Chiral Hydroxy Metabolite of Mebendazole: Analytical and Semi-Preparative High-Performance Liquid Chromatography Resolution and Chiroptical Properties.

Mebendazole (MBZ) is a benzimidazole carbamate anthelmintic used worldwide for the treatment and prevention of parasitic disorders in animals and humans. A large number of in vivo and in vitro studies have demonstrated that MBZ also has anticancer activity in multiple types of cancers. After oral administration, the phenylketone moiety of MBZ is rapidly reduced to the hydroxyl group to form the chiral hydroxy metabolite (MBZ-OH). To the best of our knowledge, there is no information in the literature on the stereochemical course of transformation and the anthelmintic and antitumor activity of individual enantiomers of MBZ-OH. In the present study, we describe in detail the direct HPLC resolution of MBZ-OH on a 100 mm × 4.6 mm Chirapak IG-3 column packed with 3 µm silica particles containing amylose (3-chloro-5-methylphenylcarbamate) as a selector. At 25 °C and using pure methanol as the mobile phase, the enantioseparation and resolution factors were 2.38 and 6.13, respectively. These conditions were scaled up at a semi-preparative scale using a 250 mm × 10 mm Chiralpak IG column to isolate multi-milligram amounts of both enantiomeric forms of the chiral metabolite. The chiroptical properties of the collected enantiomers were determined and, through a theoretical study, were related to the more stable conformations of MBZ-OH. The first and second eluted enantiomers were dextrorotatory and levorotatory, respectively, in dimethylformamide solution. Finally, by recording the retention factors of the enantiomers as the water content in the water-acetonitrile mobile phases was progressively varied, U-shaped retention maps were generated, indicating a dual and competitive hydrophilic interaction liquid chromatography and reversed-phase liquid chromatography retention mechanism on the Chirapak IG-3 chiral stationary phase.

2H-chromene and 7H-furo-chromene derivatives selectively inhibit tumour associated human carbonic anhydrase IX and XII isoforms.

Tumour associated carbonic anhydrases (CAs) IX and XII have been recognised as potential targets for the treatment of hypoxic tumours. Therefore, considering the high pharmacological potential of the chromene scaffold as selective ligand of the IX and XII isoforms, two libraries of compounds, namely 2H-chromene and 7H-furo-chromene derivatives, with diverse substitution patterns were designed and synthesised. The structure of the newly synthesised compounds was characterised and their inhibitory potency and selectivity towards human CA off target isoforms I, II and cancer-associated CA isoforms IX and XII were evaluated. Most of the compounds inhibit CA isoforms IX and XII with no activity against the I and II isozymes. Thus, while the potency was influenced by the substitution pattern along the chromene scaffold, the selectivity was conserved along the series, confirming the high potential of both 2H-chromene and 7H-furo-chromene scaffolds for the design of isozyme selective inhibitors.

Kinetic Study on the Base-Catalyzed Imine-Enamine Tautomerism of a Chiral Biologically Active Isoxazoline Derivative by HPLC on Amylose Tris(3,5-dimethylphenylcarbamate) Chiral Stationary Phase.

Isoxazoline is a nitrogen- and oxygen-containing five-membered heterocyclic scaffold with diverse biological profiles such as antimicrobial, fungicidal, anticancer, antiviral, analgesic and anti-inflammatory activity. Accordingly, the use of this peculiar structural framework in drug discovery is a successful strategy for the development of new drug candidates. Here, a chiral saccharin/isoxazoline hybrid was considered to investigate the tendency of the imine moiety of the heterocyclic ring to tautomerize to the enamine form in the presence of a basic catalyst. The pseudo-first-order rate constants for the base-catalyzed tautomerization process were measured in different solvents and at different temperatures by off-column kinetic experiments based on the amylose (3,5-dimethylphenylcarbamate)-type chiral stationary phase. The kinetic results obtained in this study may be a useful aid in the perspective of designing experimental conditions to control the stereointegrity of these types of pharmacologically active compounds and drive their synthesis toward the preferred, imine or enamine, tautomer.

Metal-organic frameworks (MOFs) as biomolecules drug delivery systems for anticancer purposes.

Metal-organic frameworks (MOF) have been studied for infinite applications. Among these, anticancer therapy has surely attracted great interest due to the intrinsic characteristics of MOFs: large surface area, tuneable porosity, remarkable biocompatibility, easy production, and the possibility to further functionalize the realized nanoparticles are the main reasons that make MOFs the ideal candidates to overcome traditional chemotherapy limits and resistance. Smart MOFs are becoming particularly relevant, as they can be activated by specific endogenous or exogenous stimuli and release their cargo only under the selected conditions. Tumor microenvironment (TME) offers the possibility to take advantage of its peculiar composition to design and build smart nanoparticles, able to selectively release the therapeutic payload only in the environment surrounding cancer cells or directly in the intracellular environment. In this review, we have summarized novel and innovative works describing anticancer MOF-based nanoparticles loaded with biomolecules published in the last three years. The reported papers have been selected with special focus on TME-responsive MOFs and the synthetic procedures employed by research groups have been reported for almost all works, to further underline the myriad of possibilities offered by these hybrid metal-organic structures.

Novel Insights on Human Carbonic Anhydrase Inhibitors Based on Coumalic Acid: Design, Synthesis, Molecular Modeling Investigation, and Biological Studies.

Human carbonic anhydrase (hCA, EC 4.2.1.1) isoforms IX and XII are overexpressed in solid hypoxic tumors, and they are considered as prognostic tools and therapeutic targets for cancer. Based on a molecular simplification of the well-known coumarin scaffold, we developed a new series of derivatives of the pyran-2-one core. The new compounds are endowed with potent and selective inhibitory activity against the tumor-related hCA isoforms IX and XII, in the low nanomolar range, whereas they are inactive against the two cytosolic off-targets hCA I and II. The compounds exhibiting the best hCA inhibition were further investigated against the breast adenocarcinoma cell line (MCF7) in hypoxic conditions, evaluating their ability to eventually synergize with doxorubicin. The compounds' biocompatibility on healthy cells was also tested and confirmed on Human Gingival Fibroblasts (HGFs). Furthermore, the possible binding mode of all compounds to the active site of the tumor-associated human CA IX was investigated by computational techniques which predicted the binding conformations and the persistency of binding poses within the active site of the enzyme, furnishing relevant data for the design of tight binding inhibitors.

Synthesis and Evaluation of Thymol-Based Synthetic Derivatives as Dual-Action Inhibitors against Different Strains of H. pylori and AGS Cell Line.

Following a similar approach on carvacrol-based derivatives, we investigated the synthesis and the microbiological screening against eight strains of H. pylori, and the cytotoxic activity against human gastric adenocarcinoma (AGS) cells of a new series of ether compounds based on the structure of thymol. Structural analysis comprehended elemental analysis and 1H/13C/19F NMR spectra. The analysis of structure-activity relationships within this molecular library of 38 structurally-related compounds reported that some chemical modifications of the OH group of thymol led to broad-spectrum growth inhibition on all isolates. Preferred substitutions were benzyl groups compared to alkyl chains, and the specific presence of functional groups at para position of the benzyl moiety such as 4-CN and 4-Ph endowed the most anti-H. pylori activity toward all the strains with minimum inhibitory concentration (MIC) values up to 4 µg/mL. Poly-substitution on the benzyl ring was not essential. Moreover, several compounds characterized by the lowest minimum inhibitory concentration/minimum bactericidal concentration (MIC/MBC) values against H. pylori were also tested in order to verify a cytotoxic effect against AGS cells with respect to 5-fluorouracil and carvacrol. Three derivatives can be considered as new lead compounds alternative to current therapy to manage H. pylori infection, preventing the occurrence of severe gastric diseases. The present work confirms the possibility to use natural compounds as templates for the medicinal semi-synthesis.

Selective inhibition of carbonic anhydrase IX and XII by coumarin and psoralen derivatives.

A small library of coumarin and their psoralen analogues EMAC10157a-b-d-g and EMAC10160a-b-d-g has been designed and synthesised to investigate the effect of structural modifications on their inhibition ability and selectivity profile towards carbonic anhydrase isoforms I, II, IX, and XII. None of the new compounds exhibited activity towards hCA I and II isozymes. Conversely, both coumarin and psoralen derivatives were active against tumour associated isoforms IX and XII in the low micromolar or nanomolar range of concentration. These data further corroborate our previous findings on analogous derivatives, confirming that both coumarins and psoralens are interesting scaffolds for the design of isozyme selective hCA inhibitors.

Synthesis and Biological Evaluation of Carvacrol-Based Derivatives as Dual Inhibitors of H. pylori Strains and AGS Cell Proliferation.

This study reports on the synthesis, structural assessment, microbiological screening against several strains of H. pylori and antiproliferative activity against human gastric adenocarcinoma (AGS) cells of a large series of carvacrol-based compounds. Structural analyses consisted of elemental analysis, 1H/13C/19F NMR spectra and crystallographic studies. The structure-activity relationships evidenced that among ether derivatives the substitution with specific electron-withdrawing groups (CF3 and NO2) especially in the para position of the benzyl ring led to an improvement of the antimicrobial activity, whereas electron-donating groups on the benzyl ring and ethereal alkyl chains were not tolerated with respect to the parent compound (MIC/MBC = 64/64 µg/mL). Ester derivatives (coumarin-carvacrol hybrids) displayed a slight enhancement of the inhibitory activity up to MIC values of 8-16 µg/mL. The most interesting compounds exhibiting the lowest MIC/MBC activity against H. pylori (among others, compounds 16 and 39 endowed with MIC/MBC values ranging between 2/2 to 32/32 µg/mL against all the evaluated strains) were also assayed for their ability to reduce AGS cell growth with respect to 5-Fluorouracil. Some derivatives can be regarded as new lead compounds able to reduce H. pylori growth and to counteract the proliferation of AGS cells, both contributing to the occurrence of gastric cancer.

Novel insights on saccharin- and acesulfame-based carbonic anhydrase inhibitors: design, synthesis, modelling investigations and biological activity evaluation.

A large library of saccharin and acesulfame derivatives has been synthesised and evaluated against four isoforms of human carbonic anhydrase, the two off-targets hCA I/II and the tumour related isoforms hCA IX/XII. Different strategies of scaffold modification have been attempted on both saccharin as well as acesulfame core leading to the obtainment of 60 compounds. Some of them exhibited inhibitory activity in the nanomolar range, albeit some of the performed changes led to either micromolar activity or to its absence, against hCA IX/XII. Molecular modelling studies focused the attention on the binding mode of these compounds to the enzyme. The proposed inhibition mechanism is the anchoring to zinc-bound water molecule. Docking studies along with molecular dynamics also underlined the importance of the compounds flexibility (e.g. achieved through the insertion of methylene group) which favoured potent and selective hCA inhibition.

Synthesis and evaluation of a large library of nitroxoline derivatives as pancreatic cancer antiproliferative agents.

Pancreatic cancer (PC) is one of the deadliest carcinomas and in most cases, which are diagnosed with locally advanced or metastatic disease, current therapeutic options are highly unsatisfactory. Based on the anti-proliferative effects shown by nitroxoline, an old urinary antibacterial agent, we explored a large library of newly synthesised derivatives to unravel the importance of the OH moiety and pyridine ring of the parent compound. The new derivatives showed a valuable anti-proliferative effect and some displayed a greater effect as compared to nitroxoline against three pancreatic cancer cell lines with different genetic profiles. In particular, in silico pharmacokinetic data, clonogenicity assays and selectivity indexes of the most promising compounds showed several advantages for such derivatives, as compared to nitroxoline. Moreover, some of these novel compounds had stronger effects on cell viability and/or clonogenic capacity in PC cells as compared to erlotinib, a targeted agent approved for PC treatment.

Inhibition of lysine acetyltransferases impairs tumor angiogenesis acting on both endothelial and tumor cells.

BACKGROUND: Understanding the signalling pathways involved in angiogenesis, and developing anti-angiogenic drugs are one of the major focuses on cancer research. Herein, we assessed the effect of CPTH6, a lysine acetyltransferase inhibitor and anti-tumoral compound, on angiogenesis-related properties of both endothelial and cancer cells. METHODS: The in vitro effect of CPTH6 on protein acetylation and anti-angiogenic properties on endothelial and lung cancer cells was evaluated via wound healing, trans-well invasion and migration, tube formation, immunoblotting and immunofluorescence. Matrigel plug assay, zebrafish embryo and mouse xenograft models were used to evaluate in vivo anti-angiogenic effect of CPTH6. RESULTS: CPTH6 impaired in vitro endothelial angiogenesis-related functions, and decreased the in vivo vascularization both in mice xenografts and zebrafish embryos. Mechanistically, CPTH6 reduced α-tubulin acetylation and induced accumulation of acetylated microtubules in the perinuclear region of endothelial cells. Interestingly, CPTH6 also affected the angiogenesis-related properties of lung cancer cells, and conditioned media derived from CPTH6-treated lung cancer cells impaired endothelial cells morphogenesis. CPTH6 also modulated the VEGF/VEGFR2 pathway, and reshaped cytoskeletal organization of lung cancer cells. Finally, anti-migratory effect of CPTH6, dependent on α-tubulin acetylation, was also demonstrated by genetic approaches in lung cancer cells. CONCLUSION: Overall, this study indicates that α-tubulin acetylation could play a role in the anti-angiogenic effect of CPTH6 and, more in general, it adds information to the role of histone acetyltransferases in tumor angiogenesis, and proposes the inhibition of these enzymes as an antiangiogenic therapy of cancer.

Metabolic profiling of different wild and cultivated Allium species based on high-resolution mass spectrometry, high-performance liquid chromatography-photodiode array detector, and color analysis.

Many plants of the genus Allium are widely cultivated and consumed for their nutraceutical and health-enhancing bioactive components effective in many metabolic and infectious diseases. In particular, Allium sativum L. (garlic), the most economically important Allium species, is known to present volatile, comparatively polar sulfur-containing compounds responsible for both the typical garlic aroma and antimicrobial property. More recently, the (moderately) polar portion of garlic metabolome, rich of polyphenols and amino acids, is gaining increasing interest as a source of antioxidants and primary nutrients. In this study, we have explored the chemical diversity of eight different hydroalcoholic extracts obtained by microwave-assisted extraction of white and red crop A. sativum and wild Allium triquetrum, Allium roseum, and Allium ampeloprasum, all originating from the Mediterranean Basin. The aim is to appraise their potential dietetic and healing value through an in-depth chemical characterization and contribute to preserve and exploit natural resources. The multimethodological method applied here is based on an untargeted metabolic profiling by means of high-resolution electrospray ionization Fourier-transform ion cyclotron resonance (ESI FT-ICR) mass spectrometry. More than 850 by ESI(+) and 450 by ESI(-) putative metabolites have been annotated covering all main classes of primary and secondary metabolites, including amino acids, alkaloids, organic and fatty acids, nucleotides, vitamins, organosulfur compounds, and flavonoids. The pigment and polyphenol components have been separated and quantified by a targeted chromatographic high-performance liquid chromatography-photodiode array detector (HPLC-PDA) and CIEL*a*b* colorimetric assay, showing characteristic yellow and red components in each extract, related to a different milieu of anthocyanins and flavonoids as assigned by high-resolution mass spectrometry (MS).

Synthesis, Antiproliferative Effect, and Topoisomerase II Inhibitory Activity of 3-Methyl-2-phenyl-1H-indoles.

A series of 3-methyl-2-phenyl-1H-indoles was prepared and investigated for antiproliferative activity on three human tumor cell lines, HeLa, A2780, and MSTO-211H, and some structure-activity relationships were drawn up. The GI50 values of the most potent compounds (32 and 33) were lower than 5 µM in all tested cell lines. For the most biologically relevant derivatives, the effect on human DNA topoisomerase II relaxation activity was investigated, which highlighted the good correlation between the antiproliferative effect and topoisomerase II inhibition. The most potent derivative, 32, was shown to induce the apoptosis pathway. The obtained results highlight 3-methyl-2-phenyl-1H-indole as a promising scaffold for further optimization of compounds with potent antiproliferative and antitopoisomerase II activities.

1,3-Dipolar Cycloaddition, HPLC Enantioseparation, and Docking Studies of Saccharin/Isoxazole and Saccharin/Isoxazoline Derivatives as Selective Carbonic Anhydrase IX and XII Inhibitors.

Two series of saccharin/isoxazole and saccharin/isoxazoline hybrids were synthesized by 1,3-dipolar cycloaddition. The new compounds showed to be endowed with potent and selective inhibitory activity against the cancer-related human carbonic anhydrase (hCA) IX and XII isoforms in the nanomolar range, while no affinity was encountered for off-targets, such as hCA I and II. Successive enantioseparation on a milligram scale of the most representative compounds led to the discovery that (S)-isomers were more potent than their corresponding (R)-enantiomers. Lastly, molecular modeling studies were conducted to define those structural requirements that were responsible for the discrimination among selected human isoforms of carbonic anhydrases. Two nanomolar hCA IX and XII inhibitors were also screened for their selective toxicity against non tumoral primary cells (fibroblasts) and against a breast adenocarcinoma cell line (MCF7) in hypoxic environment. The efficacious combination of these compounds with doxorubicin on MCF7 cells was demonstrated after 72 h of treatment.

MAO inhibitors and their wider applications: a patent review.

INTRODUCTION: Monoamine oxidase (MAO) inhibitors, after the initial 'golden age', are currently used as third-line antidepressants (selective MAO-A inhibitors) or clinically enrolled as co-adjuvants for neurodegenerative diseases (selective MAO-B inhibitors). However, the research within this field is always increasing due to their pivotal role in modulating synaptic functions and monoamines metabolism. AREAS COVERED: In this paper, MAO inhibitors (2015-2017) are disclosed ordering all the patents according to their chemical scaffold. Structure-activity relationships (SARs) are extrapolated for the most investigated chemotypes (coumarins, pyrazole/oxazepinones, (hetero)arylamides). 108 Compounds are divided into two main groups: newly synthesized molecules and naturally-occurring metabolites. Finally, new therapeutic options are outlined to ensure a more complete view on the potential of these inhibitors. EXPERT OPINION: New proposed MAO inhibitors are endowed with a marked isoform selectivity, with innovative therapeutic potential toward other targets (gliomas, inflammation, muscle dystrophies, migraine, chronic pain, pseudobulbar affect), and with a promising ability to address multi-faceted pathologies such as Alzheimer's disease. The increasing number of patents is analyzed collecting data from 2002 to 2017.

Synthesis, biological evaluation and quantitative structure-active relationships of 1,3-thiazolidin-4-one derivatives. A promising chemical scaffold endowed with high antifungal potency and low cytotoxicity.

With reference to recent studies reporting on the various biological properties of the thiazolidinone scaffold, we synthesized more than a hundred compounds characterized by a 1,3-thiazolidin-4-one nucleus derivatised at the C2 with a hydrazine bridge linked to (cyclo)aliphatic or hetero(aryl) moieties, and their N-benzylated derivatives. These molecules were assayed as potential anti-Candida agents and they were shown to possess comparable, and in some cases higher biological activity than well-established topical and systemic antimycotic drugs (i.e. clotrimazole, fluconazole, ketoconazole, miconazole, tioconazole, amphotericin B). Compounds endowed with the lowest MICs underwent further testing in order to assess their cytotoxic effect (CC50) on Hep2 cells, which demonstrated their relative safety. Finally, QSAR and 3-D QSAR models were used to predict putative chemical modifications of the 1,3-thiazolidin-4-one scaffold in order to design new and potential more active compounds against Candida spp.

Neem oil nanoemulsions: characterisation and antioxidant activity.

The aim of the present work is to develop nanoemulsions (NEs), nanosized emulsions, manufactured for improving the delivery of active pharmaceutical ingredients. In particular, nanoemulsions composed of Neem seed oil, contain rich bioactive components, and Tween 20 as nonionic surfactant were prepared. A mean droplet size ranging from 10 to 100 nm was obtained by modulating the oil/surfactant ratio. Physicochemical characterisation was carried out evaluating size, ζ-potential, microviscosity, polarity and turbidity of the external shell and morphology, along with stability in simulated cerebrospinal fluid (CSF), activity of Neem oil alone and in NEs, HEp-2 cell interaction and cytotoxicity studies. This study confirms the formation of NEs by Tween 20 and Neem oil at different weight ratios with small and homogenous dimensions. The antioxidant activity of Neem oil alone and in NEs was comparable, whereas its cytotoxicity was strongly reduced when loaded in NEs after interaction with HEp-2 cells.

Protective Effects Induced by Microwave-Assisted Aqueous Harpagophytum Extract on Rat Cortex Synaptosomes Challenged with Amyloid ß-Peptide.

Harpagophytum procumbens is a plant species that displays anti-inflammatory properties in multiple tissues. The iridoid glycosides arpagoside, harpagide, and procumbide appear to be the most therapeutically important constituents. In addition, harpagoside treatment exerted neuroprotective effects both in vitro and in vivo. Considering these findings, the aim of the present work is to explore the possible protective role of the previously described microwave-assisted aqueous extract of H. procumbens on rat hypothalamic (Hypo-E22) cells, and in rat cortex challenged with amyloid ß-peptide (1-40). In this context, we assayed the protective effects induced by H. procumbens by measuring the levels of malondialdehyde, 3-hydroxykynurenine (3-HK), brain-derived neurotrophic factor, and tumor necrosis factor-α, 3-HK. Finally, we evaluated the effects of H. procumbens treatment on cortex levels of dopamine, norepinephrine, and serotonin. H. procumbens extract was well tolerated by Hypo-E22 cells and upregulated brain-derived neurotrophic factor gene expression but down-regulated tumor necrosis factor-α gene expression. In addition, the extract reduced amyloid ß-peptide stimulation of malondialdehyde and 3-HK and blunted the decrease of dopamine, norepinephrine, and serotonin, in the cortex. In this context, our work supports further studies for the evaluation and confirmation of Harpagophytum in the management of the clinical symptoms related to Alzheimer's disease. Copyright © 2017 John Wiley & Sons, Ltd.

Optimization of Aqueous Extraction and Biological Activity of Harpagophytum procumbens Root on Ex Vivo Rat Colon Inflammatory Model.

Harpagophytum procumbens has a long story of use for the treatment of inflammatory diseases. Considering both the antiinflammatory effects of H. procumbens in multiple tissues and the stability of harpagoside in artificial intestinal fluid, the aim of the present study was to explore the possible protective role of a microwave-assisted aqueous Harpagophytum extract (1-1000 µg/mL) on mouse myoblast C2C12 and human colorectal adenocarcinoma HCT116 cell lines, and isolated rat colon specimens challenged with lipopolysaccharide (LPS), a validated ex vivo model of acute ulcerative colitis. In this context, we evaluated the effects on C2C12 and HCT116 viability, and on LPS-induced production of serotonin (5-HT), tumor necrosis factor (TNF)-α, prostaglandin (PG)E2 and 8-iso-prostaglandin (8-iso-PG)F2α . Harpagophytum extract was well tolerated by C2C12 cells, while reduced HCT116 colon cancer cell viability. On the other hand, Harpagophytum extract reduced H2 O2 -induced (1 mM) reactive oxygen species (ROS) production, in both cell lines, and inhibited LPS-induced colon production of PGE2 , 8-iso-PGF2α , 5-HT and TNFα. Concluding, we demonstrated the efficacy of a microwave-assisted Harpagophytum aqueous extract in modulating the inflammatory, oxidative stress and immune response in an experimental model of inflammatory bowel diseases (IBD), thus suggesting a rational use of Harpagophytum in the management and prevention of ulcerative colitis in humans. Copyright © 2017 John Wiley & Sons, Ltd.

Open saccharin-based secondary sulfonamides as potent and selective inhibitors of cancer-related carbonic anhydrase IX and XII isoforms.

A large number of novel secondary sulfonamides based on the open saccharin scaffold were synthesized and evaluated as selective inhibitors of four different isoforms of human carbonic anhydrase (hCA I, II, IX and XII, EC 4.2.1.1). They were obtained by reductive ring opening of the newly synthesized N-alkylated saccharin derivatives and were shown to be inactive against the two cytosolic off-target hCA I and II (Kis > 10 µM). Interestingly, these compounds inhibited hCA IX in the low nanomolar range with Kis ranging between 20 and 298 nM and were extremely potent inhibitors of hCA XII isoenzyme (Kis ranging between 4.3 and 432 nM). Since hCA IX and XII are the cancer-related isoforms recently validated as drug targets, these results represent an important goal in the development of new anticancer candidates. Finally, a computational approach has been performed to better correlate the biological data to the binding mode of these inhibitors.