Wilma Quaglia

Bis­(pyrazol-1-yl)­acetic acid (HC­(pz)₂COOH) and bis­(3,5-dimethyl-pyrazol-1-yl)­acetic acid (HC­(pz^Me2)₂COOH) were conjugated with amantadine to yield the ligands L^Ad and L^2Ad, respectively. These chelating ligands were used for the synthesis of silver complexes 1–5, whose electronic and molecular structures were investigated by X-ray Photoelectron Spectroscopy (XPS) and Near Edge X-ray Absorption Fine Structure (NEXAFS) spectroscopy. The coordination geometry of the noble metal ion was assessed by combining X-ray Absorption Spectroscopy (XAS) data analysis with Density Functional Theory (DFT) modeling. The structures of the ligand L^Ad and of the complex [Ag­(L^2Ad)­(PPh₃)]­NO₃·0.5CH₃CN (3a) were determined by single-crystal X-ray diffraction analysis. The bis­(pyrazolyl)­acetamide ligand crystallizes in the I2/a space group and is similar to the 3,5-dimethyl analogue. The complex crystallizes in the P-1 space group and is the first reported example of a tetra-coordinated silver­(I) complex incorporating a tridentate bis­(pyrazolyl)­acetate ligand. The cytotoxicity potential of the new phosphane Ag­(I) complexes was tested on various human cancer cell lines, together with that of the two unfunctionalized analogs [Ag­(L^2OiPr)­(PPh₃)]­NO₃ (6) and [Ag­(L^2OiPr)­(PTA)]­NO₃ (7), synthesized and studied for useful comparison. Notably, the novel complexes exhibited pronounced cytotoxic effects against the human-derived pancreatic cancer BxPC-3 cell line, both in 2D monolayers and in 3D spheroid models. Mechanistic studies revealed efficient intracellular accumulation of the complexes, likely favored by increased lipophilicity, and inhibition of thioredoxin reductase (TrxR), ultimately disrupting redox homeostasis and promoting oxidative stress, which are key contributors to their antiproliferative effects.

This paper reports the synthesis, structural characterization, and biological evaluation of a novel series of Cu^I and Cu^II complexes supported by an amantadine-functionalized bis(pyrazol-1-yl)acetate ligand (L^Ad) as potential anticancer agents for the treatment of glioblastoma (GBM). Comprehensive spectroscopic and structural investigations, including SR-XPS, XANES/EXAFS, and DFT modeling, confirmed the successful coordination of L^Ad to copper centers in both oxidation states, affording well-defined molecular architectures with distinct coordination geometries. Among the synthesized compounds, the Cu^I complexes bearing triphenylphosphine co-ligands (compounds 4 and 5) exhibited the strongest cytotoxicity against U87 MG and LN18 GBM cell lines, showing IC₅₀ values lower than those of cisplatin. These complexes induced a pronounced redox imbalance through reactive oxygen species (ROS) overproduction and glutathione (GSH) depletion, leading to G2/M cell cycle arrest and cell death. Flow cytometry and Western blot analyses demonstrated that cell death occurs via caspase-dependent apoptosis in LN18 cells, as evidenced by PARP cleavage, downregulation of Bcl-xL, release of cytochrome c, and mitochondrial translocation of Bax. Altogether, these findings highlight the potential of lipophilic amantadine-functionalized Cu^I complexes as promising anticancer candidates targeting glioma cells through mitochondrial dysfunction and redox-mediated pathways.

Background/Objectives: D4R antagonists have recently been suggested as potential therapeutic alternatives to the standard treatments of glioblastoma (GBM). In this study, new piperidine-based ligands, analogs of the potent and selective D4R compounds 77-LH-28-1 (7) and its 4-benzyl analog 8, were synthesized and studied to investigate the effects produced by variations in the distances between the pharmacophoric features on the D4R affinity and selectivity. Methods: All the new compounds 9–20 were evaluated for their radioligand binding affinity at D2-like receptor subtypes and the results were rationalized by docking studies and molecular dynamics (MD) simulations. The functional profiles of the most interesting derivatives were assessed at D4R Go and Gi protein and β-arrestin by BRET assay and their potential anticancer activity was determined in GBM cell lines. Results: Radioligand binding results highlighted that the derivatives bearing a terminal butyl chain showed structure–activity relationships different from those with a benzyl terminal. From functional studies performed on the best derivatives 12 and 16, the response profiles of both compounds were more robust in antagonist mode, with derivative 16 showing higher antagonist potency than 12 across all three transducers. Interestingly, 12 and 16 dose-dependently decreased the cell viability of GBM cells, inducing cell death and cell cycle arrest, promoting an increase in ROS production, causing mitochondrial dysfunction, and significantly inhibiting colony formation. Conclusions: The promising biological profiles of 12 and 16 make them new lead candidates that warrant further investigation to gain a better understanding of the mechanism behind their antitumor activity and better evaluate their potential for GBM treatment.

The dopamine D4 receptor (D4R) has recently been proposed as an emerging target for treating glioblastoma (GBM). In this article, new piperazine ligands, analogues of the potent and selective D4R lead compounds 9 and 10, were prepared and evaluated for their affinity at D2-like receptor subtypes. The most promising results were obtained by replacing the N4-phenyl terminal of 9 with a naphthyl group. Indeed, α-naphthyl derivative 15 proved to have four times higher affinity for D4R than lead 9, whereas β-naphthyl compound 16 was about tenfold more selective for D4R than 9. These compounds behaved as D4R antagonists in both Gi/Go activation and β-arrestin2 recruitment assays. Interestingly, both decreased cell viability dose-dependently and altered the cell cycle of U87 MG, T98G, and U251 MG human GBM cell lines after 48 h treatment, inducing an increase in ROS levels and time-dependent mitochondrial depolarization.

A new series of muscarinic acetylcholine receptor (mAChR) ligands obtained by inserting different substituents in position 2 of the potent 6,6‐diphenyl‐1,4‐dioxane antagonists 4 and 5 was designed and synthesized to investigate the influence of steric bulk on the mAChR affinity. Specifically, the insertion of a 2‐methyl group, affording compounds 6 and 9, resulted as the most favorable modification in terms of affinity for all muscarinic subtypes. As supported by computational studies performed on the hM₁ receptor, this substituent may contribute to stabilize the ligand within the binding site by favoring the formation of stable interactions between the cationic head of the ligand and the residue D105. The increase of steric bulk, obtained by replacing the methyl group with an ethyl (7 and 10) and especially a phenyl substituent (8 and 11), caused a marked decrease of mAChR affinity, demonstrating the crucial role played by the steric bulk of the 2‐substituent in the mAChR interaction. The most intriguing result was obtained with the tertiary amine 9, which, surprisingly, showed two different pK_i values for all mAChRs, with preferential subpicomolar affinities for the M₁, M₃, and M₄ subtypes. Interestingly, biphasic curves were also observed with both the eutomer (S)‐(–)‐9 and the distomer (R)‐( + )‐9.

The new ligand L^2Ad, obtained by conjugating the bifunctional species bis(3,5-dimethylpyrazol-1-yl)-acetate and the drug amantadine, was used as a chelator for the synthesis of new Cu complexes 1-5. Their structures were investigated by synchrotron radiation-induced X-ray photoelectron spectroscopy (SR-XPS), near-edge X-ray absorption fine structure (NEXAFS) spectroscopy, and by combining X-ray absorption fine structure (XAFS) spectroscopy techniques and DFT modeling. The structure of complex 3 was determined by single-crystal X-ray diffraction analysis. Tested on U87, T98, and U251 glioma cells, Cu(II) complex 3 and Cu(I) complex 5 decreased cell viability with IC₅₀ values significantly lower than cisplatin, affecting cell growth, proliferation, and death. Their effects were prevented by treatment with the Cu chelator tetrathiomolybdate, suggesting the involvement of copper in their cytotoxic activity. Both complexes were able to increase ROS production, leading to DNA damage and death. Interestingly, nontoxic doses of 3 or 5 enhanced the chemosensitivity to Temozolomide.

In the framework of the design, synthesis and testing of a library of copper complexes and nanostructured assemblies potentially endowed with antitumor and antiviral activity and useful for several applications, from drugs and related delivery systems to the development of biocidal nanomaterials, we present the detailed spectroscopic investigation of the molecular and electronic structure of copper‐based coordination compounds and of a new conjugated system obtained by grafting Cu(I) complexes to gold nanorods. More in detail, the electronic and molecular structures of two Cu complexes and one AuNRs/Cu‐complex adduct were investigated by X‐ray photoelectron spectroscopy (XPS), synchrotron‐induced XPS (SR‐XPS) and near edge X‐ray absorption spectroscopy (NEXAFS) in solid state, and the local structure around copper ion was assessed by X‐ray absorption spectroscopy (XAS) both in solid state and water solution for the AuNRs/Cu‐complex nanoparticles. The proposed multi‐technique approach allowed to properly define the coordination geometry around the copper ion, as well as to ascertain the molecular structures of the coordination compounds, their stability and modifications upon interaction with gold nanoparticles, by comparing solid state and liquid phase data.

Orexin‐A and orexin‐B, also named hypocretin‐1 and hypocretin‐2, are two hypothalamic neuropeptides highly conserved across mammalian species. Their effects are mediated by two distinct G protein‐coupled receptors, namely orexin receptor type 1 (OX1‐R) and type 2 (OX2‐R), which share 64% amino acid identity. Given the wide expression of OX‐Rs in different central nervous system and peripheral areas and the several pathophysiological functions in which they are involved, including sleep‐wake cycle regulation (mainly mediated by OX2‐R), emotion, panic‐like behaviors, anxiety/stress, food intake, and energy homeostasis (mainly mediated by OX1‐R), both subtypes represent targets of interest for many structure–activity relationship (SAR) campaigns carried out by pharmaceutical companies and academies. However, before 2017 the research was predominantly directed towards dual‐orexin ligands, and limited chemotypes were investigated. Analytical characterizations, including resolved structures for both OX1‐R and OX2‐R in complex with agonists and antagonists, have improved the understanding of the molecular basis of receptor recognition and are assets for medicinal chemists in the design of subtype‐selective ligands. This review is focused on the medicinal chemistry aspects of small molecules acting as dual or subtype selective OX1‐R/OX2‐R agonists and antagonists belonging to different chemotypes and developed in the last years, including radiolabeled OX‐R ligands for molecular imaging. Moreover, the pharmacological effects of the most studied ligands in different neuropsychiatric diseases, such as sleep, mood, substance use, and eating disorders, as well as pain, have been discussed. Poly‐pharmacology applications and multitarget ligands have also been considered.