Structure-function analysis of STING activation by c[G(2',5')pA(3',5')p] and targeting by antiviral DMXAA.

Binding of dsDNA by cyclic GMP-AMP (cGAMP) synthase (cGAS) triggers formation of the metazoan second messenger c[G(2',5')pA(3',5')p], which binds the signaling protein STING with subsequent activation of the interferon (IFN) pathway. We show that human hSTING(H232) adopts a "closed" conformation upon binding c[G(2',5')pA(3',5')p] and its linkage isomer c[G(2',5')pA(2',5')p], as does mouse mSting(R231) on binding c[G(2',5')pA(3',5')p], c[G(3',5')pA(3',5')p] and the antiviral agent DMXAA, leading to similar "closed" conformations. Comparing hSTING to mSting, 2',5'-linkage-containing cGAMP isomers were more specific triggers of the IFN pathway compared to the all-3',5'-linkage isomer. Guided by structural information, we identified a unique point mutation (S162A) placed within the cyclic-dinucleotide-binding site of hSTING that rendered it sensitive to the otherwise mouse-specific drug DMXAA, a conclusion validated by binding studies. Our structural and functional analysis highlights the unexpected versatility of STING in the recognition of natural and synthetic ligands within a small-molecule pocket created by the dimerization of STING.

RCHY1 and OPTN are required for melanophagy, selective autophagy of melanosomes.

Melanosomes are specific organelles dedicated to melanin synthesis and accumulation in melanocytes. Autophagy is suggestively involved in melanosome degradation, although the potential underlying molecular mechanisms remain elusive. In selective autophagy, autophagy receptors and E3-ligases are the key factors conferring cargo selectivity. In B16F10 cells, ß-mangostin efficiently induced melanosome degradation without affecting other organelles such as mitochondria, peroxisomes, and the endoplasmic reticulum. Among various autophagy receptors, optineurin (OPTN) contributes TANK-binding kinase 1 (TBK1)-dependently to melanosome degradation and its knockdown inhibited ß-mangostin-mediated melanosome degradation. OPTN translocation to melanosomes was dependent on its ubiquitin-binding domain. Moreover, OPTN-mediated TBK1 activation and subsequent TBK1-mediated S187 OPTN phosphorylation were essential for melanosome degradation. ß-mangostin increased K63-linked melanosome ubiquitination. Finally, the E3-ligase RCHY1 knockdown inhibited the melanosome ubiquitination required for OPTN- and TBK1-phosphorylation as well as melanosome degradation. This study suggests that melanophagy, melanosome-selective autophagy, contributes to melanosome degradation, and OPTN and RCHY1 are an essential autophagy receptor and a E3-ligase, respectively, conferring cargo selectivity in melanophagy.

The total xanthones extracted from Gentianella acuta alleviates HFpEF by activating the IRE1α/Xbp1s pathway.

Heart failure with preserved ejection fraction (HFpEF) is a clinical syndrome characterized by pulmonary and systemic congestion resulting from left ventricular diastolic dysfunction and increased filling pressure. Currently, however, there is no evidence on effective pharmacotherapy for HFpEF. In this study, we aimed to investigate the therapeutic effect of total xanthones extracted from Gentianella acuta (TXG) on HFpEF by establishing an high-fat diet (HFD) + L-NAME-induced mouse model. Echocardiography was employed to assess the impact of TXG on the cardiac function in HFpEF mice. Haematoxylin and eosin staining, wheat germ agglutinin staining, and Masson's trichrome staining were utilized to observe the histopathological changes following TXG treatment. The results demonstrated that TXG alleviated HFpEF by reducing the expressions of genes associated with myocardial hypertrophy, fibrosis and apoptosis. Furthermore, TXG improved cardiomyocyte apoptosis by inhibiting the expression of apoptosis-related proteins. Mechanistic investigations revealed that TXG could activate the inositol-requiring enzyme 1α (IRE1α)/X-box-binding protein 1 (Xbp1s) signalling pathway, but the knockdown of IRE1α using the IRE1α inhibitor STF083010 or siRNA-IRE1α impaired the ability of TXG to ameliorate cardiac remodelling in HFpEF models. In conclusion, TXG alleviates myocardial hypertrophy, fibrosis and apoptosis through the activation of the IRE1α/Xbp1s signalling pathway, suggesting its potential beneficial effects on HFpEF patients.

A targeting nanoplatform for chemo-photothermal synergistic therapy of small-cell lung cancer.

The precise delivery of drugs to tumor sites and the thermoresistance of tumors remain major challenges in photothermal therapy (PTT). Somatostatin receptor 2 (SSTR2) is proposed as an ideal target for the precise treatment of SCLC. We developed a targeting nano-drug delivery system comprising anti-SSTR2 monoclonal antibody (MAb) surface-modified nanoparticles co-encapsulating Cypate and gambogic acid (GA). The formed SGCPNs demonstrated excellent monodispersity, physiological stability, preferable biocompatibility, and resultant efficient photothermal conversion efficacy. SGCPNs were quickly internalized by SSTR2-overexpressing SCLC cells, triggering the release of GA under acidic and near-infrared (NIR) laser irradiation environments, leading to their escape from lysosomes to the cytosol and then diffusion into the nucleus. SGCPNs can not only decrease the cell survival rate but also inhibit the activity of heat shock protein 90 (HSP90). SGCPNs can be precisely delivered to xenograft tumors of SSTR2-positive SCLC in vivo. Upon NIR laser irradiation, therapy of SGCPNs showed significant tumor regression. In conclusion, SGCPNs provide a new chemo-photothermal synergistic treatment strategy for targeting SCLC.

Multiple Treatment of Triple-Negative Breast Cancer Through Gambogic Acid-Loaded Mesoporous Polydopamine.

Triple-negative breast cancer (TNBC) is a highly heterogeneous subtype of breast cancer, characterized by aggressiveness and high recurrence rate. As monotherapy provides limited benefit to TNBC patients, combination therapy emerges as a promising treatment approach. Gambogic acid (GA) is an exceedingly promising anticancer agent. Nonetheless, its application potential is hampered by low drug loading efficiency and associated toxic side effects. To overcome these limitations, using mesoporous polydopamine (MPDA) endowed with photothermal conversion capabilities is considered as a delivery vehicle for GA. Meanwhile, GA can inhibit the activity of heat shock protein 90 (HSP90) to enhance the photothermal effect. Herein, GA-loaded MPDA nanoparticles (GA@MPDA NPs) are developed with a high drug loading rate of 75.96% and remarkable photothermal conversion performance. GA@MPDA NPs combined with photothermal treatment (PTT) significantly inhibit the tumor growth, and effectively trigger the immunogenic cell death (ICD), which thereby increase the number of activated effector T cells (CD8+ T cells and CD4+ T cells) in the tumor, and hoist the level of immune-inflammatory cytokines (IFN-γ, IL-6, and TNF-α). The above results suggest that the combination of GA@MPDA NPs with PTT expected to activate the antitumor immune response, thus potentially enhancing the clinical therapeutic effect on TNBC.

Biosynthesis of polyprenylated xanthones in Hypericum perforatum roots involves 4-prenyltransferase.

Polyprenylated xanthones are natural products with a multitude of biological and pharmacological activities. However, their biosynthetic pathway is not completely understood. In this study, metabolic profiling revealed the presence of 4-prenylated 1,3,5,6-tetrahydroxyxanthone derivatives in St. John's wort (Hypericum perforatum) root extracts. Transcriptomic data mining led to the detection of 5 variants of xanthone 4-prenyltransferase (HpPT4px) comprising 4 long variants (HpPT4px-v1 to HpPT4px-v4) and 1 short variant (HpPT4px-sh). The full-length sequences of all 5 variants were cloned and heterologously expressed in yeast (Saccharomyces cerevisiae). Microsomes containing HpPT4px-v2, HpPT4px-v4, and HpPT4px-sh catalyzed the addition of a prenyl group at the C-4 position of 1,3,5,6-tetrahydroxyxanthone; 1,3,5-trihydroxyxanthone; and 1,3,7-trihydroxyxanthone, whereas microsomes harboring HpPT4px-v1 and HpPT4px-v3 additionally accepted 1,3,6,7-tetrahydroxyxanthone. HpPT4px-v1 produced in Nicotiana benthamiana displayed the same activity as in yeast, while HpPT4px-sh was inactive. The kinetic parameters of HpPT4px-v1 and HpPT4px-sh chosen as representative variants indicated 1,3,5,6-tetrahydroxyxanthone as the preferred acceptor substrate, rationalizing that HpPT4px catalyzes the first prenylation step in the biosynthesis of polyprenylated xanthones in H. perforatum. Dimethylallyl pyrophosphate was the exclusive prenyl donor. Expression of the HpPT4px transcripts was highest in roots and leaves, raising the question of product translocation. C-terminal yellow fluorescent protein fusion of HpPT4px-v1 localized to the envelope of chloroplasts in N. benthamiana leaves, whereas short, truncated, and masked signal peptides led to the disruption of plastidial localization. These findings pave the way for a better understanding of the prenylation of xanthones in plants and the identification of additional xanthone-specific prenyltransferases.

Nucleobase-Coupled Xanthones with Anti-ROS Effects from Marine-Derived Fungus Aspergillus sydowii.

A MS/MS-based molecular networking approach compared to the Global Natural Product Social Molecular Networking library, in association with genomic annotation of natural product biosynthetic gene clusters within a marine-derived fungus, Aspergillus sydowii, identified a suite of xanthone metabolites. Chromatographic techniques applied to the cultured fungus led to the isolation of 11 xanthone-based alkaloids, dubbed sydoxanthones F-M. The structures of these alkaloids were elucidated using extensive spectroscopic data, including electronic circular dichroism and single-crystal X-ray diffraction data for configurational assignments. Among these analogues, sydoxanthones F-K exhibit structure features typical of nucleobase-coupled xanthones, with sydoxanthone H being an N-bonded xanthone dimer. Notably, (±)sydoxanthones F (1a/1b), (±)sydoxanthones H (3b/3a), and (±)sydoxanthones J (5b/5a) are enantiomeric pairs, while sydoxanthones G (2), I (4), and K (6) are stereoisomers of 1, 3, and 5, respectively. Furthermore, (+)sydoxanthone H (3a) demonstrated significant rescue of cell viability in H2O2-injuried SH-SY5Y cells by inhibiting reactive oxygen species production, suggesting its potential for neuroprotection.

Novel xanthone derivatives as potent sirtuin 2 inhibitors.

Six amino derivatives of xanthone were obtained via chemical synthesis. Biochemical studies revealed their SIRT2 inhibitory activity ranging from 48.5 % (compound 4, 5-chloro-2-((4-(3-methoxyphenyl)piperazin-1-yl)methyl)-9H-xanthen-9-one hydrochloride) to 93.2 % (compound 3, 5-chloro-2-(((2-methoxyphenethyl)amino)methyl)-9H-xanthen-9-one hydrochloride). The structure-activity analysis showed favourable properties of secondary amines relative to tertiary piperazine derivatives. The tested compounds do not possess additional SIRT1 activating activity and no antioxidant activity (DPPH in vitro assay). Comprehensive analysis of the lipophilicity of the obtained compounds was also performed. For compound 3 potential molecular targets and similar active compounds were predicted in order to facilitate further research in this group of compounds.

Neutral rhodol-based dyes expressing localization in mitochondria.

Neutral rhodol-based red emitters are shown to efficiently localize in mitochondria, as demonstrated by confocal microscopy and co-localization studies. A simple model is proposed to explain the localization mechanism of neutral molecules. The model takes into account the strong coupling between the molecular dipole moment and the electric field of the inner mitochondrial membrane.

Caged xanthone derivatives to promote mitochondria-mediated apoptosis in breast cancer cells.

Caged xanthones represent a class of natural secondary metabolites exhibiting significant potential as antitumor agents. These compounds are characterized by their distinct cage-like structures, which offer novel and compelling frameworks for drug design. Nonetheless, there exists a dearth of research focused on the structural modification of these compounds, particularly in relation to their cage-like architectures. This study aims to address this gap by introducing an innovative synthetic method for constructing a novel caged structure that incorporates a widely employed maleimide group. Drawing upon the well-established synthetic approach for dihydroxanthones previously developed within our research group, we successfully synthesized 13 new caged xanthones using the Diels-Alder reaction. Subsequently, we evaluated their anti-proliferative activity against HepG2, A549, and MDA-MB-231 cell lines. The results revealed that compound 10i exhibited IC50 values of 15.86 µM ± 1.29, 19.27 µM ± 1.58, and 12.96 µM ± 0.09 against these cell lines, respectively. Further investigations into the mechanism of action of 10i demonstrated its ability to induce G2/M cell cycle arrest and initiate mitochondria-mediated apoptosis in breast cancer cells.

Identification of small molecule inhibitors of penicillin-binding protein 2a of methicillin-resistant Staphylococcus aureus for the therapeutics of bacterial infection.

The penicillin binding protein 2a (PBP2a) is a key enzyme associated with bacterial cell wall synthesis and bacterial infection. Therefore, targeting PBPa2 offers a promising approach for the therapeutics of bacterial resistance and infection. This study presents a comprehensive analysis of alpha-mangostin as a potential inhibitor of PBPa2. Molecular docking simulations revealed a strong binding affinity between alpha-mangostin and PBP2a, with an affinity score of -6.01 kcal/mol. Notably, alpha-mangostin formed a preferential hydrogen bond with THR216 of PBP2a, alongside several other polar and hydrophobic interactions. ADME and Toxicity predictions indicated that alpha-mangostin possesses favourable pharmacokinetic properties, suggesting its potential as a therapeutic agent. PASS analysis further highlighted its broad range of favourable biological properties. SwissTargetPrediction analysis reinforced these findings, indicating alpha-mangostin's association with various biological processes. Cell toxicity assays demonstrated that alpha-mangostin had no significant impact on the viability of HEK-293 cells, suggesting its potential safety for further development. The IC50 value for alpha-mangostin was found to be 33.43µM. Fluorescence-based binding assays showed that alpha-mangostin effectively inhibited PBP2a activity in a concentration-dependent manner, supporting its role as an inhibitor. In conclusion, the results suggest alpha-mangostin as a promising candidate for inhibiting PBP2a. Further,  extensive studies are warranted to explore its clinical applications.

Mangiferin: A natural bioactive immunomodulating glucosylxanthone with potential against cancer and rheumatoid arthritis.

Mangiferin is a naturally occurring glucosylxanthone that has shown promising immunomodulatory effects. It is generally isolated from the leaves, peels, bark, and kernels of Mangifera indica Linn. Mangiferin is like a miraculous natural bioactive molecule that has an immunomodulatory function that makes it a potential therapeutic candidate for the treatment of rheumatoid arthritis (RA) and cancer. The anticancer activity of mangiferin acts by blocking NF-κB, as well as regulating the ß-catenin, EMT, MMP9, MMP2, LDH, ROS, and NO, and also by the activation of macrophages. It has no cytotoxic effect on grown chondrocytes and lowers matrix metalloproteinase levels. Additionally, it has a potent proapoptotic impact on synoviocytes. The precise molecular mechanism of action of mangiferin on RA and malignancies is still unknown. This comprehensive review elaborates on the immunomodulatory effect of mangiferin and its anticancer and anti-RA activity. This also explained the total synthesis of mangiferin and its in vitro and in vivo screening models.

Mangiferin activates the nuclear factor erythroid 2-related factor pathway to protect SOD1-G93A induced NSC-34 motor neurons from oxidative stress and apoptosis.

One of the main factors in the pathophysiology of amyotrophic lateral sclerosis is oxidative stress. Mangiferin (MF), a natural plant polyphenol, has anti-inflammatory and antioxidant effects. The aim of our study was to investigate the protective effects and mechanisms of MF in the hSOD1-G93A ALS cell model. Our result revealed that MF treatment reduced the generation of reactive oxygen species (ROS) and malondialdehyde (MDA), decreased oxidative damage, and reduced apoptosis. Additionally, it was observed that MF significantly increased the synthesis of the antioxidant genes hemeoxygenase-1 and NAD(P)H: quinone oxidoreductase 1, which are downstream of the Nrf2 signaling pathway, and increased the expression and activation of nuclear factor erythroid 2-related factor 2 (Nrf2). Nrf2 knockdown greatly promoted apoptosis, which was reversed by MF treatment. To summarize, MF promoted the Nrf2 pathway and scavenged MDA and ROS to protect the ALS cell model.

Caged Xanthones and Biphenyls Isolated from the Tropical Plant Garcinia lateriflora.

Four cytotoxic heptacyclic caged-xanthones [gambogefic acids B-E (1-4)], a cytotoxic hexacyclic caged-xanthone [garcilatelic acid (5)], and four biphenyl derivatives [garcilatelibiphenyls A-D (6-9)] were newly isolated in a phytochemical study of a 50% MeOH/CH2Cl2 extract of Garcinia lateriflora (Clusiaceae). The isolated compounds were evaluated for antiproliferative activity against five human tumor cell lines including a vincristine-resistant line. The new caged-xanthones displayed potent activity with IC50 values from 0.5 to 6.7 µM against all tested tumor cell lines.

Xanthone Inhibitors of Unfolded Protein Response Isolated from Calophyllum caledonicum.

The unfolded protein response (UPR) is a key component of fungal virulence. The prenylated xanthone γ-mangostin isolated from Garcinia mangostana (Clusiaceae) fruit pericarp, has recently been described to inhibit this fungal adaptative pathway. Considering that Calophyllum caledonicum (Calophyllaceae) is known for its high prenylated xanthone content, its stem bark extract was fractionated using a bioassay-guided procedure based on the cell-based anti-UPR assay. Four previously undescribed xanthone derivatives were isolated, caledonixanthones N-Q (3, 4, 8, and 12), among which compounds 3 and 8 showed promising anti-UPR activities with IC50 values of 11.7 ± 0.9 and 7.9 ± 0.3 µM, respectively.

Structure-Guided Discovery of Diverse Cytotoxic Dimeric Xanthones/Chromanones from Penicillium chrysogenum C-7-2-1 and Their Interconversion Properties.

Xanthone-chromanone homo- or heterodimers are regarded as a novel class of topoisomerase (Topo) inhibitors; however, limited information about these compounds is currently available. Here, 14 new (1-14) and 6 known tetrahydroxanthone chromanone homo- and heterodimers (15-20) are reported as isolated from Penicillium chrysogenum C-7-2-1. Their structures and absolute configurations were unambiguously demonstrated by a combination of spectroscopic data, single-crystal X-ray diffraction, modified Mosher's method, and electronic circular dichroism analyses. Plausible biosynthetic pathways are proposed. For the first time, it was discovered that tetrahydroxanthones can convert to chromanones in water, whereas chromone dimerization does not show this property. Among them, compounds 5, 7, 8, and 16 exhibited significant cytotoxicity against H23 cell line with IC50 values of 6.9, 6.4, 3.9, and 2.6 µM, respectively.

Looking for Actives in the Haystack: Merging HRMS2-Based Molecular Networking, Chemometrics, and 13C NMR-Based Dereplication Approaches.

The identification of bioactive natural products (NPs) in complex mixtures has become an important subject of contemporary NP research. In an attempt to address this challenge, the present work proposes an integrated strategy that combines tandem mass spectrometry (MS2)-based molecular networking (MN), a partial least-squares (PLS) chemometric model, as well as 13C NMR-based dereplication using MixONat software. In addition, an advanced glycation end product (AGEs) assay was used for activity evaluation. The approach was implemented on a Garcinia parvifolia bark extract that comprised a high content of prenylated xanthones and had previously shown a notable inhibitory effect on AGE formation. As a main result, the proposed strategy permitted the identification of potentially active metabolites within complex mixtures and their annotation with a higher level of confidence by NMR data. Overall, this comprehensive approach provides a powerful and efficient solution for the targeting and annotating of active compounds in complex NP mixtures.

Activity of the Caged Xanthone Morellic Acid against Vancomycin-Resistant Enterococcus Infection by Targeting the Bacterial Membrane.

Vancomycin-resistant Enterococcus (VRE) is an important nosocomial opportunistic pathogen that is associated with multidrug resistance. Here, we demonstrate that morellic acid inhibits VRE by restoring its sensitivity to vancomycin and ampicillin with low drug resistance and efficient biofilm clearance effects. Morellic acid binds to inner membrane phospholipids, such as phosphatidylethanolamine (PE), phosphatidylglycerol (PG), and cardiolipin (CL) of VRE, such that the fluidity and proton-motive force (PMF) interfere with the damaged inner membrane, causing intracellular reactive oxygen species (ROS) accumulation and bacterial death. Transcriptional analyses supported this effect on inner membrane-related pathways such as fatty acid biosynthesis and glycerophospholipid metabolism. Moreover, morellic acid significantly eliminated residual bacteria in the spleen, liver, kidneys, and abdominal effusion in mice. Our findings indicate the potential applications of morellic acid as an antibacterial agent or adjuvant for treating VRE infections.

Deciphering molecular mechanisms underlying the inhibition of ß-glucuronidase by xanthones from Centaurium spicatum.

Herein, we scrutinized the inhibitory potential of five xanthones and a flavonoid, sourced from Centaurium spicatum, against ß-glucuronidase activity. The results showed that gentisin and azaleatin emerged as the most potent inhibitors, with significantly lower IC50 values of 0.96 ± 0.10 and 0.57 ± 0.04 µM, respectively. The evaluation of enzyme kinetics unveiled that the isolated xanthones manifested inhibition of ß-glucuronidase through a mixed inhibition mode, whereas azaleatin exhibited a noncompetitive inhibition mechanism. The findings from molecular docking analysis unveiled that the compounds under investigation, particularly azaleatin, displayed comparatively diminished binding affinities towards ß-glucuronidase. Furthermore, the tested drugs were shown to occupy a common binding site as the employed reference drug. Our comprehensive Molecular Dynamics (MD) simulations analysis revealed consistent trajectories for the investigated drugs, wherein azaleatin and gentisin demonstrated notable stabilization of energy levels. Analysis of various MD parameters revealed that drugs with the lowest IC50 values maintained relatively stable interactions with ß-glucuronidase. These drugs were shown to exert notable alterations in their conformation or flexibility upon complexation with the target enzyme. Conversely, the flexibility and accessibility of ß-glucuronidase was reduced upon drug binding, particularly with azaleatin and gentisin, underscoring the stability of the drug-enzyme complexes. Analysis of Coul-SR and LJ-SR interaction energies unveiled consistent and stable interactions between certain isolated drugs and ß-glucuronidase. Azaleatin notably displayed the lowest average Coul-SR interaction energy, suggesting strong electrostatic interactions with the enzyme's active site and significant conformational variability during simulation. Remarkably, LJ-SR interaction energies across different xanthones complexes were more negative than their Coul-SR counterparts, emphasizing the predominant role of van der Waals interactions, encompassing attractive dispersion and repulsive forces, in stabilizing the drug-enzyme complexes rather than electrostatic interactions.

Mangiferin alleviates diabetic pulmonary fibrosis in mice via inhibiting endothelial-mesenchymal transition through AMPK/FoxO3/SIRT3 axis.

Diabetes mellitus results in numerous complications. Diabetic pulmonary fibrosis (DPF), a late pulmonary complication of diabetes, has not attracted as much attention as diabetic nephropathy and cardiomyopathy. Mangiferin (MF) is a natural small molecular compound that exhibits a variety of pharmacological effects including anti-inflammatory, anti-cancer, anti-diabetes, and anti-fibrosis effects. In this study, we investigated whether long-term diabetes shock induces DPF, and explored whether MF had a protective effect against DPF. We first examined the lung tissues and sections of 20 diabetic patients obtained from discarded lung surgical resection specimens and found that pulmonary fibrosis mainly accumulated around the pulmonary vessels, accompanied by significantly enhanced endothelial-mesenchymal transition (EndMT). We established a mouse model of DPF by STZ injections. Ten days after the final STZ injection, the mice were administered MF (20, 60 mg/kg, i.g.) every 3 days for 4 weeks, and kept feeding until 16 weeks and euthanized. We showed that pulmonary fibrotic lesions were developed in the diabetic mice, which began around the pulmonary vessels, while MF administration did not affect long-term blood glucose levels, but dose-dependently alleviated diabetes-induced pulmonary fibrosis. In human umbilical vein endothelial cells (HUVECs), exposure to high glucose (33.3 mM) induced EndMT, which was dose-dependently inhibited by treatment with MF (10, 50 µM). Furthermore, MF treatment promoted SIRT3 expression in high glucose-exposed HUVECs by directly binding to AMPK to enhance the activity of FoxO3, which finally reversed diabetes-induced EndMT. We conclude that MF attenuates DPF by inhibiting EndMT through the AMPK/FoxO3/SIRT3 axis. MF could be a potential candidate for the early prevention and treatment of DPF.