Antiproliferative and Anti-Inflammatory Activities of Deprungsith Formulation and Its Bioactive Compounds Against Mild Psoriasis and Potential of Metabolic Herb-Drug Interactions.

Psoriasis is an incurable, chronic and auto-immune skin disorder with a global prevalence rate of approximately 2-3%. The study investigated the antipsoriasis activities of Deprungsith formulation and its bioactive components and their potential for inhibitory activities on human cytochrome P450 (CYP450). HaCaT and peripheral blood mononuclear cells (PBMCs) from healthy volunteers (n = 9) and psoriasis patients (n = 10) were exposed to Deprungsith formulation (Thai traditional medicine for psoriasis consisting of 16 plants), ethyl p-methoxycinnamate (EPMC), ligustilide and cyclosporin for 24 and 48 h. The antiproliferative, cell apoptosis and cell cycle arrest activities were evaluated using MTT assay and flow cytometry, respectively. The pro-inflammatory cytokine mRNA expression levels were measured using real-time polymerase chain reaction (RT-PCR). The CYP450 inhibitory effect was investigated using a bioluminescent-based CYP450 assay. Deprungsith formulation and the bioactive compounds inhibited HaCaT cells and PBMCs with weak to moderate potencies. EPMC and ligustilide combination produced an additive effect. Most substances arrested cell transition at sub-G1 and S phases, leading to early and late apoptosis induction. With prolonged exposure (48 h), all test substances decreased PBMCs necrosis. The mRNA expression of all pro-inflammatory cytokines was downregulated. Deprungsith formulation, EPMC, ligustilide and ferulic acid inhibited CYP1A2, CYP2C9, CYP2D6 and CYP3A4 activities with weak to moderate potencies. Deprungsith formulation and bioactive components induced cell apoptosis by inhibiting cell transition at specific cell cycle phases, which was correlated with the mRNA downregulation of interleukin (IL-6, IL-12p19, IL-23) and tumor necrosis factor (TNF-α). There is a low risk of potential adverse drug reactions and toxicity due to CYP450 interaction when Deprungsith formulation is concurrently administered with modern medicines.

Validation of an electrochemical sensor based on gold nanoparticles as a point-of-care test for quantitative determination of glycated hemoglobin.

Monitoring the level of glycated hemoglobin (HbA1c) has become the gold standard measure for diabetes mellitus (DM) diagnosis and control, used in conjunction with fasting blood glucose (FBG) and oral glucose tolerance test. This study aimed to investigate the applicability of a newly developed nanoparticle-based electrochemical sensor-multiwalled nanotubes incorporated with gold nanoparticles (POCT-HbA1cMWCNTs/AuNPs)-used as a routine point-of-care test (POCT) for detection of HbA1c for the diagnosis of DM. Finger-prick and venous blood samples were collected from 108 DM and 98 non-DM subjects to determine HbA1c and total hemoglobin by POCT-HbA1cMWCNTs/AuNPs compared with the standard HPLC method. The performance of the POCT-HbA1cMWCNTs/AuNPs was evaluated using the standard cut-off HbA1c level of >6.5%. The test's sensitivity, specificity, positive predictive value, and negative predictive value were 100.00%, 90.32%, 87.23%, and 100.00%, respectively. The probability of DM diagnosis in a subject with HbA1c >6.5% (positive predictive value) was 87.23% (82/94). The accuracy of the POCT-HbA1cMWCNTs/AuNPs was 94.18%, with a %DMV (deviation from the mean value) of 0.25%. The results indicate satisfactory assay performance and applicability of the POCT-HbA1cMWCNTs/AuNPs for diagnosis of DM using the cut-off criteria of HbA1c >6.5.

Apoptotic and Anti-metastatic Effects of Atractylodes lancea (Thunb.) DC. in a Hamster Model of Cholangiocarcinoma.

OBJECTIVES: Cholangiocarcinoma (CCA) is a highly aggressive tumor with a greater risk of distant metastasis. The promising anti-CCA activity and safety profile of Atractylodes lancea (AL) have previously been reported in a series of in vitro, in vivo and clinical studies. The present study investigated the effect of AL extract on apoptosis and metastasis signaling pathways in the Opisthorchis viverrini/dimethylnitrosamine (OV/DMN)-induced CCA hamster model. MATERIALS AND METHODS: Hamster liver tissues were obtained from the four groups (n = 5 per group), i.e., (i) 5-FU treated CCA (40 µg/mL); (ii) CCA; (iii) AL-treated CCA (5,000 mg/kg), and (iv) normal hamsters. Total RNA was isolated, and the expression levels of apoptosis-related and metastasis-related genes were determined by qRT-PCR analysis. RESULTS: The expression levels of p16, caspase-3, caspase-8, caspase-9, Apaf-1, p53 and Eef1a1 were downregulated, while that of the remaining genes were upregulated in CCA hamsters compared with normal hamsters. AL treatment increased the expression of p16, caspase-9, caspase-3, Apaf-1, p53 and E-cadherin and decreased the expression of cyclin D1, cdk4, Bax, Akt/PKB, Bcl-2, Mfge-8, Lass4, S100A6, TGF-ß, Smad-2, Smad-3, Smad-4, MMP-9, and N-cadherin. The expression of Eef1a1 was unchanged. CONCLUSION: The anti-CCA activity of AL in OV/DMN-induced CCA hamsters could be due to the induction of cell cycle arrest at the G1 phase and activation of the apoptosis pathway, resulting in cancer cell death. The activation of the apoptosis pathway mainly involved the intrinsic pathway (activation of caspase-3 and caspase-9 through p53 and Mfge-8 modulation and downregulation of anti-apoptotic genes Akt and Bcl-2). In addition, AL could also inhibit the canonical TGF-ß signaling pathway, MMP-9 and N-cadherin to suppress tumor metastasis.

Association between ABCB1 Polymorphisms and Artesunate-Mefloquine Treatment Responses of Patients with Falciparum Malaria on the Thailand-Myanmar Border.

A decrease in the clinical efficacy of a 3-day artesunate-mefloquine combination treatment was reported in the areas of multidrug-resistant Plasmodium falciparum along the Thailand-Myanmar border. The current study investigated the possible contribution of genetic polymorphisms of the three major genes encoding drug efflux transporters, ABCB1, ABCG2, and ABCC1, to responses to the aforementioned treatment in 91 patients with acute uncomplicated falciparum malaria residing along the Thailand-Myanmar border. Patients carrying homozygous mutant genotype ABCB1 c.1236C>T (TT) were found to have a three-times higher chance of successful treatment with this combination compared with other genotypes (CC and CT). Furthermore, whole blood mefloquine concentrations in these patients with the TT genotype were significantly lower than those of patients carrying the CC genotype. Patients with heterozygous mutant genotype (CT), however, were three-times more likely to experience treatment failure. No significant association was found with the ABCG2 and ABCC1 gene polymorphisms. The results suggest that ABCB1 c.1236C>T polymorphisms could be useful genetic markers for predicting responses to the 3-day artesunate-mefloquine treatment; however, studies using larger sample sizes in different malaria-endemic areas are necessary to confirm this finding. This study highlights the impact of pharmacogenetic factors on antimalarial treatment responses and the basis for the application of control policies in various malaria-endemic areas.

A Systematic Review of Drug Metabolism Studies of Plants With Anticancer Properties: Approaches Applied and Limitations.

BACKGROUND AND OBJECTIVES: Hepatic drug metabolism is a key influence on the efficacy and safety of medicines from both chemical and natural product sources. Studies of the metabolism of synthetic compounds, herbal medicines/supplements, and herb-derived bioactive compounds are therefore challenging. The aim of the present review is to provide a summary of the approaches/techniques that are currently being employed to investigate different aspects of the metabolism of herbs and herb-derived compounds (reaction phenotyping, metabolite profiling, metabolic clearance prediction, metabolic/pharmacokinetic drug interactions, and metabolism-related pharmacokinetic studies), including their limitations. METHODS: A thorough search of the PubMed database was performed using the terms 'Cancer' AND 'Cytochrome P450 (CYP)' OR 'Phase I metabolism' OR 'Phase II metabolism' AND 'Natural product' OR 'Herbal medicine' OR 'Herbal product' OR 'Herb-derived compound.' RESULTS: Most of the studies (84 studies, 83.2%) retrieved during the search investigated metabolic/pharmacokinetic drug interactions. Three (3.0%), 7 (6.9%), 6 (5.9%), and 1 (1.0%) study involved metabolism-related pharmacokinetic studies, reaction phenotyping, metabolite profiling, and prediction of metabolic clearance, respectively. CONCLUSIONS: Various studies reported conflicting results, with the results depending on the nature of the herb investigated (extracts or active constituents) and the biochemical tool (subcellular fractions, cells, or recombinant enzymes) and study system (in vitro/in vivo/ex vivo/clinical) applied. Each approach/system has its own advantages and disadvantages. Selecting the most appropriate approaches/systems allows us to extract the most meaningful and clinically relevant information on the metabolic pathways (the metabolites generated and the enzymes involved) and the potential drug interactions of herb-derived compounds for cancer therapy and prevention. Human primary hepatocytes are the best model that can be applied in any metabolic study. Human liver microsomes (HLMs) are a useful biochemical tool for preliminary drug metabolism studies. Recombinant microsomes that express specific enzymes and CYP-isoform-specific monoclonal antibodies are useful tools for enzyme inhibition studies.

Polymorphisms of genes encoding drug transporters or cytochrome P450 enzymes and association with clinical response in cancer patients: a systematic review.

PURPOSE: Not all patients respond well to cancer chemotherapy. One of the most important factors contributing to treatment response (efficacy and toxicity) is genetic determinant. The current systematic review aims to provide current status of the information on the genetic contribution of genes encoding drug transport proteins and drug metabolizing enzyme, cytochrome P450 (CYP), and relationship with clinical outcomes of cancer chemotherapy. METHODS: The literature search was performed through PubMed and ScienceDirect databases. One hundred and four research articles that fulfilled the inclusion criteria and had none of the exclusion criteria were included in the analysis. RESULTS: Various studies reported conflicting results for the polymorphisms of the major genes and association with treatment outcomes in patients with various types of cancer. Nevertheless, among the investigated gene polymorphisms, it appears that the 1236C>T, 3435C>T and 2677 G>T/A SNPs of the drug transporter gene ABCB1 were the most promising determinants of clinical outcomes. Although both 1236C>T and 3435C>T polymorphism are synonymous SNPs, several studies have demonstrated that not all synonymous SNPs are silent. Therefore, using the haplotype (1236C>T, 2677G>T, and 3435C>T) analysis rather than a single SNP may be a more useful approach for phenotype prediction. Some of the patients with variants of CYP genes were associated with unsatisfactory treatment response (efficacy and toxicity), suggesting that these variants may be associated with either reduction or absence of CYP enzyme activity. CONCLUSIONS: The controversial results may be due to several factors including difference in populations studied, sample size, tumor sites and stages, chemotherapeutic drug regimens, and evaluation parameters for efficacy and/or toxicity. Before the information can be successfully applied to individualized cancer chemotherapy, further studies should be focused on these promising genetic markers and their association with clinical outcomes using standardized protocols.

Chronic exposure to cadmium is associated with a marked reduction in glomerular filtration rate.

BACKGROUND: Urinary 20-hydroxyeicosatetraenoic acid (20-HETE) has been associated with hypertension in women with elevated urinary cadmium (Cd) excretion rates. The present study investigates the urinary Cd and 20-HETE levels in relation to the estimated glomerular filtration rate (eGFR) and albumin excretion in men and women. METHODS: A population-based, cross-sectional study, which included 225 women and 84 men aged 33-55 years, was conducted in a rural area known to be polluted with Cd. RESULTS: In all subjects, lower eGFR values were associated with higher urinary Cd excretion (P = 0.030), and tubulopathy markers N-acetyl-ß-d-glucosaminidase (P < 0.001) and ß2-microglobulin (ß2-MG) (P < 0.001). On average, the hypertensive subjects with the highest quartile of urinary Cd had eGFR values of 12 and 17 mL/min/1.73 m2 lower than that in the hypertensive (P = 0.009) and normotensive subjects (P < 0.001) with the lowest quartile of urinary Cd, respectively. In men, urinary albumin was inversely associated with 20-HETE (ß = -0.384, P < 0.001), while showing a moderately positive association with systolic blood pressure (SBP) (ß = 0.302, P = 0.037). In women, urinary albumin was not associated with 20-HETE (P = 0.776), but was associated with tubulopathy, reflected by elevated urinary excretion of ß2-MG (ß = 0.231, P = 0.002). CONCLUSIONS: Tubulopathy is a determinant of albumin excretion in women, while 20-HETE and SBP are determinants of urinary albumin excretion in men. Associations of chronic exposure to Cd with marked eGFR decline and renal tubular injury seen in both Cd-exposed men and women add to mounting research data that links Cd to the risk of developing chronic kidney disease.

Pharmacogenetic relevant polymorphisms of CYP2C9, CYP2C19, CYP2D6, and CYP3A5 in Bhutanese population.

Background Marked differences among genotype frequencies (Caucasians, Asians, and Africans) have been observed in cytochrome P450 (CYP) genes. Data on the frequency of pharmacogenetic relevant polymorphisms in Bhutanese population is absent. This study aimed to investigate the frequencies of pharmacogenetic relevant polymorphisms of CYP2C9 (*2 and *3), CYP2C19 (*2 and *3), CYP2D6 (*10), and CYP3A5 (*3) in Bhutanese population. Methods Genotyping was performed in 443 DNA samples using polymerase chain reaction-restriction fragment length polymorphism. Results For CYP2C9, allele frequencies of *2 and *3 variants were 0.339% and 0%, respectively. For CYP2C19, frequencies of *2 and *3 variants were 30.135% and 15.689%, respectively. Allele frequencies of CYP2D6*10 and CYP3A5*3 were 21.332% and 77.314%, respectively. Allele frequencies of CYP2C9*2 are similar to most Asians while CYP2C9*3 was absent. CYP2C19*2 showed a close resemblance to Japanese and Burmese, while CYP2C19*3 is near to Japanese and Korean. CYP2D6*10 is noticeably lower than other Asians. CYP3A5*3 is similar to East Asians (Chinese, Japanese, and Korean). Conclusions The Bhutanese population is polymorphic for these CYP genes, except for CYP2C9*3. Similar to other populations, genetic testing for these genes may, therefore, be helpful to obtain the benefit from pharmacological treatments and prevent adverse drug reactions.

Is renal tubular cadmium toxicity clinically relevant?

BACKGROUND: Exposure to cadmium (Cd) has been associated with the development of hypertension, especially in women, but the mechanism of such an association is not understood. We hypothesize that Cd exposure alters renal production of 20-hydroxyeicosatetraenoic acid (20-HETE), which plays an indispensable role in renal salt balance and blood pressure control. METHODS: We examined long-term Cd exposure in relation to urinary 20-HETE excretion levels, tubular dysfunction and blood pressure measures, using data from a population-based, cross-sectional study that included 115 normotensive and 110 hypertensive women, 33-55 years of age, who lived in Cd contamination areas in Thailand. RESULTS: The mean [standard deviation (SD)] blood Cd level of the study subjects was 3.57 (3.3) µg/L, while the mean (SD) urinary Cd and urinary 20-HETE levels were 0.58 (0.47) µg/g creatinine and 1651 (4793) pg/mL, respectively. Elevated 20-HETE levels were associated with a 90% increase in prevalence odds of hypertension (P = 0.029), four times greater odds of having higher urinary Cd levels (P = 0.030) and a 53% increase in odds of having higher levels of tubular dysfunction (P = 0.049), evident from an increase in urinary excretion of ß2-microglobulin. In normotensive subjects, an increase in urinary 20-HETE levels from tertile 1 to tertile 3 was associated with a systolic blood pressure increase of 6 mmHg (95% confidence interval 0.3-12, P = 0.040). CONCLUSIONS: This is the first report that links urinary 20-HETE levels to blood pressure increases in Cd-exposed women, thereby providing a plausible mechanism for associated development of hypertension.

The stress response of human proximal tubule cells to cadmium involves up-regulation of haemoxygenase 1 and metallothionein but not cytochrome P450 enzymes.

Enzymes of the cytochrome P450 (CYP) super-family are implicated in cadmium (Cd) -induced nephrotoxicity, however, direct evidence is lacking. This study investigated the endogenous expression of various CYP proteins together with the stress-response proteins, heme oxygenase-1 (HO-1) and metallothionein (MT) in human kidney sections and in cadmium-exposed primary cultures of human proximal tubular epithelial cells (PTC). By immunohistochemistry, the CYP members 2B6, 4A11 and 4F2 were prominently expressed in the cortical proximal tubular cells and to a lesser extent in distal tubular cells. Low levels of CYPs 2E1 and 3A4 were also detected. In PTC, in the absence of Cd, CYP2E1, CYP3A4, CYP4F2 and MT were expressed, but HO-1, CYP2B6 and CYP4A11 were not detected. A range of cadmium concentrations (0-100µM) were utilized to induce stress conditions. MT protein was further induced by as little as 0.5µM cadmium, reaching a 6-fold induction at 20µM, whereas for HO-1, a 5µM cadmium concentration was required for initial induction and at 20µM cadmium reached a 15-fold induction. The expression of CYP2E1, CYP3A4, and CYP4F2 were not altered by any cadmium concentrations tested at 48h. Cadmium caused a reduction in cell viability at concentrations above 10µM. In conclusion although cultured PTC, do express CYP proteins, (CYP2E1, CYP3A4, and CYP4F2), Cd-induced cell stress as indicted by induction of HO-1 and MT does not alter expression of these CYP proteins at 48h.

Expression of metallothionein isoforms in peripheral blood leukocytes from Thai population residing in cadmium-contaminated areas.

Metallothionein (MT) is a group of proteins with high cadmium (Cd) affinity and with a potential role in Cd transportation and detoxification. The aim of the present study was to investigate the relationship between MT (MT-1A, MT-2A, and MT-3 isoforms) gene expression level in peripheral blood leukocytes and Cd-associated renal injury in non-occupational exposed Thai population. The study was conducted in adult subjects residing in Cd-contaminated areas of Mae Sot District, Thailand. The basal levels of MT-1A, MT-2A, and MT-3 mRNA expression were determined in leukocytes by quantitative RT-PCR. MT-1A and MT-2A expressions, particularly MT-1A, were found to be significantly increased with elevated levels of blood and urinary Cd levels. In subjects with high urinary Cd levels, negative correlations between MT-1A and microalbumin, and between MT-2A and ß(2)-MG, were observed. These results suggest that MT gene expression may reflect susceptibility of the exposed population to Cd-induced renal dysfunction. MT-1A mRNA expression in leukocytes might be developed as a potential biomarker of Cd exposure and Cd-induced renal dysfunction.

Study on the association between environmental cadmium exposure, cytochrome P450-mediated 20-HETE, heme-oxygenase-1 polymorphism and hypertension in Thai population residing in a malaria endemic areas with cadmium pollution.

The aims of the study were to investigate (i) the effects of environmental cadmium (Cd) on hypertension, biological markers of renal dysfunction and renal cytochrome P450-mediated arachidonate metabolism; and (ii) the association between genetic polymorphism of heme oxygenase-1 (HO-1) and hypertension and Cd-induced renal injury in the exposed Thai population. The study was conducted in adult subjects residing in Cd-contaminated malaria endemic areas of Mae Sot District, Thailand. All subjects were randomly selected and consistently distributed for sex, age and residential areas. Blood and urinary Cd levels were not significantly different between the case (hypertensive) and control (matched-pair normotensive) groups. While other renal dysfunction biomarkers were comparable between the two groups, urinary microalbumin, urinary 20-hydroxy-5,8,11,14-eicosatetraenoic acid (20-HETE) and serum creatinine were siginificantly higher in the hypertensive group. Only N-acetyl-ß-glucosaminidase (NAG) showed positive correlation with Cd in hypertensive and normotensive group. With respect to heme oxygenase-1 (HO-1) polymorphism, the frequencies of (GT)(n) alleles were similar in both case and control groups. The frequency of SL genotype was significantly higher in the control group, whereas the frequency of ML genotype was significantly higher in the case group. Although no significant difference between 20-HETE and NAG levels in various HO-1 genotypes was found, a trend of increase in 20-HETE and NAG levels was observed in subjects carrying longer (GT)(n) repeats. Results from the present study provide no clear evidence on the direct effects of environmental Cd on high blood pressure development in the non-occupational exposed Thai population. Furthermore, the indirect effect of Cd through HO-1 (genetic polymorphism and prevalence of long GT(n) repeats) and 20-HETE was inconclusive. Based on the data obtained in the present investigation further studies should be performed which use a larger sample size and effectively control for confounding. This should provide more definitive evidence of the relationship between Cd exposure and high blood pressure.

Urinary and blood cadmium levels in relation to types of food and water intake and smoking status in a Thai population residing in cadmium-contaminated areas in Mae Sot.

Human exposure to cadmium (Cd) produces a wide variety of toxic effects involving many organs and systems, but the kidney is the main organ affected among long-term Cd-exposed people. In the general population, the primary sources of Cd exposure are cigarette smoke and food (shellfish, offal and certain vegetables). The aims of the study were to investigate the association between urinary and blood Cd levels and personal habits relating to Cd intake (consumption of food stuff, water and tobacco smoking), levels of renal biomarkers in the urine or serum of 314 Thai subjects (85 males, 229 females) who resided in Cd-contaminated areas of Mae Sot District, Tak Province, Thailand. Based on the cut-off levels of 1 microg/g creatinine and 5 microg/l for urinary and blood Cd levels, respectively, nearly all subjects had urinary Cd levels lower than cut-off values for urine and blood (88.2 and 77.7%, respectively). Binary logistic backward stepwise regression analysis with five covariates (gender, residential areas, consumption of bamboo or chicken, and smoking status), and eight covariates (residential areas, consumption of beans, pork, fish or liver, types and sources of rice consumed and smoking status) best predicted urinary and blood Cd levels, respectively. For renal biomarkers, N-acetyl-beta-glucosaminidase (NAG) best predicted both urinary and blood Cd with good accuracy. A larger sample size with equal distribution of subjects with low (< 2 microg/g creatinine) and high (> 2 microg/g creatinine) urinary Cd levels should be studied to obtain the regression equation that would best predict Cd body burden.

In vitro antimalarial interactions between mefloquine and cytochrome P450 inhibitors.

The treatment and control of malaria is becoming increasingly difficult due to resistance of Plasmodium falciparum strains resistance to commonly used antimalarials. Combination therapy is currently the strategy for combating multi-drug resistant falciparum malaria, through exploiting phamacodynamic synergistic effect and delaying the emergence of drug resistance. The objective of the present study was to investigate antimalarial activity of inhibitors of cytochrome P450 (CYP) enzyme including their interactions with the antimalarial mefloquine against chloroquine-resistant (K1) and chloroquine-sensitive (3D7) P. falciparum clones in vitro. Results showed IC(50) (drug concentration which produces 50% schizont maturation inhibition) values [mean (range)] of mefloquine against K1 and 3D7 clones to be 8.6 (8.0-9.3) and 12.1 (10.5-13.8) nM, respectively. The corresponding values for the IC(50) of quinidine were 32.2 (31.9-32.5) and 28.7 (28.4-29.0) nM, and for ketoconazole were 3.9 (3.7-4.1) and 4.8 (4.6-5.1) microM, respectively. Analysis of isobologram revealed a trend of decreasing of fraction IC(50) (FIC), which indicates synergistics of the either quinidine or ketoconazole with mefloquine for both chloroquine-resistant and chloroquine-sensitive clones.