Clock and ATF4 transcription system regulates drug resistance in human cancer cell lines.

The mechanisms underlying cellular drug resistance have been extensively studied, but little is known about its regulation. We have previously reported that activating transcription factor 4 (ATF4) is upregulated in cisplatin-resistant cells and plays a role in cisplatin resistance. Here, we find out a novel relationship between the circadian transcription factor Clock and drug resistance. Clock drives the periodical expression of many genes that regulate hormone release, cell division, sleep-awake cycle and tumor growth. We demonstrate that ATF4 is a direct target of Clock, and that Clock is overexpressed in cisplatin-resistant cells. Furthermore, Clock expression significantly correlates with cisplatin sensitivity, and that the downregulation of either Clock or ATF4 confers sensitivity of A549 cells to cisplatin and etoposide. Notably, ATF4-overexpressing cells show multidrug resistance and marked elevation of intracellular glutathione. The microarray study reveals that genes for glutathione metabolism are generally downregulated by the knockdown of ATF4 expression. These results suggest that the Clock and ATF4 transcription system might play an important role in multidrug resistance through glutathione-dependent redox system, and also indicate that physiological potentials of Clock-controlled redox system might be important to better understand the oxidative stress-associated disorders including cancer and systemic chronotherapy.

Preventive effects of saponins from the Pueraria lobata root on in vitro immunological liver injury of rat primary hepatocyte cultures.

Preventive effects of nine saponins obtained from Puerariae Radix (the roots of Pueraria lobata) on in vitro immunological liver injury in primary cultured rat hepatocytes were studied. Although all tested saponins showed hepatoprotective action, the levels of activity of individual saponins differed. Structure-activity relationships for the sapogenol moiety suggested that the hydroxy group at C-29 would reduce the hepatoprotective activity while the hydroxy group at C-21 could enhance the hepatoprotective activity. Furthermore, structure-activity relationships for the sugar moiety suggested that the oxygen-bearing group at C-5" would enhance the hepatoprotective activity, though the configuration of the hydroxy group at C-3" could be less important for hepatoprotective activity.

Structure-hepatoprotective relationships study of soyasaponins I-IV having soyasapogenol B as aglycone.

As a part of our study on the leguminous plants, we investigated the constituents of the aerial parts of Glycine soya. We isolated and identified four known saponins, soyasaponins I, II, III, and IV which have the same aglycone, soyasapogenol B. As a part of our studies concerning hepatoprotective drugs, we also examined the hepatoprotective actions of these saponins towards immunologically induced liver injury on primary cultured rat hepatocytes. The action of soyasaponin II was almost comparable with that of soyasaponin I, whereas those of soyasaponin III and IV were more effective than soyasaponins I and II. This means that the disaccharide group shows greater action than the trisaccharide group. Furthermore, the saponin having a hexosyl unit shows a slightly greater action than that of the pentosyl unit in each disaccharide group or trisaccharide group. Structure-activity relationships suggest that the sugar moiety linked at C-3 may play an important role in hepatoprotective actions of soybean saponins.

Kaikasaponin III and soyasaponin I, major triterpene saponins of Abrus cantoniensis, act on GOT and GPT: influence on transaminase elevation of rat liver cells concomitantly exposed to CCl4 for one hour.

The antihepatotoxic activities of soyasaponin I and kaikasaponin III, triterpenoidal saponins isolated from Abri Herba, the whole plant of Abrus cantoniensis, were studied on liver injury induced by CCl4 in primary cultured rat hepatocytes. The antihepatotoxic activities of these saponins and glycyrrhizin (positive control) were demonstrated by measuring the levels of glutamic pyruvic transaminase (GPT) and glutamic oxaloacetic transaminase (GOT). Soyasaponin I inhibited the elevation of GOT and GPT activities. The activities were comparable to those of glycyrrhizin. On the other hand, kaikasaponin III was more effective than soyasaponin I and glycyrrhizin. Kaikasaponin III showed the antihepatotoxic activity at less than 100 micrograms/ml. Furthermore, the highest activity was observed even in the lower doses (50, 100 micrograms/ml). However, soyasaponin I and kaikasaponin III showed some toxicity at the highest dose (500 micrograms/ml), though glycyrrhizin did not show toxicity at any dose.

Preventive effects of saponins from puerariae radix (the root of Pueraria lobata Ohwi) on in vitro immunological injury of rat primary hepatocyte cultures.

The preventive effects of saponins from Puerariae Radix toward in vitro immunological liver injury using an antiserum against the rat liver plasma membranes on primary cultured rat hepatocytes were studied. Crude saponin from Puerariae Radix inhibited the elevation of alanine aminotransferase (ALT) activity at the dose of 90 micrograms/ml. The inhibition was stronger than that of glycyrrhizin, which was a positive control drug. The representative saponins in this drug, soyasaponin I and kudzusaponin SA3, were also more effective than glycyrrhizin, although their effects were weaker than that of crude saponin at the lower doses (90, 200 micrograms/ml). At 500 micrograms/ml, kudzusaponin SA3 showed antihepatotoxic activity equal to that of crude saponin.

Oleanene-type triterpene glycosides from puerariae radix. IV. Six new saponins from Pueraria lobata.

From Puerariae Radix, the root of Pueraria lobata (Leguminosae), six new oleanene-type triterpene glycosides, called kudzusaponins A1 (1), A2 (2), Ar (3), SA4 (5), and SB1 (6) were isolated together with kudzusaponin A3 (7), soyasaponins SA3 (8), and I (9). The structures of 1-6 were determined to be 3-O-alpha-L-rhamnopyranosyl- (1-->2)-beta-D-arabinopyranosyl-(1-->2)beta-D-glucuronopyranosy l kudzusapogenol A 22-O-beta-D-xylopyranoside, 3-O-beta- D-galactopyranosyl-(1-->2)-beta-D-glucuronopyranosyl kudzusapogenol A, 3-O-beta-D-glucopyranosyl-(1-->2)-beta-D-glucuronopyranosyl kudzusapogenol A, 3-O-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-glucopyranosyl-(1-->2)-beta- D-glucoronopyranosyl kudzusapogenol A, 3-O-beta-D-glucuronopyranosyl[(1-->2)-beta-D-glucuronopyranosyl soyasapogenol A22-O-alpha-L-arabinopyranoside,and 3-O-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-galactopyranosyl- (1-->2)-beta- D-glucuronopyranosyl (beta-fabatriosyl) soyasapogenol B 22-O-alpha-L-arabinopyranoside, respectively.

Oleanene-type triterpene glycosides from puerariae radix. III. Three new saponins from Pueraria thomsonii.

From the root of Pueraria thomsonii (Leguminosae), three new oleanane-type triterpene glycosides, named kudzusaponin B1, acetyl-kaikasaponin III and acetyl-soyasaponin I were isolated, together with soyasaponin I (4) and subproside V (5). Their structures were determined to be 3-O-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-galactopyranosyl-(1-->2) -beta-D-glucuronopyranosyl kudzusapogenol B (1), 3-O-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-galactopyranosyl-(1-->2) -beta-D-glucuronopyranosyl sophoradiol 22-O-acetate (2) and 3-O-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-galactopyranosyl-(1-->2) -beta-D-glucuronopyranosyl soyasapogenol B 22-O-acetate (3), respectively.

Oleanene-type triterpene glycosides from Puerariae Radix. II. Isolation of saponins and the application of tandem mass spectrometry to their structure determination.

A continuing study of the ingredients of Puerariae Radix, the roots of Pueraria lobata (WILLD.) OHWI, which is one of the most important crude drugs, has resulted in the first isolation of four new oleanene-type triterpene glycosides, named kudzusaponins SA1, SA2, SA3 and C1 (1-4). Their structures were determined to be 3-O-beta-D-galactopyranosyl-(1-->2)-beta-D-glucuronopyranosyl soyasapogenol A (1), 3-O-beta-galactopyranosyl-(1-->2)-beta-D-glucuronopyranosyl soyasapogenol A 22-O-alpha-L-arabinopyranoside (2), 3-O-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-galactopyranosyl-(1-->2)-D- glucuronopyranosyl kudzusapogenol C 21-O-beta-D-glucopyranoside (4), respectively. The usefulness of tandem mass spectrometry in the structural determination of oleanene-type triterpene bisdesmosides is also discussed.