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The opportunistic human fungal pathogen Candida albicans produces and releases into the surrounding medium extracellular vesicles (EVs), which are involved in some processes as communication between fungal cells and host–pathogen interactions during infection. Here, we have conducted the isolation of EVs produced by a clinical isolate of C. albicans during biofilm formation and proved their effect towards the ability of the Gram-negative bacterial pathogen Klebsiella pneumoniae to adhere to HaCaT cells and form a biofilm in vitro. The results represent the first evidence of an antagonistic action of fungal EVs against bacteria.

Based on compelling preclinical evidence concerning the progress of our novel ruthenium-based metallotherapeutics, we are focusing research efforts on challenging indications for the treatment of invasive neoplasms such as the triple-negative breast cancer (TNBC). This malignancy mainly afflicts younger women, who are black, or who have a BRCA1 mutation. Because of faster growing and spreading, TNBC differs from other invasive breast cancers having fewer treatment options and worse prognosis, where existing therapies are mostly ineffective, resulting in a large unmet biomedical need. In this context, we benefited from an experimental model of TNBC both in vitro and in vivo to explore the effects of a biocompatible cationic liposomal nanoformulation, named HoThyRu/DOTAP, able to effectively deliver the antiproliferative ruthenium(III) complex AziRu, thus resulting in a prospective candidate drug. As part of the multitargeting mechanisms featuring metal-based therapeutics other than platinum-containing agents, we herein validate the potential of HoThyRu/DOTAP liposomes to act as a multimodal anticancer agent through inhibition of TNBC cell growth and proliferation, as well as migration and invasion. The here-obtained preclinical findings suggest a potential targeting of the complex pathways network controlling invasive and migratory cancer phenotypes. Overall, in the field of alternative chemotherapy to platinum-based drugs, these outcomes suggest prospective brand-new settings for the nanostructured AziRu complex to get promising goals for the treatment of metastatic TNBC.

Ruthenium(III) complexes are very promising candidates as metal-based anticancer drugs, and several studies have supported the likely role of human serum proteins in the transport and selective delivery of Ru(III)-based compounds to tumor cells. Herein, the anticancer nanosystem composed of an amphiphilic nucleolipid incorporating a Ru(III) complex, which we named DoHuRu, embedded into the biocompatible cationic lipid DOTAP, was investigated as to its interaction with two human serum proteins thought to be involved in the mechanism of action of Ru(III)-based anticancer drugs, i.e., human serum albumin (HSA) and human transferrin (hTf). This nanosystem was studied in comparison with the simple Ru(III) complex named AziRu, a low molecular weight metal complex previously designed as an analogue of NAMI-A, decorated with the same ruthenium ligands as DoHuRu but devoid of the nucleolipid scaffold and not inserted in liposomal formulations. For this study, different spectroscopic techniques, i.e., Fluorescence Spectroscopy and Circular Dichroism (CD), were exploited, showing that DoHuRu/DOTAP liposomes can interact with both serum proteins without affecting their secondary structures.

Heparin plays multiple biological roles depending on the availability of active sites strongly influenced by the conformation and the structure of polysaccharide chains. Combining different components at the molecular scale offers an extraordinary chance to easily tune the structural organization of heparin required for exploring new potential applications. In fact, the combination of different material types leads to challenges that cannot be achieved by each single component. In this study, hybrid heparin/silica nanoparticles were synthesized, and the role of silica as a templating agent for heparin supramolecular organization was investigated. The effect of synthesis parameters on particles compositions was deeply investigated by Fourier Transform Infrared Spectroscopy (FTIR) and Thermogravimetric Analysis (TGA). Transmission Electron Microscopy (TEM) reveals a different supramolecular organization of both components, leading to amazing organic-inorganic nanoparticles with different behavior in drug encapsulation and release. Furthermore, favorable biocompatibility for healthy human dermal fibroblasts (HDF) and tumor HS578T cells has been assessed, and a different biological behavior was observed, ascribed to different surface charge and morphology of synthesized nanoparticles.

Lipid structural diversity strongly affects biomembrane chemico-physical and structural properties in addition to membrane-associated events. At high concentrations, cholesterol increases membrane order and rigidity, while polyunsaturated lipids are reported to increase disorder and flexibility. How these different tendencies balance in composite bilayers is still controversial. In this study, electron paramagnetic resonance spectroscopy, small angle neutron scattering, and neutron reflectivity were used to investigate the structural properties of cholesterol-containing lipid bilayers in the fluid state with increasing amounts of polyunsaturated omega-3 lipids. Either the hybrid 1-stearoyl-2-docosahexaenoyl-sn-glycero-3-phosphocholine or the symmetric 1,2-docosahexaenoyl-sn-glycero-3-phosphocholine were added to the mixture of the naturally abundant 1-palmitoyl-2-oleyl-sn-glycero-3-phosphocholine and cholesterol. Our results indicate that the hybrid and the symmetric omega-3 phospholipids affect the microscopic organization of lipid bilayers differently. Cholesterol does not segregate from polyunsaturated phospholipids and, through interactions with them, is able to suppress the formation of non-lamellar structures induced by the symmetric polyunsaturated lipid. However, this order/disorder balance leads to a bilayer whose structural organization cannot be ascribed to either a liquid ordered or to a canonical liquid disordered phase, in that it displays a very loose packing of the intermediate segments of lipid chains.

The integration of nuclear imaging analysis with nanomedicine has tremendously grown and represents a valid and powerful tool for the development and clinical translation of drug delivery systems. Among the various types of nanostructures used as drug carriers, nanovesicles represent intriguing platforms due to their capability to entrap both lipophilic and hydrophilic agents, and their well-known biocompatibility and biodegradability. In this respect, here we present the development of a labelling procedure of POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine)-based liposomes incorporating an ad hoc designed lipophilic NOTA (1,4,7-triazacyclononane-1,4,7-triacetic acid) analogue, derivatized with an oleic acid residue, able to bind the positron emitter gallium-68(III). Based on POPC features, the optimal conditions for liposome labelling were studied with the aim of optimizing the Ga(III) incorporation and obtaining a significant radiochemical yield. The data presented in this work demonstrate the feasibility of the labelling procedure on POPC liposomes co-formulated with the ad hoc designed NOTA analogue. We thus provided a critical insight into the practical aspects of the development of vesicles for theranostic approaches, which in principle can be extended to other nanosystems exploiting a variety of bioconjugation protocols.

In this work CeO2 nanoparticles (CeO2-NPs) were synthesized through the thermal decomposition of Ce(NO3)3·6H2O, using as capping agents either octylamine or oleylamine, to evaluate the effect of alkyl chain length, an issue at 150 °C, in the case of octylamine and at 150 and 250 °C, in the case of oleylamine, to evaluate the effect of the temperature on NPs properties. All the nanoparticles were extensively characterized by a multidisciplinary approach, such as wide-angle X-ray diffraction, transmission electron microscopy, dynamic light scattering, UV-Vis, fluorescence, Raman and FTIR spectroscopies. The analysis of the experimental data shows that the capping agent nature and the synthesis temperature affect nanoparticle properties including size, morphology, aggregation and Ce3+/Ce4+ ratio. Such issues have not been discussed yet, at the best of our knowledge, in the literature. Notably, CeO2-NPs synthesized in the presence of oleylamine at 250 °C showed no tendency to aggregation and we made them water-soluble through a further coating with sodium oleate. The obtained nanoparticles show a less tendency to clustering forming stable aggregates (ranging between 14 and 22 nm) of few NPs. These were tested for biocompatibility and ROS inhibiting activity, demonstrating a remarkable antioxidant activity, against oxidative stress.

Water-in-oil (W/O) nanoemulsions stabilized by phospholipids (PLs) are increasingly exploited in a wide spectrum of applications, from pharmaceuticals to food and cosmetic formulations. In this work, we report the design and optimization of an innovative emulsion based on a mixture of phosphoethanolamine (PE) and phosphoglycerol (PG) PLs, inspired by the composition of the inner leaflet of a bacterial outer membrane. Using the natural oil squalene as the continuous organic phase, no additional emulsion stabilizer is needed. On the other hand, a small amount of Span 80 is required when dodecane is used. The obtained nanoemulsions are stable for at least two hours, thus allowing the droplet size and distribution to be characterized by Dynamic Light Scattering (DLS) and the lipid layer structure and dynamics to be analyzed by Electron Paramagnetic Resonance (EPR) spectroscopy. The results indicate that squalene shallowly intercalates among the lipid tail termini, being unable to deeply penetrate the adsorbed lipid monolayer. The altered lipid dynamics are proposed to be the reason for the enhanced emulsion stability, this paving the way to future implementations and possible applications.

In this review we have showcased the preclinical development of original amphiphilic nanomaterials designed for ruthenium-based anticancer treatments, to be placed within the current metallodrugs approach leading over the past decade to advanced multitarget agents endowed with limited toxicity and resistance. This strategy could allow for new options for breast cancer (BC) interventions, including the triple-negative subtype (TNBC) with poor therapeutic alternatives. BC is currently the second most widespread cancer and the primary cause of cancer death in women. Hence, the availability of novel chemotherapeutic weapons is a basic requirement to fight BC subtypes. Anticancer drugs based on ruthenium are among the most explored and advanced next-generation metallotherapeutics, with NAMI-A and KP1019 as two iconic ruthenium complexes having undergone clinical trials. In addition, many nanomaterial Ru complexes have been recently conceived and developed into anticancer drugs demonstrating attractive properties. In this field, we focused on the evaluation of a Ru(III) complex—named AziRu—incorporated into a suite of both zwitterionic and cationic nucleolipid nanosystems, which proved to be very effective for the in vivo targeting of breast cancer cells (BBC). Mechanisms of action have been widely explored in the context of preclinical evaluations in vitro, highlighting a multitarget action on cell death pathways which are typically deregulated in neoplasms onset and progression. Moreover, being AziRu inspired by the well-known NAMI-A complex, information on non-nanostructured Ru-based anticancer agents have been included in a precise manner.

NU172—a 26-mer oligonucleotide able to bind exosite I of human thrombin and inhibit its activity—was the first aptamer to reach Phase II clinical studies as an anticoagulant in heart disease treatments. With the aim of favoring its functional duplex-quadruplex conformation and thus improving its enzymatic stability, as well as its thrombin inhibitory activity, herein a focused set of cyclic NU172 analogues—obtained by connecting its 5′- and 3′-extremities with flexible linkers—was synthesized. Two different chemical approaches were exploited in the cyclization procedure, one based on the oxime ligation method and the other on Cu(I)-assisted azide-alkyne cycloaddition (CuAAC), affording NU172 analogues including circularizing linkers with different length and chemical nature. The resulting cyclic NU172 derivatives were characterized using several biophysical techniques (ultraviolet (UV) and circular dichroism (CD) spectroscopies, gel electrophoresis) and then investigated for their serum resistance and anticoagulant activity in vitro. All the cyclic NU172 analogues showed higher thermal stability and nuclease resistance compared to unmodified NU172. These favorable properties were, however, associated with reduced—even though still significant—anticoagulant activity, suggesting that the conformational constraints introduced upon cyclization were somehow detrimental for protein recognition. These results provide useful information for the design of improved analogues of NU172 and related duplex-quadruplex structures.

The great advances in the studies on metal complexes for the treatment of different cancer forms, starting from the pioneering works on platinum derivatives, have fostered an increasingly growing interest in their properties and biomedical applications. Among the various metal-containing drugs investigated thus far, ruthenium(III) complexes have emerged for their selective cytotoxic activity in vitro and promising anticancer properties in vivo, also leading to a few candidates in advanced clinical trials. Aiming at addressing the solubility, stability and cellular uptake issues of low molecular weight Ru(III)-based compounds, some research groups have proposed the development of suitable drug delivery systems (e.g., taking advantage of nanoparticles, liposomes, etc.) able to enhance their activity compared to the naked drugs. This review highlights the unique role of Ru(III) complexes in the current panorama of anticancer agents, with particular emphasis on Ru-containing nanoformulations based on the incorporation of the Ru(III) complexes into suitable nanocarriers in order to enhance their bioavailability and pharmacokinetic properties. Preclinical evaluation of these nanoaggregates is discussed with a special focus on the investigation of their mechanism of action at a molecular level, highlighting their pharmacological potential in tumour disease models and value for biomedical applications.