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.

Antiviral Effect of Stenocline ericoides DC. and Stenocline inuloides DC., Two Flavonoid-Rich Endemic Plants from Madagascar, against Dengue and Zika Viruses.

Dengue and Zika viruses are identified as the most medically important arthropod-borne viral pathogens. Over the past 20 years, the global dengue incidence has dramatically increased with epidemics of severe dengue where the case fatality rate can reach up to 20% in untreated patients. The association between Zika virus infection and severe congenital anomalies was first reported in 2015. Today no specific antiviral therapies are available for dengue and Zika virus infections, accentuating the need of adapted antiviral strategies based on medicinal plant drug discovery. Plants are a potential source of antiviral phytocompounds which act primarily by blocking virus entry in the host-cell. In the present study, we evaluated whether crude extracts from Stenocline ericoides DC. and Stenocline inuloides DC., two endemic plants from Madagascar, may have antiviral effects against dengue and Zika viruses. We showed that S. ericoides has virucidal action whereas S. inuloides inhibits the early steps of virus infection with a non-cytotoxic effect in human cells. The administration of S. ericoides and S. inuloides extracts in zebrafish had no effect on the behavior of animals at the active doses against dengue and Zika viruses, suggesting the absence of adverse effects at these doses. LC-HRMS2 and molecular networking analyses revealed the richness of these two plants in polyphenols and flavonoid with the presence of clusters of phytocompounds specific to each Stenocline species. Consequently, S. ericoides and S. inuloides represent potential sources for natural and safe antiviral phytocompounds against flaviviruses of medical concern.

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.

Drimane Derivatives as the First Examples of Covalent BH3 Mimetics that Target MCL-1.

Drimane sesquiterpenoid dialdehydes are natural compounds with antiproliferative properties. Nevertheless, their mode of action has not yet been discovered. Herein, we demonstrate that various drimanes are potent inhibitors of MCL-1 and BCL-xL, two proteins of the BCL-2 family that are overexpressed in various cancers, including lymphoid malignancies. Subtle changes in their structure significantly modified their activity on the target proteins. The two most active compounds are MCL-1 selective and bind in the BH3 binding groove of the protein. Complementary studies by NMR spectroscopy and mass spectrometry analyses, but also synthesis, showed that they covalently inhibit MCL-1 though the formation of a pyrrole adduct. In addition, cytotoxic assays revealed that these two compounds show a cytotoxic selectivity for BL2, a MCL-1/BCL-xL-dependent cell line and induce apoptosis.

Isolation of Picrotoxanes from Austrobuxus carunculatus Using Taxonomy-Based Molecular Networking.

A unique collection of 292 extracts from 107 New Caledonian Euphorbiaceae species sensu lato was profiled by LC-MS2 and the metabolite content organized by molecular networking. Based on the assumption that taxon-specific molecules are more likely to be structurally novel, taxonomic data were mapped on spectral networks to detect genus-specific clusters. Using this approach, a group of compounds unique to the genus Austrobuxus was highlighted. The subsequent MS-guided purification of the fruit EtOAc extract of Austrobuxus carunculatus led to the isolation of 13 new monolactone and "norditerpene" picrotoxanes (2-14), along with the known tutin (1). The structures of the new compounds were elucidated by HRESIMS and NMR spectroscopic data analysis, and the absolute configurations of compounds 1, 3, 7, 11, 12, and 14 were determined by single-crystal X-ray diffraction analysis. The relative and absolute configurations of compounds 4 and 5 were ascertained by chemical transformation of compound 3. The absolute configurations of other members of the series have been proposed on the basis of biogenetic considerations and specific rotation values of similar sign and magnitude. Compounds 1-14 were evaluated for their antiproliferative activities against HCT116 colon, U87-MG glioblastoma, and A549 lung human cancer cell lines. Compounds bearing an acyl chain at C-2 (i.e., 2, 4, and 13) showed IC50 values in the micromolar range for the three cell lines used.

N-myristoyltransferases inhibitory activity of ellagitannins from Terminalia bentzoë (L.) L. f. subsp. bentzoë.

N-myristoylation (Myr) is an eukaryotic N-terminal co- or post-translational protein modification in which the enzyme N-myristoyltransferase (NMT) transfers a fatty acid (C14:0) to the N-terminal glycine residues of several cellular key proteins. Depending on the cellular context, NMT may serve as a molecular target in anticancer or anti-infectious therapy, and drugs that inhibit this enzyme may be useful in the treatment of cancer or infectious diseases. As part of an on-going project to identify natural Homo sapiens N-myristoyltransferase 1 inhibitors (HsNMT1), two ellagitannins, punicalagin (1) and isoterchebulin (2), along with eschweilenol C (3) and ellagic acid (4) were isolated from the bark of Terminalia bentzoë (L.) L. f. subsp. bentzoë. Their structures were determined by means of spectroscopic analyses and comparison with literature data. Punicalagin (1) and isoterchebulin (2) showed significant inhibitory activity towards HsNMT1, and also against Plasmodium falciparum NMT (PfNMT) both in vitro and in cellulo, opening alternative paths for new NMT inhibitors development. This is the first report identifying natural products from a botanical source as inhibitors of HsNMT and PfNMT.

Endiandric acid analogues from Beilschmiedia ferruginea as dual inhibitors of Bcl-xL/Bak and Mcl-1/Bid interactions.

A rapid screening by (1)H and (1)H-(13)C HSQC NMR spectroscopy of EtOAc extracts of Endiandra and Beilschmiedia species allowed the selection of Beilschmiedia ferruginea leaves and flowers extract for a chemical investigation, leading to the isolation of 11 new tetracyclic endiandric acid analogues, named ferrugineic acids A-K (1-11). Their structures were determined by 1D and 2D NMR spectroscopic analysis in combination with HRMS data. These compounds were assayed for Bcl-xL and Mcl-1 binding affinities. Ferrugineic acids B, C, and J (2, 3, and 10) exhibited significant binding affinity for both antiapoptotic proteins Bcl-xL (Ki = 19.2, 12.6, and 19.4 µM, respectively) and Mcl-1 (Ki = 14.0, 13.0, and 5.2 µM, respectively), and ferrugineic acid D (4) showed only significant inhibiting activity for Mcl-1 (Ki = 5.9 µM).

Cytotoxic sesquiterpenoids from Winteraceae of Caledonian rainforest.

One secobutanolide, two butanolides and six drimane sesquiterpenoids were isolated from the bark and leaves of Zygogynum pancheri and Zygogynum acsmithii (Winteraceae) along with six known drimanes, isodrimanial, 1beta-O-p-methoxy-E-cinnamoyl-bemadienolide, 7-ketoisodrimenin, drimenin, polygodial and 1beta-E-cinnamoyl-6alpha-hydroxypolygodial. Their structures were elucidated through analysis of spectroscopic data. Drimane sesquiterpenoids with a dialdehyde function exhibited significant inhibitory activities in the in vitro cytotoxic assays against KB, HL60 and HCT116 cancer cell lines.