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Over the past decade, metal complexes based on N -heterocyclic carbenes (NHCs) have attracted great attention due to their wide and exciting applications in material sciences and medicinal chemistry. In particular, the gold-based complexes are the focus of research efforts for the development of new anticancer compounds. Literature data and recent results, obtained by our research group, reported the design, the synthesis and the good anticancer activity of some silver and gold complexes with NHC ligands. In particular, some of these complexes were active towards some breast cancer cell lines. Considering this evidence, here we report some new Au-NHC complexes prepared in order to improve solubility and biological activity. Among them, the compounds 1 and 6 showed an interesting anticancer activity towards the breast cancer MDA-MB-231 and MCF-7 cell lines, respectively. In addition, in vitro and in silico studies demonstrated that they were able to inhibit the activity of the human topoisomerases I and II and the actin polymerization reaction. Moreover, a downregulation of vimentin expression and a reduced translocation of NF-kB into the nucleus was observed. The interference with these vital cell structures induced breast cancer cells' death by triggering the extrinsic apoptotic pathway.

A series of highly functionalized pyrazole derivatives has been prepared by a one-pot, versatile and regioselective procedure. Pyrazoles 1-29 were tested in cell-based assay to assess their antiproliferative activity against a panel of tumour cells. Additionally, the cytotoxicity of prepared compounds was evaluated against normal human fibroblasts. The antiproliferative activity of the synthesized molecules emerged to be affected by the nature of the substituents of the pyrazole scaffold and derivatives 21-23 proved to inhibit the growth of melanoma and cervical cancer cells. Compound 23 was identified as the most active derivative and docking simulations predicted its ability to interact with estrogen receptors., (© 2022 Wiley-VCH GmbH.)

Background: Cancer tissue is characterized by low oxygen availability triggering neo angiogenesis and metastatisation. Accordingly, oxidation is a possible strategy for counteracting cancer progression and relapses. Previous studies used ozone gas, administered by invasive methods, both in experimental animals and clinical studies, transiently decreasing cancer growth. This study evaluated the effect of ozonized oils (administered either topically or orally) on cancer, exploring triggered molecular mechanisms., Methods: In vitro, in lung and glioblastoma cancer cells, ozonized oils having a high ozonide content suppressed cancer cell viability by triggering mitochondrial damage, intracellular calcium release, and apoptosis. In vivo, a total of 115 cancer patients (age 58 ± 14 years; 44 males, 71 females) were treated with ozonized oil as complementary therapy in addition to standard chemo/radio therapeutic regimens for up to 4 years., Results: Cancer diagnoses were brain glioblastoma, pancreas adenocarcinoma, skin epithelioma, lung cancer (small and non-small cell lung cancer), colon adenocarcinoma, breast cancer, prostate adenocarcinoma. Survival rate was significantly improved in cancer patients receiving HOO as integrative therapy as compared with those receiving standard treatment only., Conclusions: These results indicate that ozonized oils at high ozonide may represent an innovation in complementary cancer therapy worthy of further clinical studies.

Drugs delivered by proper carriers enter into the cells much more rapidly and carry out their action much more promptly than in the free forms. A high drug concentration can be sustained for longer periods of time at the target site in the cell. In in vivo conditions, this would translate into a reduction of systemic toxicity, dosage and frequency of dosing. Dendritic polymers significantly affect drug delivery in terms of reaching the target site, modifying the bio-distribution of the drug, and enhancing the efficacy of different drugs including anticancer compounds. 2-({[2-({[(2-tolyl)amino]carbonothioyl}oxy)ethyl]amino}carbo-nyl)benzoic acid 1 is a thiocarbamate derivative belonging to an already reported class of non-nucleoside HIV-1 reverse transcriptase inhibitors. In in vitro assay it showed no cytotoxic effects but was endowed with very low solubility and poor activity against wild-type HIV-1 (EC 50  = 27 μM). With the aim at improving its water solubility, 1 has been successfully incorporated inside non-toxic amino acids-modified core-shell hetero dendrimers. IR, NMR, zeta potential, mean size of particles, buffer capacity and in vitro release profile of prepared materials were reported. All dendriplexes were evaluated in cell-based assays to assess their cytotoxic profile. The obtained complexes, which harmonize a peripheral polycationic character and a buffer capacity which presuppose efficient cells penetration and increased residence time with a not PAMAM structured biodegradable scaffold, were well water-soluble and could rationally appear as a promising set of prodrugs for safe in vivo administrations., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Several enzymes are involved in the energy production, becoming a possible target for new anti-cancer drugs. In this paper, we used biochemical and in silico studies to evaluate the effects of two guanidine molecules, (Boc) 2 -creatine and metformin, on creatine kinase, an enzyme involved in the regulation of intracellular energy levels. Our results show that both drugs inhibit creatine kinase activity; however, (Boc) 2 -creatine displays a competitive inhibition, while metformin acts with a non-competitive mechanism. Moreover, (Boc) 2 -creatine is able to inhibit the activity of hexokinase with a non-competitive mechanism. Considering that creatine kinase and hexokinase are involved in energy metabolism, we evaluated the effects of (Boc) 2 -creatine and metformin on the ATP/AMP ratio and on cellular proliferation in healthy fibroblasts, human breast cancer cells (MDA-MB-468), a human neuroblastoma cell line (SH-SY5Y), a human Hodgkin lymphoma cell line (KMH2). We found that healthy fibroblasts were only partially affected by (Boc) 2 -creatine, while both ATP/AMP ratio and viability of the three cancer cell lines were significantly decreased. By inhibiting both creatine kinase and hexokinase, (Boc) 2 -creatine appears as a promising new agent in anticancer treatment. Further research is needed to understand what types of cancer cells are most suitable to treatment by this new compound. J. Cell. Biochem. 118: 2700-2711, 2017. © 2017 Wiley Periodicals, Inc., (© 2017 Wiley Periodicals, Inc.)

Chemo-resistance, which is the main obstacle in cancer therapy, is caused by the onset of drug-resistant cells in the heterogeneous cell population in cancer tissues. MicroRNAs regulate gene expression at the post-transcriptional level, and they are involved in many different biological processes, including cell proliferation, differentiation, metabolism, stress response, and apoptosis. The aberrant expression of microRNAs plays a major pathogenic role from the early stages of the carcinogenesis process. Recently, microRNAs have been reported to play an important role in inducing resistance to anti-cancer drugs. Specific microRNA alterations occur selectively in cancer cells, rendering these cells resistant to various chemotherapeutic agents. For example, resistance to 5-fluorouracil is mediated by alterations in miR-21, miR-27a/b, and miR-155; the sensitivity to Docetaxel is influenced by miR-98, miR-192, miR-194, miR-200b, miR-212, and miR-424; and the resistance to Cisplatin is mediated by miR-let-7, miR-15, miR-16 miR-21 and miR-214. Chemo-resistant cancer cells are characterized by altered functions in enzymes that are involved in microRNA maturation, primarily including Dicer, as demonstrated in ovarian cancer, oral squamous cell carcinoma, breast cancer and cervical cancer. Based on the evidence reviewed in this paper, various strategies have been developed to artificially re-establish microRNA expression in resistant cells, thus restoring chemo-sensitivity. These strategies employ synthetic analogs, anti-microRNA oligonucleotides, locked nucleic acid, microRNA sponges, drugs that inhibit DNA methylation or histone deacetylation, and the introduction of microRNA mimics. The ability to modulate microRNA expression is a promising strategy for overcoming the problem of drug resistance in cancer treatment., Competing Interests: None.

Purpose: By a scaffold shortening strategy, a small series of retinoidal amides fenretinide (4-HPR) analogs have been synthesized from α, β-ionones and tested for their antiproliferative and differentiating activities, and antioxidant effect., Methods: The antiproliferative activity and triggering of apoptosis of our short retinoids were evaluated by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and 4'-6-diamidino-2-phenylindole staining and microscope evaluation after 3- or 6-day exposure, while their differentiating activity was established by the analysis of the expression of the CD11b marker of differentiation in treated HL60 target cells and by the superoxide production assayed colorimetrically by the nitro blue tetrazolium-reducing activity assay. Finally, the antioxidant activity was determined by the 2,2'-azino-bis-(3-ethylbenzthiazoline-6-sulphonic acid) diammonium salt radical cation decolourisation assay utilizing the antioxidant Trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid) as reference (Trolox equivalent antioxidant capacity, or TEAC). Docking analysis was performed to study the binding features to the Retinoic Acid Receptor alpha (RARα)., Results: While no pharmacologically relevant antiproliferative activity was evidenced, some of our short retinoids showed a differentiating and antioxidant activity similar to that of 4-HPR. In particular, compound 2b6 displayed a scavenging activity two times more efficient than 4-HPR itself. Finally, the docking analysis showed that these short retinoids, like 4-HPR, bind to the RARα protein with good fitness scores., Conclusion: Our data could pave the way for the design of new potent and less toxic antioxidant and differentiating compounds related to 4-HPR.

Natural or synthetic carbazole derivatives have recently attracted the attention of the scientific world because of their multiple biological activity, leading to an increase of designed, synthesized and studied analogues. In this paper, four 1,4-dimethylcarbazole derivatives, analogues of Ellipticine, have been investigated for their ability to block cancer cells growth, with low effects on the proliferation of normal cells. DNA topoisomerases inhibition assays, docking simulations, stability studies and effects on a membrane model are reported. Particularly, compounds 2 and 3 have been found thermally stable and able to inhibit, strongly and selectively, the human DNA topoisomerase II. These properties confer a good and broad antitumoral activity in vitro, with very low cytotoxic effect on the proliferation of normal cell lines and without damaging, in contrast with Ellipticine, the cell membrane model. The presented outcomes set the most active compounds as good candidates for pre-clinical studies useful in cancer treatment., (Copyright © 2016 Elsevier B.V. All rights reserved.)

G protein-coupled receptors (GPCRs) are cell surface proteins mainly involved in signal transmission; however, they play a role also in several pathophysiological conditions. Chemically heterogeneous molecules like peptides, hormones, lipids, and neurotransmitters activate second messengers and induce several biological responses by binding to these seven transmembrane receptors, which are coupled to heterotrimeric G proteins. Recently, additional molecular mechanisms have been involved in GPCR-mediated signaling, leading to an intricate network of transduction pathways. In this regard, it should be mentioned that diverse GPCR family members contribute to the adaptive cell responses to low oxygen tension, which is a distinguishing feature of several illnesses like neoplastic and cardiovascular diseases. For instance, the G protein estrogen receptor, namely G protein estrogen receptor (GPER)/GPR30, has been shown to contribute to relevant biological effects induced by hypoxia via the hypoxia-inducible factor (HIF)-1α in diverse cell contexts, including cancer. Likewise, GPER has been found to modulate the biological outcome of hypoxic/ischemic stress in both cardiovascular and central nervous systems. Here, we describe the role exerted by GPCR-mediated signaling in low oxygen conditions, discussing, in particular, the involvement of GPER by a hypoxic microenvironment.

Estrogens influence multiple physiological processes and are implicated in many diseases as well. Cellular responses to estrogens are mainly mediated by the estrogen receptors (ER)α and ERβ, which act as ligand-activated transcription factors. Recently, a member of the G protein-coupled receptor (GPCR) superfamily, namely GPER/GPR30, has been identified as a further mediator of estrogen signalling in different pathophysiological conditions, including cancer. Today, computational methods are commonly used in all areas of health science research. Among these methods, virtual ligand screening has become an established technique for hit discovery and optimization. The absence of an established three-dimensional structure of GPER promoted studies of structure-based drug design in order to build reliable molecular models of this receptor. Here, we discuss the results obtained through the structure-based virtual ligand screening for GPER, which allowed the identification and synthesis of different selective agonist and antagonist moieties. These compounds led significant advances in our understanding of the GPER function at the cellular, tissue, and organismal levels. In particular, selective GPER ligands were critical toward the evaluation of the role elicited by this receptor in several pathophysiological conditions, including cancer. Considering that structure-based approaches are fundamental in drug discovery, future research breakthroughs with the aid of computer-aided molecular design and chemo-bioinformatics could generate a new class of drugs that, acting through GPER, would be useful in a variety of diseases as well as in innovative anticancer strategies.

Background: G-protein coupled estrogen receptor (GPER) is involved in numerous intracellular physiological and pathological events including cancer cell migration and proliferation. Its characterization is yet incomplete due to the limited number of specific ligands., Results: Two novel selective GPER antagonists, based on a benzo[b]pyrrolo[1,2-d][1,4]oxazin-4-one structure, have been designed and synthesized. Their binding to the receptor was confirmed by a competition assay, while the antagonist effects were ascertained by their capability to prevent the ligand-stimulated action of GPER. The transcription mediated by the classical estrogen receptor was not influenced, demonstrating selectivity for GPER., Conclusion: These novel compounds may be considered useful leads toward the dissection of the GPER signaling and the development of new pharmacological treatments in breast cancer.

A key feature of many cancers is the capacity and the propensity to metabolize glucose to lactic acid at a very high rate even in the presence of oxygen. This characteristic was first discovered in 1924 by Otto Heinrich Warburg. Hexokinase, the first enzyme in the glycolytic pathway, not only improves the cell's energy supply in malignant cells, but also protects cancer cells against apoptosis through direct interaction with mitochondria and with the Voltage Dependent Anion Channel 1 (VDAC1). The rupture of HK:VDAC1 protein complex provides a therapeutic opportunity, as this association appears to protect tumor cells from mitochondrial outer membrane permeabilization, an event that marks the point of no return in multiple pathways leading to cell death. In the absence of a crystallographic structure and in order to perform an in silico screening of possible small molecules able to inhibit the protein association, we are presenting a computational model of HK-I:VDAC1 complex. It appears as evident how the first 15 N-terminal residues of HK-I interact with the inner part of the barrel of VDAC1 and not with the outside walls, within the mitochondrial membrane as previously believed. This finding is in agreement with the existence of a secondary ATP binding site in the same N-terminal region of HK-I which seems to have a crucial role in HK-I interaction with VDAC1. This evidence appears to be in accord also with the high levels of ATP that are found in cancer cells. Eventually such arrangements may contribute to stabilize the tertiary structure of VDAC1 while shielding from pro-apoptotic factor binding, protecting in a synergic way the tumoral cell from programmed death., (Copyright © 2011 © Elsevier B.V. and Mitochondria Research Society. All rights reserved. Published by Elsevier B.V. All rights reserved.)

We report herein the reversal of multidrug resistance-1 (MDR1) in A2780/DX3 cells by the two nifedipine-like compounds 1 and 2 that are part of a library of 1,4-dihydropyridines (1,4-DHPs) calcium-channel modulators bearing in C-4 a different substituted imidazo[2,1-b]thiazole system. By methylthiazol tetrazolium (MTT) assay, cytofluorimetry, and fluorescence microscopy we evaluated their ability to reverse MDR in our cell system. Moreover, together with compound 3 (the diltiazem-like 8-(4-chlorophenyl)-5-methyl-8-[(2Z)-pent-2-en-1-yloxy]-8H-[1,2,4]oxadiazolo[3,4-c][1,4]thiazin-3-one) we analyzed their ability to potentiate the triggering of apoptosis after exposure to doxorubicin, through the nuclear morphological analysis after 4',6-diamidino-2-phenylindole (DAPI), the fluorescein isothiocyanate (FITC)-Annexin-V/propidium iodide (PI) staining and the caspase activity determination. Our results demonstrate that compounds 1 and 2, at concentrations showing a very low (5%) or absent inhibition of cell proliferation, in combination with doxorubicin enhance its antiproliferative activity (from 30% to 54% IC(50) reduction) in A2780/DX3 cells through an increase of doxorubicin intracellular accumulation. These compounds together with compound 3, which has already been demonstrated to act as a potent inhibitor of MDR1 function, were also able to significantly potentiate the activation of the apoptosis machinery triggered by the exposure to doxorubicin. In conclusion, our results identify two new molecules structurally related to the calcium-channel blocker nifedipine, but characterized by a very low LTCC blockers activity, able to potentiate the antiproliferative and apoptotic activities of doxorubicin through an increase of its intracellular concentration likely caused by the inhibition of MDR1 function.

Estrogens are structurally related steroids that regulate important physiological processes. 17beta-estradiol (E2) is reversibly oxidized to estrone (E1) and both E2 and E1 can be irreversibly converted to estriol (E3), which also originates directly from androstenedione. The action of E2 has been traditionally explained by the binding to the estrogen receptor (ER) alpha and ER beta, however the G protein-coupled receptor (GPR) 30 has been recently involved in the rapid signaling triggered by estrogens. Although the role of E2 in the development of breast cancer has been largely documented, the contribution of E3 still remains to be completely evaluated. Here, we demonstrate for the first time that E3 acts as a GPR30 antagonist since it was able to inhibit the GPR30-mediated responses such as the rapid ERK activation, the up-regulation of target genes like c-fos and connective tissue growth factor, the proliferative effects observed in ER-negative SkBr3 cells., ((c) 2010 Elsevier Ireland Ltd. All rights reserved.)

The interpretation of solution hydrodynamic data in terms of macromolecular structural parameters is not a straightforward task. Over the years, several approaches have been developed to cope with this problem, the most widely used being bead modeling in various flavors. We report here the implementation of the SOMO (SOlution MOdeller; Rai et al. in Structure 13:723-734, 2005) bead modeling suite within one of the most widely used analytical ultracentrifugation data analysis software packages, UltraScan (Demeler in Modern analytical ultracentrifugation: techniques and methods, Royal Society of Chemistry, UK, 2005). The US-SOMO version is now under complete graphical interface control, and has been freed from several constraints present in the original implementation. In the direct beads-per-atoms method, virtually any kind of residue as defined in the Protein Data Bank (e.g., proteins, nucleic acids, carbohydrates, prosthetic groups, detergents, etc.) can be now represented with beads whose number, size and position are all defined in user-editable tables. For large structures, a cubic grid method based on the original AtoB program (Byron in Biophys J 72:408-415, 1997) can be applied either directly on the atomic structure, or on a previously generated bead model. The hydrodynamic parameters are then computed in the rigid-body approximation. An extensive set of tests was conducted to further validate the method, and the results are presented here. Owing to its accuracy, speed, and versatility, US-SOMO should allow to fully take advantage of the potential of solution hydrodynamics as a complement to higher resolution techniques in biomacromolecular modeling.

Mutational analysis of the GNPTAB gene was performed in 46 apparently unrelated patients with mucolipidosis IIalpha/beta or IIIalpha/beta, characterized by the mistargeting of multiple lysosomal enzymes as a consequence of a UDP-GlcNAc-1-phosphotransferase defect. The GNPTAB mutational spectrum comprised 25 distinct mutant alleles, 22 of which were novel, including 3 nonsense mutations (p.Q314X, p.R375X, p.Q507X), 5 missense mutations (p.I403T, p.C442Y, p.C461G, p.Q926P, p.L1001P), 6 microduplications (c.749dupA, c.857dupA, c.1191_1194dupGCTG, c.1206dupT, c.1331dupG, c.2220_2221dupGA) and 8 microdeletions (c.755_759delCCTCT, c.1399delG, c.1959_1962delTAGT, c.1965delC, c.2550_2554delGAAAA, c.3443_3446delTTTG, c.3487_3490delACAG, c.3523_3529delATGTTCC). All micro-duplications/deletions were predicted to result in the premature termination of translation. A novel exonic SNP (c.303G>A; E101E) was identified which is predicted to create an SFRS1 (SF2/ASF) binding site that may be of potential functional/clinical relevance. This study of mutations in the GNPTAB gene, the largest yet reported, extends our knowledge of the mutational heterogeneity evident in MLIIalpha/beta/MLIIIalpha/beta.

Niemann-Pick C, the autosomal recessive neuro-visceral disease resulting from a failure of cholesterol trafficking within the endosomal-lysosomal pathway, is due to mutations in NPC1 or NPC2 genes. We characterized 34 unrelated patients including 32 patients with mutations in NPC1 gene and two patients in NPC2 gene. Overall, 33 distinct genotypes were encountered. Among the 21 unpublished NPC1 alleles, 15 were due to point mutations resulting in 13 codon replacements (p.C100S, p.P237L, p.R389L, p.L472H, p.Y634C, p.S636F, p.V780G, p.Q921P, p.Y1019C, p.R1077Q, p.L1102F, p.A1187V, and p.L1191F) and in two premature stop codons (p.R934X and p.Q447X); a new mutant carried two in cis mutations, p.[L648H;M1142T] and four other NPC1 alleles were small deletions/insertions leading both to frame shifts and premature protein truncations (p.C31WfsX26, p.F284LfsX26, p.E1188fsX54, and p.T1205NfsX53). Finally, the new intronic c.464-2A>C change at the 3' acceptor splice site of intron 4 affected NPC1 messenger RNA processing. We also found a new NPC2 mutant caused by a change of the first codon (p.M1L). The novel missense mutations were further investigated by two bioinformatics approaches. Panther proein classification system computationally predicted the detrimental effect of all new missense mutations occurring at evolutionary conserved positions. The other bioinformatics approach was based on prediction of structural alterations induced by missense mutations on the NPC1 atomic models. The in silico analysis predicted protein malfunctioning and/or local folding alteration for most missense mutations. Moreover, the effects of the missense mutations (p.Y634C, p.S636F, p.L648H, and p.V780G) affecting the sterol-sensing domain (SSD) were evaluated by docking simulation between the atomic coordinates of SSD model and cholesterol.

Integrin-dependent adhesion and signaling are regulated by conformational changes whose details remain controversial. Crystallography revealed bent shapes for resting and primed integrin ectodomains, whereas large, ligand-induced rearrangements in other constructs suggested extension, "opening," and tail separation. We have used experimental/computed hydrodynamics to discriminate among different alpha(v)beta(3) and alpha(IIb)beta(3) atomic models built on X-ray, NMR, and EM data. In contrast with X-ray structures and EM maps, hydrodynamics indicate that resting integrins are already extended. Furthermore, the hydrodynamics of an alpha(v)beta(3) ectodomain-fibronectin fragment complex support opening via additional head region conformational changes (hybrid domain swing-out), but without tail separation. Likewise, frictional changes induced by priming agents in full-length alpha(IIb)beta(3) correlate well with the swing-out coupled to a simple transmembrane helix shift in an extended, electron tomography-based model. Extension and immediate tail separation are then uncoupled from head region rearrangements following activation, thus underscoring integrins' delicate, finely tuned plasticity.

beta2-microglobulin, the light chain component of the major histocompatibility complex I, is involved in the development of DRA, an amyloid deposition disease occurring in man. Specifically, the beta2-microglobulin component, dissociated form the complex heavy chain, gives rise to amyloidogenic deposits in the joints of patients exposed to long dialysis periods. beta2-microglobulin three-dimensional structure is based on an antiparallel beta-barrel fold, with immunoglobulin domain topology, displaying structural flexibility in the crystal and NMR structures so fare determined. The structural bases of amyloidogenic potential in beta2-microglobulin can be related to local unfolding, to the tendency to aggregate laterally through non-compensated beta-strands, and partly also to its trend towards N-terminal proteolytic degradation. Such trends emerge quite clearly from inspection of a limited number of crystal structures of beta2-microglobulin as an isolated chain, separated form the major histocompatibility complex I heavy chain.

Maturation of prokaryotic hydrogenase involves several protein factors, among which is the accessory protein HypF, which hosts the consensus sequence of acylphosphatases and a sequence motif common to proteins catalyzing O-carbamoylations. The specific functions of HypF are largely unknown, although it has been observed that CN(-) and CO ligands at the hydrogenase Ni,Fe active centre originate from carbamoylphosphate. The HypF N-terminal domain (91 residues, acylphosphatase-like domain) has been crystallized in two different crystal forms belonging to the orthorhombic P2(1)2(1)2(1) space group (unit-cell parameters a = 35.5, b = 59.8, c = 87.6 A) and to the rhombohedral space group R32 (unit-cell parameters a = b = 58.1, c = 155.6 A in the hexagonal setting).