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treated with PQS. Cell viability assays, flow cytometric analysis of the cell cycle and FACS based apoptosis assays were performed after treatment. Western blot and quantitative real-time PCR analyses were performed to demonstrate the expression level of multiple cancer-related genes such as p53, p21, STAT3, bcl2, DRD2 and FANCD2 as well as TMEM79. Microarray-based gene expression analysis was used to identify prostate specific gene expression patterns and to explore novel biomarkers for potential clinical use in prostate cancer diagnostics.
The inhibitory effects on the proliferation of DU145 showed that PQS (>150 ^g/ml) inhibited the
viability of human prostate cancer cells. Cell cycle arrest assay showed that PQS induced cell cycle arrest at the G0/G1 phase. PQS treatment up-regulated the expression of p53, p21 and TMEM79, down-regulated the expression of DRD2, FANCD2, STAT3 and bcl2. The transmembrane protein TMEM79 was found to be highly expressed in normal prostate cells but lowly expressed in prostate cancer cells.
Our results indicated that TMEM79 could be a novel biomarker candidate for prostate cancer diagnosis. PQS might be an effective herbal remedy playing a complementary or alternative role for treating prostate cancer in the future.
ENZYME INHIBITORY ACTIVITIES OF PLANT EXTRACTS UNDER THE PHILIPPINE'S DRUG DISCOVERY PROGRAM
© Leomel Argulla, Jose Manuel Gutierrez, Mavis Colleen P. Fabian, Joseph Sasotona, Jasmin Tutor and Christine C. Hernandez
Institute of Chemistry, University of the Philippines, Diliman
The Philippines has more than 20,000 endemic species and it is one of the 17 countries that contain two-thirds of the entire biological diversity of the Earth. The country's rich collection of endemic and native plant species presents is a valuable source of plant-derived bioactive agents for human ailments like hypertension, uricosuria, asthma, and gastric ulcer.
The country has a high incidence of hypertension, asthma, and gout. According to the second National Nutrition and Health Survey, the prevalence of asthma in adult Filipinos is relatively high. Inhibition of lipoxygenase (LOX) may diminish leukotriene- mediated inflammatory response and other respiratory conditions particulary asthma [1]. Hypertension is still a growing health problem in the Philippines. If left untreated, it will lead to more serious health problems like cardiovascular diseases. One way of lowering blood pressure is the use of angiotensin converting enzyme (ACE) inhibitors [2]. Gout, while not necessarily a cause of death, causes episodic and severe pain in the joints due to increase in uric acid. Around 1.6 million Filipinos are suffering from gout. Xanthine oxidase (XO) catalyzes the oxidation of hypoxanthine to xanthine and of xanthine to uric acid [3]. Overproduction of uric acid leads to condition of hyperuricemia which is linked to gout [4].
Plants were collected from the different regions of the Philippines. Plants used for this study are those with reported use as a traditional medicine or those plants that are endemic in a region. Plant extracts were collected and were screened to test their XO, ACE and LOX inhibitory activities. The activity of 15-lipoxygenase is observe as it catalyzed the reaction between oxygen and linoleic acid. The increase in absorbance at 234 nm is due to the formation of the product 13-hydroperoxyoctadecadienoic acid from the reaction
of oxygen and linoleic acid. The xanthine oxidase inhibitory activity was assessed spectrophotometrically using a 96-well plate reader under aerobic conditions. Absorption increments were monitored every 30 seconds for 10 minutes at 295 nm to monitor the rate of formation of uric acid. Allopurinol was used as positive control and xanthine oxidase inhibitory activity was expressed as percentage inhibition. A colorimetric method was optimized in a 96 well quartz microplate to determine the inhibitory action of the extracts on ACE. This assay was based on the amount of hippuric acid (HA) released by the action of ACE on histidine-histidyl-leucine (HHL) substrate. The released HA from the substrate is mixed with pyridine and BSC resulting to a yellow colored complex with A max at 410nm.
Plant extracts that showed > 50% inhibition in all trials were considered active. Among the 2,395 extracts tested, 420 (18%) were active for anti-gout activity, 910 (38%) were active for anti-hypertension activity, 358 (15%) were active for anti-inflammation activity. Tradescantia fluminensis inhibited LO at 87.2%. T. fluminensis was partitioned with ethyl acetate and hexane and their IC50 values were determined at 8.72^g/ml and 98.04 ^g/ml, respectively [5]. A pure compound isolated from Antigononleptopus inhibited XO at 77.45% with IC50 at 1.79 p.g/mL. For comparison purposes, the IC50 value of allopurinol under the same experimental conditions was 1.00 ^g/mL [6]. The ethyl acetate extract of Moringaoleifera inhibited ACE at 64.23% while the positive control captopril at 87.57%. A pure compound isolated from the ethyl acetate extracts inhibited ACE at 95.85% which is even higher than the positive control (unpublished data).
Acknowledgement: Philippine Council for Health Research and Development, Department of Science and Technology for funding.
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References:
1. Young R, 1999. Eur J Med Chem, 34:671-685.
2. Chobanian A, Bakris G, Black H, Cushman W, Green L, Izzo J, Jones D, Materson B, Oparil S, Wright Jr J, Roccella E, 2003. Hypertension, 42:1206-1252.
3. Rundes R, Wyngaarden J, Hitchings G, Elion G, 1969. Annu Rev Pharmacol, 9:345-362.
4. Harris M, Siegel L, Alloway J, 1999. Am Fam Physician, 59:925-934.
5. Alaba C, Chichioco-Hernandez C, 2014. Asian Pac J Trop Biomed, 4(3):184-188.
6. Apaya M, Chichioco-Hernandez C, 2017. Arn Phar-macog Mag, 10(S3):501-505.
ACUTE AND SUB-ACUTE ORAL TOXICITY STUDY OF CHINESE-KNOTWEED [FALLOPIAMULTIFLORA (THUNB.) HARALDSON] EXTRACT IN RATS
© Heo J.D., Choi J.I., Hwang K.H., Moon Y.G., Kim J.H., Lee J.H., Tak T.K., Kim W.S.
Gyeongnam Biological Resource Research Center, Korea Institute of Toxicology, Jinju, Republic of Korea
In traditional oriental therapy, Chinese-knotweed (CK) is used as a tonic and an anti-aging remedy such as female climacteric, hair loss and pre mature greying of hiar. The present study aims to evaluate the safety of CK extract for the development of health food or natural therapeutics. We performed acute and sub-acute oral administration (2 weeks repeated) in SD rats. In subacute study,the CK was administered to male and female
SD rats at oral doses of 2000 mg/kg (acute study) and 0, 250, 500 and 1000 mg/kg (sub-acute study). Experimental animals were monitored for the mortality, body weight changes, food intake, clinical signs and gross findings during observation or administration period. Sub-acute toxicity study was also accompanied by hematological and biochemical analyses. In the above studies, we concluded that CK has weak toxicity in SD rats.
Table 1. Mortality in Acute Toxicity Study
Group Days on Test
Sex Dose (mg/kg) 0 1 2 3 4 hrs 1 2 3 4 5 6 7 8 9
Male 2000 T 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Female 2000 T 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Group Days on Test Final Mortality (Dead/used)
Sex Dose (mg/kg) 10 11 12 13 14
Male 2000 T 0 0 0 0 0 0/5
Female 2000 T 0 0 0 0 0 0/5
Figure 1. Body Weight Changes in Sub-acute Toxicity Study
Obzory po kliniceskoj farmacologii i lekarstvennoj terapii [Reviews of clinical pharmacology and drug therapy]
vol. 15/2Q17/suppLeMEnt 1
