The Chemistry and Biology of Lycopodium Alkaloids.

Lycopodium alkaloids (LAs), a class of natural alkaloids have same biogenesis and similar structure characteristics, obtained from Lycopodiales. Which comprises 388 different species and these fascinating organisms have been identified as remnants of prehistoric ferns, with early fossils dating back as far as 300 million years. LAs usually are tricyclic or tetracyclic compounds with C16N or C16N2 skeleton. But then there are also have a few C11N, C15N, C15N2, C22N2, and C27N3 skeleton. LAs have attracted much scientific attention because of their important biological activities related to acetylcholinesterase and unique structural characteristics. From 1881 to December 2023, there are 593 LAs from 49 species of Lycopodiales have been reported. Because the total amount of LAs is nearly five times that of 1994, the classification and group allocation of some newly isolated LAs is often challenging and not unambiguous by Ayer's simple classification. This review makes a more systematic and detailed classification for it and provides extensive coverage of naturally occurring LAs discovered from 1881 to December 2023. Until now, there is no comprehensively summary of biological activity of the LAs. This review is the first time covered the biological activity of the all LAs.

Lycopodium alkaloids from Huperzia serrata and their cholinesterase inhibitory activities.

Huperzia serrata, belonging to the Lycopodiaceae family, has been traditionally utilized for the management of treating rheumatic numbness, arthritic pain, dysmenorrhea, and contusions. This plant is a rich source of lycopodium alkaloids, some of which have demonstrated notable cholinesterase inhibitory activity. The objective of this study was to identify lycopodium alkaloids with cholinesterase inhibitory properties from H. serrata. The structures of these alkaloids were elucidated by HRESIMS, NMR (including a 1H-15N HMBC experiment), ECD methods and single-crystal X-ray diffraction. The inhibitory activities against acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) were assessed using a modified Ellman's method. Consequently, sixteen lycopodium alkaloids (1-16), including ten previously undescribed ones named huperradines A-G and huperradines I-K (1-7 and 9-11), along with one previously undescribed naturally occurring compound, huperradine H (8), were isolated from H. serrata. Among these, compounds 7 and 1 exhibited potent and moderate AChE inhibition, with IC50 values of 0.876 ± 0.039 µM and 13.125 ± 0.521 µM, respectively. Our results suggest that huperradine G (7) may be a promising lead compound for the development of new AChE inhibitors for Alzheimer's disease.

Lycocasine A, a Lycopodium Alkaloid from Lycopodiastrum casuarinoides and Its Acid-Sensing Ion Channel 1a Inhibitory Activity.

A novel Lycopodium alkaloid, lycocasine A (1), and seven known Lycopodium alkaloids (2-8), were isolated from Lycopodiastrum casuarinoides. Their structures were determined through NMR, HRESIMS, and X-ray diffraction analysis. Compound 1 features an unprecedented 5/6/6 tricyclic skeleton, highlighted by a 5-aza-tricyclic[6,3,1,02,6]dodecane motif. In bioactivity assays, compound 1 demonstrated weak inhibitory activity against acid-sensing ion channel 1a.

In vitro and vivo anti-tumor activity and mechanisms of the new cryptotanshinone derivative 11 against hepatocellular carcinoma.

Global burden of hepatocellular carcinoma (HCC) is increasing. Chemotherapy and immunotherapy are the prevailing options for therapy. Developing new therapeutic strategies for HCC patients is still highly desirable. Recent studies demonstrate that cryptotanshinone is capable of inhibiting tumor growth in HCC and induces antitumor immunity in vitro. In our previous research, we discovered a new cryptotanshinone derivative 11 as an effective immunoregulatory enzyme indoleamine 2, 3-dioxygenase 1 (IDO1) inhibitor. This study aims to evaluate its in vitro and in vivo antitumor activity against hepatocellular carcinoma. 11 displayed robust anti-proliferative activity against HCC cell lines and promoted apoptosis of HCC cell line through the mitochondrial-mediated apoptotic pathway. In H22 tumor-bearing mice models, 11 exhibited significant in vivo anti-tumor activity with different administration routes. And no obvious toxicity was observed. RNA-seq analysis demonstrated the differential expressed genes and alteration of key pathways associated with immune responses after administration of 11. Up-regulation of anti-tumor cytokines and down-regulation of cytokines that promote tumor growth were indicated and further validated. Our study demonstrates that 11 exhibits promising anti-tumor activity both in vitro and in vivo against hepatocellular carcinoma cancer. It is a lead compound for HCC immunotherapy and is worthy for further development.

Taxodin A, a Cytotoxic C30 Terpenoid from Taxodium mucronatum with a Unique Skeleton.

Taxodin A (1), a unique C30 terpenoid featuring an unprecedented skeleton composed of an abietane-type diterpene and a menthane-type monoterpene, was obtained from the leaves and branches of Taxodium mucronatum. The structure and absolute configuration of compound 1 was unequivocally established by the combination of extensive spectroscopic analyses and X-ray single-crystal diffraction analysis. Compound 1 exhibited potent cytotoxic activities against A549, SMMC-7721, MDA-MB-231, and SW480 cell lines with IC50 values of 15.35±0.73, 8.49±0.35, 17.53±0.79, 18.93±0.60 µM, respectively.

Unified Strategy Enables the Collective Syntheses of Structurally Diverse Indole Alkaloids.

Natural products and their analogues are significant sources of therapeutic lead compounds. However, synthetic strategies for generating large collections of these molecules remain a significant challenge. The most difficult step in their synthesis is the design of a common intermediate that can be easily transformed into natural products belonging to different families. This study demonstrates the evolution of synthetic tactics designed to assemble the functionalized piperidines present in indole alkaloids from a common intermediate. More importantly, we also report a previously unknown Ir- and Er-catalyzed dehydrogenative spirocyclization reaction that enables direct access to spirocyclic oxindole alkaloids. As a practical application, the asymmetric total syntheses of 29 natural alkaloids belonging to different families were accomplished by following a uniform synthetic route. The proposed methodology extends the capability of the iridium-catalyzed dehydrogenative coupling reaction to the realm of indole-alkaloid synthesis and provides new opportunities for the efficient preparation of natural product-like molecules.

Discovery of α-Obscurine Derivatives as Novel Cav3.1 Calcium Channel Blockers.

Voltage-gated calcium channels (VGCCs), particularly T-type calcium channels (TTCCs), are crucial for various physiological processes and have been implicated in pain, epilepsy, and cancer. Despite the clinical trials of TTCC blockers like Z944 and MK8998, none are currently available on the market. This study investigates the efficacy of Lycopodium alkaloids, particularly as natural product-based TTCC blockers. We synthesized eighteen derivatives from α-obscurine, a lycodine-type alkaloid, and identified five derivatives with significant Cav3.1 blockade activity. The most potent derivative, compound 7, exhibited an IC50 value of 0.19±0.03 µM and was further analyzed through molecular docking, revealing key interactions with Cav3.1. These findings provide a foundation for the structural optimization of Cav3.1 calcium channel blockers and present compound 7 as a promising lead compound for drug development and a tool for chemical biology research.

Lycopodium Alkaloids from Huperzia goebelii.

One new fawcettimine-type Lycopodium alkaloid, hupertimine F (1), together with five known (2-6) Lycopodium alkaloids were isolated from Huperzia goebelii. The structure of 1 was elucidated by 1D and 2D NMR spectra, HRESIMS, and X-ray diffraction. Structurally, 1 represents the fourth example of Lycopodium alkaloids characterized by a 5/5/5/5/6 pentacyclic ring system with a 1-aza-7-oxabicyclo[2.2.1]heptane moiety. These known compounds 2, 3, 5, and 6 were isolated from H. goebelii for the first time. Compounds 1-6 were evaluated for acetylcholinesterase, butyrylcholinesterase and monoamine oxidase B inhibitory activities in vitro.

Casuattimines A-N, fourteen new Lycopodium alkaloids from Lycopodiastrum casuarinoides with Cav3.1 channel inhibitory activity.

Two new dimeric Lycopodium alkaloids, casuattimines A and B (1 and 2), along with twelve previously undescribed Lycopodium alkaloids, casuattimines C-N (3-14), and eight known Lycopodium alkaloids, were isolated from Lycopodiastrum casuarinoides. Casuattimines A and B (1 and 2) are the first two ether-linked Lycopodium alkaloid dimers. Casuattimines C and D (3 and 4) are unique Lycopodium alkaloids characterized by a long fatty acid chain. Structural elucidation was achieved through HRESIMS, NMR, and electronic circular dichroism (ECD) calculations. In addition, the absolute configurations of compounds 7, 13, and 14 were determined by single crystal X-ray diffraction. Compounds 1, 2, and 4 demonstrated notable Cav3.1 channel inhibitory activities presenting IC50 values of 10.75 ± 1.02 µM, 9.33 ± 0.79 µM, and 7.14 ± 0.86 µM, respectively. The dynamics of compound 4 against the Cav3.1 channel and preliminary structure-activity relationships of these active Lycopodium alkaloids were also discussed.

Discovery and Biomimetic Semisynthesis of Spirophyllines A-D from Uncaria rhynchophylla.

Spirophyllines A-D (1-4), four new spirooxindole alkaloids all characterized by the spiro[pyrrolidin-3,3'-oxindole] core and a rare isoxazolidine ring, were isolated from Uncaria rhynchophylla. Their structures were determined by spectroscopic methods and confirmed by X-ray crystallography. Based on the biomimetic semisynthesis strategy, compounds 1-8 were synthesized in three steps via the key reactions of 1,3-dipolar cycloaddition and Krapcho decarboxylation from corynoxeine. Interestingly, compound 3 showed moderate inhibitory activity against the Kv1.5 potassium channel (IC50 = 9.1 µM).

Structurally modified Cyclovirobuxine-D Buxus alkaloids as effective analgesic agents through Cav3.2 T-Type calcium channel inhibition.

Cyclovirobuxine-D (CVB-D) is a Buxus alkaloid and a major active constituent in the Chinese medicinal herb Buxus microphylls. Traditionally, the natural alkaloid cyclovirobuxine-D has a long history of use as a traditional Chinese medicine for cardiovascular diseases as well as to treat a wide variety of medical conditions. As we found that CVB-D inhibited T-type calcium channels, we designed and synthesized a variety of fragments and analogues and evaluated them for the first time as new Cav3.2 inhibitors. Compounds 2-7 exhibited potency against Cav 3.2 channels, and two of them were more active than their parent molecules. As a result of the in vivo experiments, both compounds 3 and 4 showed significantly reduced writhes in the acetic acid-induced writhing test. Studies of molecular modeling have identified possible mechanism(s) of Cav3.2 binding. Moreover, the relationship between structure and activity was studied in a preliminary manner. Our results indicated that compounds 3 and 4 could play an important role in the discovery and development of novel analgesics.

Iridoid Glycosides and Flavonoids Isolated from the Twigs and Leaves of Callicarpa nudiflora and Their Anti-Inflammatory Activities.

A new iridoid glycoside, named 6'-O-trans-feruloyl-8-epiloganic acid, together with fifteen known compounds were isolated from the twigs and leaves of Callicarpa nudiflora, a traditional Chinese medicine to treat inflammatory-related diseases. Their structures were identified by comprehensive spectroscopic analysis and comparison with reported data. Bioassay results revealed that twelve of the isolates could obviously inhibit nitric oxide (NO) production in lipopolysaccharide (LPS)-induced RAW 264.7 cell lines with IC50 values from 0.64 to 38.72 µM. Among them, compounds 1 (3.27 µM), 6 (5.23 µM), 13 (1.56 µM) and 14 (0.64 µM) exhibited significantly higher activities than that of the positive control (27.13 µM). Additionally, it was supposed that the presence of the carboxy group at the C-4 position of iridoid glycosides and glycosylation at C-3 position of flavonoids might impact their inhibitory activities against NO production.

Design, synthesis and biological evaluation of a new class of Hsp90 inhibitors vibsanin C derivatives.

Hsp90, an ATP-dependent chaperone that is essential for a wide range of protein assembly and folding processes, has long been recognized as a potential target for cancer. Hsp90 has more recently been identified as having a significant pathogenic role in viral infection, neurodegenerative disease, and inflammation, therefore, the development of the agents to inhibit the chaperone could potentially treat such intractable diseases. Here, on the basis of primary structure-activity relationships and docking analysis, a series of novel vibsanin C analogues with an emphasis on the C18 position was first designed, synthesized and biologically evaluated. The most effective Hsp90 inhibitory activity among these analogues was demonstrated by 29 and 31, with IC50 values of 0.39 and 0.27 µM respectively. Direct interaction between Hsp90 and its inhibitors were further confirmed. Mechanism studies indicated that 29 promoted HL-60 cell apoptosis by mitochondrial-mediated apoptosis pathway. In addition, 29 suppressed tumor growth in the H22 tumor-bearing mice model and revealed low acute toxicity in mice (LD50 > 500 mg/kg), suggesting its potential for further investigations.

Development and structure-activity relationships of tanshinones as selective 11ß-hydroxysteroid dehydrogenase 1 inhibitors.

11ß-Hydroxysteroid dehydrogenase 1 (11ß-HSD1) represents a promising drug target for metabolic syndrome, including obesity and type 2 diabetes. Our initial screen of a collection of natural products from Danshen led to the identification of tanshinones as the potent and selective 11ß-HSD1 inhibitors. To improve the druggability and explore the structure-activity relationships (SARs), more than 40 derivatives have been designed and synthesized using tanshinone IIA and cryptotanshinone as the starting materials. More than 10 derivatives exhibited potent in vitro 11ß-HSD1 inhibitory activity and good selectivity over 11ß-HSD2 across human and mouse species. Based on the biological results, SARs were further discussed, which was also partially rationalized by a molecular docking model of 1 bound to the 11ß-HSD1. Remarkably, compounds 1, 17 and 30 significantly inhibited 11ß-HSD1 in 3T3-L1 adipocyte and in livers of ob/ob mice, which merits further investigations as anti-diabetic agents. This study not only provides a series of novel selective 11ß-HSD1 inhibitors with promising therapeutic potentials in metabolic syndromes, but also expands the boundaries of the chemical and biological spaces of tanshinones.

Alisol B Alleviates Hepatocyte Lipid Accumulation and Lipotoxicity via Regulating RARα-PPARγ-CD36 Cascade and Attenuates Non-Alcoholic Steatohepatitis in Mice.

Non-alcoholic steatohepatitis (NASH) is a common chronic liver disease worldwide, with no effective therapies available. Discovering lead compounds from herb medicine might be a valuable strategy for the treatment of NASH. Here, we discovered Alisol B, a natural compound isolated from Alisma orientalis (Sam.), that attenuated hepatic steatosis, inflammation, and fibrosis in high-fat diet plus carbon tetrachloride (DIO+CCl4)-induced and choline-deficient and amino acid-defined (CDA)-diet-induced NASH mice. RNA-seq showed Alisol B significantly suppressed CD36 expression and regulated retinol metabolism in NASH mice. In mouse primary hepatocytes, Alisol B decreased palmitate-induced lipid accumulation and lipotoxicity, which were dependent on CD36 suppression. Further study revealed that Alisol B enhanced the gene expression of RARα with no direct RARα agonistic activity. The upregulation of RARα by Alisol B reduced HNF4α and PPARγ expression and further decreased CD36 expression. This effect was fully abrogated after RARα knockdown, suggesting Alisol B suppressed CD36 via regulating RARα-HNF4α-PPARγ cascade. Moreover, the hepatic gene expression of RARα was obviously decreased in murine NASH models, whereas Alisol B significantly increased RARα expression and decreased CD36 expression, along with the downregulation of HNF4α and PPARγ. Therefore, this study showed the unrecognized therapeutic effects of Alisol B against NASH with a novel mechanism by regulating RARα-PPARγ-CD36 cascade and highlighted Alisol B as a promising lead compound for the treatment of NASH.

Four Highly Oxygenated Sesquiterpenoids from the Fruits of Illicium micranthum Dunn.

Four highly oxygenated sesquiterpenoids, illimicranolides A (1) and B (2), and illicinolides E (3) and F (4), were obtained from the fruits of Illicium micranthum Dunn, as well as one known analog, illicinolide B (5). The chemical structures of 1-4 were determined comprehensively by 1D (1 H and 13 C) and 2D (HMBC, HSQC, 1 H-1 H COSY, and ROESY) NMR, and HR-ESI-MS data. Structurally, compound 1 was an unprecedented sesquiterpenoid with a 5/5/6/5-fused tetracyclic ring system and was the first seco-prezizaane sesquiterpenoid featuring a 11,8-γ-lactone ring. Compounds 3 and 4 were the fifth and sixth examples of illicinolide-type sesquiterpenoids. Moreover, compound 1 demonstrated neurotrophic activity of NGF-induced PC12 cells with differentiation rate of 10.34 % at a concentration of 10 µM.

Hupertimines A-E, Fawcettimine-Type Lycopodium Alkaloids from Huperzia serrata.

Five new fawcettimine-type Lycopodium alkaloids, hupertimines A-E (1-5), were discovered from the whole plant of Huperzia serrata, along with two known alkaloids, 8α-hydroxyphlegmariurine B (6) and 8ß-hydroxyphlegmariurine B (7). The structures of 1-7 were identified through HR-MS, IR, 1 H, 13 C, and 2D NMR, and single-crystal X-ray diffraction analysis. Structurally, compound 1 was the fourth example of Lycopodium alkaloid with an ether linkage between C-5 and C-13 and 2 was the third example of Lycopodium alkaloid with a 5/5/5/5/6 pentacyclic ring system and featuring a 1-aza-7-oxabicyclo[2.2.1]heptane unit. Compounds 1-7 were tested for their BACE1 inhibitory activity. In addition, the correct 1 H- and 13 C-NMR data for 7 were reported in current study.

Eleven undescribed alkaloids from the rhizomes of Sinomenium acutum and their IDO1 and TDO inhibitory activities.

Eleven previously undescribed alkaloids, named sinometumines A-K, along with three known alkaloids, were isolated from the rhizomes of Sinomenium acutum. The chemical structures of these unreported compounds were established using extensive spectroscopic methods (IR, UV, HRESIMS, and NMR), and their absolute configurations were determined by single crystal X-ray diffraction analyses and calculated electronic circular dichroism spectroscopy (ECD). Sinometumine D was the first aporphine-type derived alkaloid inner salt with a rearranged dibenzofuran ring backbone. Sinometumine E was a rare protoberberine-type alkaloid with a complex 6/6/6/6/6/6 hexacyclic skeleton. This was the first report of alkaloids with these two skeletons isolated from S. acutum. All isolates were evaluated for their inhibitory activities against indoleamine 2,3-dioxygenase 1 (IDO1) and tryptophan 2,3-dioxygenase (TDO). Lysicamine possessed noteworthy inhibitory activities as an IDO1/TDO dual inhibitor with IC50 values of 6.22 ± 0.26 µM and 23.76 ± 2.93 µM, respectively, and liriodenine revealed moderate dual inhibition with IC50 values of 31.65 ± 4.44 µM and 15.64 ± 0.26 µM. The intermolecular interactions and binding modes between lysicamine and IDO1/TDO were elaborated by molecular docking studies.

Spiroligustolides A and B: Two pairs of enantiomeric spiro-orthoester-containing phthalide dimers as Cav3.1 calcium channel inhibitors from Ligusticum Chuanxiong Hort.

Two pairs of unprecedented enantiomeric phthalide dimers, spiroligustolides A (1a/1b) and B (2a/2b), featuring a unique spiroorthoster linkage between two monomeric units to form a 5/6/5/6/6-fused ring system, were isolated from the roots of Ligusticum chuanxiong. The structures and relative configurations of 1 and 2 were determined by HR-ESI-MS, IR, and NMR spectroscopic data, coupled with single-crystal X-ray diffraction analysis, and the absolute configurations of 1a, 1b, 2a, and 2b were established by comparing the experimental and calculated electronic circular dichroism (ECD) data. Plausible biosynthetic pathway for 1 and 2 was proposed. Moreover, compounds 1, 1b, and 2b showed remarkable inhibitory activities on Cav3.1 calcium channel with IC50 values of 8.34, 7.08, and 8.60 µM, respectively.

Discovery and biological evaluation of tanshinone derivatives as potent dual inhibitors of indoleamine 2, 3-dioxygenase 1 and tryptophan 2, 3-dioxygenase.

Indoleamine 2, 3-dioxygenase 1 (IDO1) and tryptophan 2, 3-dioxygenase (TDO), catalyzing the first and rate-limiting step of tryptophan-kynurenine (Trp-Kyn) metabolism pathway, are the appealing targets for cancer immunotherapy. A few dual IDO1/TDO inhibitors are reported in literature. However, small-molecule IDO1 and TDO inhibitors are not yet available for clinical use. Here, we report synthetic analogues of the naturally occurring terpenoids tanshinone IIA and crytotanshinone, and their IDO1/TDO inhibitory activities using enzymatic and cellular assays. The most potent compound 30 was further characterized with respect to the direct interaction, inhibition kinetics, and different binding modes against IDO1 and TDO through surface plasmon resonance (SPR), enzyme kinetics, and spectroscopic analysis approaches, respectively. Preliminary mechanistic studies showed that 30 significantly promoted cell apoptosis through the potential mitochondria-mediated Bcl-2/Bax pathway. IDO1-overexpressing HeLa cells, mimicking cancer cells, were sensitive to 30 treatments. These results provide further insights for new clinical application of tanshinones, the main component of traditional herbal medicine.