👤 Thilak Babu L

Two polypyridyl ruthenium(II) complexes [Ru(DIP)₂(BIP)](PF₆)₂ (DIP = 4,7-diphenyl-1,10-phenanthrolie, BIP = 2-(1,1'-biphenyl-4-yl)-1H-imidazo[4,5-f][1,10]phenanthroline, Ru1) and [Ru(DIP)₂(CBIP)](PF₆)₂ (CBIP = 2-(4'-chloro-1,1'-biphenyl-4-yl)-1H-imidazo[4,5-f][1,10]phenanthroline, Ru2) were synthesized. The cytotoxic activities in vitro of Ru1, Ru2 toward B16, A549, HepG2, SGC-7901, HeLa, BEL-7402, non-cancer LO2 were investigated using MTT method (3-(4,5-dimethylthiazole)-2,5-diphenltetraazolium bromide). Unexpectedly, Ru1, Ru2 can't prevent these cancer cells proliferation. To improve the anti-cancer effect, we used liposomes to entrap the complexes Ru1, Ru2 to form Ru1lipo, Ru2lipo. As expectation, Ru1lipo and Ru2lipo exhibit high anti-cancer efficacy, especially, Ru1lipo (IC₅₀ 3.4 ± 0.1 μM), Ru2lipo (IC₅₀ 3.5 ± 0.1 μM) display strong ability to block the cell proliferation in SGC-7901. The cell colony, wound healing, and cell cycle distribution show that the complexes can validly inhibit the cell growth at G2/M phase. Apoptotic studied with Annex V/PI doubling method showed that Ru1lipo and Ru2lipo can effectively induce apoptosis. Reactive oxygen species (ROS), malondialdehyde, glutathione and GPX4 demonstrate that Ru1lipo and Ru2lipo improve ROS and malondialdehyde levels, inhibit generation of glutathione, and finally result in a ferroptosis. Ru1lipo and Ru2lipo interact on the lysosomes and mitochondria and damage mitochondrial dysfunction. Additionally, Ru1lipo and Ru2lipo increase intracellular Ca²⁺ concentration and induce autophagy. The RNA-sequence and molecular docking were performed, the expression of Bcl-2 family was investigated by Western blot analysis. Antitumor in vivo experiments confirm that 1.23 mg/kg, 2.46 mg/kg of Ru1lipo possesses a high inhibitory rate of 53.53% and 72.90% to prevent tumor growth, hematoxylin-eosin (H&E) results show that Ru1lipo doesn't cause chronic organ damage and strongly promotes the necrosis of solid tumor. Taken together, we conclude that Ru1lipo and Ru2lipo cause cell death through the following pathways: autophagy, ferroptosis, ROS-regulated mitochondrial dysfunction, and blocking the PI3K/AKT/mTOR. Show less

In this article, four new Ru(II) complexes [Ru(dmbpy)₂(TFBIP)](PF₆)₂ (dmbpy = 4,4'-dimethyl-2,2'-bipyridine, TFPIP = 2-(4'-trifluoromethyl)-[1,1'-biphenyl]-4-yl)-1H-imidazo[4,5-f][1,10]phenanthroline) (Ru1), [Ru(bpy)₂(TFBIP)](PF₆)₂ (bpy = 2,2'-bipyridine) (Ru2), [Ru(phen)₂(TFBIP)](PF₆)₂ (phen = 1,10-phenanthroline) (Ru3) and [Ru(dmp)₂(TFBIP)](PF₆)₂ (dmp = 2,9-dimethyl-1,10-phenanthroline) (Ru4) were synthesized and characterized by elemental analysis, HRMS, IR, ¹H NMR, ¹³C NMR and ¹⁹F NMR. The in vitro anticancer effect of the complexes on HepG2, A549, B16, HeLa, BEL-7402 and non-cancer LO2 cells was screened using 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) method. The results illustrate that the complexes display moderate anticancer activity. Apoptotic assay with Annexin V/PI double staining method indicated that complexes induce apoptosis in HepG2 cells. Also, the complexes interfere with the mitochondrial functions, accompanied by the production of intracellular ROS as well as a reduction of mitochondrial membrane potential. The results obtained from the western blot demonstrated that the complexes upregulate pro-apoptotic Bax and downregulate anti-apoptotic Bcl-2, which further activates caspase 3 and promotes the cleavage of PARP. RNA-sequence showed that the complexes upregulate the expression of 40 genes and downregulate 66 genes. Antitumour in vivo demonstrated that Ru1 inhibits the tumor growth with a high inhibitory rate of 51.19%. Taken together, these results revealed that complexes Ru1, Ru2, Ru3 and Ru4 induce cell death in HepG2 cells via autophagy and a ROS-mediated mitochondrial apoptotic pathway. Show less

Title: Potent Half-Sandwich 16-/18-Electron Iridium(III) and Ruthenium(II) Anticancer Complexes with Readily Available Amine-Imine Ligands. Abstract: The synthesis and biological evaluation of stable 16-electron half-sandwich complexes have remained scarce. We herein present the different coordination modes (16-electron or 18-electron) between half-sandwich iridium(III) complexes and ruthenium(II) complexes derived from the same amine-imine ligands chelating hybrid sp3-N/sp2-N donors. The 16-electron iridium(III) and 18-electron ruthenium(II) complexes with different counteranions were obtained and identified by various techniques. The promising cytotoxicity of these complexes against A549 lung cancer cells, cisplatin-resistant A549/DPP cells, cervical carcinoma HeLa cells, and human hepatocellular liver carcinoma HepG2 cells was observed with IC50 values ranging from 5.4 to 16.3 μM. Moreover, these complexes showed a certain selectivity (selectivity index: 2.1-3.7) toward A549 cells and BEAS-2B normal cells. The variation of metal center, counteranion, 16/18-electron coordination mode, and ligand substituents showed little influence on the cytotoxicity and selectivity of these complexes. The mechanism of action study showed that these complexes could target mitochondria, induce the depolarization of the mitochondrial membrane, and promote the generation of intracellular reactive oxygen species (ROS). Further, the induction of cell apoptosis and the perturbation of the cell cycle in the G0/G1 phase were also observed for these complexes. Overall, it seems that the redox mechanism dominated the anticancer efficacy of these complexes. Show less

The biological efficacy of half-sandwich platinum group organometallic complexes of the formula [(η⁵-Cp^x)/(η⁶-arene)M(XY)Cl]^0/+ (XY = bidentate ligands; Cp^x = functionalized cyclopentadienyl; M = Ir, Rh, Ru, Os) has received considerable attention due to the significance of the metal center, chelating ligand, and Cp^x/arene moieties in defining their anticancer potency and selectivity. With a facile access to the BIAN-derived imine-amine ligands using alkylaluminum as the reductant, we herein described the preparation and characterization of 16 half-sandwich Ir(III), Rh(III), and Ru(II) complexes chelating the hybrid sp²-N/sp³-N donor ligand. A nonplanar five-member metallacycle was confirmed by X-ray single-crystal structures of Ir1-Ir3, Ir7, Rh1, Ru1, and Ru4. The attempt to prepare imine-amido complexes using a base as the deprotonating agent led to the mixture of imine-amine complexes, within which the leaving group Cl^- was displaced, and 16-electron imine-amido complexes without Cl^-. The half-sandwich imine-amine complexes in this system underwent rapid hydrolysis in aqueous solution, exhibited weak photoluminescence, and showed the ability of binding to CT-DNA and BSA. The cytotoxicity of all imine-amine complexes against A549 lung cancer cell lines, HeLa cervical cancer cell lines, and 4T1 mouse breast cancer cells was determined by an MTT assay. The IC₅₀ values of these complexes were in a range of 5.71-67.28 μM. Notably, most of these complexes displayed improved selectivity toward A549 cancer cells versus noncancerous BEAS-2B cells in comparison with the corresponding α-diimine complexes chelating the sp²-N/sp²-N donor ligand, which have been shown no selectivity in our previous report. The anticancer selectivity of these complexes appeared to be related to the redox-based mechanism including the catalytic oxidation of NADH to NAD⁺, reactive oxygen species (ROS) generation, and depolarization of the mitochondrial membrane. Further, inducing apoptosis of these complexes in A549 cancer cells and BEAS-2B normal cells also correlated with their anticancer selectivity, indicating the apoptosis mode of cell death in this system. In addition, these complexes could enter A549 cells via energy-dependent pathway and were able to impede the in vitro migration of A549 cells. Show less

In recent years, the studies of the ruthenium(II) complexes on anticancer activity have been paid great attention, many Ru(II) complexes possess high anticancer efficiency. In this paper, three ligands CPIP (2-(4-chlorophenyl)-1H-imidazo[4,5-f][1,10]phenanthroline), DCPIP (2-(3,4-dichlorophenyl)-1H-imidazo[4,5-f][1,10]phenanthroline), TCPIP (2-(2,3,5-trichlorophenyl)-1H-imidazo[4,5-f][1,10]phenanthroline) and their three ruthenium (II) complexes [Ru(dip)₂(CPIP)](PF₆)₂ (1, dip = 4,7-diphenyl-1,10-phenanthroline), [Ru(dip)₂(DCPIP)](PF₆)₂ (2) and [Ru(dip)₂(TCPIP)](PF₆)₂ (3) were synthesized and characterized. 3-(4,5-dimethylthiazole-2-yl)-2,5-biphenyl tetrazolium bromide (MTT) assay was used to investigate in vitro cytotoxicity of complexes against various cancer cells. The results showed that complexes 1-3 exhibited pronounced cytotoxic effect on B16 cells with low IC₅₀ values of 7.2 ± 0.1, 11.7 ± 0.6 and 1.2 ± 0.2 μM, respectively. The 3D model demonstrated that the complexes can validly prevent the cell proliferation. Apoptosis determined using Annexin V-FITC/PI double staining revealed that complexes 1-3 can effectively induce apoptosis in B16 cells. The intracellular localization of 1-3 in the mitochondria, the levels of intracellular reactive oxygen species (ROS), the opening of mitochondrial permeability transition pore as well as the decline of mitochondrial membrane potential were investigated, which demonstrated that the complexes 1-3 led to apoptosis via a ROS-mediated mitochondrial dysfunction pathway. The RNA-sequence indicated that the complexes upregulate the expression of 74 genes and downregulate the expression of 81 genes. The molecular docking showed that the complexes interact with the proteins through hydrogen bond, π-cation and π-π interaction. The results show that ruthenium(II) complexes 1, 2 and 3 can block tumor cell growth and induce cell death through autophagy and ROS-mediated mitochondrial dysfunction pathways. Show less

The main challenge of cancer chemotherapy is the resistance of tumor cells to oxidative damage. Herein, we proposed a novel antitumor strategy: cyclic metal‑ruthenium (Ru) complexes mediate reductive damage to kill tumor cells. We designed and synthesized Ru(II) complexes with β-carboline as ligands: [Ru (phen)₂(NO₂-Ph-βC)](PF₆) (RuβC-7) and [Ru(phen)₂(1-Ph-βC)](PF₆) (RuβC-8). In vitro experimental results showed that RuβC-7 and RuβC-8 can inhibit cell proliferation, promote mitochondrial abnormalities, and induce DNA damage. Interestingly, RuβC-7 with SOD activity could reduce intracellular reactive oxygen species (ROS) levels, while RuβC-8 has the opposite effect. Accordingly, this study identified the reductive damage mechanism of tumor apoptosis, and may provide a new ideas for the design of novel metal complexes. Show less

A new ligand DFIP (2-(dibenzo[b,d]furan-3-yl)-1H-imidazo[4,5-f][1,10]phenanthroline) and its two complexes iridium(III) [Ir(ppy)₂(DFIP)](PF₆) (ppy = 2-phenylpyridine, Ir1) and ruthenium(II) [Ru(bpy)₂(DFIP)](PF₆)₂ (bpy = 2,2'-bipyridine, Ru1) were synthesized and characterized. The anticancer effects of the two complexes on A549, BEL-7402, HepG2, SGC-7901, HCT116 and normal LO2 cells were tested by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method. Complex Ir1 shows high cytotoxic activity on A549, BEL-7402, SGC-7901 and HepG2, Ru1 exhibits moderate anticancer activity toward A549, BEL-7402 and SGC-7901 cells. The IC₅₀ values of Ir1 and Ru1 toward A549 are 7.2 ± 0.1 and 22.6 ± 1.4 μM, respectively. The localization of complexes Ir1 and Ru1 in the mitochondrial, intracellular accumulation of reactive oxygen species (ROS) levels, and the changes of mitochondrial membrane potential (MMP) and cytochrome c (cyto-c) were investigated. Apoptosis and cell cycle were detected by flow cytometry. Immunogenic cell death (ICD) was used to detect the effects of Ir1 and Ru1 on the A549 using a confocal laser scanning microscope. The expression of apoptosis-related proteins was detected by western blotting. Ir1 and Ru1 can increase the intracellular ROS levels and release cyto-c, reduce the MMP, leading to the apoptosis of A549 cells and blocking the A549 cells at the G0/G1 phase. Additionally, the complexes caused a decrease of the expression of polyADP-ribose polymerase (PARP), caspase 3, Bcl-2 (B-cell lymphoma-2), PI3K (phosphoinositide-3 kinase) and upregulated the expression of Bax. All these findings indicated that the complexes exert anticancer efficacy to induce cell death through immunogenic cell death, apoptosis, and autophagy. Show less

Four novel PSs (photosensitizers) of nitrogen-heterocyclic ruthenium polypyridyl complexes Ru(dip)₂(o-pipppz)(PF₆)₂ (Ru1) (dip = 4,7-diphenyl-1,10-phenanthroline; o-pipppz = 1-(4-aldehydephenyl)-3-(pyridazyl-2-yl)-1H-pyrazole), Ru(dip)₂(o-pipp) (PF₆)₂ (Ru2) (o-pipp = 1-(4-aldehydephenyl)-3-(pyrid-2-yl)-1H-pyrazole), Ru(dip)₂(m-pipp)(PF₆)₂ (Ru3) (m-pipp = 1-(4-aldehydephenyl)-3-(pyrid-3-yl)-1H-pyrazole) and Ru(dip)₂(p-pipp)(PF₆)₂ (Ru4) (p-pipp = 1-(4-aldehydephenyl)-3-(pyrid-4-yl)-1H-pyrazole) were reported, and the photodynamic activities of these complexes were studied on 2D and 3D HeLa cancer models. The longest visible absorption wavelength of these complexes was approximately 622 nm. The four Ru(II) complexes show preferable photodynamic activity and low dark toxicity (0.2-0.4 μM) in vitro against 2D HeLa tumor cells. These complexes exhibit very high singlet oxygen quantum yields in methanol (0.70-0.95), TPA cross-sections (7-31 GM), and high penetration depth. Thus, Ru1-Ru4 were utilized as one-photon and two-photon absorbing photosensitizers in both monolayer cells and 3D multicellular spheroids (MCSs). Among them, Ru2 revealed a higher singlet oxygen yield (0.95), a larger TPA cross-section (31 GM), and the strongest phototoxicity (EC₅₀ = 0.20 μM). Moreover, flow cytometry shows that the four Ru(II) complexes can induced cell death mainly through apoptosis upon singlet oxygen-dependent reaction. Show less

Title: The ruthenium complex assists in nuclear targeting and selective killing of tumor cells. Abstract: In clinical studies, the toxicity of platinum-based antitumor drugs limits their use. DNA is the most widely studied target of metal-based complexes. Thus, nuclear targeting and selective killing have become the purpose of ruthenium complex design. We synthesized a carboline derivative and its ruthenium complex, NBD and NBD-Ru, and characterized their properties. UV spectra were used to monitor their stability. Transmission electron microscopy and dynamic light scattering were used to identify the self-assembly properties. Inductively coupled plasma mass spectrometry was used to assay the distribution of the Ru complexes in cells with or without transferrin. Besides, the tumor cell killing activities with or without transferrin were detected by MTT assay. An imaging flow cytometer was applied to observe the fluorescence further to identify the cellular distribution. The effects of NBD and NBD-Ru on DNA and the cell cycle were also measured. In vivo, the antitumor and antimetastatic activities of NBD and NBD-Ru were assessed in S180 and LLC tumor-bearing mice. We found that introducing Ru improved the solubility and stability, enabling NBD-Ru to self-assemble into nanoparticles with the EPR effect. At the same time, binding affinity with transferrin increased significantly after complexation, meaning NBD-Ru could target and selectively kill tumors via Tf/TfR pathway. More interestingly, ruthenium assisted the complex in achieving nuclear penetration, which can kill tumor cells by interacting with DNA. In vivo experiments further verified our conclusion in vitro. NBD-Ru could inhibit not only the growth of a primary tumor but also lung metastasis, which was related to the killing effect of the complex on tumor cells (Ki67) and inhibition of neovascularization (CD31). At the same time, the systemic toxicity of the ruthenium complex in vivo was reduced because of the targeting effect, and the biosafety was improved. In conclusion, we found that ruthenium assisted in nuclear targeting and selective killing in vitro and in vivo. Show less

The toxicity of and resistance to platinum complexes as cisplatin, oxaliplatin or carboplatin calls for the replacement of these therapeutic agents in clinical settings. We have previously identified a set of half sandwich-type osmium, ruthenium and iridium complexes with bidentate glycosyl heterocyclic ligands exerting specific cytostatic activity on cancer cells but not on non-transformed primary cells. The apolar nature of the complexes, conferred by large, apolar benzoyl protective groups on the hydroxyl groups of the carbohydrate moiety, was the main molecular feature to induce cytostasis. We exchanged the benzoyl protective groups to straight chain alkanoyl groups with varying length (3 to 7 carbon units) that increased the IC₅₀ value as compared to the benzoyl-protected complexes and rendered the complexes toxic. These results suggest a need for aromatic groups in the molecule. The pyridine moiety of the bidentate ligand was exchanged for a quinoline group to enlarge the apolar surface of the molecule. This modification decreased the IC₅₀ value of the complexes. The complexes containing [(η⁶-p-cymene)Ru(II)], [(η⁶-p-cymene)Os(II)] or [(η⁵-Cp*)Ir(III)] were biologically active unlike the complex containing [(η⁵-Cp*)Rh(III)]. The complexes with cytostatic activity were active on ovarian cancer (A2780, ID8), pancreatic adenocarcinoma (Capan2), sarcoma (Saos) and lymphoma cell lines (L428), but not on primary dermal fibroblasts and their activity was dependent on reactive oxygen species production. Importantly, these complexes were cytostatic on cisplatin-resistant A2780 ovarian cancer cells with similar IC₅₀ values as on cisplatin-sensitive A2780 cells. In addition, the quinoline-containing Ru and Os complexes and the short chain alkanoyl-modified complexes (C3 and C4) proved to be bacteriostatic in multiresistant Gram-positive Enterococcus and Staphylococcus aureus isolates. Hereby, we identified a set of complexes with submicromolar to low micromolar inhibitory constants against a wide range of cancer cells, including platinum resistant cells and against multiresistant Gram-positive bacteria. Show less

Three ruthenium arene complexes, namely {[(η⁶-p-cymene)Ru(Cl)]₂(dpb)}(PF₆)₂ (1), [(η⁶-p-cymene)Ru(dpb)Cl](PF₆) (2) and [(η⁶-p-cymene) Ru(dpb)py](PF₆) (3) (dpb = 2,3-bis(2-pyridyl)benzo-quinoxaline, py = pyridine), were synthesized and their antitumor properties were introduced. Complexes 1-3 were characterized by ¹H NMR, MS, and elemental analysis. As a binuclear ruthenium structure, the absorption of metal ligand electron transfer (MLCT) of 1 extended to 700 nm. Complex 1 was significantly hydrolyzed under dark conditions. The cytotoxicity in vitro study showed that complexes 1 and 2 are more toxic to human lung cancer cells (A549) and human cervial cancer cells (Hela) than cisplatin. Moreover, there was almost no cross-resistance between complex 1-2 and cisplatin. Under the irradiation at 478 nm, complexes 1-3 all produced singlet oxygen (¹O₂), and the ¹O₂ quantum yield of complex 1 in PBS is the highest among complexes 1-3. Complex 1 also produced ¹O₂ under 600 nm light irradiation. DNA gel electrophoresis showed that 1 caused the photocleavage of plasmid DNA. The hydrolysis rate of complex 1 was accelerated under light (λ > 600 nm). And the phototoxicity of complex 1 to Hela cells under light (λ > 600 nm) was much greater than its dark toxicity, which may be due to its generation of ¹O₂ and the promotion of its hydrolysis under long-wave light irradiation. Show less

The potential use of Ru(II) complexes as photosensitizers (PSs) in photodynamic therapy (PDT) has gained significant attention. In comparison with fluorophores with aggregation-caused quenching (ACQ), fluorophores with aggregation-induced emission (AIE) characteristics exhibit sustained fluorescence and dispersibility in aqueous solutions. PSs with AIE characteristics have received much attention in recent years. Herein, we reported two novel biotin-conjugated Ru(II) polypyridyl complexes (Ru1 and Ru2) with AIE characteristics. When exposed to 460 nm (10 mW cm^-2) light, Ru1 and Ru2 exhibited outstanding photostability and photocatalytic activity. Ru1 and Ru2 could efficiently generate singlet oxygen and induce pUC19 DNA photolysis when exposed to 460 nm light. Interestingly, both Ru1 and Ru2 also functioned as catalysts for NADH oxidation when exposed to 460 nm light. The presence of biotin fragments in Ru1 and Ru2 enhanced the specific uptake of these complexes by tumor cells. Both complexes showed minimal toxicity to selected cells in the dark. Nevertheless, the phototoxicity of both complexes significantly increased upon 460 nm light irradiation for 15 min. Further experiments revealed that Ru2 primarily accumulated in mitochondria and might bind to mitochondrial DNA. Under 460 nm light irradiation, Ru2 induced the generation of reactive oxygen species (ROS) and NADH depletion disrupting intracellular redox homeostasis in A549 cells, activating the mitochondrial apoptosis pathway resulting in up-regulation of apoptotic marker caspase-3, effectively damaged A549 cell DNA and arrested A549 cell cycle in the S phase. In vivo anti-tumor experiments were conducted to assess the effects of Ru2 on tumor growth in A549 tumor-bearing mice. The results showed that Ru2 effectively inhibited tumor growth under 460 nm light irradiation conditions. These findings indicate that Ru2 has great potential as a targeted photosensitizer for mitochondrial targeting imaging and photodynamic therapy of tumors. Show less

The rapid efflux of Pt-based chemotherapeutics by cancer cells is one of the major causes of drug resistance in clinically available drugs. Therefore, both the high cellular uptake as well as adequate retention efficiency of an anticancer agent are important factors to overcome drug resistance. Unfortunately, rapid and efficient quantification of metallic drug concentration in individual cancer cells still remains a tricky problem. Herein, with the help of newly developed single cell inductively coupled plasma mass spectrometry (SC-ICP-MS), we have found that the well-known Ru(II)-based complex, Ru3, displayed remarkable intracellular uptake and retention efficiency in every single cancer cell with high photocatalytic therapeutic activity to overcome cisplatin resistance. Moreover, Ru3 has shown sensational photocatalytic anticancer properties with excellent in-vitro and in-vivo biocompatibility under light exposure. Show less

Six half-sandwich Ru(II) complexes (Ru1-Ru6), integrated with 5-phenyl-2-(pyridin-2-yl)-2,4-dihydro-3H-pyrazol-3-one (PDPO1-PDPO6) ligands, were synthesized and spectroscopically characterized. The structure of Ru3 that crystallized as a monoclinic crystal with P21/c space group was further confirmed by X-ray single crystal diffraction. Prototropic tautomerism within the complexes transformed OH-form ligands to NH-form, forming a hydrogen bond (Cl₁---H-N₃). The complexes and ligands' cytotoxicity was assessed against several cancerous (HepG2, A549, MCF-7) and normal Vero cell lines. Relative to the ligands and Cisplatin, complexes (Ru2, Ru3, Ru5, Ru6) exhibited potent cytotoxicity against cancer cells, with IC₅₀ values from 2.05 to 15.69 μM/L, excluding Ru1 and Ru4. Specifically, Ru2, Ru3, and Ru5 demonstrated superior anti-HepG2 properties. Compared to Cisplatin, Ru2 and Ru5 were less toxic to Vero cells, highlighting their enhanced selectivity in toxicity. Structure-activity relationship (SAR) studies indicated that t-butyl substitution (in Ru2) or -Cl (in Ru5) on the benzene ring significantly improved the selective toxicity. These complexes manifested substantial lipophilicity, cellular uptake, and were quickly hydrolyzed to Ru-H₂O species. Roughly positive correlations were observed between hydrolysis rate, lipophilicity, cellular uptake, and anticancer activities. Ru2, investigated specifically, induced apoptosis in HepG2 cells at concentrations of 10 and 20 μM/L through ROS-mediated mitochondrial dysfunction and G0/G1phase arrest, associated with altered P21, cyclin D, and CDK4 expression levels. Show less

Metallodrugs represent a combination of multifunctionalities that are present concomitantly and can act differently on diverse biotargets. Their efficacy is often related to the lipophilic features exhibited both by long carbo-chains and the phosphine ligands. Three Ru(II) complexes containing hydroxy stearic acids (HSAs) were successfully synthesized in order to evaluate possible synergistic effects between the known antitumor activity of HSA bio-ligands and the metal center. HSAs were reacted with [Ru(H)₂CO(PPh₃)₃] selectively affording O,O-carboxy bidentate complexes. The organometallic species were fully characterized spectroscopically using ESI-MS, IR, UV-Vis, and NMR techniques. The structure of the compound Ru-12-HSA was also determined using single crystal X-ray diffraction. The biological potency of ruthenium complexes (Ru-7-HSA, Ru-9-HSA, and Ru-12-HSA) was studied on human primary cell lines (HT29, HeLa, and IGROV1). To obtain detailed information about anticancer properties, tests for cytotoxicity, cell proliferation, and DNA damage were performed. The results demonstrate that the new ruthenium complexes, Ru-7-HSA and Ru-9-HSA, possess biological activity. Furthermore, we observed that the Ru-9-HSA complex shows increased antitumor activity on colon cancer cells, HT29. Show less

To enhance the cytoselective behavior of the complexes, we intended to develop a CuAAC "click"-derived synthetic protocol for the preparation of 2-(2-(4-(4-(pyridin-2-yl)-1H-1,2,3-triazol-1-yl)butoxy)phenyl)benzo[d]thiazole-based Ru(II)/Ir(III)/Re(I) complexes, and their cytotoxicity against three different cancer cell lines (MCF-7, HeLa, and U87MG) in consort with one normal cell line (HEK-293) was evaluated. In our detailed investigations, the significant cytotoxic nature of the Ru(II) complex 7a compared to Ir(III) and Re(I) complexes (7b and 7c, respectively) was observed. Complex 7a was capable of MCF-7 cell apoptosis via the inhibition of both S- and G2/M-phase cell cycle arrest in association with a substantial quantity of ROS production and DNA intercalation. Show less

A second-generation series of biscyclometalated 2-(5-aryl-thienyl)-benzimidazole and -benzothiazole Ir(III) dppz complexes [Ir(C^N)₂(dppz)]⁺, Ir1-Ir4, were rationally designed and synthesized, where the aryl group attached to the thienyl ring was p-CF₃C₆H₄ or p-Me₂NC₆H₄. These new Ir(III) complexes were assessed as photosensitizers to explore the structure-activity correlations for their potential use in biocompatible anticancer photodynamic therapy. When irradiated with blue light, the complexes exhibited high selective potency across several cancer cell lines predisposed to photodynamic therapy; the benzothiazole derivatives (Ir1 and Ir2) were the best performers, Ir2 being also activatable with green or red light. Notably, when irradiated, the complexes induced leakage of lysosomal content into the cytoplasm of HeLa cancer cells and induced oncosis-like cell death. The capability of the new Ir complexes to photoinduce cell death in 3D HeLa spheroids has also been demonstrated. The investigated Ir complexes can also catalytically photo-oxidate NADH and photogenerate ¹O₂ and/or ^•OH in cell-free media. Show less

Half-sandwich iridium(III) complexes show potential value in the anticancer field. However, complexes with favorable luminescence performance are rare, which limits further investigation of the anticancer mechanism. In this paper, 10 triphenylamine-modified fluorescent half-sandwich iridium(III) pyridine complexes {[(η⁵-Cp^x)Ir(L)Cl₂]} (Ir1-Ir10) were prepared and showed potential antiproliferative activity, effectively inhibiting the migration of A549 cells. Ir6, showing the best activity among these complexes, exhibited excellent fluorescence performance (absolute fluorescence quantum yield of 15.17%) in solution. Laser confocal detection showed that Ir6 followed an energy-dependent cellular uptake mechanism, specifically accumulating in mitochondria (Pearson co-localization coefficient of 0.95). A Western blot assay further confirmed the existence of a mitochondrial apoptotic channel. Additionally, Ir6 could arrest the cell cycle at the G₂/M phase, catalyze NADH oxidation, reduce the mitochondrial membrane potential, induce an increase in the level of intracellular reactive oxygen species, and exhibit a mechanism of oxidation. An in vivo antitumor assay confirmed that Ir6 can effectively inhibit tumor growth and is safer than cisplatin. Show less

Conventional photodynamic therapy mainly causes a therapeutic effect on the primary tumor through the localized generation of reactive oxygen species, while metastatic tumors remain poorly affected. Complementary immunotherapy is effective in eliminating small, non-localized tumors distributed across multiple organs. Here, we report the Ir(iii) complex Ir-pbt-Bpa as a highly potent immunogenic cell death inducing photosensitizer for two-photon photodynamic immunotherapy against melanoma. Ir-pbt-Bpa can produce singlet oxygen and superoxide anion radicals upon light irradiation, causing cell death by a combination of ferroptosis and immunogenic cell death. In a mouse model with two physically separated melanoma tumors, although only one of the primary tumors was irradiated, a strong tumor reduction of both tumors was observed. Upon irradiation, Ir-pbt-Bpa not only induced the immune response of CD8⁺ T cells and the depletion of regulatory T cells, but also caused an increase in the number of the effector memory T cells to achieve long-term anti-tumor immunity. Show less

Title: Mitochondria-targeted cyclometalated iridium (III) complexes: Dual induction of A549 cells apoptosis and autophagy. Abstract: In this study, we synthesized 4 cyclometalated iridium complexes using N-(1,10-phenanthrolin-5-yl)picolinamide (PPA) as the main ligand, denoted as [Ir(ppy)2PPA]PF6 (ppy = 2-phenylpyridine, Ir1), [Ir(bzq)2PPA]PF6 (bzq = benzo[h]quinoline, Ir2), [Ir(dfppy)2PPA]PF6 (dfppy = 2-(3,5-difluorophenyl)pyridine, Ir3), and [Ir(thpy)2PPA]PF6 (thpy = 2-(thiophene-2-yl)pyridine, Ir4). Compared to cisplatin and oxaliplatin, all four complexes exhibited significant anti-tumor activity. Among them, Ir2 demonstrated higher cytotoxicity against A549 cells, with an IC50 value of 1.6 ± 0.2 μM. The experimental results indicated that Ir2 primarily localized in the mitochondria, inducing a large amount of reactive oxygen species (ROS) generation, that decreased in mitochondrial membrane potential (MMP), reduced ATP production, and further impaired mitochondrial function, leading to cytochrome c release. Additionally, Ir2 caused cell cycle arrest at the S phase and induced apoptosis through the AKT-mediated signaling pathway. Further investigations revealed that Ir2 could simultaneously induce both apoptosis and autophagy in A549 cells, with the latter acting as a non-protective mechanism that promoted cell death. More importantly, Ir2 exhibited low toxicity to both normal LO2 cells in vitro and zebrafish embryos in vivo. Consequently, these newly developed Ir(III) complexes show great potential in the development of novel and low-toxicity anticancer agents. Show less

Title: Iridium(III) complexes inhibit the proliferation and migration of BEL-7402 cells through the PI3K/AKT/mTOR signaling pathway. Abstract: Iridium(III) complexes are largely studied as anti-cancer complexes due to their excellent anti-cancer activity. In this article, two new iridium(III) complexes [Ir(piq)2(THPIP)]PF6 (THPIP = 2,4-di-tert-butyl-6-(1H-imidazo[4,5-f][1,10]phenanthrolin-2-yl)phenol, piq = deprotonated 1-phenylisoquinoline) (Ir1) and [Ir(bzq)2(THPIP)]PF6 (bzq = deprotonated benzo[h]quinolone) (Ir2) were synthesized. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays showed that complex Ir1 exhibits moderate activity (IC50 = 29.9 ± 4.6 μM) and Ir2 shows high cytotoxicity (IC50 = 9.8 ± 1.8 μM) against BEL-7402 cells. Further studies on the mechanism showed that Ir1 and Ir2 induced apoptosis by changing the mitochondrial membrane potential, Ca2+ release, ROS accumulation, and cell cycle arrest at the S phase. The complexes can effectively inhibit cell colony formation and migration. The expression of B-cell lymphoma-2 (Bcl-2) family proteins, PI3K (phosphatidylinositol 3-kinase), AKT (protein kinase B), mTOR (mammalian target of rapamycin), and p-mTOR was studied by immunoblotting. Complexes Ir1 and Ir2 downregulated the expression of anti-apoptotic protein Bcl-2 and increased the expression of autophagy-related proteins of Beclin-1 and LC3-II. Further experiments showed that the complexes inhibited the production of glutathione (GSH) and increased the amounts of malondialdehyde (MDA). Fluorescence of HMGB1 was significantly increased. We also investigated the effect of the complexes on the expression of genes using RNA-sequence analysis, we further calculated the lowest binding energies between the complexes and proteins using molecular docking. Taken together, the above results indicated that complexes Ir1 and Ir2 induce apoptosis in BEL-7402 cells through a ROS-mediated mitochondrial dysfunction and inhibition of the PI3K/AKT/mTOR signaling pathway. Show less

Title: Design, synthesis and biological evaluation of liposome entrapped iridium(III) complexes toward SGC-7901 cells. Abstract: In this study, two new iridium(III) polypyridyl complexes [Ir(bzq)2(DIPH)](PF6) (bzq = deprotonated benzo[h]quinoline, DIPH = 4-(2,5-dibromo-4-(1H-imidazo[4,5-f][1,10]phenanthrolim-2-yl)-4-hydroxybutan-2-one) (Ir1) and [Ir(piq)2(DIPH)](PF6) (piq = deprotonated 1-phenylisoquinoline) (Ir2) were synthesized and characterized by elemental analysis, HRMS, 1H and 13C NMR. The cytotoxic activity of Ir1, Ir2, Ir1lipo and Ir2lipo against cancer cells SGC-7901, HepG2, A549, HeLa, B16 and normal NIH3T3 cells in vitro was evaluated using 3-(4,5-dimethylthiazole-2-yl)-2,5-biphenyl tetrazolium bromide (MTT) method. Ir1 and Ir2 showed no cytotoxic activity, but their liposome-entrapped Ir1 (Ir1lipo) and Ir2 (Ir2lipo) showed significant cellular activity, especially sensitive to SGC-7901 with IC50 values of 4.7 ± 0.2 and 12.4 ± 0.5 μM, respectively. The cellular uptake, endoplasmic reticulum (ER) localization, autophagy, tubulin polymerization, glutathione (GSH), malondialdehyde (MDA) and release of cytochrome c were investigated to explore the mechanisms of apoptosis. The calreticulin (CRT), heat shock protein 70 (HSP70), high mobility group box 1 (HMGB1) were also explored. Western blotting showed that Ir1lipo and Ir2lipo inhibited PI3K (phosphoinositide-3 kinase), AKT (protein kinase B), p-AKT and activated Bcl-2 (B-cell lymphoma-2) protein and apoptosis-regulated factor caspase 3 (cysteinyl aspartate specific proteinase-3) and cleaving PARP (poly ADP-ribose polymerase). The results demonstrated that Ir1lipo and Ir2lipo induce cell apoptosis through targeting the endoplasmic reticulum (ER), cause oxidative stress damage, inhibiting PI3K/AKT signaling pathway, immunogenic cell death (ICD) and inhibit the cell growth at G2/M phase. Show less

Title: Targeted liposomes encapsulated iridium(III) compound greatly enhance anticancer efficacy and induce cell death via ferroptosis on HepG2 cells. Abstract: In this study, ligands 2-phenyl-1H-imidazo[4,5-f][1,10]phenanthroline (PIP), 2-(2-nitrophenyl)-1H-imidazo[4,5-f][1,10]phenanthroline (NPIP), 2-(2-nitronaphthalen-1-yl)-1H-imidazo[4,5-f][1,10]phenanthroline (NNIP) and their iridium(III) metal compounds [Ir(ppy)2(PIP)](PF6) (ppy = 2-phenylpyridine, 1a), [Ir(ppy)2(NPIP)](PF6) (1b), [Ir(ppy)2(NNIP)](PF6) (1c) were designed and synthesized. The anti-cancer activities of 1a, 1b and 1c on BEL-7402, HepG2, SK-Hep1 and non-cancer LO2 were detected using MTT method. 1a shows moderate, 1b and 1c display low or no anti-cancer activities. To elevate the anti-cancer effectiveness, encapsulating the compounds 1a, 1b and 1c into the ordinary or targeted liposomes to produce 1alip, 1blip, 1clip, or targeted 1aTlip, 1bTlip and 1cTlip. The IC50 values of 1alip, 1blip, 1clip, 1aTlip, 1bTlip and 1cTlip against HepG2 cells are 7.9 ± 0.1, 8.6 ± 0.2, 16.9 ± 0.5, 5.9 ± 0.2, 7.3 ± 0.1 and 9.7 ± 0.7 μM, respectively. Specifically, the anti-tumor activity assays in vivo found that the inhibitory rates are 23.24 % for 1a, 61.27 % for 1alip, 76.06 % for 1aTlip. It is obvious that the targeted liposomes entrapped iridium(III) compound greatly enhance anti-cancer efficacy. Additionally, 1alip, 1blip and 1clip or targeted 1aTlip, 1bTlip and 1cTlip can effectively restrain the cell colony and proliferation in the G0/G1 period. 1alip, 1blip, 1clip, 1aTlip, 1bTlip and 1cTlip can increase reactive oxygen species (ROS) concentration, arouse a decline in the mitochondrial membrane potential and promote Ca2+ release. RNA-sequence was applied to examine the signaling pathways. Taken together, the liposomes or targeted liposomes encapsulated compounds trigger cell death by way of apoptosis, autophagy, ferroptosis, disruption of mitochondrial function and PI3K/AKT/mTOR signaling pathways. Show less

Title: Synthesis, characterisation and biological evaluation of monometallic Re(I) and heterobimetallic Re(I)/Fe(II) complexes with a 1,2,3-triazolyl pyridine chelating moiety. Abstract: Bioorganometallic complexes have attracted considerable interest and have shown promise for potential application in the treatment and diagnosis of cancer, as well as bioimaging agents, some acting as theranostic agents. The series of novel ferrocene, benzimidazo[1,2-a]quinoline and fluorescein derivatives with bidentate pyridyl-1,2,3-triazole and 2,2'-dipyridylamine and their tricarbonylrhenium(I) complexes was prepared and fully characterised by NMR, single-crystal X-ray diffraction, UV-Vis and fluorescence spectroscopy in biorelevant conditions. The fluorescein and benzimidazo[1,2-a]quinoline ligands and their complexes with Re(I) showed interactions with ds-DNA/RNA and HSA, characterised by thermal denaturation measurements, fluorimetric and circular dichroism titrations. The binding constants revealed that addition of Re(I) increases the affinity of fluorescein but decreases the affinity of benzimidazo[1,2-a]quinoline. The complexation of Re(I) had the opposite effect on fluorescein and benzimidazo[1,2-a]quinoline ligands' fluorimetric sensitivity upon biomacromolecule binding, Re(I) fluorescein complex emission being strongly quenched by DNA/RNA or HSA, while emission of Re(I) benzimidazo[1,2-a]quinolone complex was enhanced, particularly for HSA, making it a promising fluorescent probe. Some mono- and heterobimetallic complexes showed considerable antiproliferative activity on colon cancer cells (CT26 and HT29), with ferrocene dipyridylamine complexes exhibiting the best inhibitory activity, comparable to cisplatin. The correlation of the cytotoxicity data with the linker type between the ferrocene and the 1,2,3-triazole ring suggests that direct binding of the metallocene to the 1,2,3-triazole is favourable for antitumor activity. The Re(I) benzimidazo[1,2-a]quinolone complex showed moderate antiproliferative activity, in contrast to the Re(I) fluorescein complex, which exhibited weak activity on CT26 cells and no activity on HT29 cells. The accumulation of the Re(I) benzimidazo[1,2-a]quinolone complex in the lysosomes of CT26 cells indicates the site of its bioactivity, thus making this complex a potential theranostic agent. Show less

Five metal-arene complexes of formula [MX₂(η⁶-p-cymene)(diR(1-pyrenyl)phosphane)] (M = Os or Ru, X = Cl or I, R = isopropyl or phenyl) and symbolized as MRX2 were synthesized and fully characterized, namely OsiPrCl2, OsiPrI2, OsPhCl2, OsPhI2 and RuPhI2. Furthermore, nine cyclometalated half-sandwich complexes of formula [MX-(η⁶-p-cymene)(k²C-diR(1-pyrenyl)phosphane)] (M = Os or Ru, X = Cl or I, R = isopropyl or phenyl) or [M(η⁶-p-cymene)(kS-dmso)(k²C-diR(1-pyrenyl)phosphane)]PF₆ (M = Os or Ru, R = isopropyl or phenyl) and symbolized as c-MRX were prepared; hence, c-OsiPrCl, c-OsiPrI, c-OsiPrdmso, c-OsPhCl, c-OsPhI, c-OsPhdmso, c-RuPhCl, c-RuPhI and c-RuPhdmso were obtained and fully characterized. The crystal structures of ten out of the fourteen complexes were solved. All complexes exhibit notable cytotoxic properties against A549 (Lung Adenocarcinoma) human cells, with IC₅₀ values ranging from 48 to 1.42 μM. In addition, complex c-OsiPrdmso shows remarkable toxic behaviours agains other cell lines, namely MCF7 (breast carcinoma), MCF10A (non-tumorigenic epithelial breast) and MDA-MB-435 (melanoma) human cells, as illustrated by IC₅₀ values of 4.36, 4.71 and 2.32 μM, respectively. Finally, it has been found that OsiPrI2 affects the cell cycle of A549 cells, impeding their replication (i.e., the cell cycle is blocked), whereas OsPhI2 (namely with phenyl groups instead of isopropyl ones) does not induce this effect. Show less

We present the synthesis and characterization of six new heteroleptic osmium(II) complexes of the type [Os(C^N)(N^N)₂]OTf (N^N = 2,2'-bipyridine and dipyrido[3,2-d:2',3'-f]quinoxaline; C^N = deprotonated methyl 1-butyl-2aryl-benzimidazolecarboxylate) with varying substituents in the R3 position of the phenyl ring of the cyclometalating C^N ligand. The new compounds are highly kinetically inert and absorb a full-wavelength range of visible light. An investigation of the antiproliferative activity of the new compounds has been performed using a panel of human cancer and noncancerous 2D cell monolayer cultures under dark conditions and green light irradiation. The results demonstrate that the new Os(II) complexes are markedly more potent than conventional cisplatin. The promising antiproliferative activity of selected Os(II) complexes was also confirmed using 3D multicellular tumor spheroids, which have the characteristics of solid tumors and can mimic the tumor tissue microenvironment. The mechanism of antiproliferative action of complexes has also been investigated and revealed that the investigated Os(II) complexes activate the endoplasmic reticulum stress pathway in cancer cells and disrupt calcium homeostasis. Show less

Title: Structurally Simple Osmium(II) Polypyridyl Complexes as Photosensitizers for Photodynamic Therapy in the Near Infrared. Abstract: Five osmium(II) polypyridyl complexes of the general formula [Os(4,7-diphenyl-1,10-phenanthroline)2 L]2+ were synthesized as photosensitizers for photodynamic therapy by varying the nature of the ligand L. Thanks to the pronounced π-extended structure of the ligands and the heavy atom effect provided by the osmium center, these complexes exhibit a high absorption in the near-infrared (NIR) region (up to 740 nm), unlike related ruthenium complexes. This led to a promising phototoxicity in vitro against cancer cells cultured as 2D cell layers but also in multicellular tumor spheroids upon irradiation at 740 nm. The complex [Os(4,7-diphenyl-1,10-phenanthroline)2 (2,2'-bipyridine)]2+ was found to be the most efficient against various cancer cell lines, with high phototoxicity indexes. Experiments on CT26 tumor-bearing BALB/c mice also indicate that the OsII complexes could significantly reduce tumor growth following 740 nm laser irradiation. The high phototoxicity in the biological window of this structurally simple complex makes it a promising photosensitizer for cancer treatment. Show less

Mechanisms of defense against ferroptosis (an iron-dependent form of cell death induced by lipid peroxidation) in cellular organelles remain poorly understood, hindering our ability to target ferroptosis in disease treatment. In this study, metabolomic analyses revealed that treatment of cancer cells with glutathione peroxidase 4 (GPX4) inhibitors results in intracellular glycerol-3-phosphate (G3P) depletion. We further showed that supplementation of cancer cells with G3P attenuates ferroptosis induced by GPX4 inhibitors in a G3P dehydrogenase 2 (GPD2)-dependent manner; GPD2 deletion sensitizes cancer cells to GPX4 inhibition-induced mitochondrial lipid peroxidation and ferroptosis, and combined deletion of GPX4 and GPD2 synergistically suppresses tumor growth by inducing ferroptosis in vivo. Mechanistically, inner mitochondrial membrane-localized GPD2 couples G3P oxidation with ubiquinone reduction to ubiquinol, which acts as a radical-trapping antioxidant to suppress ferroptosis in mitochondria. Taken together, these results reveal that GPD2 participates in ferroptosis defense in mitochondria by generating ubiquinol. Show less

CCAAT/enhancer binding protein β (C/EBPβ) is a basic leucine zipper (bZIP) family transcription factor, which is upregulated or overactivated in many cancers, resulting in a gene expression profile that drives oncogenesis. C/EBPβ dimerization regulates binding to DNA at the canonical TTGCGCAA motif and subsequent transcriptional activity, suggesting that disruption of dimerization represents a powerful approach to inhibit this previously "undruggable" oncogenic target. Here we describe the mechanism of action and antitumor activity of ST101, a novel and selective peptide antagonist of C/EBPβ that is currently in clinical evaluation in patients with advanced solid tumors. ST101 binds the leucine zipper domain of C/EBPβ, preventing its dimerization and enhancing ubiquitin-proteasome dependent C/EBPβ degradation. ST101 exposure attenuates transcription of C/EBPβ target genes, including a significant decrease in expression of survival, transcription factors, and cell-cycle-related proteins. The result of ST101 exposure is potent, tumor-specific in vitro cytotoxic activity in cancer cell lines including glioblastoma, breast, melanoma, prostate, and lung cancer, whereas normal human immune and epithelial cells are not impacted. Further, in mouse xenograft models ST101 exposure results in potent tumor growth inhibition or regression, both as a single agent and in combination studies. These data provide the First Disclosure of ST101, and support continued clinical development of ST101 as a novel strategy for targeting C/EBPβ-dependent cancers. Show less

While ruthenium arene complexes have been widely investigated for their medicinal potential, studies on homologous compounds containing a tridentate tris(1-pyrazolyl)methane ligand are almost absent in the literature. Ruthenium(II) complex 1 was obtained by a modified reported procedure; then, the reactions with a series of organic molecules (L) in boiling alcohol afforded novel complexes 2-9 in 77-99% yields. Products 2-9 were fully structurally characterized. They are appreciably soluble in water, where they undergo partial chloride/water exchange. The antiproliferative activity was determined using a panel of human cancer cell lines and a noncancerous one, evidencing promising potency of 1, 7, and 8 and significant selectivity toward cancer cells. The tested compounds effectively accumulate in cancer cells, and mitochondria represent a significant target of biological action. Most notably, data provide convincing evidence that the mechanism of biological action is mediated by the inhibiting of mitochondrial calcium intake. Show less