 μM for HeLa, and 9.4 μM for HuH-7, compared to cisplatin with IC50 values of 18.9 μM, 17.68 μM, and 24 μM respectively. Notably, complex 2 demonstrated particularly promising cytotoxicity against all tested cancerous cell lines. Fluorescent staining analysis such as acridine orange/ethidium bromide (AO-EB) and HOECHST 33342 revealed cell death mechanisms involving membrane disintegration and nuclear condensation following treatment with complex 2. Further studies were conducted to measure reactive oxygen species (ROS) levels using the dichlorodihydrofluorescein diacetate (DCFH-DA) assay, and mitochondrial membrane potential (MMP) was assessed using the JC-1 dye assay. These studies demonstrated that complex 2 increased ROS levels, decreased membrane potential, and promoted mitochondrial dysfunction-mediated cell death pathways. Additionally, flow cytometry analysis, utilizing dual staining of Annexin V-FITC and propidium iodide (PI), was employed to quantitatively study apoptosis induction. Show less

Compounds capable of light-triggered cytotoxicity are appealing potential therapeutics, because they can provide spatial and temporal control over cell killing to reduce side effects in cancer therapy. Two simple homoleptic Ru(II) polypyridyl complexes with almost-identical photophysical properties but radically different physiochemical properties were investigated as agents for photodynamic therapy (PDT). The two complexes were identical, except for the incorporation of six sulfonic acids into the ligands of one complex, resulting in a compound carrying an overall -4 charge. The negatively charged compound exhibited significant light-mediated cytotoxicity, and, importantly, the negative charges resulted in radical alterations of the biological activity, compared to the positively charged analogue, including complete abrogation of toxicity in the dark. The charges also altered the subcellular localization properties, mechanism of action, and even the mechanism of cell death. The incorporation of negative charged ligands provides a simple chemical approach to modify the biological properties of light-activated Ru(II) cytotoxic agents. Show less

Introduction: Ruthenium(II) complexes have emerged recently as candidates for anti-cancer therapy, where activity is related to lipohilicity, cellular localization, and specific interactions with biomolecules. Methods: In this work, two novel complexes were synthesized and are reported based on the [Ru(phen)₂(dipyrido[3,2-f:2',3'-h]quinoxaline]²⁺ framework. Results: Compared to the parent complex, annealing of cyclopenteno and cyclohexeno rings to the extended ligand substantially increased cytotoxicity towards a number of cancer cell lines, and induced apoptosis. The complexes localize in the nuclei of cancer cells and co-locate with DAPI on DNA. DNA binding studies show that both complexes bind strongly to DNA and one complex intercalates DNA like the parent, whilst the other appears to have multiple modes of interaction. Discussion: It is likely that the increased lipophilicity of the novel complexes is a key factor for increasing their cytotoxicity, rather than their DNA binding mode. Show less

Platinum-based complexes are one of the most successful chemotherapeutic agents having a significant ground in cancer chemotherapy despite their side effects. During the past few decades, Ru(II) complexes have been emerging as efficient alternatives owing to their promising activities against platinum-resistant cancer. The pathway of action, lipophilicity, and cytotoxicity of a Pt or Ru complex may be tuned by varying the attached ligands, the coordination mode, and the leaving group. In this work, we report a family of Pt(II) and Ru(II) complexes (1-5) of three N,O and N,N donor-based trimethoxyanilines containing Schiff bases with the general formula [Pt^II(L)(DMSO)Cl], [Ru^II(L)(p-cymene)Cl], [Ru^II(L)(p-cymene)Cl]⁺, and [Pt^II(L)Cl₂]. All of the complexes are characterized by different analytical techniques. ¹H NMR and electrospray ionization mass spectrometry (ESI-MS) data suggest that the N,O-coordinated Pt(II) complexes undergo slower aquation compared to the Ru(II) analogues. The change of the coordination mode to N,N causes the Ru complexes to be more inert to aquation. The N,O-coordinating complexes show superiority over N,N-coordinating complexes by displaying excellent in vitro antiproliferative activity against different aggressive cancer cells, viz., triple-negative human metastatic breast adenocarcinoma MDA-MB-231, human pancreatic carcinoma MIA PaCa-2, and hepatocellular carcinoma Hep G2. In vitro cytotoxicity studies suggest that Pt(II) complexes are more effective than their corresponding Ru(II) analogues, and the most cytotoxic complex 3 is 10-15 times more toxic than the clinical drugs cisplatin and oxaliplatin against MDA-MB-231 cells. Cellular studies show that all of the N,O-coordinated complexes (1-3) initiate disruption of the microtubule network in MDA-MB-231 cells in a dose-dependent manner within 6 h of incubation and finally lead to the arrest of the cell cycle in the G2/M phase and render apoptotic cell death. The disruption of the microtubule network affects the agility of the cytoskeleton rendering inhibition of tyrosine phosphorylation of vascular endothelial growth factor receptor 2 (VEGFR2), a key step in angiogenesis. Complexes 1 and 2 inhibit VEGFR2 phosphorylation in a dose-dependent fashion. Among the Pt(II) and Ru(II) complexes, the former displays higher cytotoxicity, a stronger effect on the cytoskeleton, better VEGFR2 inhibition, and strong interaction with the model nucleobase 9-ethylguanine (9-EtG). Show less

Ruthenium complexes with bioactive ligands are becoming promising substitutes for platinum complexes due to their precise action against various cancers. In the present study, the synthesis of three new arene Ru(ii) complexes containing new carbazole-based hydrazone ligands of general formula [(η6-benzene)Ru(L)Cl] (1-3; L = carbazolone benzhydrazone ligands), and their anticancer properties are described. The structural characterization of the ligands and their ruthenium complexes has been realized with the aid of elemental analysis, IR, UV-vis, NMR and HR-MS techniques. The molecular structures of all three complexes have been elucidated by single crystal X-ray crystallography and reveal the existence of pseudo-octahedral geometry around the ruthenium. The in vitro cancer cell growth inhibition property of the complexes against A549 (lung carcinoma), A2780 (ovarian adenocarcinoma) and non-cancerous 16HBE (human lung bronchial epithelium) cells were examined by MTT assay. All the complexes display good cytotoxicity towards both of these types of cancer cell compared to the standard drug cisplatin, with low IC50 values. Remarkably, complex 3, which contains an electron-donating substituent, induces a significant reduction of viability in A2780 cells. The inhibition capacity of the complexes towards A2780 cells proliferation was further confirmed using 5-ethynyl-2-deoxyuridine (EdU) assay via minimal DNA synthesis. The result of the acridine orange-ethidium bromide (AO-EB) fluorescent staining assay establishes that the cytotoxicity of the complexes was mediated by apoptosis in cancer cells. Furthermore, flow cytometry using Annexin V-FITC/propidium iodide (PI) double staining determines the quantitative discrimination of early apoptosis by the externalization of phosphatidylserine. In addition, cell cycle distribution indicates that the complexes block the cell cycle progression in the S-phase. The outcome of our investigation shows the promising scope and potency of tailored arene ruthenium complexes for precise cancer chemotherapy beyond platinum drugs. Show less

Title: Impact of aliphatic acyl and aromatic thioamide substituents on the anticancer activity of Ru(II)- Abstract: Six different acylthiourea ligands (L1-L6) and their corresponding Ru(II)-p-cymene complexes (P1-P6) were designed to explore the structure-activity relationship of the complexes upon aliphatic chain and aromatic conjugation on the C- and N-terminals, respectively. The compounds were synthesized and adequately characterized using various analytical and spectroscopic techniques. The structures of P2-P6, solved using single crystal X-ray diffraction (XRD), confirmed the neutral monodentate coordination of the S atoms of the acylthiourea ligands to Ru(II) ions. In silico studies showed an increase of lipophilicity for the ligands with an increase in alkyl chain length or aromatic conjugation at the C- or N-terminal, respectively. Subsequently, mitogen-activated protein kinases (MAPK) were predicted as one of the primary targets for the complexes, which showed good binding affinity towards extracellular signal-regulated kinases (ERK1, ERK2 and ERK5), c-Jun N-terminal kinase (JNK) and p38 of the MAPK pathway. Henceforth, the complexes were tested for their anticancer activity in lung carcinoma (A549) and cisplatin-resistant lung carcinoma (cisA549R) cells and human umbilical vein epithelial normal cells (HUVEC). Interestingly, an increase in chain length or aromatic conjugation led to an increase in the activity of the complexes, with P5 (7.73 and 13.04 μM) and P6 (6.52 and 14.45 μM) showing the highest activity in A549 and cisA549R cells, which is better than the positive control, cisplatin (8.72 and 44.28 μM). Remarkably, we report the highest activity yet observed for complexes of the type [(η6-p-cymene)RuIICl2(S-acylthiourea)] in the tested cell lines. Aqueous solution studies showed that complexes P5 and P6 are rapidly hydrolyzed to produce solely aquated species that remained stable for 24 h. Staining assays and flow cytometric analyses of P5 and P6 in A549 cells revealed that the complexes induced apoptosis and arrested the cell cycle predominantly in the S phase. In vivo studies demonstrated the higher toxicity of cisplatin and a comparatively higher survival rate of mice injected with the most active complex P6. Histological analyses revealed that treatment with P6 at high doses of up to 8 mg kg-1 did not cause any palpable damage to the tested organs. Show less

The facile modification of the ligands in organometallic Ru(II)-arene complexes offers more opportunities to optimize their pharmacological profiles. Herein, three Ru(II)-arene complexes containing a glutathione S-transferase (GST) inhibitor (NBDHEX) in chelate ligand have been designed and synthesized in this study. In vitro results indicated that the ligation with NBDHEX significantly increased the activities and selectivities of the organometallic Ru(II)-arene complexes against tumor cells, especially complex 3, which was the most active compound among the tested compounds. DFT calculations and hydrolysis results demonstrated that complex 3 with more alkyl groups in the arene ligand has increased electron density at the Ru(II) center as compared with complexes 1 and 2, thus resulting in the improved hydrolysis rate, which may be responsible for its higher anticancer activity. Further studies showed that complexes 1-3 can cause the loss of the mitochondrial membrane potential and upregulate the expression of Bcl-2 and Bax in A549 cells, suggesting that complexes 1-3-induced cell death may be mediated via the mitochondrial apoptotic pathway. Thus, these findings suggested that simultaneous modification of the chelate ligands and arene rings in the organometallic Ru(II)-arene complexes is an effective way to improve their pharmacological properties. Show less

Title: Pyrene-based fluorescent Ru(II)-arene complexes for significant biological applications: catalytic potential, DNA/protein binding, two photon cell imaging and Abstract: Ruthenium complexes are being studied extensively as anticancer drugs following the inclusion of NAMI-A and KP1019 in phase II clinical trials for the treatment of metastatic phase and primary tumors. Herein, we designed and synthesized four organometallic Ru(II)-arene complexes [Ru(η6-p-cymene)(L)Cl] (1), [Ru(η6-benzene)(L)Cl] (2), [Ru(η6-p-cymene)(L)N3] (3) and [Ru(η6-benzene)(L)N3] (4) [HL = (E)-N'-(pyren-1-ylmethylene)thiopene-2-carbohydrazide] that have anticancer, antimetastatic and two-photon cell imaging abilities. Moreover, in the transfer hydrogenation of NADH to NAD+, these compounds also display good catalytic activity. All the complexes, 1-4, are well characterized by spectroscopic techniques (NMR, mass, FTIR, UV-vis and fluorescence). The single crystal X-ray diffraction technique proved that the ligand L coordinates through an N,O-bidentate chelating fashion in the solid-state structures of complexes 1 and 2. The stability study of the complexes was performed through UV-visible spectroscopy. The cytotoxicities of all the complexes were screened through MTT assay and the results revealed that the complexes have potential anticancer activity against various cancerous cells (HeLa, MCF7 and A431). Studies with spectroscopic techniques revealed that complexes 1-4 exhibit strong interactions with biological molecules i.e. proteins (HSA and BSA) and CT-DNA. The density functional theory (DFT-D) method has been employed in the present study to know the interaction between DNA and complexes by calculating the HOMO and LUMO energy. A plausible mechanism for NADH oxidation has also been explored and the DFT calculations are found to be in accord with the experimental observation. Furthermore, we have investigated intracellular reactive oxygen species (ROS) generation capabilities in the MCF7 breast cancer cell line. The Hoechst/PI dual staining method confirmed the apoptosis mode of cell death. Meanwhile, complexes 1-4 show capabilities to prevent the metastasis phase of cancer cells by inhibiting cell migration. Show less

The use of organic compounds with known medicinal properties in the synthesis of metal-based complexes is an important alternative to improve the biological activity of metal-based drugs. The reaction of [M(arene)Cl₂]₂ (M = Ru, arene = p-cymene and M = Ir, arene = pentamethylcyclopentadienyl, cp*) with avobenzone (1-(4-tert-butylphenyl)-3-(4-methoxyphenyl)propane-1,3-dione, AVBH) and KOH in methanol leads to the formation of the neutral complexes [Ru(p-cymene)(AVB)Cl] 1 and [Ir(cp*)(AVB)Cl] 2 (cp* = pentamethylcyclopentadienyl). Subsequent reaction of 1 and 2 with pyridyl derivative-BODIPY ligands, BDP and BDPCC (BODIPY = boron dipyrromethene, BDP = 4-dipyridine boron dipyrromethene, BDPCC = 4-ethynylpyridine boron dipyrromethene) in methanol gives a series of four new dicationic supramolecules: [Ru₂(p-cymene)₂(AVB)₂BDP][2CF₃SO₃] 3, [Ir₂(cp*)₂(AVB)₂BDP][2CF₃SO₃] 4, [Ru₂(p-cymene)₂(AVB)₂BDPCC][2CF₃SO₃] 5 and [Ir₂(cp*)₂(AVB)₂BDPCC][2CF₃SO₃] 6. The synthesized complexes are fully characterized using multiple analytical techniques, including elemental analysis, ¹H NMR, ¹³C NMR, ¹⁹F NMR (NMR = Nuclear Magnetic Resonance), Infrared Radiation (IR), Electrospray Ionization-Mass Spectrometry (ESI-MS), Ultraviolet-visible (UV-Vis) and fluorescence spectroscopy. The structures of these complexes are further rationalized using density functional theory (DFT) calculations. The antiproliferative activity of the neutral and dinuclear cationic complexes is evaluated in vitro in different human cancer cell lines. These complexes are found to be active against different cancer cell lines with half maximal inhibitory concentration (IC₅₀) values between 1 and 5 μM. Complexes 5 and 6 displayed the lowest IC₅₀ values in all the cell lines studied. The activity of 5 and 6 is comparable to that of the well-known chemotherapy drug doxorubicin. Detailed biophysical studies indicate that complexes 5 and 6 exhibit very good Deoxyribonucleic acid (DNA) binding properties, causing the unwinding of the double helix, which is a probable reason for their high cytotoxicity. Show less

[Ru(tBu2bpy)2(2-appt)](PF6)2 [1.(PF6)2, tBu2bpy = 4,4'-di-tert-butyl-2,2'-bipyridine, 2-appt = 2-amino-4-phenylamino-6-(2-pyridyl)-1,3,5-triazine] and [Re(CO)3(2-appt)Cl] (2) were prepared and characterized by X-ray crystal analysis. The binding of 1.(PF6)2 and 2 to calf thymus DNA (ct DNA) led to increases in the DNA melting temperature (Delta Tm = +12 degrees C), modest hypochromism (29% and 5% of the absorption bands at lambda max = 450 and 376 nm, respectively), and insignificant shifts in the absorption maxima. The binding constants of 1.(PF6)2 and 2 with ct DNA, as determined by absorption titration, are (8.9 +/- 0.5) x 104 and (3.6 +/- 0.1) x 104 dm3 mol-1, respectively. UV-vis absorption titration, DNA melting studies, and competition dialysis using synthetic oligonucleotides [poly(dA-dT)2 and poly(dG-dC)2] revealed that 1.(PF6)2 and 2 exhibit a binding preference for AT sequences. A modeling study on the interaction between 1 or 2 and B-DNA revealed that the minor groove is the most favored binding site and an extensive hydrogen-bonding network is formed. As determined by MTT assays, 1.(PF6)2 and 2 exhibited moderate cytotoxicities toward several human cancer cell lines (KB-3-1, HepG2, and HeLa), as well as a multi-drug-resistant cancer cell line (KB-V-1). According to confocal microscopic and flow cytometric studies, 1.(PF6)2 and 2 induced apoptosis (50-60%) in cancer cells with <5% necrosis detected. Show less

A new ligand DBHIP and its two ruthenium (II) complexes [Ru(bpy)(2)(DBHIP)](ClO(4))(2) (1) and [Ru(phen)(2)(DBHIP)](ClO(4))(2) (2) have been synthesized and characterized. The binding behaviors of the two complexes to calf thymus DNA were investigated by absorption spectra, viscosity measurements, thermal denaturation and photoactivated cleavage. The DNA-binding constants for complexes 1 and 2 have been determined to be 8.87+/-0.27 x 10(4)M(-1) (s=1.83) and 1.32+/-0.31 x 10(5)M(-1) (s=1.84). The results suggest that these complexes interact with DNA through intercalative mode. The cytotoxicity of DBHIP, complexes 1 and 2 has been evaluated by MTT assay. The apoptosis assay was carried out with acridine orange/ethidium bromide (AO/EB) staining methods. The studies on the mechanism of photocleavage demonstrate that superoxide anion radical (O(2)(-)) and singlet oxygen ((1)O(2)) may play an important role. Show less

Four ruthenium(II) complexes with the formula [Ru(η(5)-C(5)H(5))(PP)L][CF(3)SO(3)], being (PP = two triphenylphosphine molecules), L = 1-benzylimidazole, ; (PP = two triphenylphosphine molecules), L = 2,2'bipyridine, ; (PP = two triphenylphosphine molecules), L = 4-Methylpyridine, ; (PP = 1,2-bis(diphenylphosphine)ethane), L = 4-Methylpyridine, , were prepared, in view to evaluate their potentialities as antitumor agents. The compounds were completely characterized by NMR spectroscopy and their crystal and molecular structures were determined by X-ray diffraction. Electrochemical studies were carried out giving for all the compounds quasi-reversible processes. The images obtained by atomic force microscopy (AFM) suggest interaction with pBR322 plasmid DNA. Measurements of the viscosity of solutions of free DNA and DNA incubated with different concentrations of the compounds confirmed this interaction. The cytotoxicity of compounds 1234 was much higher than that of cisplatin against human leukemia cancer cells (HL-60 cells). IC(50) values for all the compounds are in the range of submicromolar amounts. Apoptotic death percentage was also studied resulting similar than that of cisplatin. Show less

The ruthenium compound KP1019 has demonstrated promising anticancer activity in a pilot clinical trial. This study aims to evaluate the intracellular uptake/binding patterns of KP1019 and its sodium salt KP1339, which is currently in a phase I-IIa study. Although KP1339 tended to be moderately less cytotoxic than KP1019, IC(50) values in several cancer cell models revealed significant correlation of the cytotoxicity profiles, suggesting similar targets for the two drugs. Accordingly, both drugs activated apoptosis, indicated by caspase activation via comparable pathways. Drug uptake determined by inductively coupled plasma mass spectrometry (ICP-MS) was completed after 1 h, corresponding to full cytotoxicity as early as after 3 h of drug exposure. Surprisingly, the total cellular drug uptake did not correlate with cytotoxicity. However, distinct differences in intracellular distribution patterns suggested that the major targets for the two ruthenium drugs are cytosolic rather than nuclear. Consequently, drug-protein binding in cytosolic fractions of drug-treated cells was analyzed by native size-exclusion chromatography (SEC) coupled online with ICP-MS. Ruthenium-protein binding of KP1019- and KP1339-treated cells distinctly differed from the platinum binding pattern observed after cisplatin treatment. An adapted SEC-SEC-ICP-MS system identified large protein complexes/aggregates above 700 kDa as initial major binding partners in the cytosol, followed by ruthenium redistribution to the soluble protein weight fraction below 40 kDa. Taken together, our data indicate that KP1019 and KP1339 rapidly enter tumor cells, followed by binding to larger protein complexes/organelles. The different protein binding patterns as compared with those for cisplatin suggest specific protein targets and consequently a unique mode of action for the ruthenium drugs investigated. Show less

[Ru(bipy)₂(dpphen)]Cl₂ (where bipy = 2,2'-bipyridine and dpphen = 2,9-diphenyl-1,10-phenanthroline) (complex 1) is a sterically strained compound that exhibits promising in vitro photocytotoxicity on an array of cell lines. Since lung adenocarcinoma cancer remains the most common lung cancer and the leading cause of cancer deaths, the current study aims to evaluate the plausible effect and uptake of complex 1 on human alveolar carcinoma cells (A549) and mesenchymal stem cells (MSC), and assess its cytotoxicity in vitro while considering its effect on cell morphology, membrane integrity and DNA damage. MSC and A549 cells showed similar rates of complex 1 uptake with a plateau at 12 h. Upon photoactivation, complex 1 exhibited selective, potent anticancer activity against A549 cells with phototoxicity index (PI) values of 16, 25 and 39 at 24, 48 and 72 h, respectively. This effect was accompanied by a significant increase in A549-cell rounding and detachment, loss of membrane integrity and DNA damage. Flow cytometry experiments confirmed that A549 cells undergo apoptosis when treated with complex 1 followed by photoactivation. In conclusion, this present study suggests that complex 1 might be a promising candidate for photochemotherapy with photoproducts that possess selective anticancer effects in vitro. These results are encouraging to probe the potential activity of this complex in vivo. Show less

Cancer stem cells (CSCs) are defined as a rare population of cancer cells related to tumor initiation and maintenance. These cells are primarily responsible for tumor growth, invasion, metastasis, recurrence, and resistance to chemotherapy. In this paper, we demonstrated the ability of Ru(II)-based complexes containing 2-thiouracil derivatives with the chemical formulas trans-[Ru(2TU)(PPh₃)₂(bipy)]PF₆ (1) and trans-[Ru(6m2TU)(PPh₃)₂(bipy)]PF₆ (2) (where 2TU = 2-thiouracil and 6m2TU = 6-methyl-2-thiouracil) to suppress liver CSCs by targeting NF-κB and Akt/mTOR signaling. Complexes 1 and 2 displayed potent cytotoxic effects on cancer cell lines and suppressed liver CSCs from HepG2 cells. Increased phosphatidylserine exposure, loss of mitochondrial transmembrane potential, increased PARP (Asp214) cleavage, DNA fragmentation, chromatin condensation and cytoplasmic shrinkage were detected in HepG2 cells treated with these complexes. Mechanistically, complexes 1 and 2 target NF-κB and Akt/mTOR signaling in HepG2 cells. Cell motility inhibition was also detected in HepG2 cells treated with these complexes. Complexes 1 and 2 also inhibited tumor progression in mice with HepG2 cell xenografts and exhibited tolerable systemic toxicity. Taken together, these results indicate that these complexes are new anti-HCC drug candidates that can suppress liver CSCs. Show less

Ruthenium-based compounds represent a class of potential antineoplastic drugs. Recently, we designed, synthesized, and identified the Ru(II)-thymine complex [Ru(PPh₃)₂(Thy)(bipy)]PF₆ (where PPh = triphenylphosphine, Thy = thymine and bipy = 2,2'-bipyridine) as a potent cytotoxic agent with the ability to bind to DNA and human and bovine serum albumins. In this study, the underlying cytotoxic mechanism of the [Ru(PPh₃)₂(Thy)(bipy)]PF₆ complex was assessed. This complex displayed potent cytotoxicity in different cancer cell lines; the morphology that is associated with apoptotic cell death, increased internucleosomal DNA fragmentation without cell membrane permeability, loss of the mitochondrial transmembrane potential, increased phosphatidylserine externalization, and caspase-3 activation were observed in human promyelocytic leukemia HL-60 cells that were treated with the complex. Moreover, pretreatment of HL-60 cells with Z-VAD(OMe)-FMK, a pan-caspase inhibitor, partially reduced the apoptosis that was induced by the complex, indicating that the apoptotic cell death occurred through a caspase-mediated pathway. In conclusion, the [Ru(PPh₃)₂(Thy)(bipy)]PF₆ complex displays potent cytotoxicity to different cancer cells and induces caspase-mediated apoptosis in HL-60 cells. Show less