Keiichi Tabata

Cytotoxic and apoptosis-inducing activities of triterpene acids from Poria cocos.

Six lanostane-type triterpene acids (1a-6a), isolated from Poria cocos , and their methyl ester (1b-6b) and hydroxy derivatives (1c-6c) were prepared. Upon evaluation of the cytotoxic activity of these compounds against leukemia (HL60), lung (A549), melanoma (CRL1579), ovary (NIH:OVCAR-3), breast (SK-BR-3), prostate (DU145), stomach (AZ521), and pancreas (PANC-1) cancer cell lines, 11 compounds (5a, 6a, 2b-5b, 1c, and 3c-6c) exhibited activity with single-digit micromolar IC(50) values against one or more cell lines. Poricotriol A (1c), a hydroxy derivative of poricoic acid A (1a), exhibited potent cytotoxicities against six cell lines with IC(50) values of 1.2-5.5 μM. Poricotriol A induced typical apoptotic cell death in HL60 and A549 cells on evaluation of the apoptosis-inducing activity by flow cytometric analysis. Western blot analysis in HL60 cells showed that poricotriol A activated caspases-3, -8, and -9, while increasing the ratio of Bax/Bcl-2. This suggested that poricotriol A induced apoptosis via both mitochondrial and death receptor pathways in HL60. On the other hand, poricotriol A did not activate caspases-3, -8, and -9, but induced translocation of apoptosis-inducing factor (AIF) from mitochondria and increased the ratio of Bax/Bcl-2 in A549. This suggested that poricotriol A induced apoptosis via the mitochondrial pathway mostly by translocation of AIF, independent from the caspase pathway in A549. Furthermore, poricotriol A was shown to possess high selective toxicity in lung cancer cells since it exhibited only weak cytotoxicity against a normal lung cell line (WI-38).

Cytotoxic and Apoptosis-Inducing Activities of Limonoids from the Seeds of Azadirachta indica (Neem)

Thirty-five limonoids, including 15 of the azadiradione type (1-15), five of the gedunin type (16-20), four of the azadirachtin type (21-24), nine of the nimbin type (25-33), and two degraded limonoids (34, 35), isolated from Azadirachta indica seed extracts, were evaluated for their cytotoxic activities against five human cancer cell lines. Seven compounds (3, 6, 7, 16, 18, 28, and 29) exhibited cytotoxic activity against one or more cell lines. Among these compounds, 7-deacetyl-7-benzoylepoxyazadiradione (7), 7-deacetyl-7-benzoylgeduin (18), and 28-deoxonimbolide (28) exhibited potent cytotoxic activity against HL60 leukemia cells with IC(50) values in the range 2.7-3.1 μM. Compounds 7, 18, and 28 induced early apoptosis in HL60 cells, observed by flow cytometry. Western blot analysis showed that compounds 7, 18, and 28 activated caspases-3, -8, and -9 in HL60 cells. This suggested that compounds 7, 18, and 28 induced apoptotic cell death in HL60 cells via both the mitochondrial- and the death receptor-mediated pathways. Futhermore, compound 7 was shown to possess high selective cytotoxicity for leukemia cells since it exhibited only weak cytotoxicity against a normal lymphocyte cell line (RPMI 1788).

Survivin as a prognostic factor for osteosarcoma patients.

Purpose: Survivin is one of the apoptosis inhibitor genes and is rarely expressed in adult tissues. However, survivin expression has been detected in various human cancers and correlations have been recognized between the level of expression of this gene in tumors and prognosis. In this study, we investigated the correlations between survivin mRNA expression in osteosarcoma tissues and clinicopathological parameters. Methods: There were 22 osteosarcoma patients in our hospital with paraffin-embedded tissues which could be extracted from biopsy specimens. We used the RT-PCR method after extracting total RNA and conducted a densitometric analysis to determine the ratio of survivin relative to h-GAPDH as an internal marker. Results: Expression of survivin mRNA was detected in all osteosarcoma samples. Patients with metastasis had high survivin mRNA levels in initial biopsy specimens (p<0.01). Moreover, there was a statistically significant difference in survivin mRNA expression between patients with and without metastasis (p<0.01). Conclusion: We concluded that high levels of survivin mRNA expression suggest poor prognosis for osteosarcoma patients.

Effects of Uncaria tomentosa Total Alkaloid and its Components on Experimental Amnesia in Mice: Elucidation Using the Passive Avoidance Test

The effects of Uncaria tomentosa total alkaloid and its oxindole alkaloid components, uncarine E, uncarine C, mitraphylline, rhynchophylline and isorhynchophylline, on the impairment of retention performance caused by amnesic drugs were investigated using a step‐down‐type passive avoidance test in mice. In this test, the retention performance of animals treated with the amnesic and test drugs before training was assessed 24 h after training.

New Cytotoxic Diarylheptanoids from the Rhizomes of Alpinia officinarum

Bioassay-guided fractionation of the cytotoxic MeOH extract from the rhizomes of Alpinia officinarum Hance led to the isolation of two new diarylheptanoids named alpinoid D (1) and E (2), together with fifteen known diarylheptanoids (3 - 17). The structures of compounds 1 and 2 were elucidated on the basis of spectroscopic data and chemical evidence. The cytotoxic activity of the isolated diarylheptanoids was evaluated against the IMR-32 human neuroblastoma cell line. Among the tested compounds, 11, 12 and 14 exhibited the most potent activities with IC (50) values of 0.83, 0.23 and 0.11 microM, respectively.

Acute and chronic intracerebroventricular morphine infusions affect long-term potentiation differently in the lateral perforant path.

Experiments were performed to investigate the effects of acute and chronic intracerebroventricular (icv) morphine infusions via osmotic minipumps on long-term potentiation (LTP) in the lateral perforant path (LPP)-granule cell synapse of the rat dentate gyrus. Although significant antinociceptive activity was observed when morphine was infused (25 nmol/microl/h) for 30 min or 1 h, the activity was not observed in rats receiving morphine chronically for 72 h, and the tail-flick latency of this group was comparable to that of rats receiving saline. LTP induction was significantly attenuated after acute morphine infusion (1 h) in LPP-granule cell synapses of the dentate gyrus. In contrast, LTP induction was augmented after chronic morphine infusion for 72 h. Bath-perfused morphine augmented the baseline population spike (PS) amplitude in rats treated with saline, whereas it attenuated the LTP induced by chronic morphine infusion. Returning the LTP to the level of saline infusion after in vitro morphine perfusion suggests that enhancement of the LTP is a withdrawal-like phenomenon. These results suggest a difference between the effects of acute and chronic intracerebroventricular morphine infusions on synaptic plasticity in the LPP-granule cell synapses of the dentate gyrus.

Pharmacological Profile of Novel Acid Pump Antagonist 7-(4-Fluorobenzyloxy)-2,3-dimethyl-1-{[(1S,2S)-2-methyl cyclopropyl]methyl}-1H-pyrrolo[2,3-d]pyridazine (CS-526)

The pharmacological profiles of the novel acid pump antagonist 7-(4-fluorobenzyloxy)-2,3-dimethyl-1-{[(1S,2S)-2-methylcyclopropyl]methyl}-1H-pyrrolo[2,3-d]pyridazine (CS-526) were investigated in terms of hog gastric H+,K+-ATPase activity, gastric acid secretion, and acute gastroesophageal lesions in comparison with other proton pump inhibitors (PPIs). CS-526 inhibited H+,K+-ATPase activity in a concentration-dependent manner, with an IC50 value of 61 nM. The inhibitory effect of CS-526 on H+,K+-ATPase activity was more potent than that of any of the other PPIs examined. The inhibitory mechanism of CS-526 on H+,K+-ATPase was a competitive antagonism to the K+ binding site of H+,K+-ATPase, and it was also a reversible inhibition. In pylorus-ligated rats, intraduodenal or oral administration of CS-526 inhibited gastric acid secretion in a dose-dependent manner, and the ID50 values were 2.8 or 0.7 mg/kg, respectively. In Heidenhain pouch dogs, intrapouch administration of CS-526 inhibited histamine-stimulated gastric acid secretion in a dose- and retention time-dependent manner. In a reflux esophagitis model, intraduodenal and oral administration of CS-526 prevented esophageal lesions with ID50 values of 5.4 and 1.9 mg/kg, respectively. Lansoprazole prevented esophagitis only by intraduodenal administration (ID50 = 2.2 mg/kg). Furthermore, CS-526 inhibited acute gastric mucosal lesions. These data demonstrate that the novel acid pump antagonist CS-526 has potent antisecretory and antiulcer effects. These findings indicate that CS-526 would have a curative effect on gastroesophageal reflux disease via its potent antisecretory and antiulcer actions.

Kuguaglycoside C, a constituent of Momordica charantia, induces caspase‐independent cell death of neuroblastoma cells

Kuguaglycoside C is a triterpene glycoside isolated from the leaves of Momordica charantia, and the biological effects of this compound remain almost unknown. We investigated the anti;?>cancer effect of kuguaglycoside C against human neuroblastoma IMR‐32 cells. In the MTT assay, kuguaglycoside C induced significant cytotoxicity against the IMR‐32 cells (IC50: 12.6 μM) after 48 h treatment. Although examination by Hoechst 33342 staining revealed that kuguaglycoside C induced nuclear shrinkage at a high concentration (100 μM), no apoptotic bodies were observed on flow cytometry. No activation of caspase‐3 or caspase‐9 was observed at the effective concentration (30 μM) of kuguaglycoside C. On the other hand, the substance significantly decreased the expression of survivin and cleaved poly (ADP‐ribose) polymerase (PARP). Kuguaglycoside C also significantly increased the expression and cleavage of apoptosis‐inducing factor (AIF). Moreover, kuguaglycoside C was found to induce caspase‐independent DNA cleavage in the dual‐fluorescence apoptosis detection assay. These results suggest that kuguaglycoside C induces caspase‐independent cell death, and is involved, at least in part, in the mechanism underlying cell necroptosis.

Antitumor-promoting effects and cytotoxic activities of dammar resin triterpenoids and their derivatives.

Nineteen known triterpenoids, 1–19, and one known sesquiterpenoid, 20, were isolated from dammar resin obtained from Shorea javanica K. & V. (Dipterocarpaceae). One of the acidic triterpenoids, dammarenolic acid (1), was converted to fourteen derivatives, namely, an alcohol, 21, an aldehyde, 22, and twelve L‐amino acid conjugates, 23–34. Compounds 1–34 were examined for their inhibitory effects on the induction of Epstein–Barr virus early antigen (EBV‐EA) by 12‐O‐tetradecanoylphorbol 13‐acetate (TPA) in Raji cells, a known primary screening test for antitumor promoters. All of the compounds tested, except for compounds 4, 5, 12–14, 16, and 17, showed inhibitory effects against EBV‐EA activation with potencies either comparable with or stronger than that of β‐carotene, a known natural antitumor promoter. In addition, (20S)‐20‐hydroxy‐3,4‐secodammara‐4(28),24‐dien‐3‐al (22) exhibited inhibitory effects on skin tumor promotion in an in vivo two‐stage mouse skin carcinogenesis test based on 7,12‐dimethylbenz[a]anthracene (DMBA) as initiator, and with TPA as promoter. Furthermore, evaluation of the cytotoxic activities of compounds 1–34 against human cancer cell lines showed that reduction (i.e., 21 and 22) or conjugation with L‐amino acids (i.e., 23–34) of compound 1 enhanced the cytotoxicity against human melanoma cell line CRL1579.

Delayed healing of chronic gastric ulcer after Helicobacter pylori infection in mice

It has been suggested that there is a close relationship between Helicobacter pylori and the onset or recurrence of gastroduodenal disease. The aim of this study was to examine the effect of H. pylori on the healing of chronic gastric ulcers induced in mice.