Published papers

The inhibition of the hedgehog (Hh) signaling pathway has emerged as an attractive anti-cancer strategy. As part of our continuing search for natural inhibitors of the Hh/GLI1 signaling pathway, we isolated three alkaloids (1-3) from Crinum asiaticum. Compounds 1 and 3 showed potent Hh/GLI1-mediated transcriptional inhibitory activity and exhibited cytotoxicity against human pancreatic (PANC1) and prostate (DU145) cancer cells. Our data revealed that compounds 1 and 3 clearly inhibited the Hh signaling pathway by down-regulating the expression of GLI-related proteins (PTCH and BCL2) in DU145 cells.

The aberrant hedgehog (Hh)/GLI signaling pathway causes the formation and progression of a variety of tumors. By screening tropical plant extracts by using our screening system, Zizyphus cambodiana was found to include Hh/GLI signaling inhibitors. Bioassay-guided fractionation of this plant extract led to the isolation of three active pentacyclic triterpenes, colubrinic acid (1), betulinic acid (2) and alphitolic acid (3), as potent inhibitors. The inhibition of GLI-related protein expression with 1 or 2 was observed in HaCaT cells with exogenous GLI1, or human pancreatic cancer cells (PANC1), which express Hh/GLI components aberrantly. The expressions of GLI-related proteins PTCH and BCL2 were clearly inhibited by 1 or 2. We also examined the cytotoxicity of these active compounds against PANC1, human prostate cancer cells (DU145) and mouse embryo fibroblast cells (C3H10T1/2). The cytotoxicity against cancer cells (PANC1 and DU145) by 1 or 2 would be caused by inhibition of the expression of the anti-apoptosis protein BCL2. These pentacyclic triterpene inhibitors showed an important relationship between Hh/GLI signaling inhibition, the decrease of BCL2, and cytotoxicity against cancer cells. (C) 2008 Elsevier Ltd. All rights reserved.

© 2019, The Japanese Society of Pharmacognosy. Upon screening compounds having Wnt signal inhibitory activity through evaluating TCF/β-catenin transcriptional (TOP) activity, eight cadinane sesquiterpenoids, including three new compounds (1–3), were isolated from wood extracts of Santalum album (Santalaceae). Structures of compounds 1–3 were elucidated by spectral data to have a cadinane skeleton with an aromatic ring. Of the eight compounds isolated, compound 4, identified as mansonone I, was found to be active against TOP, having an IC50 of 1.2 μM.

Copyright © 2018 American Chemical Society. A coculture method with a pathogenic actinomycete of the genus Nocardia and an animal cell line was designed to reconstruct and emulate the initial infection state, and a new cyclic nonapeptide, named nocarjamide (1), was obtained by coculture of Nocardia tenerifensis IFM 10554T and the mouse macrophage-like cell line J774.1 in a modified Czapek-Dox medium. Nocarjamide (1) exhibited Wnt signal-activating effects.

© 2017, The Japanese Society of Pharmacognosy and Springer Japan KK, part of Springer Nature. A new amide, named dehydropropylpantothenamide (1), was obtained by a co-culture of Nocardia tenerifensis IFM 10554T in the presence of the mouse macrophage-like cell line J774.1 in modified Czapek-Dox (mCD) medium. Compound 1 was synthesized from d-pantothenic acid calcium salt in 6 steps. The absolute configuration of natural compound 1 was determined by comparisons of the optical rotation and CD spectra of synthetic 1. In the present study, a new method for producing secondary metabolites was demonstrated using a “co-culture” in which the genus Nocardia was cultured in the presence of an animal cell line.

Screening with a cell-based luciferase assay was conducted to identify bioactive natural products which inhibit Wnt signaling activity-guided separation of an MeOH extract of Bauhinia malabarica (Caesalpiniaceae) leaves yielded five compounds, which were identified as beta-sitosterol (1), quercetin (2), 6,8-C-dimethyl kaempferol-3-O-rhamnopyranoside (3), hyperin (4), and 6,8-C-dimethyl kaempferol-3-methyl ether (5). The tested compounds 1, 3, and 5 exhibited Wnt signaling inhibitory activity, with IC50 values of 0.77, 0.74, and 16.6 mu M, respectively.

The myxomycetes (true slime molds) are an unusual group of primitive organisms that may be assigned to one of the lowest classes of eukaryotes. We studied the chemical constituents of a laboratory culture of plasmodia as well as field-collected fruit bodies of myxomycetes, and isolated various compounds with unique chemical structures and bioactivities. We recently also performed screening studies of natural products targeting signaling pathways related to basic studies of diseases and life sciences. Among myxomycetes constituents, we found several bioactive compounds that have effects on signaling pathways, including a cycloanthranilylproline with TRAIL resistance-overcoming activity, peptide lactone derivatives with Wnt signal inhibitory activity, a naphthoquinone pigment with Hes1 dimerization inhibition activity, and bisindole alkaloids with hedgehog signal inhibitory activity. Here we describe our recent results of their isolation, structure elucidation, synthesis, and findings in bioactivity studies.

Natural products from actinomycetes are important and valuable sources for drug discovery and the development of biological tools. The present review describes our recent study on the isolation of new natural products mainly possessing heterocyclic and aromatic ring structures with biological effects on cancer related cellular pathways such as tumor necrosis factor-alpha-related apoptosis-inducing ligand (TRAIL) and Wnt signaling.

TRAIL is a potent inducer of apoptosis in most cancer cells, but not in normal cells, and therefore has deserved intense interest as a promising agent for cancer therapy. In the search for bioactive natural products for overcoming TRAIL-resistance, we previously reported a number of active compounds. In our screening program on natural resources targeting overcoming TRAIL-resistance, activity-guided fractionation of the MeOH extract of Datura stramonium leaves led to the isolation of three alkaloids - scopolamine (1), trigonelline (2), and tyramine (3). Compounds 1, 2, and 3 exhibited TRAIL-resistance overcoming activity at 50, 150, and 100 mu M, respectively in TRAIL-resistant AGS cells.

The Wnt signal pathway modulates numerous biological processes, and its aberrant activation is related to various diseases. Therefore, inhibition of the Wnt signal may provide an effective (or efficient) strategy for these diseases. Cell-based luciferase assay targeting the Wnt signal (TOP assay) revealed that Hibiscus ficulneus extract inhibited the Wnt signal. The activity-guided isolation of the MeOH extract of H. ficulneus stems yielded four known (1-4) lignans along with myriceric acid (5). Compounds 1-4 potently inhibited the Wnt signal with TOPflash IC50 values of 1.0, 4.5, 6.3, and 1.9 mu M, respectively. Compound 1 exhibited cytotoxicity against both Wnt-dependent (HCT116) and Wnt-independent (RKO) cells. Western blot analysis showed that 1 decreased the expression of full, cytosolic and nuclear b-catenin along with c-myc in STF/293 cells. Our results suggested that 1 may have inhibited the Wnt signal by decreasing beta-catenin levels. (C) 2015 Elsevier Ltd. All rights reserved.

TRAIL is a promising anticancer agent due to its unique ability to kill tumor cells selectively without harming non-malignant cells. However, some cancer cells are reported to be TRAIL-resistant. Searching potent TRAIL agonist to overcome resistance is an important strategy for its successful use as an anticancer agent. As a part of our continuous research on natural inhibitors, activity-guided fractionation of the MeOH extract of Boesenbergia pandurata rhizomes afforded three (1-3) new isopimarane diterpenes, boesenberols I-K, together with two (4-5) known compounds. The structures of these new compounds were elucidated using HRESIMS, IR, 1D and 2D NMR spectroscopy. All compounds showed TRAIL-resistance overcoming activity. (C) 2017 Elsevier Ltd. All rights reserved.

TRAIL is a potent and selective inducer of apoptosis in most cancer cells while sparing normal cells, which makes it an attractive target for the development of new cancer therapies. In a screening program on natural resources with the ability to abrogate TRAIL resistance, the bioassay guided fractionation of Boesenbergia pandurata rhizomes resulted in the isolation of 17 pimarane diterpenes and a monoterpene. Among these, compounds 1-8, named boesenberols A-H, are new pimarane diterpenes. All compounds exhibited TRAIL-resistance-overcoming activity in TRAIL-resistant AGS cells. Subtoxic doses of the major compound 9 sensitized AGS cells to TRAIL-induced apoptosis by up-regulating apoptosis-inducing proteins, such as DR4, DRS, p53, Fas, CHOP, Bak, and cleaved caspases-3,-8, and -9, and down-regulating the levels of cell survival proteins, such as Bcl-2, c-FLIP, and GSK-3 beta, in TRAIL-resistant AGS cells. Furthermore, compound 9 did not decrease the viability of noncancerous (HEK293) cells at concentrations up to 30 mu M.

Wnt signaling plays key roles in embryonic development and various human diseases. Activity-guided testing to isolate Wnt signaling inhibitors from the methanol extract of Calotropis gigantea (Asclepiadaceae) exudutes identified six Wnt inhibitory cardenolides (1-6), of which 1, 3, 5, and 6 exhibited potent TCF/-catenin inhibitory activities (IC50 0.7-3.6 nM). Calotropin (1) inhibited Wnt signaling by decreasing both nuclear and cytosolic -catenin in a dose-dependent manner, and promoted degradation of -catenin by increasing the phosphorylation of -catenin at Ser45 through casein kinase 1 (CK1). Moreover, 1 significantly increased CK1 protein and mRNA levels. The results suggest that 1 inhibits the Wnt signaling pathway by increasing CK1 protein levels. To the best of our knowledge, calotropin is the first small molecule to increase CK1 levels.

A new bis-aporphine alkaloid, cerasoidine (1), was isolated from the root extract of Polyalthia cerasoides together with the known bis-aporphine bidebiline E (2) during screening for compounds with Wnt signal inhibitory activities. The structure of cerasoidine (1) was established by X-ray analysis and shown by chiral HPLC analyses and electronic circular dichroism to be a 57:43 mixture of R(-)- and S(+)-atropisomers. Bidebiline E (2) exhibited inhibition of transcriptional activity of TCF/beta-catenin with an IC50 value of 20.2 mu M and was also found to inhibit Wnt signaling by decreasing nuclear beta-catenin.