Evaluation of prednisolone and prednisolone sodium succinate concentrations in human plasma and inner ear perilymph during cochlear implantation 24 h after intratympanic injection.

BACKGROUND: The use of intratympanic (IT) steroids has drastically increased over the past 10-15 years to manage many otological pathologies. OBJECTIVES: This study aimed to compare the concentrations of prednisolone and prednisolone sodium succinate (SS) in the plasma and inner ear perilymph of participants who underwent cochlear implantation 24 h after IT injection. MATERIALS AND METHODS: It was a prospective comparative randomized study. Twenty participants received an IT injection of prednisolone SS ∼24 h before the cochlear implantation. The other five participants received an IT saline injection and represented the control group. Perilymph and blood were sampled during the cochlear implantation surgery. RESULTS: Both prednisolone and prednisolone SS were still present in perilymph ∼24 h after the IT administration. Only prednisolone was present in the blood plasma of seven participants (35%). CONCLUSION: IT injection of prednisolone SS resulted in high perilymph concentrations of prednisolone and prednisolone SS, which could stay in the perilymph for at least 24 h. Using a mini-endoscope during the IT injection may effectively detect barriers infront of the round window membrane, increasing the drug concentration in the inner ear. SIGNIFICANCE: IT injection is an effective method for delivering prednisolone to the inner ear.

Hydromethanolic Extracts from Adansonia digitata L. Edible Parts Positively Modulate Pathophysiological Mechanisms Related to the Metabolic Syndrome.

Metabolic syndrome includes a cluster of risk factors for many pathological conditions, including hyperglycemia, abdominal obesity, hyperlipidemia, and hypertension. Adansonia digitata L. (also known as baobab) is used in traditional African Medicine and recent studies showed that it improves the metabolism of carbohydrates and lipids. The aim of this study is to investigate the mechanisms of action associated with the beneficial effects of extracts from the edible parts of baobab (fruit pulp, leaves, raw and toasted seeds), evaluating their inhibitory activity against: alpha-amylase, alpha-glucosidase, angiotensin-converting enzyme, 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase, and pancreatic lipase. Baobab fruit pulp and leaf extracts resulted to be the most active ones and were then tested on the differentiation process of SW-872 human liposarcoma cells to mature adipocytes. The addition of these latter extracts did not affect triglyceride accumulation, indicating a neutral impact on this parameter. The findings here reported help to explain the growing amount of evidence on the biological properties of baobab and provide suggestions about their use in food and nutraceutical fields.

Multi Dynamic Extraction: An Innovative Method to Obtain a Standardized Chemically and Biologically Reproducible Polyphenol Extract from Poplar-Type Propolis to Be Used for Its Anti-Infective Properties.

Antimicrobial activity is a well-known property of propolis, making it a candidate for antimicrobial surfaces in biomedical devices. Nevertheless, large-scale use of propolis as an anti-infective agent is limited by the heterogeneity of its chemical composition and consequent variation in antimicrobial activity. The aim of this study was to demonstrate that the multi dynamic extraction (M.E.D.) method produces standardized polyphenolic mixtures from poplar-type propolis, with reproducible chemical composition and anti-microbial activity, independently from the chemical composition of the starting raw propolis. Three raw propolis samples, from Europe, America, and Asia, were analyzed for their polyphenol chemical composition by means of HPLC-UV and then combined to obtain three mixtures of propolis, which werme submitted to the M.E.D. extraction method. The chemical composition and the antimicrobial activity of M.E.D. propolis against bacteria and fungi were determined. The three M.E.D. propolis showed similar chemical compositions and antimicrobial activities, exhibiting no relevant differences against antibiotic-susceptible and antibiotic-resistant strains. The batch-to-batch reproducibility of propolis extracts obtained with the M.E.D. method encourages the design of drugs alternative to traditional antibiotics and the development of anti-infective surface-modified biomaterials.

Antidepressive effects of a chemically characterized maqui berry extract (Aristotelia chilensis (molina) stuntz) in a mouse model of Post-stroke depression.

Mood disorders occur in 30% of stroke patients, and of these post-stroke depression (PSD) is the most significant. This study aimed to evaluate the antidepressive-like effects and in vivo antioxidant activity of a chemically characterized maqui berry (Aristotelia chilensis (Molina) Stuntz) extract obtained from an optimized extraction method, on a murine PSD model. The extraction process was optimized to maximize anthocyanin content, and the phytochemical profile of the extract was evaluated using a multi-methodological approach including a liquid chromatographic method coupled with mass spectrometry and nuclear magnetic resonance spectroscopy. The antidepressive-like activity was investigated through despair swimming and tail suspension tests. The in vivo antioxidant activity was evaluated in mouse brain tissue by measuring the activity of antioxidant enzymes and lipid peroxidation products. A number of compounds have been first identified in maqui berry here, including malvidin-glucoside, GABA, choline and trigonelline. Moreover, the results showed that the antidepressive-like activity exerted by the extract, which was found to restore normal mouse behavior in both despair swimming and tail suspension tests, could be linked to its antioxidant activity, leading to the conclusion that maqui berries might be useful for supporting pharmacological therapy of PSD by modulating oxidative stress.

Investigational drugs targeting cyclin-dependent kinases for the treatment of cancer: an update on recent findings (2013-2016).

INTRODUCTION: Cell cycle and gene transcription are under the control of cyclin-dependent kinases (CDKs), whose activity depends on the binding with cyclins. Deregulated CDK activities have been reported in a majority of human cancers, representing potential therapeutic targets. AREAS COVERED: This review provides preclinical and clinical (phase I/II) updates of promising therapeutic compounds targeting CDKs published between 2013 and 2016 EXPERT OPINION: First generation pan-CDK inhibitors showed marked toxicity in clinical trials and most compounds were discontinued. Despite their failure was ascribed also to inadequate patient selection rules, novel pan-CDK inhibitors have entered clinical trials with still poorly defined selection strategies. The most interesting results have been obtained with dual CDK4/6 inhibitors and through a more accurate evaluation of predictive biomarkers, suggesting the usefulness of CDK inhibitors for personalized treatment. The increased knowledge on the roles of CDKs in cell cycle and gene transcription suggests to review also the anticancer potential of first generation CDK inhibitors by defining more appropriate rules for patients engagement. Recent findings has highlighted CDK8 as a novel target for cancer treatment. Indeed some biomarkers for CDK8 inhibition sensitivity have already been proposed. CDK8 inhibition is also supposed to prevent cancer metastasis.

Identification of noncovalent proteasome inhibitors with high selectivity for chymotrypsin-like activity by a multistep structure-based virtual screening.

Noncovalent proteasome inhibitors introduce an alternative mechanism of inhibition to that of covalent inhibitors, e.g. carfilzomib, used in cancer therapy. A multistep hierarchical structure-based virtual screening (SBVS) of the 65,375 NCI lead-like compound library led to the identification of two compounds (9 and 28) which noncovalently inhibited the chymotrypsin-like (ChT-L) activity (Ki = 2.18 and 2.12 µM, respectively) with little or no effects on the other two major proteasome proteolytic activities, trypsin-like (T-L) and post-glutamyl peptide hydrolase (PGPH) activities. A subsequent hierarchical similarity search over the full NCI database with the most active tripeptide-based inhibitor 9 resulted in the discovery of the ß5/ß6-specific tripeptide derivative 38 that noncovalently binds the ChT-L site (Ki = 0.42 µM). The solution structure of 9 and 38 was solved by (1)H NMR spectroscopy and the binding mode of the inhibitors was elucidated by docking experiments using the yeast 20S proteasome. Compound 38 (IC50 = 26.7 µM) is slightly more potent than 9 (IC50 = 34.3 µM) at inhibiting survival of dexamethasone-resistant (MM.1R) human multiple myeloma cells. The identified ligand thus provides valuable insights for the future structure-based design of subtype-specific proteasome inhibitors.

Novel non-peptide small molecules preventing IKKß/NEMO association inhibit NF-κB activation in LPS-stimulated J774 macrophages.

Nuclear Factor-κB (NF-κB) is a transcription factor regulating several genes involved in important physiological and pathological processes. NF-κB has been found constitutively activated in many inflammatory/immune diseases. In addition, a positive correlation between persistent activation of NF-κB and tumor promotion has been demonstrated. Since the IKK (IκB kinase) activation is an indispensable component of all pro-inflammatory signaling pathways leading to NF-κB activation, considerable efforts have been done in order to develop novel anti-inflammatory therapeutics targeting IKK. Association of the IKK complex relies on critical interactions between the C-terminus NBD (NEMO binding domain) of the catalytic subunits IKKα and IKKß, and the regulatory subunit NEMO (NF-κB Essential Modulator). Thus, this IKK/NEMO interacting region provides an attractive target to prevent the IKK complex formation and NF-κB activation. In this regard, we have identified non-peptide small molecule disruptors of IKKß/NEMO complex through a structure-based virtual screening (SBVS) of the NCI chemical library. Phenothiazine 22 and its close analogues (22.2, 22.4 and 22.10) were able to reduce nitrite production and iNOS mRNA expression in J774 murine macrophages stimulated with LPS for 24h. These effects were associated with a reduced NF-κB/DNA binding activity as well as a decreased expression of phosphorylated IKKß, IκBα and NF-κB/p65 in these cells. These observations suggest that compound 22 and its three structural analogues by inhibiting IKKß/NEMO association mediate the blockage of NF-κB signaling pathway and may prove effective in treatment of diseases in which the IKK/NF-κB pathway is dysregulated.

Amorfrutins are efficient modulators of peroxisome proliferator-activated receptor gamma (PPARγ) with potent antidiabetic and anticancer properties: a patent evaluation of WO2014177593 A1.

INTRODUCTION: PPARγ is an essential regulator of lipid, glucose, and insulin metabolism. PPARγ full agonists, such as thiazolidinediones, are the mainstay drugs for the treatment of type 2 diabetes; however, undesirable clinical side effects have contributed to poor compliance with therapy and limited their full therapeutic potential. In the last few years, many efforts have been made in the discovery and development of selective PPARγ modulators (SPPARγMs) as safer alternatives to PPARγ full agonists. AREAS COVERED: This application claims the plant-derived amorfrutins or their synthetic analogs as SPPARγMs with potential to exhibit glucose-lowering effects without provoking side effects associated with full PPARγ activation. Specifically, the in vivo glucose-lowering properties of the high-affinity SPPARγM amorfrutin B are described. Moreover, examples of this class of compounds exhibit interesting antiproliferative activities. EXPERT OPINION: The patent (WO2014177593 A1) under discussion proposes enriching functional food products or phytomedical extracts with safe licorice extracts, containing sufficient amounts of amorfrutins, with the ultimate goal of inhibiting the early development of disorders such as insulin resistance. Interestingly, some example compounds show anticancer properties in colon, prostate, and breast malignancies. However, further in vivo investigations of the claimed compounds for these specific indications will be necessary to definitively support their clinical applications.

Ligand-based chemoinformatic discovery of a novel small molecule inhibitor targeting CDC25 dual specificity phosphatases and displaying in vitro efficacy against melanoma cells.

CDC25 phosphatases are important regulators of the cell cycle and represent promising targets for anticancer drug discovery. We recently identified NSC 119915 as a new quinonoid CDC25 inhibitor with potent anticancer activity. In order to discover more active analogs of NSC 119915, we performed a range of ligand-based chemoinformatic methods against the full ZINC drug-like subset and the NCI lead-like set. Nine compounds (3, 5-9, 21, 24, and 25) were identified with Ki values for CDC25A, -B and -C ranging from 0.01 to 4.4 µM. One of these analogs, 7, showed a high antiproliferative effect on human melanoma cell lines, A2058 and SAN. Compound 7 arrested melanoma cells in G2/M, causing a reduction of the protein levels of CDC25A and, more consistently, of CDC25C. Furthermore, an intrinsic apoptotic pathway was induced, which was mediated by ROS, because it was reverted in the presence of antioxidant N-acetyl-cysteine (NAC). Finally, 7 decreased the protein levels of phosphorylated Akt and increased those of p53, thus contributing to the regulation of chemosensitivity through the control of downstream Akt pathways in melanoma cells. Taken together, our data emphasize that CDC25 could be considered as a possible oncotarget in melanoma cells and that compound 7 is a small molecule CDC25 inhibitor that merits to be further evaluated as a chemotherapeutic agent for melanoma, likely in combination with other therapeutic compounds.

Discovery of new small molecules inhibiting 67 kDa laminin receptor interaction with laminin and cancer cell invasion.

The 67 kDa laminin receptor (67LR) is a non-integrin receptor for laminin (LM) that derives from a 37 kDa precursor (37LRP). 67LR expression is increased in neoplastic cells and correlates with an enhanced invasive and metastatic potential. We used structure-based virtual screening (SB-VS) to search for 67LR inhibitory small molecules, by focusing on a 37LRP sequence, the peptide G, able to specifically bind LM. Forty-six compounds were identified and tested on HEK-293 cells transfected with 37LRP/67LR (LR-293 cells). One compound, NSC47924, selectively inhibited LR-293 cell adhesion to LM with IC50 and Ki values of 19.35 and 2.45 µmol/L. NSC47924 engaged residues W176 and L173 of peptide G, critical for specific LM binding. Indeed, NSC47924 inhibited in vitro binding of recombinant 37LRP to both LM and its YIGSR fragment. NSC47924 also impaired LR-293 cell migration to LM and cell invasion. A subsequent hierarchical similarity search with NSC47924 led to the identification of additional four compounds inhibiting LR-293 cell binding to LM: NSC47923, NSC48478, NSC48861, and NSC48869, with IC50 values of 1.99, 1.76, 3.4, and 4.0 µmol/L, respectively, and able to block in vitro cancer cell invasion. These compounds are promising scaffolds for future drug design and discovery efforts in cancer progression.

Structural development studies of PPARs ligands based on tyrosine scaffold.

PPARs are nuclear receptors with a critical physiological role in lipid and glucose metabolism. As part of our effort to develop new and selective PPAR agonists containing stilbene and its bioisoster phenyldiazene, novel analogs were synthesized starting from tyrosine and evaluated as PPAR agonists. We tested the effects of phenyloxazole replacement of GW409544, a well-known PPARα/γ dual agonist, with stilbene or phenyldiazene moiety, spaced by an ether bridge to tyrosine portion. These structural modifications provided potent and selective PPARγ agonists. Molecular docking studies performed on these new compounds complemented the experimental results and allowed to gain some insights into the nature of binding of the ligands.

Development of novel selective peptidomimetics containing a boronic acid moiety, targeting the 20S proteasome as anticancer agents.

This paper describes the design, synthesis, and biological evaluation of peptidomimetic boronates as inhibitors of the 20S proteasome, a validated target in the treatment of multiple myeloma. The synthesized compounds showed a good inhibitory profile against the ChT-L activity of 20S proteasome. Compounds bearing a ß-alanine residue at the P2 position were the most active, that is, 3-ethylphenylamino and 4-methoxyphenylamino (R)-1-{3-[4-(substituted)-2-oxopyridin-1(2H)-yl]propanamido}-3-methylbutylboronic acids (3 c and 3 d, respectively), and these derivatives showed inhibition constants (Ki ) of 17 and 20 nM, respectively. In addition, they co-inhibited post glutamyl peptide hydrolase activity (3 c, Ki=2.57 µM; 3 d, Ki=3.81 µM). No inhibition was recorded against the bovine pancreatic α-chymotrypsin, which thus confirms the selectivity towards the target enzyme. Docking studies of 3 c and related inhibitors into the yeast proteasome revealed the structural basis for specificity. The evaluation of growth inhibitory effects against 60 human tumor cell lines was performed at the US National Cancer Institute. Among the selected compounds, 3 c showed 50% growth inhibition (GI50) values at the sub-micromolar level on all cell lines.

Identification of a new series of amides as non-covalent proteasome inhibitors.

Proteasome inhibition has emerged as an important therapeutic strategy for the treatment of multiple myeloma (MM) and some forms of lymphoma, with potential application in other types of cancers. 20S proteasome consists of three different catalytic activities known as chymotrypsin-like (ChT-L), trypsin-like (T-L), and, post-glutamyl peptide hydrolyzing (PGPH) or caspase-like (C-L), which are located respectively on the ß5, ß2, and ß1 subunits of each heptameric ß rings. Currently a wide number of covalent proteasome inhibitors are reported in literature; however, the less widely investigated non-covalent inhibitors might be a promising alternative to employ in therapy, because of the lack of all drawbacks and side-effects related to irreversible inhibition. In the present work we identified a series of amides, two of which (1b and 1f) are good candidates to non-covalent inhibition of the chymotrypsin-like activity of the ß5 proteasome subunit. The non-covalent binding mode was corroborated by docking simulations of the most active inhibitors 1b, 1f and 2h into the yeast 20S proteasome crystal structure.

Novel inhibitors of signal transducer and activator of transcription 3 signaling pathway: an update on the recent patent literature.

INTRODUCTION: The signal transducer and activator of transcription 3 (STAT3) is a transcription factor that plays a key role in normal cell growth and is constitutively activated in about 70% of solid and hematological cancers. Thus, the development of potent and selective inhibitors that target STAT3 is of interest especially in the cancer therapeutic area. AREAS COVERED: This review updates new patents claiming STAT3 inhibitors and their uses published from 2011 to 2013. Pre-2011 patents have been extensively covered in previous reviews. Comments on the context of each chemical series are given where applicable to orientate the readers on the bewildering array of molecular designs now available. EXPERT OPINION: The growing number of preclinical studies in numerous malignances as well as the first clinical trials of STAT3 inhibitors suggest that STAT3 remains a valid target for the treatment of human cancers as well as inflammatory diseases and/or autoimmune disorders. So, the future looks bright for patients because many new drugs are being developed and now combinations of STAT3 inhibitors with other targeted agents can diminish the resistance to traditional chemotherapy. These advances are expected to lead to further significant progress improving patient outcomes and quality of life.

Development of peptidomimetic boronates as proteasome inhibitors.

Proteasome inhibition has emerged over the past decade as an effective therapeutic approach for the treatment of hematologic malignancies. It is a multicatalytic complex, whose proteolytic activity relies in three types of subunits: chymotrypsin-like (ß5), trypsin-like (ß2) and caspase-like (ß1). Most important for the development of effective antitumor agents is the inhibition of the ß5 subunits. In this context, the dipeptide boronate bortezomib (Velcade(®)) represents the first proteasome inhibitor approved by the FDA and the lead compound in drug discovery. This paper describes the synthesis and biological evaluation of a series of conformationally constrained pseudopeptide boronates (1-3) structurally related to bortezomib. The synthesized compounds showed a promising inhibitory profile by blocking primarily the chymotrypsin-like activity of the proteasome with Ki values in submicromolar/micromolar range. These compounds also resulted quite selective since no significant inhibition was recorded in the test against bovine pancreatic α-chymotrypsin. The obtained results were rationalized by means of docking experiments based on a model of the crystal structure of bortezomib bound to the yeast 20S proteasome providing essential insights for further optimization of this class of inhibitors.

Discovery of new small molecules targeting the vitronectin-binding site of the urokinase receptor that block cancer cell invasion.

Besides focusing urokinase (uPA) proteolytic activity on the cell membrane, the uPA receptor (uPAR) is able to bind vitronectin, via a direct binding site. Furthermore, uPAR interacts with other cell surface receptors, such as integrins, receptor tyrosine kinases, and chemotaxis receptors, triggering cell-signaling pathways that promote tumor progression. The ability of uPAR to coordinate binding and degradation of extracellular matrix (ECM) and cell signaling makes it an attractive therapeutic target in cancer. We used structure-based virtual screening (SB-VS) to search for small molecules targeting the uPAR-binding site for vitronectin. Forty-one compounds were identified and tested on uPAR-negative HEK-293 epithelial cells transfected with uPAR (uPAR-293 cells), using the parental cell line transfected with the empty vector (V-293 cells) as a control. Compounds 6 and 37 selectively inhibited uPAR-293 cell adhesion to vitronectin and the resulting changes in cell morphology and signal transduction, without exerting any effect on V-293 cells. Compounds 6 and 37 inhibited uPAR-293 cell binding to vitronectin with IC50 values of 3.6 and 1.2 µmol/L, respectively. Compounds 6 and 37 targeted S88 and R91, key residues for uPAR binding to vitronectin but also for uPAR interaction with the fMLF family of chemotaxis receptors (fMLF-Rs). As a consequence, compounds 6 and 37 impaired uPAR-293 cell migration toward fetal calf serum (FCS), uPA, and fMLF, likely by inhibiting the interaction between uPAR and FPR1, the high affinity fMLF-R. Both compounds blocked in vitro ECM invasion of several cancer cell types, thus representing new promising leads for pharmaceuticals in cancer.

Discovery of a novel small molecule inhibitor targeting the frataxin/ubiquitin interaction via structure-based virtual screening and bioassays.

Friedreich's ataxia (FRDA) is an autosomal recessive neuro- and cardiodegenerative disorder for which there are no proven effective treatments. FRDA is caused by decreased expression and/or function of the mitochondrial protein frataxin. Here, we report findings that frataxin is degraded via the ubiquitin-proteasomal pathway and that it is ubiquitinated at residue K(147) in Calu-6 cells. A theoretical model of the frataxin-K(147)/Ub complex, constructed by combining bioinformatics interface predictions with information-driven docking, revealed a hitherto unnoticed, potential ubiquitin-binding domain in frataxin. Through structure-based virtual screening and cell-based assays, we discovered a novel small molecule (compound (+)-11) able to prevent frataxin ubiquitination and degradation. (+)-11 was synthesized and tested for specific binding to frataxin by an UF-LC/MS based ligand-binding assay. Follow-up scaffold-based searches resulted in the identification of a lead series with micromolar activity in disrupting the frataxin/Ub interaction. This study also suggests that frataxin could be a potential target for FRDA drug development.

Discovery of new inhibitors of Cdc25B dual specificity phosphatases by structure-based virtual screening.

Cell division cycle 25 (Cdc25) proteins are highly conserved dual specificity phosphatases that regulate cyclin-dependent kinases and represent attractive drug targets for anticancer therapies. To discover more potent and diverse inhibitors of Cdc25 biological activity, virtual screening was performed by docking 2.1 million compounds into the Cdc25B active site. An initial subset of top-ranked compounds was selected and assayed, and 15 were found to have enzyme inhibition activity at micromolar concentration. Among these, four structurally diverse inhibitors with a different inhibition profile were found to inhibit human MCF-7, PC-3, and K562 cancer cell proliferation and significantly affect the cell cycle progression. A subsequent hierarchical similarity search with the most active reversible Cdc25B inhibitor found led to the identification of an additional set of 19 ligands, three of which were confirmed as Cdc25B inhibitors with IC(50) values of 7.9, 4.2, and 9.9 µM, respectively.

Conformational switch of a flexible loop in human laminin receptor determines laminin-1 interaction.

The 37/67-kDa human laminin receptor(LamR) is a cell surface protein that interacts with molecules located in the extra-cellular matrix. In particular, interactions between LamR and laminins play a major role in mediating changes in the cellular environment that affect cell adhesion, neurite outgrowth, tumor growth and metastasis. The exact interaction mode of laminin-1 and LamR is not fully understood. Laminin-1 is thought to bind to LamR through interaction with the so-called peptide G (residues 161­180) and the C-terminal helix (residues 205­229). Here we performed 100-ns atomistic force field based molecular dynamics simulations to explore the structure and dynamics of LamR related to laminin-1 interactions. Our main finding is that loop 188­197 in the C-terminal region is highly flexible. It undergoes a major change resulting in a conformational switch that partially solvent exposes the R180 residue in the final part of the G peptide. So, R180 could contribute to laminin-1 binding. Projection of the simulations along the first two principal components also confirms the importance of this conformational switch in the LamR. This may be a basic prerequisite to clarify the key structural determinants of the interaction of LamR with laminin-1.

Synthesis, biological evaluation and molecular investigation of fluorinated peroxisome proliferator-activated receptors α/γ dual agonists.

PPARs are transcription factors that govern lipid and glucose homeostasis and play a central role in cardiovascular disease, obesity, and diabetes. Thus, there is significant interest in developing new agonists for these receptors. Given that the introduction of fluorine generally has a profound effect on the physical and/or biological properties of the target molecule, we synthesized a series of fluorinated analogs of the previously reported compound 2, some of which turned out to be remarkable PPARα and PPARγ dual agonists. Docking experiments were also carried out to gain insight into the interactions of the most active derivatives with both receptors.