Imine‐N‐Heterocyclic Carbenes as Versatile Ligands in Ruthenium(II) p‐Cymene Anticancer Complexes: A Structure–Activity Relationship Study

A family of novel imine‐N‐heterocyclic carbene ruthenium(II) complexes of the general formula [(η⁶‐p‐cymene)Ru(C^N)Cl]PF₆⁻ (where C^N is an imine‐N‐heterocyclic carbene chelating ligand with varying substituents) have been prepared and characterized. In this imine‐N‐heterocyclic carbene chelating ligand framework, there are three potential sites that can be modified, which distinguishes this class of ligand and provides a body of flexibilities and opportunities to tune the cytotoxicity of these ruthenium(II) complexes. The influence of substituent effects of three tunable domains on the anticancer activity and catalytic ability in converting coenzyme NADH to NAD⁺ is investigated. This family of complexes displays an exceedingly distinct anticancer activity against A549 cancer cells, despite their close structural similarity. Complex 9 shows the highest anticancer activity in this series against A549 cancer cells (IC₅₀=14.36 μm ), with an approximately 1.5‐fold better activity than the clinical platinum drug cisplatin (IC₅₀=21.30 μm ) in A549 cancer cells. Mechanistic studies reveal that complex 9 mediates cell death mainly through cell stress, including cell cycle arrest, inducing apoptosis, increasing intracellular reactive oxygen species (ROS) levels, and depolarization of the mitochondrial membrane potential (MMP). Furthermore, lysosomal damage is also detected by confocal microscopy.     

Synthesis,cytotoxicity and anti‐metastatic properties of new pyridyl–thiazole arene ruthenium(II) complexes

A series of novel ruthenium(II)–cymene complexes ( 1 – 8 ) containing substituted pyridyl–thiazole ligands, [Ru(η⁶‐p‐cymene)(L)Cl]Cl (L = N,N‐chelating derivatives), have been synthesized and characterized using elemental analysis, infrared, ¹H NMR and ¹³C NMR spectroscopies and mass spectrometry. All these complexes not only display marked cytotoxicity in vitro against three different human cancer cell lines (HeLa, A549 and MDA‐MB‐231), but also exhibit promising anti‐metastatic activity at sub‐cytotoxic concentrations. Cell cycle analysis shows that the ruthenium(II) complex‐induced growth inhibition was mainly caused by S‐phase cell cycle arrest. Further protein level analysis suggests that compound 5 may exert antitumor activity via a p53‐independent mechanism.     

Novel microwave synthesis of half‐sandwich [(η6‐C6H6)Ru] complexes and an evaluation of the biological activity and biochemical reactivity

We have used a novel microwave‐assisted method to synthesize a pair of half‐sandwich ruthenium–arene–thiosemicarbazone complexes of the type [(η⁶‐C₆H₆Ru(TSC)Cl]PF₆. The thiosemicarbazone (TSC) ligands are 2‐(anthracen‐9‐ylmethylene)hydrazinecarbothioamide and 2‐(anthracen‐9‐ylmethylene)‐N‐ethylhydrazinecarbothioamide derived from 9‐anthraldehyde. The complexes are moderately strong binders of DNA, with binding constants of 10⁴ m ^?1. They are also strong binders of human serum albumin, having binding constants of the order of 10⁵ m ^?1. The complexes show some in vitro anticancer activity against human colon cancer cells, Caco‐2 and HCT‐116, with positive therapeutic indices. They did not show any activity as antibacterial agents against the organisms that were studied. Copyright © 2013 John Wiley & Sons, Ltd.     

Heteroscorpionate‐based heteroleptic copper(II) complexes: Antioxidant,molecular docking and in vitro cytotoxicity studies

Three new heteroscorpionate ligands, (2‐hydroxyphenyl)bis(imidazol‐1‐yl)methane (HL¹), (4‐diethylamino‐2‐hydroxyphenyl)bis(imidazol‐1‐yl)methane (HL²) and (5‐bromo‐2‐hydroxyphenyl)bis(imidazol‐1‐yl)methane (HL³), and their heteroleptic copper(II) complexes of the type [Cu(L^1–3)diimine]ClO₄ ( 1 – 6 ; where diimine =2,2′‐bipyridyl or 1,10‐phenanthroline) have been synthesized and characterized using spectroscopic methods. The molecular structure of ligand HL¹ was determined by single‐crystal X‐ray diffraction. UV–visible, electron paramagnetic resonance and theoretical studies suggest a distorted square pyramidal geometry around copper(II) ion. Analyses of highest occupied and lowest unoccupied molecular orbitals have been used to explain the charge transfer taking place within the complexes. The antioxidant activities of the heteroscorpionate ligands and their heteroleptic copper(II) complexes were determined using ABTS, DPPH and H₂O₂ free radical scavenging assays with respect to standard antioxidant ascorbic acid. In molecular docking studies, the complexes showed π–π, hydrogen bonding, van der Waals and electrostatic interactions with fibroblast growth factor receptor kinase. In vitro cytotoxicity activities of ligands and copper(II) complexes were examined on human breast adenocarcinoma (MCF‐7), cervical (HeLa) and lung (A549) cancer cell lines and normal human dermal fibroblast cell line using MTT assay. Complex 4 exhibited higher anticancer activity than the other complexes against all three cancer lines, being more potent than the standard drug cisplatin.     

Histone‐Deacetylase‐Targeted Fluorescent Ruthenium(II) Polypyridyl Complexes as Potent Anticancer Agents

Histone deacetylases inhibitors (HDACis) have gained much attention as a new class of anticancer agents in recent years. Herein, we report a series of fluorescent ruthenium(II) complexes containing N¹‐hydroxy‐N⁸‐(1,10‐phenanthrolin‐5‐yl)octanediamide ( L ), a suberoylanilide hydroxamic acid (SAHA) derivative, as a ligand. As expected, these complexes show interesting chemiphysical properties, including relatively high quantum yields, large Stokes shifts, and long emission lifetimes. The in vitro inhibitory effect of the most effective drug, [Ru(DIP)₂ L ](PF₆)₂ ( 3 ; DIP: 4,7‐diphenyl‐1,10‐phenanthroline), on histone deacetylases (HDACs) is approximately equivalent in activity to that of SAHA, and treatment with complex 3 results in increased levels of the acetylated histone H3. Complex 3 is highly active against a panel of human cancer cell lines, whereas it shows relatively much lower toxicity to normal cells. Further mechanism studies show that complex 3 can elicit cell cycle arrest and induce apoptosis through mitochondria‐related pathways and the production of reactive oxygen species. These data suggest that these fluorescent ruthenium(II)–HDACi conjugates may represent a promising class of anticancer agents for potential dual imaging and therapeutic applications targeting HDACs.     

Design,synthesis,characterization,cytotoxic and structure activity relationships of novel Ru(Ⅱ) complexes

Platinum containing compounds have shown antineoplastic potential, but their clinical applications have been limited by high toxicity. Ruthenium containing complexes have long been known to be well suited for biological applications, and have long been utilized as replacements to popular platinum based-drugs. Here, we report a novel series of ruthenium(II) arene compounds bearing thiosemicarbazone and isonicotinylhydrazone ligands with potent anticancer activity their structure activity relationships and apoptosis was studied. The cytotoxic activity of the new ruthenium(II) arene compounds has been evaluated in several cell lines (Molt 4/C8, L1210, CEM, HL60 and BEL7402). Among them, ten complexes were found to be excellent in vitro growth inhibitory activity against various cell lines with IC50 in the sub-micromolar range.     

Voltammetric detection of avidin and biotin by biotin labeled with cysteine

An avidin-biotin assay was developed from a voltammetric procedure using biotin labeled with cysteine. Mercury(II) as a marker was used to detect avidin and biotin, because the oxidation wave of mercury decreases when the cysteine part of labeled biotin(LB) complexes with mercury(II).The formation of the mercury(II)-cysteine complex is suppressed when the LB binds to the biotin site of avidin. Accordingly, the concentration of avidin can be estimated from the increasing mercury peak current. Detection of biotin is also carried out by a competitive reaction of biotin and the LB to the binding site on avidin, where the addition of biotin decreases the peak current of mercury. Limits of detection for avidin and biotin were in the 10^–9 mol/L range. The length of the spacer between the cysteine and biotin was investigated. It was observed that the strength of binding increased with increasing length of spacer. Size considerations rules out steric influences, so it is suggested that the binding constant depends on hydrophobic interactions in the binding site.     

Self‐Assembling Hydrogels Crosslinked Solely by Receptor–Ligand Interactions: Tunability,Rationalization of Physical Properties,and 3D Cell Culture

We report a novel, noncovalent hydrogel system crosslinked solely by receptor–ligand interactions between biotin and avidin. The simple hydrogel synthesis and functionalization together with the widespread use of biotinylated ligands in biosciences make this versatile system suitable for many applications. The gels possess a range of tunable physical properties, including stiffness, lifetime, and swelling. The erosion rates, unexpectedly fast compared to the kinetic parameters for biotin–avidin, are explored in terms of stretching tensions on the polymers, a concept well‐known on the single‐molecule level, but largely unexplored in supramolecular systems. As proof of utility, the gels were functionalized with different peptide sequences to control human mesenchymal stromal cell morphology in 3D culture.     

Maltol‐Derived Ruthenium–Cymene Complexes with Tumor Inhibiting Properties: The Impact of Ligand–Metal Bond Stability on Anticancer Activity In Vitro

Organometallic ruthenium–arene compounds bearing a maltol ligand have been shown to be nearly inactive in in vitro anticancer assays, presumably due to the formation of dimeric Ru^II species in aqueous solutions. In an attempt to stabilize such complexes, [Ru(η⁶‐p‐cymene)(XY)Cl] (XY=pyrones or thiopyrones) complexes with different substitution pattern of the (thio)pyrone ligands have been synthesized, their structures characterized spectroscopically, and their aquation behavior investigated as well as their tumor‐inhibiting potency. The aquation behavior of pyrone systems with electron‐donating substituents and of thiopyrone complexes was found to be significantly different from that of the maltol‐type complex reported previously. However, the formation of the dimer can be excluded as the primary reason for the inactivity of the complex because some of the stable compounds are not active in cancer cell lines either. In contrast, studies of their reactivity towards amino acids demonstrate different reactivities of the pyrone and thiopyrone complexes, and the higher stability of the latter probably renders them active against human tumor cells.     

Preparation of Hyaluronic Acid Micro-Hydrogel by Biotin–Avidin-Specific Bonding for Doxorubicin-Targeted Delivery

Hyaluronic acid is a naturally ionic polysaccharide with cancer cell selectivity. It is an ideal candidate material for delivery of anticancer agents. In this study, hyaluronic acid (HA) micro-hydrogel loaded with anticancer drugs was prepared by the biotin–avidin system approach. Firstly, carboxyl groups on HA were changed into amino groups with adipic acid dihydrazide (ADH) to graft with biotin by 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride named as HA–biotin. When HA–biotin solution mixed with doxorubicin hydrochloride (DOX·HCl) was blended with neutravidin, the micro-hydrogels would be formed with DOX loading. If excess biotin was added into the microgel, it would be disjointed, and DOX will be released quickly. The results of the synthesis procedure were characterized by ¹H-NMR and FTIR; ADH and biotin have been demonstrated to graft on the HA molecule. A field emission scanning electron microscope was used to observe morphologies of HA micro-hydrogels. Furthermore, the in vitro DOX release results revealed that the release behaviors can be adjusted by adding biotin. Therefore, the HA micro-hydrogel can deliver anticancer drugs efficiently, and the rate of release can be controlled by biotin-specific bonding with the neutravidin. Consequently, the micro-hydrogel will perform the promising property of switching in the specific site in cancer therapy.     

Organometallic chemistry, biology and medicine: ruthenium arene anticancer complexes

Our work has shown that certain ruthenium(II) arene complexes exhibit promising anticancer activity in vitro and in vivo. The complexes are stable and water-soluble, and their frameworks provide considerable scope for optimising the design, both in terms of their biological activity and for minimising side-effects by variations in the arene and the other coordinated ligands. Initial studies on amino acids and nucleotides suggest that kinetic and thermodynamic control over a wide spectrum of reactions of Ru(II) arene complexes with biomolecules can be achieved. These Ru(II) arene complexes appear to have an altered profile of biological activity in comparison with metal-based anticancer complexes currently in clinical use or on clinical trial.     

Am(m)ines Make the Difference: Organoruthenium Am(m)ine Complexes and Their Chemistry in Anticancer Drug Development

With the aim of systematically studying fundamental structure–activity relationships as a basis for the development of Ru^II arene complexes (arene=p‐cymene or biphenyl) bearing mono‐, bi‐, or tridentate am(m)ine ligands as anticancer agents, a series of ammine, ethylenediamine, and diethylenetriamine complexes were prepared by different synthetic routes. Especially the synthesis of mono‐, di‐, and triammine complexes was found to be highly dependent on the reaction conditions, such as stoichiometry, temperature, and time. Hydrolysis and protein‐binding studies were performed to determine the reactivity of the compounds, and only those containing chlorido ligands undergo aquation or form protein adducts. These properties correlate well with in vitro tumor‐inhibiting potency of the compounds. The complexes were found to be active in anticancer assays when meeting the following criteria: stability in aqueous solution and low rates of hydrolysis and binding to proteins. Therefore, the complexes least reactive to proteins were found to be the most cytotoxic in cancer cells. In general, complexes with biphenyl as arene ligand inhibited the growth of tumor cells more effectively than the cymene analogues, consistent with the increase in lipophilicity. This study highlights the importance of finding a proper balance between reactivity and stability in the development of organometallic anticancer agents.     

Cyclometalated Ruthenium(II) Anthraquinone Complexes Exhibit Strong Anticancer Activity in Hypoxic Tumor Cells

Hypoxia is the critical feature of the tumor microenvironment that is known to lead to resistance to many chemotherapeutic drugs. Six novel ruthenium(II) anthraquinone complexes were designed and synthesized; they exhibit similar or superior cytotoxicity compared to cisplatin in hypoxic HeLa, A549, and multidrug‐resistant (A549R) tumor cell lines. Their anticancer activities are related to their lipophilicity and cellular uptake; therefore, these physicochemical properties of the complexes can be changed by modifying the ligands to obtain better anticancer candidates. Complex 1 , the most potent member of the series, is highly active against hypoxic HeLa cancer cells (IC₅₀=0.53 μM ). This complex likely has 46‐fold better activity than cisplatin (IC₅₀=24.62 μM ) in HeLa cells. This complex tends to accumulate in the mitochondria and the nucleus of hypoxic HeLa cells. Further mechanistic studies show that complex 1 induced cell apoptosis during hypoxia through multiple pathways, including those of DNA damage, mitochondrial dysfunction, and the inhibition of DNA replication and HIF‐1α expression, making it an outstanding candidate for further in vivo studies.     

Organometallic ruthenium and iridium phosphorus complexes: Synthesis,cellular imaging,organelle targeting and anticancer applications

The use of metal complexes containing phosphorus ligands as anticancer agents has not been well studied. In this work, eight novel half‐sandwich Ir^III and Ru^II compounds with P^P‐chelating ligands have been synthesized and fully characterized, and alongside two crystal structures were reported. All eight complexes displayed highly potent antiproliferative activity, up to nine times more potent than the clinical anticancer drug cisplatin towards A549 lung cancer cells. Complex Ir1 , which has a simpler structure and highly potent antiproliferative activity, was selected to investigate in further mechanistic studies. No hydrolysis and nucleobase binding occurred for complex Ir1 . In order to elucidate subcellular localization, the self‐luminescence of the complex Ir1 was utilized. Ir1 can specifically target lysosomes and facilitate excessive production of reactive oxygen species, resulting in lysosomal membrane permeabilization in A549 cells. Release of cathepsin B and changes in the mitochondria membrane potential also contributed to the observed cytotoxicity of Ir1 , which demonstrated an anticancer action mechanism that was different from that of cisplatin. The favorable results from biological and chemical research demonstrated that these types of complexes hold significant theranostic potential.     

Unmasking the Action of Phosphinous Acid Ligands in Nitrile Hydration Reactions Catalyzed by Arene–Ruthenium(II) Complexes

The catalytic hydration of benzonitrile and acetonitrile has been studied by employing different arene–ruthenium(II) complexes with phosphinous (PR₂OH) and phosphorous acid (P(OR)₂OH) ligands as catalysts. Marked differences in activity were found, depending on the nature of both the P‐donor and η⁶‐coordinated arene ligand. Faster transformations were always observed with the phosphinous acids. DFT computations unveiled the intriguing mechanism of acetonitrile hydration catalyzed by these arene–ruthenium(II) complexes. The process starts with attack on the nitrile carbon atom of the hydroxyl group of the P‐donor ligand instead of on a solvent water molecule, as previously suggested. The experimental results presented herein for acetonitrile and benzonitrile hydration catalyzed by different arene–ruthenium(II) complexes could be rationalized in terms of such a mechanism.     

Ruthenium(II) complexes containing 4‐methoxybenzhydrazone ligands: synthesis,characterization, DNA binding,DNA cleavage,radical scavenging and in vitro cytotoxic activity

Three ruthenium(II) hydrazone complexes of composition [RuCl(CO)(PPh₃)₂L] were synthesized from the reactions of [RuHCl(CO)(PPh₃)₃] with hydrazones derived from 4‐methoxybenzhydrazide and 4‐formylbenzoic acid (HL¹), 4‐methylbenzaldehyde (HL²) and 2‐bromobenzaldehyde (HL³). The synthesized hydrazone ligands and their metal complexes were characterized using elemental analysis and infrared, UV–visible, NMR (¹H, ¹³C and ³¹P) and mass spectral techniques. The hydrazone ligands act as bidentate ones, with O and N as the donor sites, and are predominantly found in the enol form in all the complexes studied. The molecular structures of the ligands HL¹, HL² and HL³ were determined using single‐crystal X‐ray diffraction. The interactions of the ligands and the complexes with calf thymus DNA were studied using absorption spectroscopy and cyclic voltammetry which revealed that the compounds could interact with calf thymus DNA through intercalation. The DNA cleavage activity of the complexes was evaluated using a gel electrophoresis assay which revealed that the complexes act as good DNA cleavage agents. In addition, all the complexes were subjected to antioxidant assay, which showed that they all possess significant scavenging activity against 2,2‐diphenyl‐2‐picrylhydrazyl, OH and NO radicals. The in vitro cytotoxic effect of the complexes examined on cancerous cell lines (HeLa and MCF‐7) showed that the complexes exhibit substantial anticancer activity. Copyright © 2016 John Wiley & Sons, Ltd.     

Tuning the anticancer activity of maltol-derived ruthenium complexes by derivatization of the 3-hydroxy-4-pyrone moiety

Organometallic ruthenium(II)-arene complexes coordinated to maltol-derived ligands were prepared and their anticancer activity against human tumor cell lines was studied. In addition, their hydrolysis behavior and reaction with 5′-GMP was tested and compared to the parent compound chlorido[2-methyl-3-(oxo-κO)-pyran-4(1H)-onato-κO4](η⁶-p-cymene)ruthenium(II) (Ru-maltol). Improved stability and in vitro anticancer activity at maintained GMP binding capability were observed, in comparison to the Ru-maltol complex.     

Novel half‐sandwich η5‐Cp *–rhodium(III) and η5‐Cp *–ruthenium(II) complexes bearing bis(phosphino)amine ligands and their use in the transfer hydrogenation of aromatic ketones

Two new half‐sandwich η⁵‐Cp*–rhodium(III) and η⁵‐Cp*–ruthenium(II) complexes have been prepared from corresponding bis(phosphino)amine ligands, thiophene‐2‐(N,N‐bis(diphenylphosphino)methylamine) or furfuryl‐2‐(N,N‐bis(diphenylphosphino)amine). Structures of the new complexes have been elucidated by multinuclear one‐ and two‐dimensional NMR spectroscopy, elemental analysis and IR spectroscopy. These Cp*–rhodium(III) and Cp*‐ruthenium(II) complexes bearing bis(phosphino)amine ligands were successfully applied to transfer hydrogenation of various ketones by 2‐propanol. Copyright © 2013 John Wiley & Sons, Ltd.     

Ruthenium–Arene–β‐Carboline Complexes as Potent Inhibitors of Cyclin‐Dependent Kinase 1: Synthesis,Characterization and Anticancer Mechanism Studies

A series of Ru^II–arene complexes ( 1 – 6 ) of the general formula [(η⁶‐arene)Ru(L)Cl]PF₆ (arene=benzene or p‐cymene; L=bidentate β‐carboline derivative, an indole alkaloid with potential cyclin‐dependent kinases (CDKs) inhibitory activities) is reported. All the complexes were fully characterized by classical analytical methods, and three were characterized by X‐ray crystallography. Hydrolytic studies show that β‐carboline ligands play a vital role in their aqueous behaviour. These complexes are highly active in vitro, with the most active complex 6 displaying a 3‐ to 12‐fold higher anticancer activity than cisplatin against several cancer cell lines. Interestingly, the complexes are able to overcome cross‐resistance to cisplatin, and show much lower cytotoxicity against normal cells. Complexes 1 – 6 may directly target CDK1, because they can block cells in the G2M phase, down‐regulate the expression of CDK1 and cyclin B1, and inhibit CDK1/cyclin B in vitro. Further mechanism studies show that the complexes can effectively induce apoptosis through mitochondrial‐related pathways and intracellular reactive oxygen species (ROS) elevation.