New Cytotoxic Sesquiterpene Lactones from the Leaves of Tithonia diversifolia and their Apoptosis Effect Evaluation in KB Cancer Cells.

From the leaves of Tithonia diversifolia,  nine sesquiterpenoids (1-9), including two new ones (1, 2) were isolated and structurally determined. Their chemical structures were elucidated by extensive analyses of HRESIMS and NMR spectral data, as well as comparison with the literature. All of the isolated compounds (except compounds 7, 8, 9) significantly exhibited cytotoxic activity against four human cancer cell lines (KB, HepG2, A549 and MCF7), with IC50 values ranging from 0.29-17.0 µM,  which was in the same range as the positive control ellipticine or even lower. Further, the apoptosis induction of two new compounds 1 and 2 were also investigated and reported. While compound 2 did not induce cell apoptosis in KB cells at test concentrations, compound 1 was found to possess anti-proliferative activity through concentration-dependently inducing cell cycle arrest at S phase, morphological changes, activation of caspase 3, and an increase in the early-stage apoptosis of KB cells at a concentration of 7.26 µM.

Targeted Isolation of Antiviral Labdane Diterpenes from the Bark of Neo-uvaria foetida (Annonaceae) using LC-MS/MS-Based Molecular Networking.

In the search of new inhibitors for human coronavirus (HCoV), we screened extracts of endemic Annonaceae plants on an assay using a cellular model of Huh-7 cells infected with the human alphacoronavirus HCoV-229E. The EtOAc bark extract of the rare Southeast Asian plant Neo-uvaria foetida exhibited inhibition of HCoV-229E and SARS-CoV-2 viruses with IC50 values of 3.8 and 7.8 µg/mL, respectively. Using LC-MS/MS and molecular networking analysis guided isolation, we discovered two new labdane-type diterpenoids, 8-epi-acuminolide (1) and foetidalabdane A (4), and three known labdane diterpenoids, acuminolide (2), 17-O-acetylacuminolide (3), and spiroacuminolide (5). A new norlabdane diterpene, 16-foetinorlabdoic acid (6), was also isolated and identified. Excluding compounds 5 and 6, all other metabolites were active against the virus HCoV-229E. Terpenoids 1 and 4 presented antiviral activity against SARS-CoV-2 with IC50 values of 63.3 and 93.5 µM, respectively, indicating lower potency. Additionally, virological assays demonstrated that compounds 1, 2, and 3 exert antiviral effects against Zika virus by specifically interfering with the late stage of its infectious cycle with IC50 values of 76.0, 31.9, and 14.9 µM, respectively.

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.

Antiviral miliusanes and isolation of an unprecedented miliusane dimer from Miliusa balansae.

In an extensive screening endeavor for anti-coronaviral compounds, we examined 824 tropical plant extracts from the Annonaceae and Rutaceae families. The screening identified an ethyl acetate extract from the aerial parts of Miliusa balansae for its potent inhibitory activity against Human coronavirus HCoV-229E. Subsequent bioassay-guided fractionation of this extract revealed two unreported miliusanes including a complex dimeric structure and seven known compounds, comprising miliusane XXXVI, (+)-miliusol, bistyryls, styryl-pyranones, and the flavonoid rhamnetin. The absolute configuration of the new dimeric miliusane was determined by X-ray crystallography and a putative biogenetic origin was proposed. Investigation of the antiviral effect of these nine phytochemicals within HCoV-229E-infected Huh-7 cells showed that (+)-miliusol and miliusane XXXVI exert antiviral activity at non-cytotoxic concentrations, with IC50 values of 1.15 µM and 19.20 µM, respectively. Furthermore, these compounds significantly inhibited SARS-CoV-2 infection in Vero cells, presenting IC50 values of 11.31 µM for (+)-miliusol and 17.92 µM for miliusane XXXVI. Additionally, both compounds exhibited a potent antiviral effect against the emergent mosquito-borne Zika virus, with IC50 values of 1.34 µM and 23.45 µM, respectively. Time-of-addition assays suggest that their mechanism of action might target later stages of the viral cycle, indicating potential modulation of specific cellular pathways. These findings reinforce the invaluable contribution of medicinal flora as reservoirs of natural antiviral agents and emphasize their prospective role in combatting viruses of medical interest.

A Fast and Efficient Molecular Networking Approach for Reactivity of Natural Products Exploration in Plant Extracts: Application to Diterpene Esters from Euphorbia dendroides.

Natural products represent a rich source of bioactive compounds, covering a large chemical space. Even if challenging, this diversity can be extended by applying chemical modifications. However, these studies generally require multigram amounts of isolated natural products and face frequent testing failures. To overcome this limitation, we propose a rapid and efficient approach that uses molecular networking (MN) to visualize the new chemical diversity generated by simple chemical modifications of natural extracts. Moreover, the strategy deployed enables the most appropriate reagents to be defined quickly upstream of a reaction on a pure compound, in order to maximize chemical diversity. This methodology was applied to the latex extract of Euphorbia dendroides to follow the reactivity toward a series of Brønsted and Lewis acids of three class of diterpene esters identified in this species: jatrophane, terracinolide, and phorbol. Through the molecular networking interpretation, with the aim to illustrate our approach, BF3·OEt2 was selected for chemical modification on isolated jatrophane esters. Three rearranged compounds (3-5) were obtained, showing that the most appropriate reagents can be selected by MN interpretation.

Chemodiverse monoterpene indole alkaloids from Kopsia teoi, inhibitory potential against α-amylase, and their molecular docking studies.

Diabetes mellitus stands as a metabolic ailment marked by heightened blood glucose levels due to inadequate insulin secretion. The primary aims of this investigative inquiry encompassed the isolation of phytochemical components from the bark of Kopsia teoi, followed by the assessment of their α-amylase inhibition. The phytochemical composition of the K. teoi culminated in the discovery of a pair of new indole alkaloids; which are 16-epi-deacetylakuammiline N(4)-methylene chloride (akuammiline) (1), and N(1)-methoxycarbonyl-11-methoxy-12-hydroxy-Δ14-17-kopsinine (aspidofractinine) (2), together with five known compounds i.e. kopsiloscine G (aspidofractinine) (3), akuammidine (sarpagine) (4), leuconolam (aspidosperma) (5), N-methoxycarbonyl-12-methoxy-Δ16, 17-kopsinine (aspidofractinine) (6), and kopsininate (aspidofractinine) (7). All compounds were determined via spectroscopic analyses. The in vitro evaluation against α-amylase showed good inhibitory activities for compounds 5-7 with the inhibitory concentration (IC50) values of 21.7 ± 1.2, 34.1 ± 0.1, and 30.0 ± 0.8 µM, respectively compared with the reference acarbose (IC50 = 34.4 ± 0.1 µM). The molecular docking outputs underscored the binding interactions of compounds 5-7 ranging from -8.1 to -8.8 kcal/mol with the binding sites of α-amylase. Consequently, the outcomes highlighted the anti-hyperglycemic attributes of isolates from K. teoi.

Five undescribed aryltetralin lignans with cytotoxic activities from the fruits of Cleistanthus eberhardtii.

Five undescribed lignans, cleiseberharnins A-D (1-4), cleiseberharside A (5) were isolated from the fruits of Cleistanthus eberhartii (Phyllanthaceae), together with six known aryltetralin lignans, cleistantoxin (6), picroburseranin (7), neocleistantoxin (8), 7-hydroxypicropolygamain (9), cleisindoside D (10), and cleisindoside A (11). Their structures and relative configurations were established by analysis of HRESIMS and NMR data, and quantum chemical calculations of JH,H coupling constants. The absolute configurations of 1-5 were determined by analysis of their experimental CD spectra and comparison with calculated electronic circular dichroism (ECD) spectra. All compounds (1-11) were evaluated for their cytotoxicity against KB, MCF-7, HepG-2, and Lu-1 human cancer cell lines. Among the tested compounds, compounds 6 and 7 showed strong activity against KB, MCF7, HepG2 and Lu-1 cell lines with IC50 values in the range of 0.02-0.62 µM. Compound 1 showed activity against three cancer cell lines KB, HepG2, and Lu-1 with IC50 values of 6.98, 7.61 and 11.75 µM, respectively. Compound 2 exhibited a selective inhibition with moderate cytotoxicity against Lu-1 with IC50 value of 15.30 µM. Compounds 4, 5 and 9 showed moderate activity against the three cancer cell lines with IC50 values in the range of 8.73-19.70 µM.

Erythrocarpines IN, new limonoids from the barks of Chisocheton erythrocarpus and their neuroprotective effects against hydrogen peroxide in NG108-15 cells.

A phytochemical study on the bark of Chisocheton erythrocarpus Hiern (Meliaceae) has led to the isolation of six new phragmalin-type limonoids named erythrocarpines I - N (1-6) along with one known limonoid, erythrocarpine F (7). Their structures were fully characterized by spectroscopic methods. The pre-treatment of NG108-15 cells with 1-5, 7 (2 h) demonstrated low to good protective effects against H2O2 exposure; 1 (83.77% ± 1.84 at 12.5 µM), 2 (69.07 ± 2.01 at 12.5 µM), 3 (80.38 ± 2.1 at 12.5 µM), 4 (62.33 ± 1.95 at 25 µM),5 (58.67 ± 1.85 at 50 µM) and 7 (66.07 ± 2.03 at 12.5 µM). Interestingly, 1 and 3 demonstrated comparable protective effects to positive control epigallocatechin gallate (EGCG) with similar cell viability capacity (≈ 80%), having achieved that at lower concentration (12.5 µM) than EGCG (50 µM). Collectively, the results suggested the promising use of 1 and 3 as potential neuroprotective agents against hydrogen peroxide-induced cytotoxicity in neuronal model.

Two new flavonoids with antimicrobial activity from the roots of Byttneria aspera Colebr. ex Wall (Malvaceae).

Two new flavonoids, 4',5,7-trihydroxy-5'-methoxy-6,8-dimethylisoflavone (1) and 2',5',7-trihydroxy-5-methoxy-6,8-dimethylflavanone (2) together with the known flavonoids 4´,5,7-trihydroxy-3´-methoxy-6.8-dimethylflavone (3), epigallocatechin (4), 4´-O-methylepicatechin (5) and quercetin (6) were isolated from the roots of Byttneria aspera. The structures of these compounds were determined by means of spectroscopic methods. Compounds 1-6 were submitted to cytotoxic activity assays against three cancer cell lines including KB, MCF7 and A549, as well as their antimicrobial activity against a panel of clinically significant microorganisms. Compound 6 showed moderate cytotoxic activity with IC50 values of 12.7, 56.9 and 17.5 µM against KB, MCF7 and A549. Interestingly, the new compounds 1 and 2 exhibits antimicrobial activity, with compound 1 displaying selective antifungal activity against Candida albicans giving an MIC value of 128 µg/mL, compared to cyclohexamide with 32 µg/mL, while compound 2 shows potent inhibition of the Gram-positive bacterium Enterococcus faecalis displaying an MIC of 64 µg/mL, compared to streptomycin with 256 µg/mL.

Structure-Based Design of a Lead Compound Derived from Natural Schweinfurthins with Antitumor Properties That Target Oxysterol-Binding Protein.

Schweinfurthins (SWs) are naturally occurring prenylated stilbenes with promising anticancer properties. They act through a novel mechanism of action similar to that of other families of natural compounds. Their known target, oxysterol-binding protein (OSBP), plays a crucial role in controlling the intracellular distribution of cholesterol. We synthesized 15 analogues of SWs and demonstrated for the first time that their cytotoxicity as well as that of natural derivatives correlates with their affinity for OSBP. Through this extensive SAR study, we selected one synthetic analogue obtained in one step from SW-G. Using its fluorescence properties, we showed that this compound recapitulates the effect of natural SW-G in cells and confirmed that it leads to cell death via the same mechanism. Finally, after pilot PK experiments, we provided the first evidence of its in vivo efficacy in combination with temozolomide in a patient-derived glioblastoma xenograft model.

N-Methyl Costaricine and Costaricine, Two Potent Butyrylcholinesterase Inhibitors from Alseodaphne pendulifolia Gamb.

Studies have been conducted over the last decade to identify secondary metabolites from plants, in particular those from the class of alkaloids, for the development of new anti-Alzheimer's disease (AD) drugs. The genus Alseodaphne, comprising a wide range of alkaloids, is a promising source for the discovery of new cholinesterase inhibitors, the first-line treatment for AD. With regard to this, a phytochemical investigation of the dichloromethane extract of the bark of A. pendulifolia Gamb. was conducted. Repeated column chromatography and preparative thin-layer chromatography led to the isolation of a new bisbenzylisoquinoline alkaloid, N-methyl costaricine (1), together with costaricine (2), hernagine (3), N-methyl hernagine (4), corydine (5), and oxohernagine (6). Their structures were elucidated by the 1D- and 2D-NMR techniques and LCMS-IT-TOF analysis. Compounds 1 and 2 were more-potent BChE inhibitors than galantamine with IC50 values of 3.51 ± 0.80 µM and 2.90 ± 0.56 µM, respectively. The Lineweaver-Burk plots of compounds 1 and 2 indicated they were mixed-mode inhibitors. Compounds 1 and 2 have the potential to be employed as lead compounds for the development of new drugs or medicinal supplements to treat AD.

New Phenolic Lipids from the Leaves of Clausena harmandiana Inhibit SARS-CoV-2 Entry into Host Cells.

Induced by the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the COVID-19 pandemic underlined the clear need for antivirals against coronaviruses. In an effort to identify new inhibitors of SARS-CoV-2, a screening of 824 extracts prepared from various parts of 400 plant species belonging to the Rutaceae and Annonaceae families was conducted using a cell-based HCoV-229E inhibition assay. Due to its significant activity, the ethyl acetate extract of the leaves of Clausena harmandiana was selected for further chemical and biological investigations. Mass spectrometry-guided fractionation afforded three undescribed phenolic lipids (1-3), whose structures were determined via spectroscopic analysis. The absolute configurations of 1 and 2 were determined by analyzing Mosher ester derivatives. The antiviral activity against SARS-CoV-2 was subsequently shown, with IC50 values of 0.20 and 0.05 µM for 2 and 3, respectively. The mechanism of action was further assessed, showing that both 2 and 3 are inhibitors of coronavirus entry by acting directly on the viral particle. Phenolic lipids from Clausena harmandiana might be a source of new antiviral agents against human coronaviruses.

Metabolic adjustments in response to ATP spilling by the small DX protein in a Streptomyces strain.

ATP wasting is recognized as an efficient strategy to enhance metabolic activity and productivity of specific metabolites in several microorganisms. However, such strategy has been rarely implemented in Streptomyces species whereas antibiotic production by members of this genus is known to be triggered in condition of phosphate limitation that is correlated with a low ATP content. In consequence, to assess the effects of ATP spilling on the primary and specialized metabolisms of Streptomyces, the gene encoding the small synthetic protein DX, that has high affinity for ATP and dephosphorylates ATP into ADP, was cloned in the integrative vector pOSV10 under the control of the strong ErmE promoter. This construct and the empty vector were introduced into the species Streptomyces albogriseolus/viridodiastaticus yielding A37 and A36, respectively. A37 yielded higher biomass than A36 indicating that the DX-mediated ATP degradation resulted into a stimulation of A37 metabolism, consistently with what was reported in other microorganisms. The comparative analysis of the metabolomes of A36 and A37 revealed that A37 had a lower content in glycolytic and Tricarboxylic Acid Cycle intermediates as well as in amino acids than A36, these metabolites being consumed for biomass generation in A37. In contrast, the abundance of other molecules indicative either of energetic stress (ADP, AMP, UMP, ornithine and thymine), of activation (NAD and threonic acid) or inhibition (citramalic acid, fatty acids, TAG and L-alanine) of the oxidative metabolism, was higher in A37 than in A36. Furthermore, hydroxyl-pyrimidine derivatives and polycyclic aromatic polyketide antibiotics belonging to the angucycline class and thought to have a negative impact on respiration were also more abundantly produced by A37 than by A36. This comparative analysis thus revealed the occurrence in A37 of antagonistic metabolic strategies, namely, activation or slowing down of oxidative metabolism and respiration, to maintain the cellular energetic balance. This study thus demonstrated that DX constitutes an efficient biotechnological tool to enhance the expression of the specialized metabolic pathways present in the Streptomyces genomes that may include cryptic pathways. Its use thus might lead to the discovery of novel bioactive molecules potentially useful to human health.

Structure elucidation of an aspidofractinine-type monoterpene indole alkaloid from Melodinus reticulatus.

The structure and complete NMR assignments of aspidoreticulofractine, an aspidofractinine N-oxide, are reported. Its structure was elucidated based on a combination of spectroscopic techniques including 1D and 2D NMR, high-resolution mass spectrometry, and electronic circular dichroism.

In vitro evaluation of NA1-115-7-loaded nanoemulsions, an MCL-1-specific inhibitor of natural origin, intended to treat B-cell lymphoproliferative disorders after oral administration.

MCL-1, an anti-apoptotic member of the BCL-2 protein family, is overexpressed in many types of cancer and contributes to chemotherapy resistance. The drimane derivative NA1-115-7 is a natural compound isolated from Zygogynum pancheri that can be considered as a very promising lead for treating MCL-1-dependent hematological malignancies. As this drug suffers from low stability in acidic conditions and poor aqueous solubility, we evaluated the potential oral use of NA1-115-7 by encapsulating it in lipid nanoemulsions (NA-NEs) prepared by spontaneous emulsification. NA-NEs showed a particle size of 41.9 ± 2.2 nm, PDI of 0.131 ± 0.016, zeta potential of -5.8 ± 3.4 mV, encapsulation efficiency of approximately 100 % at a concentration of 24 mM. The stability of NA-1-115-7 was sixfold higher than that of the unencapsulated drug in simulated gastric fluid. NA-NEs significantly restored apoptosis and halved the effective doses of NA1-115-7 on BL2, a Burkitt lymphoma cell line, without toxicity in normal cells. Such a drug-delivery system appears to be particularly interesting for the oral administration of NA1-115-7, as it improves its solubility and stability, as well as efficacy, by reducing the therapeutic dose, making it possible to further consider in-vivo studies of this promising drug in BL2 xenografted mice.

Synthesis and Anti-Leishmanial Properties of Quinolones Derived from Zanthosimuline.

Quinoline derivatives and especially quinolones are considered as privileged structures in medicinal chemistry and are often associated with various biological properties. We recently isolated a series of original monoterpenyl quinolones from the bark of Codiaeum peltatum. As this extract was found to have a significant inhibitory activity against a Leishmania species, we decided to study the anti-leishmanial potential of this type of compound. Leishmaniasis is a serious health problem affecting more than 12 million people in the world. Available drugs cause harmful side effects and resistance for some of them. With the aim of finding anti-leishmanial compounds, we developed a synthetic strategy to access natural quinolones and analogues derived from zanthosimuline. We showed the versatility of this natural compound toward cyclization conditions, leading to various polycyclic quinolone-derived structures. The natural and synthetic compounds were evaluated against amastigote forms of Leishmania infantum. The results obtained confirmed the interest of this family of natural compounds but also revealed promising activities for some intermediates deriving from zanthosimuline. Following the same synthetic strategy, we then prepared 14 new analogues. In this work, we identified two promising molecules with good activities against intramacrophage L. infantum amastigotes without any cytotoxicity. We also showed that slight changes in amide functional groups affect drastically their anti-parasitic activity.

NA1-115-7, from Zygogynum pancheri, is a new selective MCL-1 inhibitor inducing the apoptosis of hematological cancer cells but non-toxic to normal blood cells or cardiomyocytes.

The overexpression of antiapoptotic members (BCL-2, BCL-xL, MCL-1, etc.) of the BCL-2 family contributes to tumor development and resistance to chemotherapy or radiotherapy. Synthetic inhibitors targeting these proteins have been developed, and some hematological malignancies are now widely treated with a BCL-2 inhibitor (venetoclax). However, acquired resistance to venetoclax or chemotherapy drugs due to an upregulation of MCL-1 has been observed, rendering MCL-1 an attractive new target for treatment. Six MCL-1 inhibitors (S64315, AZD-5991, AMG-176, AMG-397, ABBV-467 and PRT1419) have been evaluated in clinical trials since 2016, but some were affected by safety issues and none are currently used clinically. There is, therefore, still a need for alternative molecules. We previously described two drimane derivatives as the first covalent BH3 mimetics targeting MCL-1. Here, we described the characterization and biological efficacy of one of these compounds (NA1-115-7), isolated from Zygogynum pancheri, a plant belonging to the Winteraceae family. NA1-115-7 specifically induced the apoptosis of MCL-1-dependent tumor cells, with two hours of treatment sufficient to trigger cell death. The treatment of lymphoma cells with NA1-115-7 stabilized MCL-1, disrupted its interactions with BAK, and rapidly induced apoptosis through a BAK- and BAX-mediated process. Importantly, a similar treatment with NA1-115-7 was not toxic to erythrocytes, peripheral blood mononuclear cells, platelets, or cardiomyocytes. These results highlight the potential of natural products for use as specific BH3 mimetics non-toxic to normal cells, and they suggest that NA1-115-7 may be a promising tool for use in cancer treatment.

A new cytotoxic compound from the leaves of Styrax annamensis Guillaumin.

New propene derivative 1-(3',4'-methylenedioxyphenyl)-2-(2''-hydroxy-5-(3'''-hydroxypropyl)-3''-methoxyphenyl)prop-2-en-1-one (1), along with three known triterpenoids ursolic acid (2), pomolic acid (3), and maslinic acid (4) were isolated from the leaves of Styrax annamensis species. All structures were assigned by spectroscopic analysis. Compound 1 showed potent cytotoxicity against four cancer cell lines (KB, HepG2, Lu, and MCF7) with the IC50 values of 3.19, 2.87, 2.33, and 2.44 µM, respectively.

A new flavonoid from the leaves of Garcinia mckeaniana Craib and α-glucosidase and acetylcholinesterase inhibitory activities.

From the ethyl acetate extract (EtOAc) of the Vietnamese Garcinia mckeaniana leaves, a new flavone 8-C-glycoside 2'',6''-di-O-acetylvitexin (1), together with six known analogs 2-7 were isolated. Their structures were determined by spectral methods and compared with literature data. In α-glucosidase inhibitory assay, the EtOAc extract and its flavone and biflavone derivatives possessed the significant IC50 range of 9.17-97.53 µM, as compared with that of the positive control acarbose (249 µM). Flavones and biflavones showed are better than flavone glycosides in both α-glucosidase and acetylcholinesterase inhibitory activities[Formula: see text].

Cytotoxic and α-Glucosidase Inhibitory Xanthones from Garcinia mckeaniana Leaves and Molecular Docking Study.

A new racemic xanthone, garmckeanin A (1), and eight known analogs 2-9 were isolated from the ethyl acetate (AcOEt) extract of the Vietnamese Garcinia mckeaniana leaves. Their structures were determined by MS and NMR spectral analyses and compared with the literature. The AcOEt extract showed good cytotoxicity against cancer cell lines KB, Lu, Hep-G2 and MCF7, with IC50 values of 5.40-8.76 µg/mL, and it also possessed α-glucosidase inhibitory activity, with an IC50 value of 9.17 µg/mL. Garmckeanin A (1) exhibited inhibition of all cancer cell lines, with an IC50 value of 7.3-0.9 µM. Allanxanthone C (5) successfully controlled KB growth, with an IC50 value of 0.54 µM, higher than that of the positive control, ellipticine (IC50 1.22 µM). Norathyriol (8) was a promising α-glucosidase inhibitor, with an IC50 value of 0.07 µM, much higher than that of the positive control, acarbose (IC50 161.0 µM). The interactions of the potential α-glucosidase inhibitors with the C- and N-terminal domains of human intestinal α-glucosidase were also investigated by molecular docking study. The results indicated that bannaxanthone D (2), garcinone E (4), bannaxanthone E (6), and norathyriol (8) exhibit higher binding affinity to the C-terminal than to the N-terminal domain through essential residues in the active sites. In particular, compound 8 could be assumed to be the most potent mixed inhibitor.