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Colorectal cancer (CRC) has been addressed in the framework of molecular, cellular biology, and biochemical traits. A new approach to studying CRC is focused on the relationship between biochemical pathways and biophysical cues, which may contribute to disease understanding and therapy development. Herein, we investigated the mechanical properties of CRC cells, namely, HCT116, HCT15, and SW620, using static and dynamic methodologies by atomic force microscopy (AFM). The static method quantifies Young's modulus; the dynamic method allows the determination of elasticity, viscosity, and fluidity. AFM results were correlated with confocal laser scanning microscopy and cell migration assay data. The SW620 metastatic cells presented the highest Young's and storage moduli, with a defined cortical actin ring with distributed F-actin filaments, scarce vinculin expression, abundant total focal adhesions (FAK), and no filopodia formation, which could explain the lessened migratory behavior. In contrast, HCT15 cells presented lower Young's and storage moduli, high cortical tubulin, less cortical F-actin and less FAK, and more filopodia formation, probably explaining the higher migratory behavior. HCT116 cells presented Young's and storage moduli values in between the other cell lines, high cortical F-actin expression, intermediate levels of total FAK, and abundant filopodia formation, possibly explaining the highest migratory behavior. Show less

Photoreactive Ru(II) complexes capable of ejecting ligands have been used extensively for photocaging applications and for the creation of "photocisplatin" reagents. The incorporation of distortion into the structure of the coordination complex lowers the energy of dissociative excited states, increasing the yield of the photosubstitution reaction. While steric clash between ligands induced by adding substituents at the coordinating face of the ligand has been extensively utilized, a lesser known, more subtle approach is to distort the coordination sphere by altering the chelate ring size. Here a systematic study was performed to alter metal-ligand bond lengths, angles, and to cause intraligand distortion by introducing a "linker" atom or group between two pyridine rings. The synthesis, photochemistry, and photobiology of five Ru(II) complexes containing CH₂, NH, O, and S-linked dipyridine ligands was investigated. All systems where stable in the dark, and three of the five were photochemically active in buffer. While a clear periodic trend was not observed, this study lays the foundation for the creation of photoactive systems utilizing an alternative type of distortion to facilitate photosubstitution reactions. Show less

Mitochondria are key regulators of many important cellular processes and their dysfunction has been implicated in a large number of human disorders. Importantly, mitochondrial function is tightly linked to their ultrastructure, which possesses an intricate membrane architecture defining specific submitochondrial compartments. In particular, the mitochondrial inner membrane is highly folded into membrane invaginations that are essential for oxidative phosphorylation. Furthermore, mitochondrial membranes are highly dynamic and undergo constant membrane remodeling during mitochondrial fusion and fission. It has remained enigmatic how these membrane curvatures are generated and maintained, and specific factors involved in these processes are largely unknown. This review focuses on the current understanding of the molecular mechanism of mitochondrial membrane architectural organization and factors critical for mitochondrial morphogenesis, as well as their functional link to human diseases. Show less

Cryo-electron microscopy studies of Escherichia coli complex I suggest a conserved mechanism of coupled proton transfers and electrostatic interactions that result in proton ejection from the complex exclusively at the distal NuoL subunit. Show less

Acute inflammation or resolved inflammation is an adaptive host defense mechanism and is self-limiting, which returns the body to a state of homeostasis. However, unresolved, uncontrolled, or chronic inflammation may lead to various maladies, including cancer. Important evidence that links inflammation and cancer is that nonsteroidal anti-inflammatory drugs (NSAIDs), such as aspirin, reduce the risk and mortality from many cancers. The fact that NSAIDs inhibit the eicosanoid pathway prompted mechanistic drug developmental work focusing on cyclooxygenase (COX) and its products. The increased prostaglandin E2 levels and the overexpression of COX-2 in the colon and many other cancers provided the rationale for clinical trials with COX-2 inhibitors for cancer prevention or treatment. However, NSAIDs do not require the presence of COX-2 to prevent cancer. In this review, we highlight the effects of NSAIDs and selective COX-2 inhibitors (COXIBs) on targets beyond COX-2 that have shown to be important against many cancers. Finally, we hone in on specialized pro-resolving mediators (SPMs) that are biosynthesized locally and, in a time, -dependent manner to promote the resolution of inflammation and subsequent tissue healing. Different classes of SPMs are reviewed, highlighting aspirin's potential in triggering the production of these resolution-promoting mediators (resolvins, lipoxins, protectins, and maresins), which show promise in inhibiting cancer growth and metastasis. Show less

Lipid-conjugated Ru(III) complexes - designed to obtain lipophilic analogues of the low molecular weight derivative AziRu, which is a NAMI-A-like anticancer agent - have been synthesized and fully characterized. A detailed biophysical investigation, including multiple, integrated techniques, allowed determining their molecular and self-assembling properties in aqueous solutions mimicking the extracellular environment, showing that our design produced a protective effect from hydrolysis of the Ru(III) complexes. In vitro biological experiments, carried out in comparison with AziRu, demonstrated that, among the novel lipophilic Ru(III) complexes synthesized, the compounds derivatized with palmitic and stearic acid, that we named PalmiPyRu and StePyRu respectively, showed attractive features and a promising antiproliferative activity, selective on specific breast cancer phenotypes. To get a deeper insight into their interactions with potential biomacromolecular targets, their ability to bind both bovine serum albumin (BSA), an abundant serum carrier protein, and some DNA model systems, including duplex and G-quadruplex structures, has been investigated by spectroscopic techniques. Inductively coupled plasma-mass spectrometry (ICP-MS) analysis of the ruthenium amount incorporated in human MCF-7 and MDA-MB-231 breast cancer cells, after incubation in parallel experiments with PalmiPyRu and AziRu, showed a markedly higher cell uptake of the lipophilic Ru(III) complex with respect to AziRu. These data confirmed that the proper lipidic tail decorating the metal complex not only favoured the formation of aggregates in the extracellular media but also improved their cell membrane penetration, thus leading to higher antiproliferative activity selective on breast cancer cells. Show less

Abstract In eukaryotes, three RNA polymerases (RNAPs) play essential roles in the synthesis of various types of RNA: namely, RNAPI for rRNA; RNAPII for mRNA and most snRNAs; and RNAPIII for tRNA and other small RNAs. All three RNAPs possess a short flexible tail derived from their common subunit RPB6. However, the function of this shared N-terminal tail (NTT) is not clear. Here we show that NTT interacts with the PH domain (PH-D) of the p62 subunit of the general transcription/repair factor TFIIH, and present the structures of RPB6 unbound and bound to PH-D by nuclear magnetic resonance (NMR). Using available cryo-EM structures, we modelled the activated elongation complex of RNAPII bound to TFIIH. We also provide evidence that the recruitment of TFIIH to transcription sites through the p62–RPB6 interaction is a common mechanism for transcription-coupled nucleotide excision repair (TC-NER) of RNAPI- and RNAPII-transcribed genes. Moreover, point mutations in the RPB6 NTT cause a significant reduction in transcription of RNAPI-, RNAPII- and RNAPIII-transcribed genes. These and other results show that the p62–RPB6 interaction plays multiple roles in transcription, TC-NER, and cell proliferation, suggesting that TFIIH is engaged in all RNAP systems. Show less

Title: Ursolic acid-piperazine-dithiocarbamate ruthenium(II) polypyridyl complexes induced necroptosis in MGC-803 cells. Abstract: Three ursolic acid-piperazine-dithiocarbamate ruthenium(II) polypyridyl complexes Ru1-Ru3 were designed and synthesized for evaluating antitumor activity. All the complexes exhibited high in vitro cytotoxicity against MGC-803, T24, HepG2, CNE2, MDA-MB-231, MCF-7, A549, and A549/DDP cell lines. Ru1, Ru2, and Ru3 were 11, 8 and 10 times, respectively, more active than cisplatin against A549/DDP. An in vivo study on MGC-803 xenograft mouse models demonstrated that representative Ru2 exhibited an effective inhibitory effect on tumor growth, showing stronger antitumor activity than cisplatin. Biological investigations suggested that Ru2 entered MGC-803 cells by a clathrin-mediated endocytic pathway, initially localizing in the lysosomes and subsequently escaping and localizing in the mitochondria. Mitochondrial swelling resulted in vacuolization, which induced vacuolation-associated cell death and necroptosis with the formation of necrosomes (RIP1-RIP3) and the uptake of propidium iodide. These results demonstrate that the potential of Ru2 as a chemotherapeutic agent to kill cancer cells via a dual mechanism represents an alternative way to eradicate apoptosis-resistant forms of cancer. Show less

Glioblastoma (GBM) is a highly malignant brain tumor characterized by a heterogeneous population of genetically unstable and highly infiltrative cells that are resistant to chemotherapy. Although substantial efforts have been invested in the field of anti-GBM drug discovery in the past decade, success has primarily been confined to the preclinical level, and clinical studies have often been hampered due to efficacy-, selectivity-, or physicochemical property-related issues. Thus, expansion of the list of molecular targets coupled with a pragmatic design of new small-molecule inhibitors with central nervous system (CNS)-penetrating ability is required to steer the wheels of anti-GBM drug discovery endeavors. This Perspective presents various aspects of drug discovery (challenges in GBM drug discovery and delivery, therapeutic targets, and agents under clinical investigation). The comprehensively covered sections include the recent medicinal chemistry campaigns embarked upon to validate the potential of numerous enzymes/proteins/receptors as therapeutic targets in GBM. Show less

AbstractProfiling approaches have been increasingly employed for the characterization of disease‐relevant phenotypes or compound perturbation as they provide a broad, unbiased view on impaired cellular states. We report that morphological profiling using the cell painting assay (CPA) can detect modulators of de novo pyrimidine biosynthesis and of dihydroorotate dehydrogenase (DHODH) in particular. The CPA can differentiate between impairment of pyrimidine and folate metabolism, which both affect cellular nucleotide pools. The identified morphological signature is shared by inhibitors of DHODH and the functionally tightly coupled complex III of the mitochondrial respiratory chain as well as by UMP synthase, which is downstream of DHODH. The CPA appears to be particularly suited for the detection of DHODH inhibitors at the site of their action in cells. As DHODH is a validated therapeutic target, the CPA will enable unbiased identification of DHODH inhibitors and inhibitors of de novo pyrimidine biosynthesis for biological research and drug discovery. Show less

Identifying plausible mechanisms for compartmentalization and accumulation of the organic intermediates of early metabolic cycles in primitive cells has been a major challenge in theories of life's origins. Here, we propose a mechanism, where positive membrane potentials elevate the concentration of the organic intermediates. Positive membrane potentials are generated by positively charged surfaces of protocell membranes due to accumulation of transition metals. We find that (i) positive membrane potentials comparable in magnitude to those of modern cells can increase the concentration of the organic intermediates by several orders of magnitude; (ii) generation of large membrane potentials destabilize ion distributions; (iii) violation of electroneutrality is necessary to induce nonzero membrane potentials; and (iv) violation of electroneutrality enhances osmotic pressure and diminishes reaction efficiency, resulting in an evolutionary driving force for the formation of lipid membranes, specialized ion channels, and active transport systems. Show less

Ferroptosis is a new iron-dependent form of programmed cell death characterized by iron accumulation and lipid peroxidation. In recent years, ferroptosis has garnered enormous interest in disease treatment research communities in pursuit to reveal the mechanism and key targets of ferroptosis because ferroptosis is closely related to the pathophysiological processes of many diseases. Recent studies have shown some key targets, such as glutathione peroxidase 4 (GPX4) and System Xc−, and several inducers and inhibitors have been developed to regulate these key targets. With the emergence of new ferroptosis targets, studies on inducers and inhibitors have made new developments. The selection and use of inducers and inhibitors are very important for related work. This paper briefly introduces important regulatory targets in the ferroptosis metabolic pathway, lists and categorizes commonly used and recently developed inducers and inhibitors, and discusses their medical application. The paper ends of with potential future research direction for ferroptosis. Show less

Osteoporosis is a disorder of bone metabolism that is extremely common in elderly patients as well as in postmenopausal women. The main manifestation is that the bone resorption capacity is greater than the bone formation capacity, which eventually leads to a decrease in bone mass, increasing the risk of fracture. There is growing evidence that inhibiting osteoclast formation and resorption ability can be effective in treating and preventing the occurrence of osteoporosis. Our study is the first time to explore the role of the mitochondrial calcium uniporter (MCU) and its inhibitor ruthenium red (RR) in bone metabolism, clarifying the specific mechanism by which it inhibits osteoclast formation in vitro and plays a therapeutic role in osteoporosis in vivo. We verified the suppressive effects of RR on the receptor activator of nuclear factor‐κB ligand (RANKL‐)‐induced differentiation and bone resorption function of osteoclasts in vitro. The reactive oxygen species (ROS) production stimulated by RANKL and the expression level of P38 MAPK/NFATc1 were also found to be inhibited by RR. Moreover, the promotion of RR on osteogenesis differentiation was investigated by alkaline phosphatase (ALP) and alizarin red S (ARS) staining and the detection of osteogenesis‐specific gene expression levels by quantitative polymerase chain reaction (qPCR) and western blotting. Moreover, in ovariectomy (OVX‐)‐induced osteoporosis models, RR can downregulate the expression and function of the MCU, relieving bone loss and promoting osteogenesis to present a therapeutic effect on osteoporosis. This new finding will provide an important direction for the study of RR and MCU in the study of bone metabolism therapy targets. Show less

Title: Increasing Anticancer Activity with Phosphine Ligation in Zwitterionic Half-Sandwich Iridium(III), Rhodium(III), and Ruthenium(II) Complexes. Abstract: The synthesis and biological assessment of neutral or cationic platinum group metal-based anticancer complexes have been extremely studied, whereas there are few reports on the corresponding zwitterionic complexes. Herein, the synthesis, characterization, and bioactivity of zwitterionic half-sandwich phosphine-imine iridium(III), rhodium(III), and ruthenium(II) complexes were presented. The sulfonated phosphine-imine ligand and a group of zwitterionic half-sandwich P,N-chelating organometallic complexes were fully characterized by nuclear magnetic resonance (NMR), mass spectrum (electrospray ionization, ESI), elemental analysis, and X-ray crystallography. The solution stability of these complexes and their spectral properties were also determined. Notably, almost all of these complexes showed enhanced anticancer activity against model HeLa and A549 cancer cells than the corresponding zwitterionic pyridyl-imine N,N-chelating iridium(III) and ruthenium(II) complexes, which have exhibited inactive or low active in our previous work. The increase in the lipophilic property and intracellular uptake levels of these zwitterionic P,N-chelating complexes appeared to be associated with their superior cytotoxicity. In addition, these complexes showed biomolecular interactions with bovine serum albumin (BSA). The flow cytometry studies indicated that the representative complex Ir1 could induce early-stage apoptosis in A549 cells. Further, confocal microscopy imaging analysis displayed that Ir1 entered A549 cells through the energy-dependent pathway, targeted lysosome, and could cause lysosomal damage. In particular, these complexes could impede cell migration in A549 cells. Show less

Elongating RNA polymerase II (Pol II) can be paused or arrested by a variety of obstacles. These obstacles include DNA lesions, DNA-binding proteins, and small molecules. Hairpin pyrrole-imidazole (Py-Im) polyamides bind to the minor groove of DNA in a sequence-specific manner and induce strong transcriptional arrest. Remarkably, this Py-Im-induced Pol II transcriptional arrest is persistent and cannot be rescued by transcription factor TFIIS. In contrast, TFIIS can effectively rescue the transcriptional arrest induced by a nucleosome barrier. The structural basis of Py-Im-induced transcriptional arrest and why TFIIS cannot rescue this arrest remain elusive. Here we determined the X-ray crystal structures of four distinct Pol II elongation complexes (Pol II ECs) in complex with hairpin Py-Im polyamides as well as of the hairpin Py-Im polyamides-dsDNA complex. We observed that the Py-Im oligomer directly interacts with RNA Pol II residues, introduces compression of the downstream DNA duplex, prevents Pol II forward translocation, and induces Pol II backtracking. These results, together with biochemical studies, provide structural insight into the molecular mechanism by which Py-Im blocks transcription. Our structural study reveals why TFIIS fails to promote Pol II bypass of Py-Im-induced transcriptional arrest. Show less

Title: Ru(II)-modified TiO Abstract: The alternations in the hypoxic and immune microenvironment are closely related to the therapeutic effect and prognosis of oral squamous cell carcinoma (OSCC). Herein, a new nanocomposite, TiO2@Ru@siRNA is constructed from a ruthenium-based photosensitizer (Ru) modified-TiO2 nanoparticles (NPs) loaded with siRNA of hypoxia-inducible factor-1α (HIF-1α). Under visible light irradiation, TiO2@Ru@siRNA can elicit both Type I and Type II photodynamic effects, which causes lysosomal damage, HIF-1α gene silencing, and OSCC cell elimination efficiently. As a consequence of hypoxia relief and pyroptosis induction, TiO2@Ru@siRNA reshapes the immune microenvironment by downregulation of key immunosuppressive factors, upregulation of immune cytokines, and activation of CD4+ and CD8+ T lymphocytes. Furthermore, patient-derived xenograft (PDX) and rat oral experimental carcinogenesis models prove that TiO2@Ru@siRNA-mediated photodynamic therapy significantly inhibits the tumor growth and progression, and markedly enhances cancer immunity. In all, this study presents an effective hypoxia-adaptive photo-immunotherapeutic nanosystem with great potential for OSCC prevention and treatment. Show less

Abstract The crystal structure of the Cu(II) binuclear complex with 5-phenyltetrazole and 4,4'-dimethyl-2,2'-bipyridine (dmbipy), [Cu2(dmbipy)2L4], has been determined. The complex crystallizes in a monoclinic space group P21/c. The distorted square-pyramidal environment of Cu(II) atoms is formed by five nitrogen atoms belonging to three 5-phenyltetrazolate anions and one dmbipy molecule. Due to the presence of the intermolecular interactions π···π and C–H···π, the binuclear molecules combine into a supramolecular layered structure. The bonding energy has been calculated in terms of the density functional theory for two models of the [Cu2(dmbipy)2L4] complex with various coordination modes of the bridging 5-phenyltetrazole. Show less

PT-112 is a novel pyrophosphate-platinum conjugate, with clinical activity reported in advanced pretreated solid tumors. While PT-112 has been shown to induce robust immunogenic cell death (ICD) in vivo but only minimally bind DNA, the molecular mechanism underlying PT-112 target disruption in cancer cells is still under elucidation. The murine L929 in vitro system was used to test whether differential metabolic status alters PT-112's effects, including cell cytotoxicity. The results showed that tumor cells presenting mutations in mitochondrial DNA (mtDNA) (L929dt and L929dt cybrid cells) and reliant on glycolysis for survival were more sensitive to cell death induced by PT-112 compared to the parental and cybrid cells with an intact oxidative phosphorylation (OXPHOS) pathway (L929 and dtL929 cybrid cells). The type of cell death induced by PT-112 did not follow the classical apoptotic pathway: the general caspase inhibitor Z-VAD-fmk did not inhibit PT-112-induced cell death, alone or in combination with the necroptosis inhibitor necrostatin-1. Interestingly, PT-112 initiated autophagy in all cell lines, though this process was not complete. Autophagy is known to be associated with an integrated stress response in cancer cells and with subsequent ICD. PT-112 also induced a massive accumulation of mitochondrial reactive oxygen species, as well as changes in mitochondrial polarization-only in the sensitive cells harboring mitochondrial dysfunction-along with calreticulin cell-surface exposure consistent with ICD. PT-112 substantially reduced the amount of mitochondrial CoQ10 in L929 cells, while the basal CoQ10 levels were below our detection limits in L929dt cells, suggesting a potential relationship between a low basal level of CoQ10 and PT-112 sensitivity. Finally, the expression of HIF-1α was much higher in cells sensitive to PT-112 compared to cells with an intact OXPHOS pathway, suggesting potential clinical applications. Show less

AbstractAuranofin is an oral gold(I) compound, initially developed for the treatment of rheumatoid arthritis. Currently, Auranofin is under investigation for oncological application within a drug repurposing plan due to the relevant antineoplastic activity observed both in vitro and in vivo tumor models. In this review, we analysed studies in which Auranofin was used as a single drug or in combination with other molecules to enhance their anticancer activity or to overcome chemoresistance. The analysis of different targets/pathways affected by this drug in different cancer types has allowed us to highlight several interesting targets and effects of Auranofin besides the already well‐known inhibition of thioredoxin reductase. Among these targets, inhibitory‐κB kinase, deubiquitinates, protein kinase C iota have been frequently suggested. To rationalize the effects of Auranofin by a system biology‐like approach, we exploited transcriptomic data obtained from a wide range of cell models, extrapolating the data deposited in the Connectivity Maps website and we attempted to provide a general conclusion and discussed the major points that need further investigation. Show less

We have recently reported a series of half-sandwich ruthenium(II) complexes with curcuminoid ligands showing excellent cytotoxic activities (particularly ionic derivatives containing PTA (PTA = 1,3,5-triaza-7-phosphaadamantane). In the present study, new members of this family of compounds have been prepared with the objective to investigate the effect of a long hydrophobic chain obtained by replacing the OH-groups, present in curcumin and bisdemethoxycurcumin, with the palmitic acid ester. We report the synthesis of ruthenium(II) and osmium(II) p-cymene derivatives containing palmitic acid curcumin ester ligands ((1E,3Z,6E)-3-hydroxy-5-oxohepta-1,3,6-triene-1,7-diyl)bis(2-methoxy-4,1-phenylene)dipalmitate (p-curcH) and ((1E,3Z,6E)-3-hydroxy-5-oxohepta-1,3,6-triene-1,7-diyl)bis(4,1-phenylene)dipalmitate (p-bdcurcH). Complexes [M(II)(cym)(p-curc)/(p-bdcurc)(Cl)] 1-4 (M = Ru or Os) are neutral, whereas [M(II)(cym)(p-curc)/(p-bdcurc)(PTA)][SO₃CF₃] 5-8 are salts obtained when the chloride ligand is replaced by the PTA ligand. Stability studies performed on 1-8 in DMSO-PBS under physiological conditions (pH = 7.4) indicate that the complexes remain intact. The complexes exhibit potent and selective cytotoxic activity against an ovarian carcinoma cell line and its cisplatin-resistant form (A2780 and A2780cis), and non-cancerous human embryonic kidney (HEK293T) cells. To define the structure-activity relationships (SAR), the compounds have been compared with other Ru(II) and Os(II) complexes with curcuminoid ligands previously reported. SAR data reveal that the bisdemethoxycurcumin complexes are generally more active and selective than analogous curcumin-containing complexes. Show less