Impact of Composition and Autoclave Sterilization on the Mechanical and Biological Properties of ECM-Mimicking Cryogels.

Cryogels represent a valid strategy as scaffolds for tissue engineering. In order to adequately support adhesion and proliferation of anchorage-dependent cells, different polymers need to be combined within the same scaffold trying to mimic the complex features of a natural extracellular matrix (ECM). For this reason, in this work, gelatin (Gel) and chondroitin sulfate (CS), both functionalized with methacrylic groups to produce CSMA and GelMA derivatives, were selected to prepare cryogel networks. Both homopolymer and heteropolymer cryogels were produced, via radical crosslinking reactions carried out at -12 °C for 2 h. All the scaffolds were characterized for their mechanical, swelling and morphological properties, before and after autoclave sterilization. Moreover, they were evaluated for their biocompatibility and ability to support the adhesion of human gingival fibroblasts and tenocytes. GelMA-based homopolymer networks better withstood the autoclave sterilization process, compared to CSMA cryogels. Indeed, GelMA cryogels showed a decrease in stiffness of approximately 30%, whereas CSMA cryogels of approximately 80%. When GelMA and CSMA were blended in the same network, an intermediate outcome was observed. However, the hybrid scaffolds showed a general worsening of the biological performance. Indeed, despite their ability to withstand autoclave sterilization with limited modification of the mechanical and morphological properties, the hybrid cryogels exhibited poor cell adhesion and high LDH leakage. Therefore, not only do network components need to be properly selected, but also their combination and ability to withstand effective sterilization process should be carefully evaluated for the development of efficient scaffolds designed for tissue engineering purposes.

Eg5 and Diseases: From the Well-Known Role in Cancer to the Less-Known Activity in Noncancerous Pathological Conditions.

Eg5 is a protein encoded by KIF11 gene and is primarily involved in correct mitotic cell division. It is also involved in nonmitotic processes such as polypeptide synthesis, protein transport, and angiogenesis. The scientific literature sheds light on the ubiquitous functions of KIF11 and its involvement in the onset and progression of different pathologies. This review focuses attention on two main points: (1) the correlation between Eg5 and cancer and (2) the involvement of Eg5 in noncancerous conditions. Regarding the first point, several tumors revealed an overexpression of this kinesin, thus pushing to look for new Eg5 inhibitors for clinical practice. In addition, the evaluation of Eg5 expression represents a crucial step, as its overexpression could predict a poor prognosis for cancer patients. Referring to the second point, in specific pathological conditions, the reduced activity of Eg5 can be one of the causes of pathological onset. This is the case of Alzheimer's disease (AD), in which Aß and Tau work as Eg5 inhibitors, or in acquired immune deficiency syndrome (AIDS), in which Tat-mediated Eg5 determines the loss of CD4+ T-lymphocytes. Reduced Eg5 activity, due to mutations of KIF11 gene, is also responsible for pathological conditions such as microcephaly with or without chorioretinopathy, lymphedema, or intellectual disability (MCLRI) and familial exudative vitreous retinopathy (FEVR). In conclusion, this review highlights the double impact that overexpression or loss of function of Eg5 could have in the onset and progression of different pathological situations. This emphasizes, on one hand, a possible role of Eg5 as a potential biomarker and new target in cancer and, on the other hand, the promotion of Eg5 expression/activity as a new therapeutic strategy in different noncancerous diseases.

Benzenesulfonamide decorated dihydropyrimidin(thi)ones: carbonic anhydrase profiling and antiproliferative activity.

In the last decades, carbonic anhydrases (CAs) have become the top investigated innovative pharmacological targets and, in particular, isoforms IX and XII have been widely studied due to the evidence of their overexpression in hypoxic tumors. The frantic race to find new anticancer agents places the quick preparation of large libraries of putative bioactive compounds as the basis of a successful drug discovery and development programme. In this context, multi-component and, in general, one-step reactions are becoming very popular and, among them, Biginelli's reaction gave clean and easy-to-isolate products. Thus, we synthesized a series of Biginelli's products (10-17a-b) and similar derivatives (20-21) bearing the benzenesulfonamide moiety, which is known to inhibit CA enzymes. Through the stopped-flow technique, we were able to assess their ability to inhibit the targeted CAs IX and XII in the nanomolar range with promising selectivity over the physiologically relevant isoforms I and II. Crystallography studies and docking simulations helped us to gain insight into the interaction patterns established in the enzyme-inhibitor complex. From a chemical similarity-based screening of in-house libraries of compounds, a diphenylpyrimidine (23) emerged. The surprisingly potent inhibitory activity of 23 for CAs IX and XII along with its strong antiproliferative effect on two (triple-negative breast cancer MDA-MB-231 and glioblastoma U87MG) cell lines laid the foundation for further investigation, again confirming the key role of CAs in cancer.

Fine-tuning of membrane permeability by reversible photoisomerization of aryl-azo derivatives of thymol embedded in lipid nanoparticles.

Responsiveness of liposomes to external stimuli, such as light, should allow a precise spatial and temporal control of release of therapeutic agents or ion transmembrane transport. Here, some aryl-azo derivatives of thymol are synthesized and embedded into liposomes from 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine to obtain light-sensitive membranes whose photo-responsiveness, release behaviour, and permeability towards Cl- ions are investigated. The hybrid systems are in-depth characterized by dynamic light scattering, atomic force microscopy and Raman spectroscopy. In liposomal bilayer the selected guests undergo reversible photoinduced isomerization upon irradiation with UV and visible light, alternately. Non-irradiated hybrid liposomes retain entrapped 5(6)-carboxyfluorescein (CF), slowing its spontaneous leakage, whereas UV-irradiation promotes CF release, due to guest trans-to-cis isomerization. Photoisomerization also influences membrane permeability towards Cl- ions. Data processing, according to first-order kinetics, demonstrates that Cl- transmembrane transport is enhanced by switching the guest from trans to cis but restored by back-switching the guest from cis to trans upon illumination with blue light. Finally, the passage of Cl- ions across the bilayer can be fine-tuned by irradiation with light of longer λ and different light-exposure times. Fine-tuning the photo-induced structural response of the liposomal membrane upon isomerization is a promising step towards effective photo-dynamic therapy.

Meta-analysis of nano-phytosomes: unleashing the potential of plant-derived compounds for advancing cancer therapy.

Nano-phytosomes are considered as an efficient drug delivery system for phytochemicals. Phytosomes enhance stability and significantly improves phytochemicals bioavailability and therapeutic efficacy. Thorough meta-analysis of 93 articles, phytochemical versus phytosomes size, charge, polydispersity index (PDI) and IC50 values were investigated. Multivariate Analysis of Covariance revealed significant phytochemicals type effects, even when accounting for cancer cell type and phospholipid type as covariates. Least Significant Difference (LSD) post hoc tests described unique attributes among various phytosomes. Flavonoid-based phytosomes exhibited larger particle sizes than others. In contrast, terpenoid-based phytosomes displayed significantly lower charges. Flavonoids demonstrated higher poly dispersity index (PDI) values than alkaloids and polyphenols. Alkaloids exhibited more extensive PDI values, while polyphenols had lower PDI values than terpenoids. Furthermore, flavonoid-containing nanoparticles exhibited higher IC50 values than terpenoids. In conclusion, nano-phytosomes offer promising prospects for revolutionising drug delivery methodologies and advancing the development of innovative therapeutic solutions in the domain of cancer therapy.

Migraine and Its Treatment from the Medicinal Chemistry Perspective.

Migraine is a disease of neurovascular origin that affects the quality of life of more than one billion people and ranks sixth among the most common diseases in the world. Migraine is characterized by a moderate or severe recurrent and throbbing headache, accompanied by nausea, vomiting, and photo-phonophobia. It usually starts in adolescence and is twice as common in women as in men. It is classified as with or without aura and has chronic or acute treatment types according to the frequency of occurrence. In acute treatment, analgesics that relieve pain in the fastest way are preferred, while there are different options in chronic treatment. While non-specific methods were used in the treatment of migraine until the 1950s, triptans, ditans, and CGRP-receptor-dependent therapies (monoclonal antibodies and gepants) started to be used in the clinic more recently. In this Review, we focus on the synthesis, side effects, and pharmacological and pharmacokinetic properties of FDA-approved drugs used in acute and preventive-specific treatment of migraine.

2-Substituted-4,7-dihydro-4-ethylpyrazolo[1,5-a]pyrimidin-7-ones alleviate LPS-induced inflammation by modulating cell metabolism via CD73 upon macrophage polarization.

Macrophage polarization towards the M1 phenotype under bacterial product-related exposure (LPS) requires a rapid change in gene expression patterns and cytokine production along with a metabolic rewiring. Metabolic pathways and redox reactions are such tightly connected, giving rise to an area of research referred to as immunometabolism. A role in this context has been paid to the master redox-sensitive regulator Nuclear factor erythroid 2-related factor 2 (Nrf2) and to the 5'-ectonucleotidase CD73, a marker related to macrophage metabolism rearrangement under pro-inflammatory conditions. In this light, a cell model of LPS-stimulated macrophages has been established and nine 4,7-dihydro-4-ethylpyrazolo[l,5-a]pyrimidin-7-ones with a potential anti-inflammatory effect have been administered. Our data highlight that two selected compounds (namely, 5 and 8) inhibit the LPS-induced Nrf2 nuclear translocation and ameliorate the activity rate of the antioxidant enzyme catalase. Additionally, the pyridine-containing compound (8) promotes the shift from the pro-inflammatory immunophenotype M1 to the pro-resolving M2 one, by downregulating CD80 and iNOS and by enhancing CD163 and TGFß1 expression. Most importantly, CD73 is modulated by these compounds as well as the lactate production. Our data demonstrate that pyrazolo[l,5-a]pyrimidine derivatives are effective as anti-inflammatory compounds. Furthermore, these pyrazolo[l,5-a]pyrimidines exert their action via CD73-related signaling and modulation of cell metabolism of activated macrophages.

Dual Inhibitors of Brain Carbonic Anhydrases and Monoamine Oxidase-B Efficiently Protect against Amyloid-ß-Induced Neuronal Toxicity, Oxidative Stress, and Mitochondrial Dysfunction.

We report here the first dual inhibitors of brain carbonic anhydrases (CAs) and monoamine oxidase-B (MAO-B) for the management of Alzheimer's disease. Classical CA inhibitors (CAIs) such as methazolamide prevent amyloid-ß-peptide (Aß)-induced overproduction of reactive oxygen species (ROS) and mitochondrial dysfunction. MAO-B is also implicated in ROS production, cholinergic system disruption, and amyloid plaque formation. In this work, we combined a reversible MAO-B inhibitor of the coumarin and chromone type with benzenesulfonamide fragments as highly effective CAIs. A hit-to-lead optimization led to a significant set of derivatives showing potent low nanomolar inhibition of the target brain CAs (KIs in the range of 0.1-90.0 nM) and MAO-B (IC50 in the range of 6.7-32.6 nM). Computational studies were conducted to elucidate the structure-activity relationship and predict ADMET properties. The most effective multitarget compounds totally prevented Aß-related toxicity, reverted ROS formation, and restored the mitochondrial functionality in an SH-SY5Y cell model surpassing the efficacy of single-target drugs.

Recent developments of agents targeting Vibrio cholerae: patents and literature data.

INTRODUCTION: Vibrio cholerae bacteria cause an infection characterized by acute diarrheal illness in the intestine. Cholera is sustained by people swallowing contaminated food or water. Even though symptoms can be mild, if untreated disease becomes severe and life-threatening, especially in low-income countries. AREAS COVERED: After a description of the most recent literature on the pathophysiology of this infection, we searched for patents and literature articles following the PRISMA guidelines, filtering the results disclosed from 2020 to present. Moreover, some innovative molecular targets (e.g., carbonic anhydrases) and pathways to counteract this rising problem were also discussed in terms of design, structure-activity relationships and structural analyses. EXPERT OPINION: This review aims to cover and analyze the most recent advances on the new druggable targets and bioactive compounds against this fastidious pathogen, overcoming the use of old antibiotics which currently suffer from high resistance rate.

Synthetic Approaches to Novel Human Carbonic Anhydrase Isoform Inhibitors Based on Pyrrol-2-one Moiety.

New dihydro-pyrrol-2-one compounds, featuring dual sulfonamide groups, were synthesized through a one-pot, three-component approach utilizing trifluoroacetic acid as a catalyst. Computational analysis using density functional theory (DFT) and condensed Fukui function explored the structure-reactivity relationship. Evaluation against human carbonic anhydrase isoforms (hCA I, II, IX, XII) revealed potent inhibition. The widely expressed cytosolic hCA I was inhibited across a range of concentrations (KI 3.9-870.9 nM). hCA II, also cytosolic, exhibited good inhibition as well. Notably, all compounds effectively inhibited tumor-associated hCA IX (KI 1.9-211.2 nM) and hCA XII (low nanomolar). Biological assessments on MCF7 cancer cells highlighted the compounds' ability, in conjunction with doxorubicin, to significantly impact tumor cell viability. These findings underscore the potential therapeutic relevance of the synthesized compounds in cancer treatment.

Immunophenotyping of hemocytes from infected Galleria mellonella larvae as an innovative tool for immune profiling, infection studies and drug screening.

In recent years, there has been a considerable increasing interest in the use of the greater wax moth Galleria mellonella as an animal model. In vivo pharmacological tests, concerning the efficacy and the toxicity of novel compounds are typically performed in mammalian models. However, the use of the latter is costly, laborious and requires ethical approval. In this context, G. mellonella larvae can be considered a valid option due to their greater ease of use and the absence of ethical rules. Furthermore, it has been demonstrated that the immune system of these invertebrates has similarity with the one of mammals, thus guaranteeing the reliability of this in vivo model, mainly in the microbiological field. To better develop the full potential of this model, we present a novel approach to characterize the hemocyte population from G. mellonella larvae and to highlight the immuno modulation upon infection and treatments. Our approach is based on the detection in isolated hemocytes from G. mellonella hemolymph of cell membrane markers typically expressed by human immune cells upon inflammation and infection, for instance CD14, CD44, CD80, CD163 and CD200. This method highlights the analogies between G. mellonella larvae and humans. Furthermore, we provide an innovative tool to perform pre-clinical evaluations of the efficacy of antimicrobial compounds in vivo to further proceed with clinical trials and support drug discovery campaigns.

Modulation of NRF2: Biological Dualism in Cancer, Targets and Possible Therapeutic Applications.

Significance: The nuclear factor erythroid 2-related factor 2 (NRF2)-Kelch-like ECH-associated protein 1 (KEAP1) system is a master regulator of redox homeostasis and cell adaptation to a variety of exogenous and endogenous stressors. Accumulating evidence from the last decade indicates that the impairment of the redox balance leads to oxidative stress (OS), a common alteration occurring in many human acute and chronic inflammatory diseases, such as cancer, diabetes, neurodegeneration, and metabolic disorders, and aging. Recent Advances: Being located at the intersection of crucial signaling pathways, NRF2 can influence several cellular functions, which extend beyond the maintenance of the redox balance and include cellular metabolism, proteostasis, mitochondrial function and inflammation. For this reason, there is a growing interest in the pharmacologic manipulation of NRF2 for therapeutic purposes, which requires the accurate knowledge of the cell context and the specific time frame both of NRF2 activation and inhibition. This appears to be an important prerequisite and reflects the extreme complexity of the NRF2 signaling, characterized by an intrinsic dualism that mediates beneficial or detrimental effects even in the same biological process. Critical Issues: Of crucial importance will be to understand whether the NRF2 activity modulation might be exploited to exert beneficial outcomes in patients suffering from pathological conditions, in which the OS and the deregulation of inflammatory processes play a crucial role. Future Directions: In this review, we discuss the dual involvement of NRF2 in aging, neurodegeneration, metabolic diseases, long-COVID-19, and carcinogenesis and we present an overview of the most recent therapeutic modulators of NRF2, particularly emphasizing on those selected for clinical trials. Antioxid. Redox Signal. 40, 636-662.

Comprehensive metabolite and biological profile of "Sulmona Red Garlic" ecotype's aerial bulbils.

"Sulmona Red Garlic" is a well-known Italian traditional product. Bulbs, used for culinary purposes, have been largely investigated for their medicinal properties whereas aerial bulbils are usually removed as waste material. Here, for the first time, chemical composition and biological properties of the hydroalcoholic extract from aerial bulbils were investigated. Complementary information on metabolite composition were obtained using both NMR based untargeted and HPLC-DAD targeted methodologies. The NMR analysis revealed the presence of sugars, organic acids, amino acids, organosulphur compounds (methiin, alliin, allicin and cycloalliin), and other secondary metabolites. In particular, methiin and alliin were identified for the first time in the NMR spectra of aerial bulbil garlic extracts. Polyphenol content was determined by HPLC-DAD analysis: catechin, chlorogenic acid, and gallic acid turned out to be the most abundant phenolics. Hydroalcoholic extract blocked cell proliferation of colon cancer cell line HCT116 with an IC50 of 352.07 µg/mL, while it was non-toxic to myoblast cell line C2C12. In addition, it caused seedling germination reduction of two edible and herbaceous dicotyledon species, namely Cichorium intybus and C. endivia. Moreover, the same extract reduced the gene expression of TNF-α (tumor necrosis factor), HIF1-α (hypoxia-inducible factor), VEGFA (vascular endothelial growth factor), and transient receptor potential (TRP) M8 (TRPM8) indicating the ability to contrast cancer development through the angiogenic pathway. Final, in silico experiments were also carried out supporting the biological effects of organosulphur compounds, particularly alliin, which may directly interact with TRPM8. The results here reported suggest the potential use of garlic aerial bulbils often considered a waste product as a source in phytotherapeutic remedies.

A Multitarget Approach against Neuroinflammation: Alkyl Substituted Coumarins as Inhibitors of Enzymes Involved in Neurodegeneration.

Neurodegenerative disorders (NDs) include a large range of diseases characterized by neural dysfunction with a multifactorial etiology. The most common NDs are Alzheimer's disease and Parkinson's disease, in which cholinergic and dopaminergic systems are impaired, respectively. Despite different brain regions being affected, oxidative stress and inflammation were found to be common triggers in the pathogenesis and progression of both diseases. By taking advantage of a multi-target approach, in this work we explored alkyl substituted coumarins as neuroprotective agents, capable to reduce oxidative stress and inflammation by inhibiting enzymes involved in neurodegeneration, among which are Carbonic Anhydrases (CAs), Monoamine Oxidases (MAOs), and Cholinesterases (ChEs). The compounds were synthesized and profiled against the three targeted enzymes. The binding mode of the most promising compounds (7 and 9) within MAO-A and -B was analyzed through molecular modeling studies, providing and explanation for the different selectivities observed for the MAO isoforms. In vitro biological studies using LPS-stimulated rat astrocytes showed that some compounds were able to counteract the oxidative stress-induced neuroinflammation and hamper interleukin-6 secretion, confirming the success of this multitarget approach.

Computational medicinal chemistry applications to target Asian-prevalent strain of hepatitis C virus.

Hepatitis C Virus (HCV), affecting millions of people worldwide, is the leading cause of liver disorder, cirrhosis, and hepatocellular carcinoma. HCV is genetically diverse having eight genotypes and several subtypes predominant in different regions of the globe. The HCV NS3/4A protease is a primary therapeutic target for HCV with various FDA-approved antivirals and several clinical developments. However, available protease inhibitors (PIs) have lower potency against HCV genotype 3 (GT3), prevalent in South Asia. In this study, the incumbent computational tools were utilized to understand and explore interactions of the HCV GT3 receptor with the potential inhibitors after the virtual screening of one million compounds retrieved from the ZINC database. The molecular dynamics, pharmacological studies, and experimental studies uncovered the potential PIs as ZINC000224449889, ZINC000224374291, and ZINC000224374456 and the derivative of ZINC000224374456 from the ZINC library. The study revealed that these top-hit compounds exhibited good binding and better pharmacokinetics properties that might be considered the most promising compound against HCV GT3 protease. Viability test, on primary healthy Human Gingival Fibroblasts (HGFs) and cancerous AGS cell line, was also carried out to assess their safety profile after administration. In addition, Surface Plasmon Resonance (SPR) was also performed for the determination of affinity and kinetics of synthesized compounds with target proteins.

Erlotinib-containing benzenesulfonamides as anti-Helicobacter pylori agents through carbonic anhydrase inhibition.

Aim: Development of dual-acting antibacterial agents containing Erlotinib, a recognized EGFR inhibitor used as an anticancer agent, with differently spaced benzenesulfonamide moieties known to bind and inhibit Helicobacter pylori carbonic anhydrase (HpCA) or the antiviral Zidovudine. Methods & materials: Through rational design, ten derivatives were obtained via a straightforward synthesis including a click chemistry reaction. Inhibitory activity against a panel of pathogenic carbonic anhydrases and antibacterial susceptibility of H. pylori ATCC 43504 were assessed. Docking studies on α-carbonic anhydrase enzymes and EGFR were conducted to gain insight into the binding mode of these compounds. Results & conclusion: Some compounds proved to be strong inhibitors of HpCA and showed good anti-H. pylori activity. Computational studies on the targeted enzymes shed light on the interaction hotspots.

Natural products as a source of new anticancer chemotypes.

INTRODUCTION: Exploring the chemical diversity and molecular mechanisms of natural products continues to be an important research area for identifying novel promising therapeutic approaches for fighting cancer. This is a complex disease and poses important challenges, which require not only targeted interventions to improve chemotherapy efficacy and tolerability, but also adjuvant strategies to counteract chemoresistance development and relapses. AREAS COVERED: After a brief description of the recent literature on the anticancer potential of natural compounds, we searched for patents following the PRISMA guidelines, filtering the results published from 2019 onwards. In addition, some relevant publications from the overall scientific literature were also discussed. EXPERT OPINION: This review comprehensively covers and analyzes the most recent advances on the anticancer mechanism of licensed natural compounds and their chemical optimization. Patentability of natural compounds was discussed according to the recent legislation in the U.S.A. and Europe.

Antimalarial Agents Targeting Plasmodium falciparum Carbonic Anhydrase: Towards Artesunate Hybrid Compounds with Dual Mechanism of Action.

Malaria continues to be a major public health challenge worldwide and, as part of the global effort toward malaria eradication, plasmodium carbonic anhydrases (CAs) have recently been proposed as potential targets for malaria treatment. In this study, a series of eight hybrid compounds combining the Artesunate core with a sulfonamide moiety were synthesized and evaluated for their inhibition potency against the widely expressed human (h) CAs I, II and the isoform from P. falciparum (PfCA). All derivatives demonstrated high inhibition potency against PfCA, achieving a KI value in the sub-nanomolar range (0.35 nM). Two Compounds showed a selectivity index of 4.1 and 3.1, respectively, against this protozoan isoform compared to hCA II. Three Derivatives showed no cytotoxic effects on human gingival fibroblasts at 50 µM with a high killing rate against both P. falciparum and P. knowlesi strains with IC50 in the sub-nanomolar range, providing a wide therapeutic window. Our findings suggest that these compounds may serve as promising leads for developing new antimalarial drugs and warrant further investigation, including activity against antimalarial-resistant strains, mode of action studies, and in vivo efficacy assessment in preclinical mouse models of malaria.

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.