Pharmaceuticals, Volume 14, Issue 12 (December 2021) – 134 articles

Garcinia indica (commonly known as kokum), belonging to the Clusiaceae family (mangosteen family), is a tropical evergreen tree distributed in certain regions of India. It has been used in culinary and industrial applications for a variety of purposes, including acidulant in curries, pickles, health drinks, wine, and butter. In particular, G. indica has been used in traditional medicine to treat inflammation, dermatitis, and diarrhea, and to promote digestion. According to several studies, various phytochemicals such as garcinol, hydroxycitric acid (HCA), cyanidin-3-sambubioside, and cyanidin-3-glucoside were isolated from G. indica, and their pharmacological activities were published. This review highlights recent updates on the various pharmacological activities of G. indica. These studies reported that G. indica has antioxidant, anti-obesity, anti-arthritic, anti-inflammatory, antibacterial, hepatoprotective, cardioprotective, antidepressant and anxiolytic effects both in vitro and in vivo. These findings, together with previously published reports of pharmacological activity of various components isolated from G. indica, suggest its potential as a promising therapeutic agent to prevent various diseases. Full article

Osteoarthritis (OA) is a common degenerative joint disorder that affects joint function, mobility, and pain. The release of proinflammatory cytokines stimulates matrix metalloproteinases (MMPs) and aggrecanase production which further induces articular cartilage degradation. Hypertrophy-like changes in chondrocytes are considered to be an important feature of OA pathogenesis. A Glycyrrhiza new variety, Wongam (WG), was developed by the Korea Rural Development Administration to enhance the cultivation and quality of Glycyrrhizae Radix et Rhizoma (licorice). This study examined the regulatory effect of WG against hypertrophy-like changes such as RUNX2, Collagen X, VEGFA, MMP-13 induction, and Collagen II reduction induced by IL-1β in SW1353 human chondrocytes. Additionally, in silico methods were performed to identify active compounds in licorice to target chondrocyte hypertrophy-related proteins. WG showed inhibitory effects against IL-1β-induced chondrocyte hypertrophy by regulating both HDAC4 activation via the PTH1R/PKA/PP2A pathway and the SOX9/β-catenin signaling pathway. In silico analysis demonstrated that 21 active compounds from licorice have binding potential with 11 targets related to chondrocyte hypertrophy. Further molecular docking analysis and in vivo studies elicited four compounds. Based on HPLC, isoliquiritigenin and its precursors were identified and quantified. Taken together, WG is a potential therapeutic agent for chondrocyte hypertrophy-like changes in OA. Full article

Transient receptor potential ankyrin 1 (TRPA1), a nonselective cation channel, contributes to several (patho)physiological processes. Smell loss is an early sign in several neurodegenerative disorders, such as multiple sclerosis, Parkinson’s and Alzheimer’s diseases; therefore, we focused on its role in olfaction and social behaviour with the aim to reveal its potential therapeutic use. The presence of Trpa1 mRNA was studied along the olfactory tract of mice by combined RNAscope in situ hybridisation and immunohistochemistry. The aversive effects of fox and cat odour were examined in parallel with stress hormone levels. In vitro calcium imaging was applied to test if these substances can directly activate TRPA1 receptors. The role of TRPA1 in social behaviour was investigated by comparing Trpa1 wild-type and knockout mice (KO). Trpa1 mRNA was detected in the olfactory bulb and piriform cortex, while its expression was weak in the olfactory epithelium. Fox, but not cat odour directly activated TRPA1 channels in TRPA1-overexpressing Chinese Hamster Ovary cell lines. Accordingly, KO animals showed less aversion against fox, but not cat odour. The social interest of KO mice was reduced during social habituation–dishabituation and social interaction, but not during resident–intruder tests. TRPA1 may contribute to odour processing at several points of the olfactory tract and may play an important role in shaping the social behaviour of mice. Thus, TRPA1 may influence the development of certain social disorders, serving as a potential drug target in the future. Full article

Gold nanoparticles have gained interest in biomedical sciences in the areas of nano-diagnostics, bio-labeling, drug delivery, and bacterial infection. In this study, we examined, for the first time, the antibacterial and antibiofilm properties of plasmonic gold nanoprisms against human pathogenic bacteria using MIC and crystal violet. In addition, the expression level of GroEL/GroES heat shock proteins was also investigated by western blot. Gold nanoparticles were characterized by TEM and EDX, which showed equilateral triangular prisms with an average edge length of 150 nm. Antibacterial activity testing showed a great effect of AuNPs against pathogenic bacteria with MICs values ranging from 50 μg/mL to 100 μg/mL. Nanoparticles demonstrated strong biofilm inhibition action with a percentage of inhibition ranging from 40.44 to 82.43%. Western blot analysis revealed that GroEL was an AuNPs-inducible protein with an increase of up to 66.04%, but GroES was down-regulated with a reduction of up to 46.81%. Accordingly, plasmonic gold nanoprisms, could be a good candidate for antibiotics substitution in order to treat bacterial infections. Full article

Aegopodium podagraria L. (goutweed), a member of the Apiaceae family, is a common perennial plant found all around the world that has been used in folk medicine since antiquity. Goutweed leaves contain polyacetylenes, essential oils, mono- and sesquiterpenes, vitamins, macro- and microelements, and phenolic compounds. In spite of its many health-promoting properties, including antioxidant effects, the plant has not been thoroughly studied. The aim of this study was to investigate the antioxidant properties of different goutweed leaf extracts and their effects on the THP-1 cell line, and also to describe the chemical characteristics of goutweed. Falcarinol and falcarindiol and essential oil were determined by gas chromatography coupled with mass spectrometry. Spectrophotometry was used to measure the total content of polyphenols and antioxidant activity–by DPPH and FRAP methods. Oxidative stress in THP-1 cells was induced via sodium fluoride. Then, goutweed leaf extracts were added to evaluate their influence on antioxidant potential (ABTS) and the activity of antioxidant enzymes. Confocal microscopy was used to visualise the production of cytoplasmic and mitochondrial reactive oxygen species (ROS) and for in vitro imaging of apoptosis. The ethanol extracts have a high total content of polyphenols, polyacetylenes, and essential oil, as well as high antioxidant potential. The main volatiles represented diverse chemical groups, which are both oxygenated derivatives of sesquiterpenes and monoterpenes. We also demonstrated positive effects of the high antioxidant potential and increased activity of antioxidant enzymes on cell cultures under severe fluoride-induced oxidative stress. Extraction at 80 ℃ and the use of ethanol as a solvent increased the antioxidant capacity of the extract. The leaves of Aegopodium podagraria may serve as a valuable source of antioxidants in the daily diet and assist in the prevention and treatment of oxidative stress-mediated conditions, e.g., inflammatory conditions, cardiovascular diseases, neurodegenerative diseases, and even obesity. Full article

The treatment of CNS disorders suffers from the inability to deliver large therapeutic agents to the brain parenchyma due to protection from the blood-brain barrier (BBB). Herein, we investigated high-frequency pulsed electric field (HF-PEF) therapy of various pulse widths and interphase delays for BBB disruption while selectively minimizing cell ablation. Eighteen male Fisher rats underwent craniectomy procedures and two blunt-tipped electrodes were advanced into the brain for pulsing. BBB disruption was verified with contrast T1W MRI and pathologically with Evans blue dye. High-frequency irreversible electroporation cell death of healthy rodent astrocytes was investigated in vitro using a collagen hydrogel tissue mimic. Numerical analysis was conducted to determine the electric fields in which BBB disruption and cell ablation occur. Differences between the BBB disruption and ablation thresholds for each waveform are as follows: 2-2-2 $\mathsf{\mu }$s (1028 V/cm), 5-2-5 $\mathsf{\mu }$s (721 V/cm), 10-1-10 $\mathsf{\mu }$s (547 V/cm), 2-5-2 $\mathsf{\mu }$s (1043 V/cm), and 5-5-5 $\mathsf{\mu }$s (751 V/cm). These data suggest that HF-PEFs can be fine-tuned to modulate the extent of cell death while maximizing peri-ablative BBB disruption. Furthermore, numerical modeling elucidated the diffuse field gradients of a single-needle grounding pad configuration to favor large-volume BBB disruption, while the monopolar probe configuration is more amenable to ablation and reversible electroporation effects. Full article

Fission yeast contains three essential β(1,3)-D-glucan synthases (GSs), Bgs1, Bgs3, and Bgs4, with non-overlapping roles in cell integrity and morphogenesis. Only the bgs4⁺ mutants pbr1-8 and pbr1-6 exhibit resistance to GS inhibitors, even in the presence of the wild-type (WT) sequences of bgs1⁺ and bgs3⁺. Thus, Bgs1 and Bgs3 functions seem to be unaffected by those GS inhibitors. To learn more about echinocandins’ mechanism of action and resistance, cytokinesis progression and cell death were examined by time-lapse fluorescence microscopy in WT and pbr1-8 cells at the start of treatment with sublethal and lethal concentrations of anidulafungin, caspofungin, and micafungin. In WT, sublethal concentrations of the three drugs caused abundant cell death that was either suppressed (anidulafungin and micafungin) or greatly reduced (caspofungin) in pbr1-8 cells. Interestingly, the lethal concentrations induced differential phenotypes depending on the echinocandin used. Anidulafungin and caspofungin were mostly fungistatic, heavily impairing cytokinesis progression in both WT and pbr1-8. As with sublethal concentrations, lethal concentrations of micafungin were primarily fungicidal in WT cells, causing cell lysis without impairing cytokinesis. The lytic phenotype was suppressed again in pbr1-8 cells. Our results suggest that micafungin always exerts its fungicidal effect by solely inhibiting Bgs4. In contrast, lethal concentrations of anidulafungin and caspofungin cause an early cytokinesis arrest, probably by the combined inhibition of several GSs. Full article

Here, we describe the synthesis, characterization, and biological activities of a series of 26 new styryl-2(3H)-benzothiazolone analogs of combretastatin-A4 (CA-4). The cytotoxic activities of these compounds were tested in several cell lines (EA.hy926, A549, BEAS-2B, MDA-MB-231, HT-29, MCF-7, and MCF-10A), and the relations between structure and cytotoxicity are discussed. From the series, compound (Z)-3-methyl-6-(3,4,5-trimethoxystyryl)-2(3H)-benzothiazolone (26Z) exhibits the most potent cytotoxic activity (IC₅₀ 0.13 ± 0.01 µM) against EA.hy926 cells. 26Z not only inhibits vasculogenesis but also disrupts pre-existing vasculature. 26Z is a microtubule-modulating agent and inhibits a spectrum of angiogenic events in EA.hy926 cells by interfering with endothelial cell invasion, migration, and proliferation. 26Z also shows anti-proliferative activity in CA-4 resistant cells with the following IC₅₀ values: HT-29 (0.008 ± 0.001 µM), MDA-MB-231 (1.35 ± 0.42 µM), and MCF-7 (2.42 ± 0.48 µM). Cell-cycle phase-specific experiments show that 26Z treatment results in G2/M arrest and mitotic spindle multipolarity, suggesting that drug-induced centrosome amplification could promote cell death. Some 26Z-treated adherent cells undergo aberrant cytokinesis, resulting in aneuploidy that perhaps contributes to drug-induced cell death. These data indicate that spindle multipolarity induction by 26Z has an exciting chemotherapeutic potential that merits further investigation. Full article

The identification of effective pharmacological tools for Alzheimer’s disease (AD) represents one of the main challenges for therapeutic discovery. Due to the variety of pathological processes associated with AD, a promising route for pharmacological intervention involves the development of new chemical entities that can restore cellular homeostasis. To investigate this strategy, we designed and synthetized SG2, a compound related to the thyroid hormone thyroxine, that shares a pleiotropic activity with its endogenous parent compound, including autophagic flux promotion, neuroprotection, and metabolic reprogramming. We demonstrate herein that SG2 acts in a pleiotropic manner to induce recovery in a C. elegans model of AD based on the overexpression of Aβ42 and improves learning abilities in the 5XFAD mouse model of AD. Further, in vitro ADME-Tox profiling and toxicological studies in zebrafish confirmed the low toxicity of this compound, which represents a chemical starting point for AD drug development. Full article

Marine pharmacology is an exciting and growing discipline that blends blue biotechnology and natural compound pharmacology together. Several sea-derived compounds that are approved on the pharmaceutical market were discovered in sponges, marine organisms that are particularly rich in bioactive metabolites. This paper was specifically aimed at reviewing the pharmacological activities of extracts or purified compounds from marine sponges that were collected in the Mediterranean Sea, one of the most biodiverse marine habitats, filling the gap in the literature about the research of natural products from this geographical area. Findings regarding different Mediterranean sponge species were individuated, reporting consistent evidence of efficacy mainly against cancer, infections, inflammatory, and neurological disorders. The sustainable exploitation of Mediterranean sponges as pharmaceutical sources is strongly encouraged to discover new compounds. Full article

The lack of medication to treat COVID-19 is still an obstacle that needs to be addressed by all possible scientific approaches. It is essential to design newer drugs with varied approaches. A receptor-binding domain (RBD) is a key part of SARS-CoV-2 virus, located on its surface, that allows it to dock to ACE2 receptors present on human cells, which is followed by admission of virus into cells, and thus infection is triggered. Specific receptor-binding domains on the spike protein play a pivotal role in binding to the receptor. In this regard, the in silico method plays an important role, as it is more rapid and cost effective than the trial and error methods using experimental studies. A combination of virtual screening, molecular docking, molecular simulations and machine learning techniques are applied on a library of natural compounds to identify ligands that show significant binding affinity at the hydrophobic pocket of the RBD. A list of ligands with high binding affinity was obtained using molecular docking and molecular dynamics (MD) simulations for protein–ligand complexes. Machine learning (ML) classification schemes have been applied to obtain features of ligands and important descriptors, which help in identification of better binding ligands. A plethora of descriptors were used for training the self-organizing map algorithm. The model brings out descriptors important for protein–ligand interactions. Full article

Hydrogen, as a medical gas, is a promising emerging treatment for many diseases related to inflammation and oxidative stress. Molecular hydrogen can be generated through hydrogen ion reduction by a metal, and magnesium-containing effervescent tablets constitute an attractive formulation strategy for oral delivery. In this regard, saccharide-based excipients represent an important class of potential fillers with high water solubility and sweet taste. In this study, we investigated the effect of different saccharides on the morphological and mechanical properties and the disintegration of hydrogen-generating effervescent tablets prepared by dry granulation. Mannitol was found to be superior to other investigated saccharides and promoted far more rapid hydrogen generation combined with acceptable mechanical properties. In further product optimization involving investigation of lubricant effects, adipic acid was selected for the optimized tablet, due to regulatory considerations. Full article

Aptamers constitute an answer for the growing need for targeted therapy development. One of the most well-known representatives of this group of compounds is thrombin binding aptamers (TBA) targeted towards thrombin. The TBA inhibitory activity is determined by its spatial arrangement, which consists of two G-tetrads linked by two shorter TT loops and one longer TGT loop and folds into a unimolecular, antiparallel G-quadruplex structure. Interesting properties of the aptamer can be further improved via the introduction of a number of chemical modifications. Herein, a comprehensive analysis of the influence of pyrrolo-2’-deoxycytidine (Py-dC) and its derivatives on TBA physicochemical and biological properties has been presented. The studies have shown that the presence of modified residues at the T7 position of the TGT loop has only minor effects on TBA thermodynamic stability without affecting its folding topology. All analyzed oligomers exhibit anticoagulant properties, but only aptamer modified with a decyl derivative of Py-dC was able to inhibit thrombin activity more efficiently than unmodified, parental compounds. Importantly, the same compound also possessed the potential to effectively restrain HeLa cell line growth. Full article

The AHCC standardized extract of cultured Lentinula edodes mycelia, and the standardized extract of Asparagus officinalis stem, trademarked as ETAS, are well known supplements with immunomodulatory and anticancer potential. Several reports have described their therapeutic effects, including antioxidant and anticancer activity and improvement of immune response. In this study we aimed at investigating the effects of a combination of AHCC and ETAS on colorectal cancer cells and biopsies from healthy donors to assess the possible use in patients with colorectal cancer. Our results showed that the combination of AHCC and ETAS was synergistic in inducing a significant decrease in cancer cell growth, compared with single agents. Moreover, the combined treatment induced a significant increase in apoptosis, sparing colonocytes from healthy donors, and was able to induce a strong reduction in migration potential, accompanied by a significant modulation of proteins involved in invasiveness. Finally, combined treatment was able to significantly downregulate LGR5 and Notch1 in SW620 cancer stem cell (CSC) colonospheres. Overall, these findings support the potential therapeutic benefits of the AHCC and ETAS combinatorial treatment for patients with colorectal cancer. Full article

High-throughput screening of drug response in cultured cell lines is essential for studying therapeutic mechanisms and identifying molecular variants associated with sensitivity to drugs. Assessment of drug response is typically performed by constructing a dose-response curve of viability and summarizing it to a representative, such as IC₅₀. However, this is limited by its dependency on the assay duration and lack of reflections regarding actual cellular response phenotypes. To address these limitations, we consider how each response-phenotype contributes to the overall growth behavior and propose an alternative method of drug response screening that takes into account the cellular response phenotype. In conventional drug response screening methods, the ranking of sensitivity depends on either the metric used to construct the dose-response curve or the representative factor used to summarize the curve. This ambiguity in conventional assessment methods is due to the fact that assessment methods are not consistent with the underlying principles of population dynamics. Instead, the suggested phenotype metrics provide all phenotypic rates of change that shape overall growth behavior at a given dose and better response classification, including the phenotypic mechanism of overall growth inhibition. This alternative high-throughput drug-response screening would improve preclinical pharmacogenomic analysis and the understanding of a therapeutic mechanism of action. Full article

The pharmacological inhibition of soluble epoxide hydrolase (sEH) has been suggested as a potential therapy for the treatment of pain and inflammatory diseases through the stabilization of endogenous epoxyeicosatrienoic acids. Numerous potent sEH inhibitors (sEHI) have been developed, however many contain highly lipophilic substituents limiting their availability. Recently, a new series of benzohomoadamantane-based ureas endowed with potent inhibitory activity for the human and murine sEH was reported. However, their very low microsomal stability prevented further development. Herein, a new series of benzohomoadamantane-based amides were synthetized, fully characterized, and evaluated as sEHI. Most of these amides were endowed with excellent inhibitory potencies. A selected compound displayed anti-inflammatory effects with higher effectiveness than the reference sEHI, TPPU. Full article

Prostate cancer (PCa) is the major cause of cancer-related death in males; however, effective treatments to prevent aggressive progression remain an unmet need. We have previously demonstrated that secreted extracellular nicotinamide phosphoribosyltransferase (eNAMPT) is a multifunctional innate immunity regulator that promotes PCa invasion. In the current study, we further investigate the therapeutic effects of an eNAMPT-neutralizing humanized monoclonal antibody (ALT-100 mAb) in preclinical PCa orthotopic xenograft models. We utilized human aggressive PCa cells (DU145 or PC3) for prostate implantation in SCID mice receiving weekly intraperitoneal injections of either ALT-100 mAb or IgG/PBS (control) for 12 weeks. Prostatic tumors and solid organs were examined for tumor growth, invasion, and metastasis and for biochemical and immunohistochemistry evidence of NFκB activation. ALT-100 mAb treatment significantly improved overall survival of SCID mice implanted with human PCa orthotopic prostate xenografts while inducing tumor necrosis, decreasing PCa proliferation and reducing local invasion and distal metastases. The ALT-100 mAb inhibits NFκB phosphorylation and signaling in PCa cells both in vitro and in vivo. This study demonstrates that eNAMPT neutralization effectively prevents human PCa aggressive progression in preclinical models, indicating its high potential to directly address the unmet need for an effective targeted therapy for patients with aggressive PCa. Full article

One year after the spread of the pandemic, we analyzed the assessment results of the quality documentation submitted to the Clinical Trials Office of the Italian Medicines Agency as part of the request for authorization of clinical trials with a COVID-19 indication. In this article, we report the classification of the documentation type, an overview of the assessment results, and the related issues focusing on the most frequently detected ones. Relevant data regarding the Investigational Medicinal Products (IMPs) tested in COVID-19 clinical trials and their quality profiles are provided in the perspective of increasing transparency and availability of information. Some criticalities that have been exacerbated by the management of clinical trials during the emergency period are highlighted. Results confirm that IMPs tested in authorized COVID-19 clinical trials are developed in agreement with the same legal requirements for quality, safety, and efficacy as for any other medicinal product in the European Union (EU). The same strong regulatory framework applies, and there is no lowering in the safety profile due to the pandemic; authorized IMPs meet the highest standards of quality. The regulatory network should capitalize on lessons learned from the emergency setting. Some take-home messages are provided that could support the regulatory framework to expand its boundaries by innovating and evolving even though remaining strong and effective. Full article

Ipomoea batatas (L.) Lam., Convolvulaceae is widely distributed in Asian areas from tropical to warm-temperature regions. Their tubers are known for their antioxidant, anti-bacterial, anti-diabetic, wound healing, anti-inflammatory, and anti-ulcer activities. The preventive and therapeutic effects of orange-fleshed sweet potato on gastric ulcers have not been investigated. In this study, the carotenoid extract (CE) of orange-fleshed sweet potato was found to protect against gastric ulcers induced by HCl/ethanol in mice. The anti-inflammatory and antioxidant activities of the carotenoid pigment extract were also evaluated as possible evidence of their protective effects. Administration of CE reduced gastric ulcers. Oral administration of CE (100 mg/kg) protected against gastric ulcers by 78.1%, similar to the positive control, sucralfate (77.5%). CE showed potent reducing power and decreased nitric oxide production in a mouse macrophage cell line, RAW 264.7, in a concentration-dependent manner. The production of the inflammatory cytokine interleukin-6 and prostaglandin E₂ was also reduced by CE in a dose-dependent manner. The high carotenoid content of orange-fleshed sweet potato could play a role in its protective effect against gastric ulcers. This result suggests the possibility of developing functional products using this nutrient-fortified material. Full article

The development of new anticancer drugs is necessary in order deal with the disease and with the drawbacks of currently applied drugs. Epigenetic dysregulations are a central hallmark of cancerogenesis and histone deacetylases (HDACs) emerged as promising anticancer targets. HDAC inhibitors are promising epigenetic anticancer drugs and new HDAC inhibitors are sought for in order to obtain potent drug candidates. The new HDAC inhibitor SF5-SAHA was synthesized and analyzed for its anticancer properties. The new compound SF5-SAHA showed strong inhibition of tumor cell growth with IC₅₀ values similar to or lower than that of the clinically applied reference compound vorinostat/SAHA (suberoylanilide hydroxamic acid). Target specific HDAC inhibition was demonstrated by Western blot analyses. Unspecific cytotoxic effects were not observed in LDH-release measurements. Pro-apoptotic formation of reactive oxygen species (ROS) and caspase-3 activity induction in prostate carcinoma and hepatocellular carcinoma cell lines DU145 and Hep-G2 seem to be further aspects of the mode of action. Antiangiogenic activity of SF5-SAHA was observed on chorioallantoic membranes of fertilized chicken eggs (CAM assay). The presence of the pentafluorothio-substituent of SF5-SAHA increased the antiproliferative effects in both solid tumor and leukemia/lymphoma cell models when compared with its parent compound vorinostat. Based on this preliminary study, SF5-SAHA has the prerequisites to be further developed as a new HDAC inhibitory anticancer drug candidate. Full article

Triple-negative breast cancer (TNBC) is characterized by the absence of estrogen receptor-α progesterone receptor and human epidermal growth factor receptor-2. Treatment for this breast cancer subtype is restricted to multidrug chemotherapy and survival pathway-based molecularly targeted therapy. The long-term treatment options are associated with systemic toxicity, spontaneous and/or acquired tumor resistance and the emergence a of drug-resistant stem cell population. These limitations lead to advanced stage metastatic cancer. Current emphasis is on research directions that identify efficacious, naturally occurring agents representing an unmet need for testable therapeutic alternatives for therapy resistant breast cancer. Chinese herbs are widely used in traditional Chinese medicine in women for estrogen related health issues and also for integrative support for cancer treatment. This review discusses published evidence on a TNBC model for growth inhibitory effects of several mechanistically distinct nontoxic Chinese herbs, most of them nutritional in nature, and identifies susceptible pathways and potential molecular targets for their efficacy. Documented anti-proliferative and pro-apoptotic effects of these herbs are associated with downregulation of RB, RAS, PI3K, and AKT signaling, modulation of Bcl-2/BAX protein expressions and increased caspase activity. This review provides a proof of concept for Chinese herbs as testable alternatives for prevention/therapy of TNBC. Full article

5-Azacitidine, a cytidine analogue used as a hypomethylating agent, is one of the main drugs for the treatment of myelodysplastic syndromes (MDSs) and acute myeloid leukemia (AML) in the elderly. However, after administration, it exhibits several limitations, including restricted diffusion and cellular internalization due to its hydrophilicity, and a rapid enzymatic degradation by adenosine deaminase. The aim of this study was to improve the drug cell diffusion and protect it from metabolic degradation via the synthesis of amphiphilic prodrugs and their potential self-assembly. Azacitidine was conjugated to two different omega-3 fatty acids, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). The carboxylic acid group of the omega-3 fatty acids was effectively conjugated to the amine group of the azacitidine base, yielding two amphiphilic prodrugs. Nanoprecipitation of the obtained prodrugs was performed and self-assemblies were successfully obtained for both prodrugs, with a mean diameter of 190 nm, a polydispersity index below 0.2 and a positive zeta potential. The formation of self-assemblies was confirmed using pyrene as a fluorescent dye, and the critical aggregation concentrations were determined: 400 µM for AzaEPA and 688 µM for AzaDHA. Additionally, the stability of the obtained self-assemblies was studied and after 5 days their final stable arrangement was reached. Additionally, cryo-TEM revealed that the self-assemblies attain a multilamellar vesicle supramolecular structure. Moreover, the obtained self-assemblies presented promising cytotoxicity on a leukemia human cell line, having a low IC₅₀ value, comparable to that of free azacitidine. Full article

The endocannabinoid system (ECS) plays an integral role in maintaining metabolic homeostasis and may affect hunger, caloric intake, and nutrient absorption. Obesity has been associated with higher levels of the endogenous cannabinoid transmitters (endocannabinoids). Therefore, the ECS is an important target in obesity treatment. Modulating the enzymes that synthesize and degrade endocannabinoids, namely fatty acid amide hydrolase (FAAH), monoacylglycerol lipase (MAGL), and diacylglycerol lipase (DAGL), may be a promising strategy to treat obesity. This review aims to synthesize all studies investigating pharmacological or genetic manipulation of FAAH, MAGL, or DAGL enzymes in association with obesity-related measures. Pharmacological inhibition or genetic deletion of FAAH tended to promote an obesogenic state in animal models, though the relationships between human FAAH polymorphisms and obesity-related outcomes were heterogeneous, which could be due to FAAH having both pro-appetitive and anti-appetitive substrates. Genetic deletion of Mgll and Dagla as well as pharmacological inhibition of DAGL tended to reduce body weight and improve metabolic state in animal studies, though the effects of Mgll manipulation were tissue-dependent. Monitoring changes in body weight in ongoing clinical trials of FAAH inhibitors may clarify whether FAAH inhibition is a potential therapeutic strategy for treatment obesity. More preclinical work is needed to characterize the role of MAGL and DAGL modulation in obesity-related outcomes. Full article

Ovarian cancer (OCa) is characterized as one of the common reasons for cancer-associated death in women globally. This gynecological disorder is chiefly named the “silent killer” due to lacking an association between disease manifestations in the early stages and OCa. Because of the disease recurrence and resistance to common therapies, discovering an effective therapeutic way against the disease is a challenge. According to documents, some popular herbal formulations, such as curcumin, quercetin, and resveratrol, can serve as an anti-cancer agent through different mechanisms. However, these herbal products may be accompanied by some pharmacological limitations, such as poor bioavailability, instability, and weak water solubility. On the contrary, using nano-based material, e.g., nanoparticles (NPs), micelles, liposomes, can significantly solve these limitations. Therefore, in the present study, we will summarize the anti-cancer aspects of these herbal and-nano-based herbal formulations with a focus on their mechanisms against OCa. Full article

Highly stable and facile one-pot copper nanoclusters (Cu NCs) coated with poly(allylamine hydrochloride) (PAH) have been synthesized for selectively sensing deferasirox (DFX) in β-thalassemia plasma. DFX is an important drug used for treating iron overloading in β-thalassemia, but needs to be monitored due to certain toxicity. In this study, the PAH-Cu NCs showed highly stable fluorescence with emission wavelengths at 450 nm. The DFX specifically interacted with the copper nanocluster to turn off the fluorescence of the PAH-Cu NCs, and could be selectively quantified through the fluorescence quenching effect. The linear range of DFX in plasma analyzed by PAH-Cu NCs was 1.0–100.0 µg/mL (r = 0.985). The relative standard deviation (RSD) and relative error (RE) were lower than 6.51% and 7.57%, respectively, showing excellent reproducibility of PAH-Cu NCs for sensing DFX in plasma. This method was also successfully applied for an analysis of three clinical plasma samples from β-thalassemia patients taking DFX. The data presented high similarity with that obtained through a capillary electrophoresis method. According to the results, the PAH-Cu NCs could be used as a tool for clinically sensing DFX in human plasma for clinical surveys. Full article

The global emergence of the COVID-19 pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has focused the entire world’s attention toward searching for a potential remedy for this disease. Thus, we investigated the antiviral activity of Agrimonia pilosa ethanol extract (APEE) against SARS-CoV-2 and it exhibited a potent antiviral activity with IC₅₀ of 1.1 ± 0.03 µg/mL. Its mechanism of action was elucidated, and it exhibited a virucidal activity and an inhibition of viral adsorption. Moreover, it presented an immunomodulatory activity as it decreased the upregulation of gene expression of COX-2, iNOS, IL-6, TNF-α, and NF-κB in lipopolysaccharide (LPS)-induced peripheral blood mononuclear cells. A comprehensive analysis of the phytochemical fingerprint of APEE was conducted using LC-ESI-MS/MS technique for the first time. We detected 81 compounds and most of them belong to the flavonoid and coumarin classes. Interestingly, isoflavonoids, procyanidins, and anthocyanins were detected for the first time in A. pilosa. Moreover, the antioxidant activity was evidenced in DPPH (IC₅₀ 62.80 µg/mL) and ABTS (201.49 mg Trolox equivalents (TE)/mg) radical scavenging, FRAP (60.84 mg TE/mg), and ORAC (306.54 mg TE/g) assays. Furthermore, the protective effect of APEE was investigated in Lipopolysaccharides (LPS)-induced acute lung injury (ALI) in mice. Lung W/D ratio, serum IL-6, IL-18, IL-1β, HO-1, Caspase-1, caspase-3, TLR-4 expression, TAC, NO, MPO activity, and histopathological examination of lung tissues were assessed. APEE induced a marked downregulation in all inflammation, oxidative stress, apoptosis markers, and TLR-4 expression. In addition, it alleviated all histopathological abnormalities confirming the beneficial effects of APEE in ALI. Therefore, APEE could be a potential source for therapeutic compounds that could be investigated, in future preclinical and clinical trials, in the treatment of patients with COVID-19. Full article

The utility of clinically available antifungals is limited by their narrow spectrum of activity, high toxicity, and emerging resistance. Antifungal drug discovery has always been a challenging area, since fungi and their human host are eukaryotes, making it difficult to identify unique targets for antifungals. Novel antifungals in clinical development include first-in-class agents, new structures for an established target, and formulation modifications to marketed antifungals, in addition to repurposed agents. Membrane interacting peptides and aromatherapy are gaining increased attention in the field. Immunotherapy is another promising treatment option, with antifungal antibodies advancing into clinical trials. Novel targets for antifungal therapy are also being discovered, allowing the design of new promising agents that may overcome the resistance issue. In this mini review, we will summarize the current status of antifungal drug pipelines in clinical stages, and the most recent advancements in preclinical antifungal drug development, with special focus on their chemistry. Full article

Pyrido[2,3-d]pyrimidin-7(8H)-ones have attracted widespread interest due to their similarity with nitrogenous bases found in DNA and RNA and their potential applicability as tyrosine kinase inhibitors. Such structures, presenting up to five diversity centers, have allowed the synthesis of a wide range of differently substituted compounds; however, the diversity at the C4 position has mostly been limited to a few substituents. In this paper, a general synthetic methodology for the synthesis of 4-substituted-2-(phenylamino)-5,6-dihydropyrido[2,3-d]pyrimidin-7(8H)-ones is described. By using cross-coupling reactions, such as Ullmann, Buchwald–Hartwig, Suzuki–Miyaura, or Sonogashira reactions, catalyzed by Cu or Pd, we were able to describe new potential biologically active compounds. The resulting pyrido[2,3-d]pyrimidin-7(8H)-ones include N-alkyl, N-aryl, O-aryl, S-aryl, aryl, and arylethynyl substituents at C4, which have never been explored in connection with the biological activity of such heterocycles as tyrosine kinase inhibitors, in particular as ZAP-70 inhibitors. Full article

The edible leaves of the mulberry (Morus alba L.) plant are used worldwide. They contain abundant polyphenolic compounds with strong anticancer properties. We previously revealed that apoptosis was mediated in p53-negative Hep3B cells, and mulberry leaf polyphenol extract (MLPE) induced autophagy in p53-transfected Hep3B cells. However, how this autophagy is induced by p53 in human hepatoma HepG2 (p53 wild type) cells remains unclear. In the current study, MLPE induced autophagy, as demonstrated by enhanced acidic vesicular organelle staining, by upregulating beclin-1, increasing LC3-II conversion, and phosphorylating AMPK. In HepG2 cells, these processes were associated with p53. Western blot also revealed phosphatidylinositol-3 kinase (PI3K), p-AKT, and fatty acid synthase (FASN) suppression in MLPE-treated cells. Moreover, treatment with the p53 inhibitor pifithrin-α (PFT-α) inhibited autophagy and increased apoptotic response in MLPE-treated HepG2 cells. PFT-α treatment also reversed MLPE-induced PI3K, p-AKT, and FASN suppression. Thus, co-treatment with MLPE and PFT-α significantly increased caspase-3, caspase-8, and cytochrome c release, indicating that p53 deficiency caused the apoptosis. In addition, rutin, a bioactive polyphenol in MLPE, may affect autophagy in HepG2 cells. This study demonstrates that MLPE is a potential anticancer agent targeting autophagy and apoptosis in cells with p53 status. Moreover, this work provides insight into the mechanism of p53 action in MLPE-induced cytotoxicity in hepatocellular carcinoma. Full article

Pancreatic cancer currently represents a severe issue for the entire world. Therefore, much effort has been made to develop an effective treatment against it. Emerging evidence has shown that icariin, a flavonoid glycoside, is an effective anti-pancreatic cancer drug. Melittin, as a natural active biomolecule, has also shown to possess anticancer activities. In the present study, with the aim to increase its effectiveness against cancerous cells, icariin-loaded bilosome-melittin (ICA-BM) was developed. For the selection of an optimized ICA-BM, an experimental design was implemented, which provided an optimized formulation with a particle size equal to 158.4 nm. After estimation of the release pattern, the anti-pancreatic cancer efficacy of this new formulation was evaluated. The MTT assay was employed for the determination of half maximal inhibitory concentration (IC₅₀), providing smaller IC₅₀ for ICA-BM (2.79 ± 0.2 µM) compared to blank-BM and ICA-Raw (free drug) against PNAC1, a human pancreatic cancer cell line isolated from a pancreatic carcinoma of ductal cell origin. Additionally, cell cycle analysis for ICA-BM demonstrated cell arrest at the S-phase and pre-G1 phase, which indicated a pro-apoptotic behavior of the new developed formulation. The pro-apoptotic and anti-proliferative activity of the optimized ICA-BM against PNAC1 cells was also demonstrated through annexin V staining as well as estimation of caspase-3 and p53 protein levels. It can be concluded that the optimized ICA-BM formulation significantly improved the efficacy of icariin against cancerous pancreatic cells. Full article