Francesca Ghirga

Gli1/DNA interaction is a druggable target for Hedgehog‐dependent tumors

Hedgehog signaling is essential for tissue development and stemness, and its deregulation has been observed in many tumors. Aberrant activation of Hedgehog signaling is the result of genetic mutations of pathway components or other Smo‐dependent or independent mechanisms, all triggering the downstream effector Gli1. For this reason, understanding the poorly elucidated mechanism of Gli1‐mediated transcription allows to identify novel molecules blocking the pathway at a downstream level, representing a critical goal in tumor biology. Here, we clarify the structural requirements of the pathway effector Gli1 for binding to DNA and identify Glabrescione B as the first small molecule binding to Gli1 zinc finger and impairing Gli1 activity by interfering with its interaction with DNA. Remarkably, as a consequence of its robust inhibitory effect on Gli1 activity, Glabrescione B inhibited the growth of Hedgehog‐dependent tumor cells in vitro and in vivo as well as the self‐renewal ability and clonogenicity of tumor‐derived stem cells. The identification of the structural requirements of Gli1/DNA interaction highlights their relevance for pharmacologic interference of Gli signaling.

Inhibition of Hedgehog-dependent tumors and cancer stem cells by a newly identified naturally occurring chemotype

Hedgehog (Hh) inhibitors have emerged as valid tools in the treatment of a wide range of cancers. Indeed, aberrant activation of the Hh pathway occurring either by ligand-dependent or -independent mechanisms is a key driver in tumorigenesis. The smoothened (Smo) receptor is one of the main upstream transducers of the Hh signaling and is a validated target for the development of anticancer compounds, as underlined by the FDA-approved Smo antagonist Vismodegib (GDC-0449/Erivedge) for the treatment of basal cell carcinoma. However, Smo mutations that confer constitutive activity and drug resistance have emerged during treatment with Vismodegib. For this reason, the development of new effective Hh inhibitors represents a major challenge for cancer therapy. Natural products have always represented a unique source of lead structures in drug discovery, and in recent years have been used to modulate the Hh pathway at multiple levels. Here, starting from an in house library of natural compounds and their derivatives, we discovered novel chemotypes of Hh inhibitors by mean of virtual screening against the crystallographic structure of Smo. Hh functional based assay identified the chalcone derivative 12 as the most effective Hh inhibitor within the test set. The chalcone 12 binds the Smo receptor and promotes the displacement of Bodipy-Cyclopamine in both Smo WT and drug-resistant Smo mutant. Our molecule stands as a promising Smo antagonist able to specifically impair the growth of Hh-dependent tumor cells in vitro and in vivo and medulloblastoma stem-like cells and potentially overcome the associated drug resistance.

The Pictet-Spengler Reaction Still on Stage.

Today, in spite of being older than a century (born in 1911), the Pictet-Spengler two component reaction (PS-2CR) is still one of the most popular reactions, not only for the synthesis of tetrahydroisoquinolines (THIQs), tetrahydro-β-carbolines (THBCs), or more complex structures containing these two privileged moieties, but also for the construction of novel scaffolds, available for structure-activity relationship (SAR) studies and/or for combinatorial libraries targeted at drug discovery. The prominence of the P-S cyclization is brought about by the inheritance from analogous enzyme-catalyzed reactions of the biogenetic pathways of natural products, mainly indole alkaloids, with a broad range of biological activities. This knowledge has been the starting point for the biomimetic synthesis or the bio-engineering production of pharmacologically important drugs. The long-lasting life of the P-S reaction depends on the discovery of its multiple facets, the modifications of its parameters and components, as well as the continuous renovation of solutions for the challenging stereochemical outcome of the transformation. This paper deals with an updated visit to the P-S reaction aiming to find the threads of the story without forgetting the numerous facets of the prism. It is organized as a theater piece, with a prologue and the main scene (namely, Act 1) where the readers can follow the parade of the two above-mentioned very recurring motifs (namely, THIQ and THBC) by moving from one step to another (a cyclization, an intramolecular attack, a stereoselective passage) to find the way out of the labyrinth of the P-S reaction.

Antioxidant Properties of Aminoethylcysteine Ketimine Decarboxylated Dimer: A Review

Aminoethylcysteine ketimine decarboxylated dimer is a natural sulfur-containing compound detected in human plasma and urine, in mammalian brain and in many common edible vegetables. Over the past decade many studies have been undertaken to identify its metabolic role. Attention has been focused on its antioxidant properties and on its reactivity against oxygen and nitrogen reactive species. These properties have been studied in different model systems starting from plasma lipoproteins to specific cellular lines. All these studies report that aminoethylcysteine ketimine decarboxylated dimer is able to interact both with reactive oxygen and nitrogen species (hydrogen peroxide, superoxide anion, hydroxyl radical, peroxynitrite and its derivatives). Its antioxidant activity is similar to that of Vitamin E while higher than other hydrophilic antioxidants, such as trolox and N-acetylcysteine.

Occurrence of enantioselectivity in nature: the case of (S)-norcoclaurine

This review article is aimed at providing a monographic overview on (S)-norcoclaurine (NC) alkaloid from three diverse points of view, collected all together for the first time: 1) the synthetic one, where the compound is seen as a target chiral molecule to be obtained in the highest optical purity and as a starting point for the development of biocatalytic asymmetric syntheses of tetrahydroisoquinoline alkaloids; 2) the chromatographic one, which addresses the HPLC separation of the two NC enantiomers; and 3) the biochemical one, for which a thorough understanding of the topology and mechanism of action of norcoclaurine synthase (NCS) enzyme is still a matter of debate. Special emphasis on the most recent studies in the field is given by discussing the results published by the main research groups who are working on NC and NCS.

Identification of a novel chalcone derivative that inhibits Notch signaling in T-cell acute lymphoblastic leukemia

Notch signaling is considered a rational target in the therapy of several cancers, particularly those harbouring Notch gain of function mutations, including T-cell acute lymphoblastic leukemia (T-ALL). Although currently available Notch-blocking agents are showing anti-tumor activity in preclinical studies, they are not effective in all the patients and often cause severe side-effects, limiting their widespread therapeutic use. Here, by functional and biological analysis of the most representative molecules of an in house library of natural products, we have designed and synthetized the chalcone-derivative 8 possessing Notch inhibitory activity at low micro molar concentration in T-ALL cell lines. Structure-activity relationships were afforded for the chalcone scaffold. Short term treatments with compound 8 resulted in a dose-dependent decrease of Notch signaling activity, halted cell cycle progression and induced apoptosis, thus affecting leukemia cell growth. Taken together, our data indicate that 8 is a novel Notch inhibitor, candidate for further investigation and development as an additional therapeutic option against Notch-dependent cancers.

Synthesis, biological evaluation and molecular modeling studies on novel quinonoid inhibitors of CDC25 phosphatases

Abstract The cell division cycle 25 phosphatases (CDC25A, B, and C; E.C. 3.1.3.48) are key regulator of the cell cycle in human cells. Their aberrant expression has been associated with the insurgence and development of various types of cancer, and with a poor clinical prognosis. Therefore, CDC25 phosphatases are a valuable target for the development of small molecule inhibitors of therapeutic relevance. Here, we used an integrated strategy mixing organic chemistry with biological investigation and molecular modeling to study novel quinonoid derivatives as CDC25 inhibitors. The most promising molecules proved to inhibit CDC25 isoforms at single digit micromolar concentration, becoming valuable tools in chemical biology investigations and profitable leads for further optimization.

Synthesis of a Double-Spanned Resorc[4]arene via Ring-Closing Metathesis and Calculation of Aggregation Propensity

Ring-closing metathesis (RCM) catalyzed by a second-generation Grubbs catalyst has been used to synthesize resorc[4]arenes 2b-5b starting from undecenyl resorc[4]arene 1b fixed in the cone conformation. X-ray diffraction analysis of the major metathesis product, 3b (50% yield), revealed a cavity-shaped architecture resembling a basket, endowed with a large intramolecular space (∼10 Å) and a strong propensity to self-assemble as a supramolecular trio of heterochiral dimers. This prompted us to investigate the aggregation propensity of basket 3b in THF/water solution by UV-visible spectroscopy. The cavitation Gibbs free-energy change (ΔΔGcav = 4.78 kcal mol(-1)) associated with the self-assembly of macrocycle 3b was calculated as a measure of the solvophobic interactions involved in the process.

Chemical, computational and functional insights into the chemical stability of the Hedgehog pathway inhibitor GANT61

Abstract This work aims at elucidating the mechanism and kinetics of hydrolysis of GANT61, the first and most-widely used inhibitor of the Hedgehog (Hh) signalling pathway that targets Glioma-associated oncogene homologue (Gli) proteins, and at confirming the chemical nature of its bioactive form. GANT61 is poorly stable under physiological conditions and rapidly hydrolyses into an aldehyde species (GANT61-A), which is devoid of the biological activity against Hh signalling, and a diamine derivative (GANT61-D), which has shown inhibition of Gli-mediated transcription. Here, we combined chemical synthesis, NMR spectroscopy, analytical studies, molecular modelling and functional cell assays to characterise the GANT61 hydrolysis pathway. Our results show that GANT61-D is the bioactive form of GANT61 in NIH3T3 Shh-Light II cells and SuFu−/− mouse embryonic fibroblasts, and clarify the structural requirements for GANT61-D binding to Gli1. This study paves the way to the design of GANT61 derivatives with improved potency and chemical stability. Graphical Abstract

Synergistic effects of trace amounts of water in the enantiodiscrimination processes by lipodex E: a spectroscopic and computational investigation.

Nuclear magnetic resonance (NMR) investigations on mixtures containing octakis(3-O-butanoyl-2,6-di-O-pentyl)-γ-cyclodextrin (Lipodex E) and each enantiomer of methyl-2-chloropropionate (MCP) ascertained the role of trace amounts of water in the enantiodiscrimination processes. Water is deeply included into the cyclodextrin and favors the formation of the inclusion complex with (S)-MCP, whereas (R)-MCP is only slightly affected, thus causing a significant increase of NMR differentiation. Molecular dynamics simulations were performed to shed light on the possible behavior of Lipodex E in different conditions (i.e., solvent, inclusion complexes), providing energetic and atomistic details that are in agreement with NMR observations.