Different transcriptome profiles between human retinoblastoma Y79 cells and an etoposide-resistant subline reveal a chemoresistance mechanism.

BACKGROUND: Retinoblastoma (RB) is the most frequent pediatric retinal tumor. In the present study, to elucidate chemoresistance mechanisms and identify potential biomarkers in RB, we utilized RNA sequencing (RNAseq) technological platforms to reveal transcriptome profiles and identify any differentially expressed genes (DEGs) between an etoposide drug-resistant subline (Y79/EDR) and parental Y79 cells. METHODS: To test whether Y79/EDR cells showed resistance to antineoplastic agents for RB, we treated the cells with etoposide, carboplatin and vincristine and analyzed them with a Cell Counting Kit-8 (CCK-8). Y79/EDR and parental Y79 cells were used for RNAseq and bioinformatics analysis to enable a genome-wide review of DEGs between the two lines using the DESeq R package (1.10.1). Then, DEG enrichment in Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways was analyzed with KOBAS software. Next, real-time quantitative reverse transcription polymerase chain reaction (real time QRT-PCR) and cytotoxicity assays were performed to experimentally and functionally validate the identified candidate biomarkers. RESULTS: Y79/EDR cells showed resistance to etoposide, carboplatin and vincristine at different concentrations. In total, 524 transcripts were differentially expressed in Y79/EDR cells based on analysis of fragments per kilobase of transcript per million fragments mapped (FPKM); among these, 57 genes were downregulated and 467 genes were upregulated in Y79/EDR cells compared to parental Y79 cells. We selected candidate DEGs, including ARHGAP9, HIST1H4H, RELN, DDIT4, HK2, STC1 and PFKFB4, for mRNA expression validation with real time QRT-PCR assays and found that the expression levels determined by real time QRT-PCR were consistent with the RNAseq data. Further studies involving downregulation of ARHGAP9 with a specific siRNA showed that ARHGAP9 altered the cellular sensitivity of Y79 cells to etoposide and carboplatin. CONCLUSION: Our initial findings provided a genomic view of the transcription profiles of etoposide-induced acquired resistance in RB. Follow-up studies indicated that ARHGAP9 might be a chemoresistance biomarker in RB, providing insight into potential therapeutic targets for overcoming acquired chemoresistance in RB. These findings can aid in understanding and overcoming chemoresistance during treatment of RB in the clinic.

TRIM28 mediates chromatin modifications at the TCRα enhancer and regulates the development of T and natural killer T cells.

T-cell receptor-α (TCRα) rearrangement in CD4(+)CD8(+) double-positive immature thymocytes is a prerequisite for production of αß T cells and invariant natural killer T cells. This developmental event is regulated by the TCRα enhancer (Eα), which induces chromatin modification and recruitment of the recombination-activating proteins Rag1 and Rag2. However, the molecular mechanism underlying the activation and long-range action of Eα remains incompletely understood. We show here that the chromatin-modifying factor TRIM28 is highly expressed in double-positive thymocytes and persistently phosphorylated at serine 473. TRIM28 binds to Eα and induces histone 3 lysine 4 trimethylation in the Eα and distant regions of the TCRα locus, coupled with recruitment of Rag proteins. T-cell-conditional ablation of TRIM28 impaired TCRα gene rearrangement and compromised the development of αß T cells and invariant natural killer T cells. These findings establish TRIM28 as a unique regulator of thymocyte development and highlight an epigenetic mechanism involving TRIM28-mediated active chromatin modification in the TCRα locus.

Poly[[di-aqua-(µ4-benzene-1,2,4,5-tetra-carboxyl-ato)tetrakis-(1H-imidazole-κN (3))dicopper(II)] N,N-di-methyl-formamide monosolvate].

The asymmetric unit of the polymeric title compound, {[Cu2(C10H2O8)(C3H4N2)4(H2O)2]·C3H7NO} n , contains two independent Cu(II) ions, each coordinated by one water mol-ecule, two imidazole N atoms and two carboxyl-ate O atoms from benzene-1,2,4,5-tetra-carboxyl-ate anions in a distorted square-pyramidal geometry. The benzene-1,2,4,5-tetra-carboxyl-ate anion bridges four Cu(II) ions, forming a polymeric sheet parallel to (010). In the crystal, extensive N-H⋯O and O-H⋯O hydrogen bonds link the polymeric sheets and di-methyl-formamide solvent mol-ecules into a three-dimensional supra-molecular structure.

Electrochemical sizing of organic nanoparticles.

The size of organic nanoparticles (NPs) can be electrochemically determined by Faradaic charge transfer when nanoparticles strike an electrode. Indigo NPs were used as a model (see distribution of the NP diameter). This strategy could be used for monitoring the size of a wide range of organic nanoparticles.

Regulation of Th17 cell differentiation and EAE induction by MAP3K NIK.

Th17 cells play an important role in mediating autoimmune diseases, but the molecular mechanism underlying Th17 differentiation is incompletely understood. We show here that NF-kappaB-inducing kinase (NIK), which is known to regulate B-cell maturation and lymphoid organogenesis, is important for the induction of Th17 cells. NIK-deficient naive CD4 T cells are attenuated in the differentiation to Th17 cells, although they are competent in committing to the other effector lineages. Consistently, NIK knockout mice are resistant to experimental autoimmune encephalomyelitis, a disease model that involves the function of Th17 cells. This phenotype was also detected in Rag2 knockout mice reconstituted with NIK-deficient T cells, confirming a T-cell intrinsic defect. We further show that NIK mediates synergistic activation of STAT3 by T-cell receptor and IL-6 receptor signals. NIK deficiency attenuates activation of STAT3 and induction of STAT3 target genes involved in Th17-commitment program. These findings establish NIK as an important signaling factor that regulates Th17 differentiation and experimental autoimmune encephalitis induction.

Sesamin protects against DSS-induced colitis in mice by inhibiting NF-κB and MAPK signaling pathways.

OBJECTIVE: To investigate the protective effects and mechanisms of sesamin (SES) on dextran sulfate sodium (DSS)-induced experimental colitis in mice. METHODS: SES (50, 100, and 200 mg kg-1) were orally administered to C57BL/6 male mice after DSS instillation. The anti-inflammatory effect of SES on colonic damage was assessed by clinical, macroscopic, microscopic, and inflammatory signaling pathways. RESULTS AND CONCLUSIONS: It could be found that bodyweight and colon length of mice treated with DSS was significantly decreased while that were increased by SES treatment. SES treatment reduced the DAI values and improved the histopathology of the colon in the DSS-treated mice. SES also reduced TNF-α, IL-1ß and IL-6 production caused by DSS. We also measured the expression of the phosphorylation of p65, IκB, p38, ERK and JNK protein and found that SES can alleviate colon damage via the NF-κB and MAPK signaling pathways. The findings of this study suggested that SES had anti-inflammatory effects on intestinal inflammation and can be used as a new therapeutic candidate for inflammatory bowel disease.

A retrospective survey of Chlamydia pneumoniae infection rates in paediatric patients from a single centre in Wuxi, China.

OBJECTIVE: To provide some epidemiological data on Chlamydia pneumoniae infection rates in paediatric patients at a single centre in Wuxi, China. METHODS: This was a retrospective analysis of serum samples from paediatric patients (<12 years) with a respiratory tract infection (RTI) and who had been admitted to the Department of Paediatrics, Wuxi No.2 People's Hospital, China, from 01 January 2015 to 31 December 2016. C. pneumoniae IgM antibodies had been analysed using enzyme-linked immunosorbent assay (ELISA). RESULTS: Of the 3866 children (2073 boys, 1793 girls) with a RTI that provided serum samples, 19% were positive for C. pneumoniae IgM antibodies. Among these children, 56% were positive for other infections. CONCLUSIONS: Children over 6 years of age with a RTI had a higher C.pneumoniae infection rate than younger children and the infection rate was more common in winter months compared with other times of the year.

Anti-inflammatory and antioxidative properties of helicid protect against CCl4 induced acute liver injury in mice.

Acute liver injury can be caused by chemicals and can lead to liver failure. We investigated the protective effect of helicid (HEL) on acute liver injury caused by CCl4 in mice. We found that ALT and AST levels as well as hepatic pathological damage in mice treated with CCl4 was increased significantly, while the effects were decreased by HEL treatment. HEL treatment increased the activity of T-SOD, GSH and CAT and reduced the level of MDA in CCl4 treated mice. HEL improved the histopathology of liver caused by CCl4. HEL also reduced TNF-α, IL-1ß and IL- 6 activity caused by CCl4. We investigated the phosphorylation of p65 and IκB protein and found that HEL can alleviate liver damage via the NF-κB signaling pathway. Our findings indicate that HEL protects against acute liver injury induced by CCl4. The protective effect of HEL appears to be due to its antioxidative and anti-inflammatory properties through the NF-κB signaling pathway.

Effect of danefukang on symptoms and biomarkers in women with endometriosis.

OBJECTIVE: The purpose of this study was to observe the efficacy of Danefukang (DEFK) soft extract for the treatment of symptoms associated with endometriosis, and its effect on quality of life, the Self-rating Anxiety Scale (SAS) and Self-rating Depression Scale (SDS) scores, and on levels of carbohydrate antigen (CA)-125, tumor necrosis factor (TNF)-α, and interleukin (IL)-6. MATERIALS AND METHODS: A total of 174 patients with endometriosis treated from January 2010 to December 2013 were randomly divided into a control group treated with mifepristone (n = 87) or DEFK (n = 87). Both groups were treated for 3 months. Symptoms, quality of life, SAS, SD scores, and levels of CA-125, TNF-α, and IL-6 were evaluated before and after treatment. RESULTS: The effectiveness rate was 93.10% in the DEFK group and 81.61% in the mifepristone control group (χ2 = 4.215, P < 0.05). Treatment with DEFK resulted in a greater improvement in quality of life, SDS, and SAS scores compared with mifepristone (all, P < 0.05). DEFK treatment also resulted in a greater decrease of CA-125, TNF-α, and IL-6 levels compared with mifepristone (all, P < 0.05). CONCLUSION: Based on the current results - improved symptoms, attenuated depression and anxiety, and reduced the levels of pro-inflammatory cytokines and CA-125 -treatment with DEFK is a meaningful option for patients with endometriosis. DEFK fills an unmet need in the pharmacologic treatment of endometriosis.

Effects of retrorsine on mouse hepatocyte proliferation after liver injury.

AIM: To study the effect of retrorsine on mouse hepatocyte proliferation. METHODS: Mice and rats were treated respectively with two injections of retrorsine (as retrosine-treated group) or saline (as non-treated group) at 2 wk intervals. They received a single injection of carbon tetrachloride (CCl4) 4 wk later. On d 0, 1, 2, 3, 4, 6, 15 after CCl4 administration, the animals were killed and their livers were excised. Hematoxylin and eosin (HE) staining and Ki-67 antibody immunohistochemical analysis of liver samples were used to evaluate the pathological changes and hepatocyte proliferation. RESULTS: In rats treated with retrorsine and CCl4, the liver displayed obvious megalocytosis, proliferation of mild bile duct,small hepatocyte-forming nodule, which were not found in liver samples from non-treated group. However,in mice treated with retrorsine combined with CCl4, the liver displayed hepatocyte degeneration and necrosis in perivenous areas.There was no obvious difference between retrorsine-treated group and non-treated group. Ki-67 immunohistochemical analysis showed that in rats treated with retrorsine, the positive hepatocytes mainly found in small hepatocyte nodules, were obviously less than those in non-treated group. The mice treated with retrorsine showed that the number of Ki-67 positive hepatocytes was very high and more than that in non-treated group. CONCLUSION: Retrorsine has no effect on mouse hepatocyte proliferation.

Establishment of the cytoplasmic incompatibility-inducing Wolbachia strain wMel in an important agricultural pest insect.

The wMel Wolbachia strain was known for cytoplasmic incompatibility (CI)-induction and blocking the transmission of dengue. However, it is unknown whether it can establish and induce CI in a non-dipteran host insect. Here we artificially transferred wMel from Drosophila melanogaster into the whitefly Bemisia tabaci. Fluorescence in situ hybridisation demonstrated that wMel had successfully transfected the new host. Reciprocal crossing was conducted with wMel-transfected and wild-type isofemale lines, indicating that wMel could induce a strong CI without imposing significant cost on host fecundity. We then determined the maternal transmission efficiency of wMel in the offspring generations, showing a fluctuating trend over a period of 12 generations. We thus detected the titre of wMel during different developmental stages and in different generations by using real-time quantitative PCR, revealing a similar fluctuating mode, but it was not significantly correlated with the dynamics of transmission efficiency. These results suggest that wMel can be established in B.tabaci, a distantly related pest insect of agricultural importance; moreover, it can induce a strong CI phenotype in the recipient host insect, suggesting a potential for its use in biological control of B. tabaci.

Effects of new 4-aryl-1,4-dihydropyridines and 4-arylpyridines on drug efflux mediated by multidrug resistance-associated protein 1.

The purpose of this study is to evaluate the effects of newly synthesized 4-aryl-1,4-dihydropyridine and pyridines on drug efflux mediated by multidrug resistance-associated protein 1 (MRP1, ABCC1). These compounds were designed to maximize inhibition of P-glycoprotein and minimize calcium channel binding activity, based on structure modifications of niguldipine. A [3H]vinblastine accumulation study was conducted in human small cell lung cancer H69AR (overexpressing MRP1) and wild type H69 cells. Five out of 16 dihydropyridines and 6 out of 9 pyridines were found to significantly increase the intracellular accumulation of vinblastine in resistant H69AR cells (p<0.01) at a concentration of 2.5 microM. Daunomycin accumulation studies, determined using a flow cytometric assay, were also performed in H69AR and human pancreatic adenocarcinoma Panc-1 cells and the results were highly correlated with those obtained from the [3H]vinblastine accumulation studies. Four compounds, which significantly increased vinblastine accumulation, were tested for their effect on daunomycin cytotoxicity in H69AR cells and found to significantly decrease the IC50 of daunomycin, confirming the accumulation study results. Compounds were also tested for their effect on intracellular glutathione (GSH) concentrations, a cosubstrate for MRP1-mediated efflux in H69AR and Panc-1 cells. No significant changes in the intracellular GSH level were observed in H69AR cells after treatment with these test compounds. However, following a 2-hr and 24-hr incubation with a dihydropyridine compound, Im, and its pyridine derivative IIm, there was a small (approximately 20%) but statistically significant decrease in intracellular GSH in Panc-1 cells. Our results indicate that some dihydropyridine and pyridine compounds in our series could inhibit MRP1-mediated transport and that GSH modulation plays a minor, if any, role in this effect.

Nano-impacts of bifunctional organic nanoparticles.

The synthesis and characterization of Oil Blue Dye nanoparticles is reported along with their use for nano-impacts experiments in aqueous solution. The latter reveal current spikes corresponding to quantitative two electron reductions due to the reduction of the quinone groups and quantitative two electron oxidations from the 1,4-phenylenediamine groups presented in the Oil Blue Dye molecules within the nanoparticles. In both cases, the oxidation or reduction leads to size distributions in good agreement with independent measurements made using dynamic light scattering showing that the redox events accompanying the nano-impacts lead to the full dissolution of the nanoparticles.

[Dispersion and analysis of odor pollution in landfill area under the enclosed operation condition].

Odor pollution of landfill site is a serious problem accompanied with the urbanization process that influences city life. Generally, odor emission points in landfill boundary are detected by experience, but the pollution intensity, distribution and variation in the scope of landfill boundary are difficulty to describe. In this research, odor emission points were disclosed with equal odor concentration curves that were delineated using electric nose and GPS instrument. The leakage of landfill gas and exhaust emission from biogas incineration torch was the main cause of the odor pollution in landfill area. Gas production evaluation suggested that the improvement of landfill gas consumption is the key point to control the odor pollution at the landfill site.

Imapct of laparoscopic myomectomy on IL-6, IL-2, and CRP in patients with uterine fibroids / 中国内镜杂志

Objective To investigate the effect of laparoscopic myomectomy for uterine fibroids. Methods From February 2015 to February 2016, 84 cases of uterine fibroids were selected, 51 patients underwent laparoscopic myomectomy (observation group), and 33 patients received abdominal myomectomy (control group), the operation time and intraoperative blood loss and so on were observed in two groups, C reactive protein (CRP), interleukin-2 (IL-2), IL-6, follicle-stimulating hormone (FSH), luteinizing hormone (LH) and antral follicle (AFC) were detected before and after operation. Results The operation time of the observation group was (74.29 ± 13.20) min, significantly higher than the control group (P < 0.05), the amount of bleeding, postoperative exhaust time, hospitalization time and intraoperative myoma rejection number were (105.39 ± 32.04) ml, (17.40 ± 2.31) h, (4.02 ± 1.32) d and (6.02 ± 1.32), significantly less than the control group (P < 0.05); The CRP and IL-6 in the observation group were (25.20 ± 5.32) mg/L and (38.04 ± 10.11) ng/ml, significantly lower than those in the control group (P < 0.05), while IL-2 was (46.13 ± 13.10) ng/ml, was significantly higher than that of the control group (P < 0.05); The incidence of postoperative complications in the observation group was 3.92％, which was significantly lower than that in the control group (P < 0.05); There was no significant difference in the levels of (FSH, LH and AFC between the two groups (P > 0.05); The postoperative recurrence rate and pregnancy rate in the observation group were 21.56％ and 54.90％, compared with the control group, there was no significant difference (P > 0.05). Conclusion Laparoscopic myomectomy is safe and effective in treatment of uterine fibroids with less stress response and no effect on ovarian function.

A high-performance liquid chromatographic method for determination of the niguldipine analogue DHP-014.

A simple and reliable reversed-phase high-performance liquid chromatography method was developed and validated for the determination of DHP-014, a niguldipine analogue with potent P-glycoprotein inhibitory and negligible calcium channel blocking properties, in rat plasma. DHP-014 and niguldipine hydrochloride (the internal standard) were extracted from rat plasma by liquid extraction using hexane. DHP-014 was then separated by HPLC on a C18 column and quantified by ultraviolet detection at 238 nm. The mobile phase consisted of acetonitrile-aqueous 5 mM phosphate buffer (65:35, v/v) containing 0.4% (v/v) triethylamine adjusted to pH 7.0. The mean extraction efficiency of DHP-014 was 109.0 +/- 12.9, 97.7 +/- 8.0 and 102.9 +/- 7.5% for DHP-014 concentrations of 10, 50 and 100 nM, respectively (n = 5). The method was linear over the concentration range 2.5-200 nM with a regression coefficient of 0.998. The limit of detection of DHP-014 in rat plasma was 1.0 nM. The intra- and inter-day coefficients of variation for DHP-014 in rat plasma were 4.7-7.9 and 6.9-9.9%, respectively. The intra- and inter-day accuracy was 98.2-99.5 and 97.9-103%, respectively. The bioanalytical technique was used to determine DHP-014 in plasma samples in a pharmacokinetic study of DHP-014 administered to female Sprague-Dawley rats.

Quantitative structure-activity relationship and quantitative structure-pharmacokinetics relationship of 1,4-dihydropyridines and pyridines as multidrug resistance modulators.

PURPOSE: The aim of this study was to develop quantitative structure-activity/pharmacokinetic relationships (QSAR/QSPKR) for a series of synthesized 1,4-dihydropyridines (DHPs) and pyridines as P-glycoprotein (P-gp) inhibitors. METHODS: Molecular descriptors of test compounds were generated by 3D molecular modeling using SYBYL and KowWin programs. Forward inclusion coupled with multiple linear regression (MLR) was used to derive a QSAR equation for Ca2+ channel binding. A multivariate statistical technique, partial least square (PLS) regression, was applied to derive a QSAR model for P-gp inhibition and QSPKR models. Cross-validation using the "leave-one-out" method was performed to evaluate the predictive performance of models. RESULTS: For Ca2+ channel binding, the MLR equation indicated a good correlation between observed and predicted values (R2 = 0.90), and cross-validation confirmed the predictive ability of the model (Q2 = 0.67). For P-gp reversal, the model obtained by PLS could account for most of the variation in P-gp inhibition (R2 = 0.76) with fair predictive performance (Q2 = 0.62). Nine structurally related 1,4-DHP drugs were used for QSPKR analysis. The models could explain the majority of the variation in clearance (R2 = 0.90), and cross-validation confirmed the prediction ability (Q2 = 0.69). CONCLUSION: QSAR/QSPKR models were developed, and the QSAR models were capable of identifying synthesized 1,4-DHPs and pyridines with potent P-gp inhibition and reduced Ca2+ channel binding. The QSPKR models provide insight into the contribution of electronic, steric, and lipophilic factors to the clearance of DHPs.

Pharmacokinetics and bioavailability of a newly synthesized dihydropyridine compound with multidrug resistance reversal activity.

(+/-)3-(3-(4,4-diphenylpiperidin-1-yl)propyl) 5-methyl 4-(3,4-dimethoxyphenyl)-2,6-dimethyl-1,4-dihydropyridine-3,5-dicarboxylate ((+/-)-DHP-014), is a new 4-aryl-1,4-dihydropyridine that can reverse multidrug resistance mediated by the ATP-binding cassette (ABC) transport proteins, P-glycoprotein, multidrug resistance-associated protein 1 and breast cancer resistance protein; it exhibits negligible calcium channel blocking activity. The objective of this work was to investigate the pharmacokinetics of this new compound in rats. Three intravenous (1, 2 and 5 mg/kg) and two oral (25 and 50 mg/kg) doses were administered to female Sprague-Dawley rats. A two-compartment model with nonlinear elimination best characterized the pharmacokinetic profiles after intravenous and oral administration in rats. The terminal half-life of (+/-)-DHP-014 increased and the systemic clearance significantly decreased at higher doses, indicating nonlinear elimination. The dose-dependent clearance is likely due to saturation of metabolism. The apparent volume of distribution of (+/-)-DHP-014 was 2.0 L/kg in rats and was unchanged with increasing intravenous doses of (+/-)-DHP-014. The estimated oral bioavailability of (+/-)-DHP-014 was 8.2%. The poor bioavailability is likely due to the poor solubility of the compound, as well as to substantial first-pass elimination.

[Biological characteristics of mesenchymal stem cells from bone marrow of patients with aplastic anemia].

To investigate the differences of proliferation capacity and phenotype properties of mesenchymal stem cells (MSCs) derived from bone marrow (BM) of aplastic anemia patients, fetuses and children, MSCs were isolated from BM of patients with aplastic anemia and expanded in vitro; MSCs derived from BM of fetuses and children were used as normal control groups, three sources of MSCs were compared by morphology, in vitro proliferation capacity, phenotype and immunocytochemistry. The results showed that MSCs could be isolated and expanded from aplastic anemia patient BM. MSCs derived from BM of aplastic anemia patients shared a similar morphology and phenotype with derived MSCs from BM of fetuses and children. However, in vitro proliferation capacity of MSCs derived from BM of aplastic anemia patients after 20 population doublings (PD) was significantly lower, compared with MSCs from BM of fetuses and children. BM MSCs derived from children and fetuses proliferated for more than 30 PD. It is concluded that BM MSCs from aplastic anemia patients appears to be normal in phenotype but their proliferation capacity is lower in comparison with control groups.

Effects of dihydropyridines and pyridines on multidrug resistance mediated by breast cancer resistance protein: in vitro and in vivo studies.

Breast cancer resistance protein (BCRP, ABCG2) is a recently identified member of the ATP-binding cassette family of cell surface transport proteins. This study was conducted to investigate the effect of a series of newly synthesized 1,4-dihydropyridines and pyridines, designed as potent P-glycoprotein inhibitors, on BCRP-mediated drug efflux both in vitro and in vivo. The effects of 25 synthesized dihydropyridines and corresponding pyridines along with 4 commercially available dihydropyridines (niguldipine, nicardipine, nifedipine, and nitrendipine) on the intracellular accumulation of the BCRP substrate mitoxantrone were evaluated in BCRP-expressing human breast cancer MCF-7/MX100 and human non-small cell lung cancer H460/MX20 cells. At a 2.5 microM concentration, 24 of 25 newly synthesized dihydropyridines and pyridines produced a significant increase of mitoxantrone accumulation in both cell lines. The most potent compound was able to enhance mitoxantrone accumulation approximately 4.5-fold, greater than that obtained with 10 microM fumitremorgin C, which is a specific BCRP inhibitor. The results from the two cell lines showed good correlation (r(2) = 0.71, p < 0.01). Niguldipine, nicardipine, and nitrendipine also demonstrated potent BCRP inhibition, whereas nifedipine had no effect. The effects of the dihydropyridine and pyridine compounds on mitoxantrone cytotoxicity paralleled their effects on mitoxantrone accumulation. Coadministration of a selected dihydropyridine compound, I(m) [DHP-014; 3-(3-(4,4-diphenylpiperidin-1-yl)propyl) 5-methyl 4-(3,4-dimethoxyphenyl)-2,6-dimethyl-1,4-dihydropyridine-3,5-dicarboxylate)] with topotecan, a good BCRP substrate and a moderate to poor P-glycoprotein substrate, resulted in significant increases in the systemic exposure and peak concentration of topotecan in Sprague-Dawley rats when oral topotecan (2 mg/kg) was combined with 20 mg/kg DHP-014. The observed increase of topotecan exposure provides proof-of-concept for in vivo inhibition of BCRP by these agents.