The Active Fraction of Polyrhachis vicina Roger (AFPR) activates ERK to cause necroptosis in colorectal cancer.

ETHNOPHARMACOLOGICAL RELEVANCE: Polyrhachis vicina Roger (P. vicina), a traditional Chinese medicinal animal, has been used to treat rheumatoid arthritis, hepatitis, cancer, and other conditions. Due to its anti-inflammatory properties, our previous pharmacological investigations have demonstrated that it is effective against cancer, depression, and hyperuricemia. Nevertheless, the key active components and targets of P. vicina in cancers are still unexplored. AIM OF THE STUDY: The study aimed to evaluate the pharmacological treatment mechanism of the active fraction of P. vicina (AFPR) in treating colorectal cancer (CRC) and to further reveal its active ingredients and key targets. METHODS: To examine the inhibitory impact of AFPR on CRC growth, tumorigenesis assays, cck-8 assays, colony formation assays, and MMP detection were utilized. The primary components of AFPR were identified by GC-MS analysis. The network pharmacology, molecular docking, qRT-PCR, western blotting, CCK-8 assays, colony formation assay, Hoechst staining, Annexin V-FITC/PI double staining, and MMP detection were performed to pick out the active ingredients and potential key targets of AFPR. The function of Elaidic acid on necroptosis was investigated through siRNA interference and the utilization of inhibitors. Elaidic acid's effectiveness to suppress CRC growth in vivo was assessed using a tumorigenesis experiment. RESULTS: Studies confirmed that AFPR prevented CRC from growing and evoked cell death. Elaidic acid was the main bioactive ingredient in AFPR that targeted ERK. Elaidic acid greatly affected the ability of SW116 cells to form colonies, produce MMP, and undergo necroptosis. Additionally, Elaidic acid promoted necroptosis predominantly by activating ERK/RIPK1/RIPK3/MLKL. CONCLUSION: According to our findings, Elaidic acid is the main active component of AFPR, which induced necroptosis in CRC through the activation of ERK. It represents a promising alternative therapeutic option for CRC. This work provided experimental support for the therapeutic application of P. vicina Roger in the treatment of CRC.

Active fraction of Polyrhachis vicina Rogers (AFPR) suppressed breast cancer growth and progression via regulating EGR1/lncRNA-NKILA/NF-κB axis.

Breast cancer (BC) is a major contributor of cancer-associated mortality in women. It is essential to find new therapeutic targets and drugs. Polyrhachis vicina Rogers is one of the Traditional Chinese Medicine (TCM). Our previous studies have shown an active fraction of Polyrhachis vicina Rogers (AFPR) has significant anti-inflammatory activity, suggesting its anti-cancer effect. Here, we aimed to explore the inhibitory effects of AFPR on BC and reveal its mechanism. The effects of AFPR on BC were examined by cell proliferation assay, wound healing assay, invasion assay and xenograft assay. Microarray sequencing, qRT-PCR, Western blot, chromatin immunoprecipitation assay and luciferase reporter assay were performed to investigate the regulation of AFPR on related genes and underlying mechanisms. As a result, AFPR suppressed BC cell growth, migration and invasion and inhibited tumor growth. LncRNA NKILA was most prominently upregulated in AFPR-treated MCF7 cells. AFPR inactivated NF-κB signaling pathway via regulating NKILA. Furthermore, AFPR regulated the expression of NKILA by inhibiting its transcript suppressor EGR1. This study firstly indicated that AFPR was a potential inhibitor of BC development via regulating EGR1/NKILA/NF-κB axis.

ABCC2 rs2273697 is associated with valproic acid concentrations in patients with epilepsy on valproic acid monotherapy.

Valproic acid (VPA), a widely used antiepileptic drug, is characterized by intensive inter-individual variability in concentration. Both efflux and influx transporters are reported to play important roles in the disposition of VPA, however, no comprehensive investigation into the association of the single nucleotide polymorphism (SNP) in ABC/SLC families with VPA concentration are reported. In the present study, we investigated the association of 12 SNPs in ABCC2, ABCC4, ABCG2, MCT1, MCT2, and OATP2B1 in 187 Chinese patients with epilepsy on VPA monotherapy with the trough concentrations of VPA. The data showed that VPA concentration in patients with ABCC2 rs2273697 AA genotype was significantly higher than that in those with GA+GG genotypes (p=0.000). The findings of the present study suggest that ABCC2 polymorphisms influence VPA concentrations in patients with epilepsy on VPA monotherapy, which may affect the treatment outcomes.

Can therapeutic drug monitoring increase the safety of Imatinib in GIST patients?

Imatinib at 400 mg daily is the standard treatment for patients affected with CML and GIST. The intervariability in plasma concentration is very significant. In many reports, a good therapeutic effect is attributed to an adequate concentration of Imatinib. However, few studies have been conducted to investigate the association between plasma concentration and side effects. Besides, no upper concentration limit of Imatinib plasma concentration detection has been established. The correlation of Imatinib trough concentrations (Cmin ) with adverse effects (AEs) was described here. Plasma samples were obtained from patients after 3 months treatment with Imatinib (steady state, n = 122). Liquid chromatography/ tandem mass spectrometry was used to determine the concentration of Imatinib and its metabolite NDI. The incidence of myelosuppression was increased significantly with the increased Imatinib trough plasma concentration. The plasma level of Imatinib and NDI in patients who developed myelosuppression are 1698.3 ± 598.6 ng/mL and 242.1 ng/mL, respectively, which were significantly higher than those in patients who did not (1327.2 ± 623.4 ng/mL, P = 1.75 × 10-4 ; 206.3 ng/mL, P = 0.006). Estimated exposure thresholds of Imatinib and NDI were 1451.6 ng/mL with ROCAUC (95%CI) of 0.693 (0.597-0.789) and 207.1 ng/mL with ROCAUC (95%CI) of 0.646 (0.546-0.745), respectively. Multivariate regression confirmed the correlation of Imatinib Cmin with myelosuppression. Other side effects such as fluid retention and rash were not found to be correlated with Imatinib concentrations. These results suggest that trough concentration of Imatinib should be taken into consideration to increase the safety of Imatinib therapy in GIST patients.

Livin serves as a prognostic marker for mid-distal rectal cancer and a target of mid-distal rectal cancer treatment.

Livin is a novel member of the inhibitor of apoptosis protein family, which has been identified to be expressed in various malignancies and is suggested to be associated with poor prognostic significance. However, no data are available concerning the significance of livin in mid-distal rectal cancer. In the present study, livin expression, and its association with clinicopathological characteristics and prognosis was examined in patients with mid-distal rectal cancer. Apoptotic susceptibility, invasion capacity and chemosensitivity of LoVo cells were investigated using small interfering RNA (siRNA)-mediated knockdown of livin. It was revealed that livin was highly expressed in mid-distal rectal cancer tissues compared with the normal rectal mucosal tissues. Livin expression was associated with pathological grade, extent of invasion (T stage) and extent of lymph node metastasis (N stage) of tumor, contributing to poor prognosis of mid-distal rectal cancer following surgery. The data suggest that aggressive surgery should be applied in patients with mid-distal rectal cancer with high expression of livin. It was also revealed that knockdown of livin by siRNA increased the apoptotic rate, suppressed invasion of LoVo cells, and decreased the half-maximal inhibitory concentration of oxaliplatin and 5-fluorouracil by ~50% in LoVo cells significantly compared with control groups. The data suggested that a combination of downregulation of livin and anticancer drugs may significantly decrease the toxicity of anticancer drugs. Taken together, the present study indicated that livin may be a promising target in clinical therapy of mid-distal rectal cancer.

Impact of the haplotypes of the human pregnane X receptor gene on the basal and St John's wort-induced activity of cytochrome P450 3A4 enzyme.

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT: Human pregnane X receptor (PXR/NR1I2) is a key regulator of cytochrome P450 3A4. To date, there are 198 reported SNPs for the human PXR/NR1I2 gene. Some of these SNPs are found to affect the inducing ability of PXR to CYP3A4. WHAT THIS STUDY ADDS: This study, for the first time, has investigated the effect of PXR haplotype on basal and St John's wort-induced CYP3A4 activity in humans. H1/H1 of the PXR gene had weaker basal transcriptional activity but greater inducible transcriptional activity to CYP3A4 than H1/H2 and H2/H2. AIMS: Human pregnane X receptor (PXR/NR1I2) is the master regulator of CYP3A4, which metabolizes >50% of drugs on the market. This study investigated the relationship between the two most frequent haplotypes [H1 (TCAGGGGCCACC) and H2 (CCGAAAACTAAT)] of PXR and basal and St John's wort (SJW)-induced CYP3A4 activity. METHODS: Ten healthy subjects carrying H1 and H2 haplotypes (three subjects with H1/H1, four with H1/H2 and three with H2/H2) entered this study. The 10 subjects did not carry CYP3A4*4, *5 and *6. All subjects were administrated a 300-mg SJW tablet three times daily for 14 days, and CYP3A4 activity was measured using nifedipine (NIF) as a probe. The plasma concentrations of NIF and dehydronifedipine (DNIF) were determined by a validated liquid chromatography/mass spectrometry/mass spectrometry method. RESULTS: After administration of SJW, the AUC(0-infinity) of NIF decreased significantly, and the AUC(0-infinity) of DNIF increased significantly (P < 0.05). For H1/H2, the AUC(0-infinity) of NIF decreased by 42.4%, and the AUC(0-infinity) of DNIF increased by 20.2%; for H2/H2, the AUC(0-infinity) of NIF decreased by 47.9%, and the AUC(0-infinity) of DNIF increased by 33.0%; for H1/H1, the AUC(0-infinity) of NIF decreased by 29.0%, yet the AUC(0-infinity) of DNIF increased by 106.7%. The increase of the AUC(0-infinity) of DNIF in H1/H1 was significantly different from the other two haplotype pairs (P < 0.05). Meanwhile, before administration of SJW, the ratio of AUC(0-infinity(DNIF))/AUC(0-infinity(NIF)) was the lowest for H1/H1 (22.1%), compared with H1/H2 (58.7%) and H2/H2 (30.0%). CONCLUSIONS: H1/H1 of the human PXR gene had weaker basal transcriptional activity but greater inducible transcriptional activity to CYP3A4 than H1/H2 and H2/H2.

Single nucleotide polymorphisms of the pregnane x receptor gene in Han Chinese and a comparison with other ethnic populations.

The pregnane X receptor (PXR/NR1I2) gene is a master regulator for a number of cytochrome P450s (CYPs) and drug transporters. This study aimed to detect the single nucleotide polymorphisms (SNPs) of the PXR gene in Han Chinese (n = 186) and to compare the frequencies of polymorphisms of the PXR gene with those in Caucasian and African Americans reported in the literature. The SNPs of the PXR gene were analyzed using the polymerase chain reaction (PCR) and direct sequencing analysis. The mutant frequencies of A11156C and T11193C in Han Chinese were 55% (95% confidence interval (CI): 0.49-0.61) and 59% (95% CI: 0.52-0.64), respectively, higher than those of Caucasian Americans (16 and 16%, respectively) and African Americans (33 and 30%, respectively). However, the reported SNPs in exons 2 and 4 (PXR*2,*3,*4,*6,*9,*10,and *11) were not detected in Han Chinese. These results indicate that there are marked differences in the mutant frequencies of A11156C and T11193C of PXR between Han Chinese and other ethnic groups. The mutant frequency in the coding region (exons 2 and 4) of PXR was very low in Han Chinese. Further studies are needed to determine the impact of common SNPs of PXR in Han Chinese and other ethnic populations on the phenotypic activity of cytochrome P450s and drug transporters transactivated by PXR.

Biotransformation and pharmacokinetics of the novel anticancer drug, SYUIQ-5, in the rat.

SYUIQ-5, a novel telomerase inhibitor, has demonstrated antitumor activity in nude mouse studies. The objective of the present study was to examine the metabolism and pharmacokinetics of SYUIQ-5 in rats. The plasma pharmacokinetics of SYUIQ-5 was nonlinear following i.p. administration at 15, 30 and 60 mg/kg. SYUIQ-5 metabolism in rat liver microsomes followed Michaelis-Menten kinetics, with Km and Vmax values of 12.3 microM and 2.01 nmol/min/mg protein, respectively. Ketoconazole significantly inhibited the metabolism of SYUIQ-5 in liver microsomes from rats pretreated with control vehicle or various inducers, whereas sulphaphenazole, ticlopidine, quinidine, and methylpyrazole had no inhibitory effects on SYUIQ-5 metabolism. Dexamethasone and beta-naphthoflavone (BNF), but not phenobarbital and ethanol, significantly induced SYUIQ-5 metabolism in rats. Alpha-naphthoflavone significantly inhibited SYUIQ-5 metabolism in liver microsomes from BNF-pretreated rats. Similar to other secondary amines, SYUIQ-5 underwent N-demethylation and O-oxygenation to at least two metabolites by rat liver microsomes. Pretreatment of rats with SYUIQ-5 at 0.1, 5 or 10 mg/kg for 5 days significantly induced the expression and activity of rat Cyp1A1/2, and induced Cyp3A1/2 expression at 10 mg/kg, but not Cyp2E1 and 2B1/2. These results indicate that that SYUIQ-5 exhibits dose-dependent pharmacokinetics in rats and it is mainly metabolized by Cyp3A1/2.

Determination of CH330331, a novel 4-anilinoquinazoline inhibitor of epidermal growth factor receptor tyrosine kinase, in human Caco-2 monolayers by high performance liquid chromatography with ultraviolet detection: application to a trans-epithelial transport study.

4-Anilinoquinazolines (e.g. Iressa and Glivec) are a class of epidermal growth factor receptor tyrosine kinase (EGFR-TK) inhibitors widely used to treat non-small cell lung cancer and other tumors. However, low clinical response rate, resistance, and host toxicity of currently available EGFR-TK inhibitors prompt the development of second generation of TK inhibitors with improved efficacy, selectivity, and less resistance. CH330331 is a recently synthesized novel 4-anilinoquinazoline analog with confirmed anticancer activity in vitro and in vivo. To predict its oral pharmacokinetic behavior and transport nature in the intestine before entering clinical trials, we have developed and validated a high performance liquid chromatographic (HPLC) method for the determination of CH330331 in Caco-2 (a human colon cancer cell line) monolayers. The developed HPLC method was sensitive and reliable, with acceptable accuracy (90-110% of nominal values) and precision (intra- and inter-assay R.S.D.<10%). The total running time was within 10 min, with acceptable separation of the target analytes. The lower limit of quantitation (LLOQ) value for CH330331 was 200 ng/ml when an aliquot of 100 microl sample was injected onto the HPLC. The validated HPLC method was applied to characterize the epithelial transport of CH330331 in Caco-2 monolayers. The transport of CH330331 across the Caco-2 monolayers from the apical to basolateral side was 8- to 10-fold higher than that from the basolateral to apical side. Co-incubation of sodium azide or MK-571, but not verapamil, significantly inhibited the apical to basolateral transport of CH330331. These findings provide initial evidence that the intestinal absorption of CH330331 is mediated by an active mechanism. Further studies are required to explore the interaction of CH330331 with ATP-binding cassette transporters and the possible influence on its pharmacokinetics and pharmacodynamics.

High performance liquid chromatography with ultraviolet detection for the determination of SYUIQ-5, a novel telomerase inhibitor for cancer therapy: application to an enzyme kinetic study in rat liver microsomes.

A sensitive assay for the determination of SYUIQ-5, a novel telomerase inhibitor and anti-tumor drug, in rat liver microsomes was developed by using high-performance liquid chromatography with ultraviolet detection. SYUIQ-5 was incubated in vitro with liver microsomes from rats pre-treated with control vehicle, beta-naphthofIavone, phenobarbital, 20% ethanol or dexamethasone. The analytes were extracted with diethyl ether and separated a C(18) 5-microm analytical column. Elution was conducted with 30 mM dipotassium hydrogen phosphate (pH 8.0)-methanol-triethylamine (30:70:0.05, v/v/v) at a flow-rate of 1.0 ml/min and the detection of UV absorbance was conducted at 278 nm. Intra-day and inter-day precision and accuracy of the method were within 10%. The mean analytical recoveries of SYUIQ-5 ranged from 78.8 to 95.3%. The linearity of the calibration curve was in the range of 1.0-80.0 microM. The lower limit of quantification (LOQ) was 1.0 microM. Kinetic analysis showed that beta-naphthofIavone and dexamethasone significantly induced SYUIQ-5 metabolism, suggesting that cytochrome P450 1A and 3A are the major contributor to SYUIQ-5 metabolism in rat liver microsomes.

A pharmacogenetic study of pregnane X receptor (NR1I2) in Han Chinese.

The pregnane X receptor (PXR/NR1I2) gene is a critical transcriptional regulator of a number of important drug metabolizing enzymes and transporters. This study was undertaken to determine the frequencies of single nucleotide polymorphisms (SNPs) and haplotypes and to detect yet unknown SNPs in the NR1I2 gene in 210 unrelated healthy Han Chinese in comparison with other ethnic groups. We also characterized the functional impact of two SNPs, -24622A>T in the 5'-untranslated region and -24446C>A in exon 1 of NR1I2, by constructing three recombinants and monitoring promoter activity using the dual luciferase reporter gene assay. Genomic DNA was isolated from peripheral leukocytes and subjected to polymerase chain reaction (PCR) amplification, followed by direct DNA sequencing. Sixteen SNPs in NR1I2 with frequencies of 0.3-90.3% were found in Han Chinese, two of which (-25439A>G in the 5'-untranslated region and 7637C>T in intron 5) are previously unknown. The mutant allelic frequencies varied from 0.3% to 90.3%. Most of the detected SNPs were located in introns. A total of 15 linkage disequilibriums were detected; and positive linkage disequilibriums were found between -24381A>C in exon 1 and -24113G>A in intron 1, and 252A>G in intron 2 and 275A>G in intron 2 (rho(2) = 1, P<0.001). A total of 42 haplotypes were inferred and the two most frequent haplotypes were H1 (TCAGGGGCCACC) and H2 (CCGAAAACTAAT) with a frequency of 15.1%. The activity of the recombinants with alleles containing the -24622A>T in the 5'-untranslated region or -24446C>A in exon 1 was 30-40% higher than that in the wild-type (reference genotype). These results indicate that there are marked ethnic differences in the frequency between Han Chinese and other ethnic groups and that alleles with -24622A>T in the 5'-untranslated region and -24446C>A in exon 1 of the NR1I2 gene result in an increased activity compared to the wild-type. Further studies are warranted to explore the clinical and toxicological impact of SNPs and haplotypes of NR1I2 in various ethnic groups.