Quantitative live cell imaging of a tauopathy model enables the identification of a polypharmacological drug candidate that restores physiological microtubule interaction.

Tauopathies such as Alzheimer's disease are characterized by aggregation and increased phosphorylation of the microtubule-associated protein tau. Tau's pathological changes are closely linked to neurodegeneration, making tau a prime candidate for intervention. We developed an approach to monitor pathological changes of aggregation-prone human tau in living neurons. We identified 2-phenyloxazole (PHOX) derivatives as putative polypharmacological small molecules that interact with tau and modulate tau kinases. We found that PHOX15 inhibits tau aggregation, restores tau's physiological microtubule interaction, and reduces tau phosphorylation at disease-relevant sites. Molecular dynamics simulations highlight cryptic channel-like pockets crossing tau protofilaments and suggest that PHOX15 binding reduces the protofilament's ability to adopt a PHF-like conformation by modifying a key glycine triad. Our data demonstrate that live-cell imaging of a tauopathy model enables screening of compounds that modulate tau-microtubule interaction and allows identification of a promising polypharmacological drug candidate that simultaneously inhibits tau aggregation and reduces tau phosphorylation.

Targeting Ovarian Cancer with Chalcone Derivatives: Cytotoxicity and Apoptosis Induction in HGSOC Cells.

Ovarian cancer ranks as the eighth most prevalent form of cancer in women across the globe and stands as the third most frequent gynecological cancer, following cervical and endometrial cancers. Given its resistance to standard chemotherapy and high recurrence rates, there is an urgent imperative to discover novel compounds with potential as chemotherapeutic agents for treating ovarian cancer. Chalcones exhibit a wide array of biological properties, with a particular focus on their anti-cancer activities. In this research, we documented the synthesis and in vitro study of a small library of chalcone derivatives designed for use against high-grade serous ovarian cancer (HGSOC) cell lines, specifically OVCAR-3, OVSAHO, and KURAMOCHI. Our findings revealed that three of these compounds exhibited cytotoxic and anti-proliferative effects against all the tested HGSOC cell lines, achieving IC50 concentrations lower than 25 µM. Further investigations disclosed that these chalcones prompted an increase in the subG1 phase cell cycle and induced apoptosis in OVCAR-3 cells. In summary, our study underscores the potential of chalcones as promising agents for the treatment of ovarian cancer.

Tetraphenylethylene-Based Photoluminescent Self-Assembled Nanoparticles: Preparation and Biological Evaluation.

The conjugation of tetraphenylethylene (TPE) with podophyllotoxin, N-desacetylthiocolchicine, and cabazitaxel through a sebacic acid linker led to the formation of fluorescent nanoparticles. Dynamic light scattering (DLS) and photoluminescence spectroscopy were used for the identification and characterization of the fluorescent nanoparticles. The biological evaluation was determined in three human ovarian (KURAMOCHI, OVCAR3, OVSAHO) and three human breast (MCF7, SKBR 3, and MDA-MB231) cancer cell lines. In the case of cabazitaxel, the nanoparticles maintained the activity of the parent drug, at the low nanomolar range, while exhibiting high blue fluorescence. The internalization of the fluorescent NPs into cells was detected using immunofluorescence assay.

Exploration of Novel Scaffolds Targeting Cytochrome b of Pyricularia oryzae.

The fulfilment of the European "Farm to Fork" strategy requires a drastic reduction in the use of "at risk" synthetic pesticides; this exposes vulnerable agricultural sectors-among which is the European risiculture-to the lack of efficient means for the management of devastating diseases, thus endangering food security. Therefore, novel scaffolds need to be identified for the synthesis of new and more environmentally friendly fungicides. In the present work, we employed our previously developed 3D model of P. oryzae cytochrome bc1 (cyt bc1) complex to perform a high-throughput virtual screening of two commercially available compound libraries. Three chemotypes were selected, from which a small collection of differently substituted analogues was designed and synthesized. The compounds were tested as inhibitors of the cyt bc1 enzyme function and the mycelium growth of both strobilurin-sensitive (WT) and -resistant (RES) P. oryzae strains. This pipeline has permitted the identification of thirteen compounds active against the RES cyt bc1 and five compounds that inhibited the WT cyt bc1 function while inhibiting the fungal mycelia only minimally. Serendipitously, among the studied compounds we identified a new chemotype that is able to efficiently inhibit the mycelium growth of WT and RES strains by ca. 60%, without inhibiting the cyt bc1 enzymatic function, suggesting a different mechanism of action.

Design, synthesis, biological evaluation and crystal structure determination of dual modulators of carbonic anhydrases and estrogen receptors.

Multi-target compounds have become increasingly important for the development of safer and more effective drug candidates. In this work, we devised a combined ligand-based and structure-based multi-target repurposing strategy and applied it to a series of hexahydrocyclopenta[c]quinoline compounds synthesized previously. The in silico analyses identified human Carbonic Anhydrases (hCA) and Estrogen Receptors (ER) as top scoring candidates for dual modulation. hCA isoforms IX and XII, and ER subtypes ER⍺ and/or ERß are co-expressed in various cancer cell types, including breast and prostate cancer cells. ER⍺ is the primary target of anti-estrogen therapy in breast cancer, and the hCA IX isoform is a therapeutic target in triple-negative breast cancer. ER⍺-mediated transcriptional programs and hCA activity in cancer cells promote favorable microenvironments for cell proliferation. Interestingly, several lines of evidence indicate that the combined modulation of these two targets may provide significant therapeutic benefits. Moving from these first results, two additional hexahydrocyclopenta[c]quinoline derivatives bearing a sulfonamide zinc binding group (hCA) and a phenolic hydroxyl (ER) pharmacophoric group placed at the appropriate locations were designed and synthesized. Interestingly, these compounds were able to directly modulate the activities of both hCA and ER targets. In cell-based assays, they inhibited proliferation of breast and prostate cancer cells with micromolar potency and cell type-selective efficacy. The compounds inhibited hCA activity with nanomolar potency and isoform-selectivity. In transactivation assays, they reduced estrogen-driven ER activity with micro-molar potency. Finally, crystal structures of the synthesized ligands in complex with the two targets revealed that the compounds bind directly to the hCA active site, as well as to the ER ligand-binding domain, providing structural explanation to the observed activity and a rationale for optimization of their dual activity. To the best of our knowledge, this work describes the design, synthesis and biological characterization of the first dual modulators of hCA and ER, laying the ground for the structure-based optimization of their multi-target activity.

Correlation between Perturbation of Redox Homeostasis and Antibiofilm Capacity of Phytochemicals at Non-Lethal Concentrations.

Biofilms are the multicellular lifestyle of microorganisms and are present on potentially every type of biotic or abiotic surface. Detrimental biofilms are generally targeted with antimicrobial compounds. Phytochemicals at sub-lethal concentrations seem to be an exciting alternative strategy to control biofilms, as they are less likely to impose selective pressure leading to resistance. This overview gathers the literature on individual phytocompounds rather than on extracts of which the use is difficult to reproduce. To the best of our knowledge, this is the first review to target only individual phytochemicals below inhibitory concentrations against biofilm formation. We explored whether there is an overall mechanism that can explain the effects of individual phytochemicals at sub-lethal concentrations. Interestingly, in all experiments reported here in which oxidative stress was investigated, a modest increase in intracellular reactive oxygen species was reported in treated cells compared to untreated specimens. At sub-lethal concentrations, polyphenolic substances likely act as pro-oxidants by disturbing the healthy redox cycle and causing an accumulation of reactive oxygen species.

Fatty Acids/Tetraphenylethylene Conjugates: Hybrid AIEgens for the Preparation of Peptide-Based Supramolecular Gels.

Aggregation-induced emissive materials are gaining particular attention in the last decades due to their wide application in different fields, from optical devices to biomedicine. In this work, compounds having these kinds of properties, composed of tetraphenylethylene scaffold combined with fatty acids of different lengths, were synthesized and characterized. These molecules were found able to self-assemble into different supramolecular emissive structures depending on the chemical composition and water content. Furthermore, they were used as N-terminus capping agents in the development of peptide-based materials. The functionalization of a 5-mer laminin-derived peptide led to the obtainment of luminescent fibrillary materials that were not cytotoxic and were able to form supramolecular gels in aqueous environment.

Design and Synthesis of Hsp90 Inhibitors with B-Raf and PDHK1 Multi-Target Activity.

The design of multi-target ligands has become an innovative approach for the identification of effective therapeutic treatments against complex diseases, such as cancer. Recent studies have demonstrated that the combined inhibition of Hsp90 and B-Raf provides synergistic effects against several types of cancers. Moreover, it has been reported that PDHK1, which presents an ATP-binding pocket similar to that of Hsp90, plays an important role in tumor initiation, maintenance and progression, participating also to the senescence process induced by B-Raf oncogenic proteins. Based on these premises, the simultaneous inhibition of these targets may provide several benefits for the treatment of cancer. In this work, we set up a design strategy including the assembly and integration of molecular fragments known to be important for binding to the Hsp90, PDHK1 and B-Raf targets, aided by molecular docking for the selection of a set of compounds potentially able to exert Hsp90-B-Raf-PDHK1 multi-target activities. The designed compounds were synthesized and experimentally validated in vitro. According to the in vitro assays, compounds 4 a, 4 d and 4 e potently inhibited Hsp90 and moderately inhibited the PDHK1 kinase. Finally, molecular dynamics simulations were performed to provide further insights into the structural basis of their multi-target activity.

Promising Non-cytotoxic Monosubstituted Chalcones to Target Monoamine Oxidase-B.

A library of monosubstituted chalcones (1-17) bearing electron-donating and electron-withdrawing groups on both aromatic rings were selected. The cell viability on human tumor cell lines was evaluated first. The compounds unable to induce detectable cytotoxicity (1, 13, and 14) were tested using the monoamine oxidase (MAO) activity assay. Interestingly, they inhibit MAO-B, acting as competitive inhibitors, with 13 and 14 showing the best profiles. In particular, 13 exhibited a potency higher than that of safinamide, taken as a reference. Docking studies and crystallographic analysis showed that in human MAO-B 13 binds with the halogen-substituted aromatic ring in the entrance cavity, similar to safinamide, whereas 14 is accommodated in the opposite way. The main conclusion of this cell biology, biochemistry, and structural study is to highlights 13 as a chalcone derivative that is worth consideration for the development of novel MAO-B-selective inhibitors for the treatment of neurodegenerative diseases.

Biological Properties of New Chiral 2-Methyl-5,6,7,8-tetrahydroquinolin-8-amine-based Compounds.

The synthesis of a small library of 8-substituted 2-methyl-5,6,7,8-tetrahydroquinoline derivatives is presented. All the compounds were tested for their antiproliferative activity in non-cancer human dermal microvascular endothelial cells (HMEC-1) and cancer cells: human T-lymphocyte cells (CEM), human cervix carcinoma cells (HeLa), human dermal microvascular endothelial cells (HMEC-1), colorectal adenocarcinoma (HT-29), ovarian carcinoma (A2780), and biphasic mesothelioma (MSTO-211H). Compounds 3a, 5a, and 2b, showing significant IC50 values against the whole panel of the selected cells, were further synthesized and tested as pure enantiomers in order to shed light on how their stereochemistry might impact on the related biological effect. The most active compound (R)-5a was able to affect cell cycle phases and to induce mitochondrial membrane depolarization and cellular ROS production in A2780 cells.

Engineered Ferritin Nanoparticles for the Bioluminescence Tracking of Nanodrug Delivery in Cancer.

The identification of a highly sensitive method to check the delivery of administered nanodrugs into the tumor cells is a crucial step of preclinical studies aimed to develop new nanoformulated cures, since it allows the real therapeutic potential of these devices to be forecast. In the present work, the ability of an H-ferritin (HFn) nanocage, already investigated as a powerful tool for cancer therapy thanks to its ability to actively interact with the transferrin receptor 1, to act as an efficient probe for the monitoring of nanodrug delivery to tumors is demonstrated. The final formulation is a bioluminescent nanoparticle, where the luciferin probe is conjugated on nanoparticle surface by means of a disulfide containing linker (Luc-linker@HFn) which is subjected to glutathione-induced cyclization in tumor cell cytoplasm. The prolonged imaging of luciferase+ tumor models, demonstrated by an in vitro and an in vivo approach, associated with the prolonged release of luciferin into cancer cells by disulfide bridge reduction, clearly indicates the high efficiency of Luc-linker@HFn for drug delivery to the tumor tissues.

A small library of chalcones induce liver cancer cell death through Akt phosphorylation inhibition.

Hepatocellular carcinoma (HCC) ranks as the fifth most common and the second deadliest cancer worldwide. HCC is extremely resistant to the conventional chemotherapeutics. Hence, it is vital to develop new treatment options. Chalcones were previously shown to have anticancer activities in other cancer types. In this study, 11 chalcones along with quercetin, papaverin, catechin, Sorafenib and 5FU were analyzed for their bioactivities on 6 HCC cell lines and on dental pulp stem cells (DPSC) which differentiates into hepatocytes, and is used as a model for untransformed control cells. 3 of the chalcones (1, 9 and 11) were selected for further investigation due to their high cytotoxicity against liver cancer cells and compared to the other clinically established compounds. Chalcones did not show significant bioactivity ([Formula: see text]) on dental pulp stem cells. Cell cycle analysis revealed that these 3 chalcone-molecules induced SubG1/G1 arrest. Akt protein phosphorylation was inhibited by these molecules in PTEN deficient, drug resistant, mesenchymal like Mahlavu cells leading to the activation of p21 and the inhibition of NF[Formula: see text]B-p65 transcription factor. Hence the chalcones induced apoptotic cell death pathway through NF[Formula: see text]B-p65 inhibition. On the other hand, these molecules triggered p21 dependent activation of Rb protein and thereby inhibition of cell cycle and cell growth in liver cancer cells. Involvement of PI3K/Akt pathway hyperactivation was previously described in survival of liver cancer cells as carcinogenic event. Therefore, our results indicated that these chalcones can be considered as candidates for liver cancer therapeutics particularly when PI3K/Akt pathway involved in tumor development.

Antiproliferative effects of chalcones on T cell acute lymphoblastic leukemia-derived cells: Role of PKCß.

In this study, a series of 20 chalcone derivatives was synthesized, and their antiproliferative activity was tested against the human T cell acute lymphoblastic leukemia-derived cell line, CCRF-CEM. On the basis of the structural features of the most active compounds, a new library of chalcone derivatives, according to the structure-activity relationship design, was synthesized, and their antiproliferative activity was tested against the same cancer cell line. Furthermore, four of these derivatives (compounds 3, 4, 8, 28), based on lower IC50 values (between 6.1 and 8.9 µM), were selected for further investigation regarding the modulation of the protein expression of RACK1 (receptor for activated C kinase), protein kinase C (PKC)α and PKCß, and their action on the cell cycle level. The cell cycle analysis indicated a block in the G0/G1 phase for all four compounds, with a statistically significant decrease in the percentage of cells in the S phase, with no indication of apoptosis (sub-G0/G1 phase). Compounds 4 and 8 showed a statistically significant reduction in the expression of PKCα and an increase in PKCß, which together with the demonstration of an antiproliferative role of PKCß, as assessed by treating cells with a selective PKCß activator, indicated that the observed antiproliferative effect is likely to be mediated through PKCß induction.

Divergent Conversion of 4-Naphthoquinone-substituted 4H-Isoxazolones to Different Benzo-fused Indole Derivatives.

4,4-Disubstituted 4H-isoxazol-5-ones bearing a 1,4-naphthoquinone moiety undergo transformation into different types of benzoindolyl products depending on the different reaction conditions. A decarboxylative ring opening/ring closure promoted by catalytic [Ru(p-cymene)2Cl2]2 yields benzo[f]indole-4,9-diones. Alternatively, hydrogenation reactions provide the conversion of 4-(1,4-naphthoquinone)-substituted isoxazol-5-ones to benzo[g]indole compounds, with the level of reduction depending on the substituents present on the ring. Starting materials have been easily prepared by the functionalization of isoxazolinones with naphthoquinone under mild conditions.

Novel 3,3-disubstituted oxindole derivatives. Synthesis and evaluation of the anti-proliferative activity.

3,3-Disubstituted oxindole derivatives bearing a nitrogen atom at the C-3 position have been synthesized starting from 3-alkyl oxindole through a metal free pathway. These derivatives have been tested in five human tumor cell lines (PC3, MCF7, SW620, MiaPaca2 and A375) and on primary cells (PBMCs) from healthy donors providing compound 6d showing a strong anticancer effect in all cancer lines on the low micromolar range.

Nanolipid-Trehalose Conjugates and Nano-Assemblies as Putative Autophagy Inducers.

The disaccharide trehalose is an autophagy inducer, but its pharmacological application is severely limited by its poor pharmacokinetics properties. Thus, trehalose was coupled via suitable spacers with squalene (in 1:2 and 1:1 stoichiometry) and with betulinic acid (1:2 stoichiometry), in order to yield the corresponding nanolipid-trehalose conjugates 1-Sq-mono, 2-Sq-bis and 3-Be-mono. The conjugates were assembled to produce the corresponding nano-assemblies (NAs) Sq-NA1, Sq-NA2 and Be-NA3. The synthetic and assembly protocols are described in detail. The resulting NAs were characterized in terms of loading and structure, and tested in vitro for their capability to induce autophagy. Our results are presented and thoroughly commented upon.

Vancomycin-Iridium (III) Interaction: An Unexplored Route for Enantioselective Imine Reduction.

The chiral structure of antibiotic vancomycin (Van) was exploited as an innovative coordination sphere for the preparation of an IrCp* based hybrid catalysts. We found that Van is able to coordinate iridium (Ir(III)) and the complexation was demonstrated by several analytical techniques such as MALDI-TOF, UV, Circular dichroism (CD), Raman IR, and NMR. The hybrid system so obtained was employed in the Asymmetric Transfer Hydrogenation (ATH) of cyclic imines allowing to obtain a valuable 61% e.e. (R) in the asymmetric reduction of quinaldine 2. The catalytic system exhibited a saturation kinetics with a calculated efficiency of Kcat/KM = 0.688 h-1mM-1.

2-Phenyloxazole-4-carboxamide as a Scaffold for Selective Inhibition of Human Monoamine Oxidase†B.

A series of 2-phenyloxazoles bearing an amide group at position 4 were designed and synthesized for evaluation as potential inhibitors of human recombinant monoamine oxidases (hrMAOs). Results of kinetics experiments demonstrated that all compounds behave as competitive MAO inhibitors, with good selectivity toward the MAO-B isoform. The most potent and selective derivatives are characterized by inhibition constant (Ki ) values in the sub-micromolar range and a good selectivity index (Ki MAO-A /Ki MAO-B >50). Some derivatives were also found to be able to inhibit MAO activity in nerve growth factor (NGF)-differentiated PC12 cells, taken as a model of neuronal cells. In particular, 2-(2-hydroxyphenyl)-N-phenyloxazole-4-carboxamide (compound 4 a) may be a promising new scaffold, exerting the highest selectivity and inhibitory effect toward MAOs in NGF-differentiated PC12 cell lysates, without compromising cell viability. Molecular docking analysis allowed a rationalization of the experimentally observed binding affinity and selectivity.

Self-assembling Releasable Thiocolchicine-Diphenylbutenylaniline Conjugates.

The design and the synthesis of new self-assembling conjugates is reported. The target compounds are characterized by the presence of a self-immolative linker that secures a controlled release induced by lipase cleavage. 4-(1,2-Diphenylbut-1-en-1-yl)aniline is used as a self-assembling inducer and amino-thiocolchicine as prototype of drug. The release of thiocolchicine derivative has been demonstrated in vitro in the presence of porcine pancreatic lipase and Celite-supported lipase. The formation of nanoparticles is confirmed by dynamic light scattering, atomic force microscopy, and fluorescence microscopy. The antiproliferative activity has been proved on two human cancer cell lines.

Chalcones and Chalcone-mimetic Derivatives as Notch Inhibitors in a Model of T-cell Acute Lymphoblastic Leukemia.

Based on hit-likeness and chemical diversity, a number of chalcones and chalcone-mimetic compounds were selected as putative Notch inhibitors. The evaluation of the antiproliferative effect combined with the inhibition of Notch1 expression in KOPTK1 cell line identified compound 18, featuring a tetrahydronaphthalene-based scaffold, as a new promising Notch-blocking agent.