Purine cyclin-dependent kinase inhibitors have been recognized as promising candidates for the treatment of various cancers; nevertheless, data regarding interaction of these substances with drug efflux transporters is still lacking. Recently, we have demonstrated inhibition of breast cancer resistance protein (ABCG2) by olomoucine II and purvalanol A and shown that these compounds are able to synergistically potentiate the antiproliferative effect of mitoxantrone, an ABCG2 substrate. In this follow up study, we investigated whether olomoucine II and purvalanol A are transported by ABCG2 and ABCB1 (P-glycoprotein). Using monolayers of MDCKII cells stably expressing human ABCB1 or ABCG2, we demonstrated that olomoucine II, but not purvalanol A, is a dual substrate of both ABCG2 and ABCB1. We, therefore, assume that pharmacokinetics of olomoucine II will be affected by both ABCB1 and ABCG2 transport proteins, which might potentially result in limited accumulation of the compound in tumor tissues or lead to drug-drug interactions. Pharmacokinetic behavior of purvalanol A, on the other hand, does not seem to be affected by either ABCG2 or ABCB1, theoretically favoring this drug in the potential treatment of efflux transporter-based multidrug resistant tumors. In addition, we observed intensive sulfatation of olomoucine II in MDCKII cell lines with subsequent active efflux of the metabolite out of the cells. Therefore, care should be taken when performing pharmacokinetic studies in MDCKII cells, especially if radiolabeled substrates are used; the generated sulfated conjugate may largely contaminate pharmacokinetic analysis and result in misleading interpretation. With regard to chemical structures of olomoucine II and purvalanol A, our data emphasize that even drugs with remarkable structure similarity may show different pharmacokinetic behavior such as interactions with ABC transporters or biotransformation enzymes.

• MeSH
• ABC transportéry metabolismus   MeSH
• biologický transport MeSH
• buněčné linie MeSH
• chemorezistence účinky léků   MeSH
• nádorové proteiny metabolismus   MeSH
• P-glykoprotein metabolismus   MeSH
• psi MeSH
• puriny farmakokinetika   MeSH
• zvířata MeSH
• Check Tag
• psi MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

ATP Binding Cassette B1 (ABCB1) is a transporter with a broad substrate specificity involved in the elimination of several carcinogens from the gut. Several polymorphic variants within the ABCB1 gene have been reported as modulators of ABCB1-mediated transport. We investigated the impact of ABCB1 genetic variants on colorectal cancer (CRC) risk. A hybrid tagging/functional approach was performed to select 28 single nucleotide polymorphisms (SNPs) that were genotyped in 1,321 Czech subjects, 699 CRC cases and 622 controls. In addition, six potentially functional SNPs were genotyped in 3,662 German subjects, 1,809 cases and 1,853 controls from the DACHS study. We found that three functional SNPs (rs1202168, rs1045642 and rs868755) were associated with CRC risk in the German population. Carriers of the rs1202168_T and rs868755_T alleles had an increased risk for CRC (P(trend) = 0.016 and 0.029, respectively), while individuals bearing the rs1045642_C allele showed a decreased risk of CRC (P(trend) = 0.022). We sought to replicate the most significant results in an independent case-control study of 3,803 subjects, 2,169 cases and 1,634 controls carried out in the North of Germany. None of the SNPs tested were significantly associated with CRC risk in the replication study. In conclusion, in this study of about 8,800 individuals we show that ABCB1 gene polymorphisms play at best a minor role in the susceptibility to CRC.

• MeSH
• alely MeSH
• dospělí MeSH
• genetická predispozice k nemoci * MeSH
• genotyp MeSH
• kolorektální nádory diagnóza   genetika   MeSH
• lidé středního věku MeSH
• lidé MeSH
• modely genetické MeSH
• P-glykoprotein genetika   MeSH
• P-glykoproteiny MeSH
• polymorfismus genetický * MeSH
• riziko MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• studie případů a kontrol MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH

The interaction between ABCB1 transporter and its substrates takes place in cell membranes but the available data precludes quantitative analysis of the interaction between transporter and substrate molecules. Further, the amount of transporter is usually expressed as a number of ABCB1 molecules per cell. In contrast, the substrate concentration in cell membranes is estimated by determination of substrate-lipid partition coefficient, as examples. In this study, we demonstrate an approach, which enables us to estimate the concentration of ABCB1 molecules within plasma membranes. For this purpose, human leukemia K562 cells with varying expression levels of ABCB1 were used: drug selected K562/Dox and K562/HHT cells with very high transporter expression, and K562/DoxDR2, K562/DoxDR1, and K562/DoxDR05 cells with gradually decreased expression of ABCB1 derived from K562/Dox cells using RNA interference technology. First, we determined the absolute amount of ABCB1 in cell lysates using immunoblotting and recombinant ABCB1 as a standard. We then determined the relative portion of transporter residing in the plasma membrane using immunohistochemistry in nonpermeabilized and permeabilized cells. These results enabled us to estimate the concentration of ABCB1 in the plasma membrane in resistant cells. The ABCB1 concentrations in the plasma membrane of drug selected K562/Dox and K562/HHT cells containing the highest amount of transporter reached millimolar levels. Concentrations of ABCB1 in the plasma membrane of resistant K562/DoxDR2, K562/DoxDR1, and K562/DoxDR05 cells with lower transporter expression were proportionally decreased.

• MeSH
• buněčná membrána metabolismus   MeSH
• buňky K562 MeSH
• fluorescenční protilátková technika MeSH
• lidé MeSH
• P-glykoproteiny genetika   metabolismus   MeSH
• RNA interference MeSH
• viabilita buněk genetika   fyziologie   MeSH
• western blotting MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND: The ABCB1 gene encodes P-glycoprotein implicated in the development of cellular drug resistance. The aim of this study was to develop high-resolution melting (HRM) analysis for determination of ABCB1 polymorphisms and evaluate their associations with clinical data of breast carcinoma patients. METHODS: HRM analysis was designed to assess five single nucleotide polymorphisms (SNPs) in ABCB1 (rs2214102, rs1128503, rs2032582, rs2032583 and rs1045642) in genomic DNA from 103 breast carcinoma patients. Results were confirmed by direct DNA sequencing. RESULTS: HRM analysis revealed distinct patterns of melting curves for the respective genotypes of all followed SNPs. Sensitivity of HRM analysis compared with direct DNA sequencing was superior (97.1% vs. 93.9%). The overall accuracy of HRM was 97.6%. The coefficients of variation in replicate experiments encompassed the range 0.002%-0.038%. On the basis of the examined SNPs, one strong haplotype block containing rs2032582 and rs1128503 SNPs was identified. Significant associations of rs2032582 SNP with tumor size, negative HER-2/neu status, and family history of breast carcinoma were found. Patients carrying the ancestral homozygous genotype (GG) in rs2214102 had significantly worse progression-free survival in comparison with carriers of the non-ancestral allele (A) in the adjuvant set (p=0.005). CONCLUSIONS: A rapid, accurate, low-cost and time-effective method for screening ABCB1 SNPs was developed. Significant associations of ABCB1 rs2032582 and rs2214102 SNPs with prognostic factors and survival of patients were found.

• MeSH
• denaturace nukleových kyselin MeSH
• jednonukleotidový polymorfismus * MeSH
• lidé středního věku MeSH
• lidé MeSH
• nádory prsu diagnóza   genetika   terapie   MeSH
• P-glykoprotein genetika   MeSH
• prognóza MeSH
• sekvenční analýza DNA metody   MeSH
• tranzitní teplota * MeSH
• výsledek terapie MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Maraviroc is a chemokine receptor 5 (CCR5) inhibitor used in the treatment of human immunodeficiency virus (HIV) that also shows therapeutic potential for several autoimmune, cancer, and inflammatory diseases that can afflict pregnant women. However, only limited information exists on the mechanisms underlying the transplacental transfer of the drug. We aimed to expand the current knowledge base on how maraviroc interacts with several placental ATP-binding cassette (ABC) efflux transporters that have a recognized role in the protection of a developing fetus: P-glycoprotein (ABCB1), breast cancer resistance protein (ABCG2), and multidrug resistance protein 2 (ABCC2). We found that maraviroc does not inhibit any of the three studied ABC transporters and that its permeability is not affected by ABCG2 or ABCC2. However, our in vitro results revealed that maraviroc shows affinity for human ABCB1 and the endogenous canine P-glycoprotein (Abcb1) expressed in Madin-Darby canine kidney II (MDCKII) cells. Perfusion of rat term placenta showed accelerated transport of maraviroc in the fetal-to-maternal direction, which suggests that ABCB1/Abcb1 facilitates in situ maraviroc transport. This transplacental transport was saturable and significantly diminished after the addition of the ABCB1/Abcb1 inhibitors elacridar, zosuquidar, and ritonavir. Our results indicate that neither ABCG2 nor ABCC2 influence maraviroc pharmacokinetic but that ABCB1/Abcb1 may be partly responsible for the decreased transplacental permeability of maraviroc to the fetus. The strong affinity of maraviroc to Abcb1 found in our animal models necessitates studies in human tissue so that maraviroc pharmacokinetics in pregnant women can be fully understood. SIGNIFICANCE STATEMENT: Antiretroviral drug maraviroc shows low toxicity and is thus a good candidate for prevention of mother-to-child transmission of human immunodeficiency virus when failure of recommended therapy occurs. Using in vitro cell-based experiments and in situ dually perfused rat term placenta, we examined maraviroc interaction with the placental ABC drug transporters ABCB1, ABCG2, and ABCC2. We demonstrate for the first time that placental ABCB1 significantly reduces mother-to-fetus transport of maraviroc, which suggests that ABCB1 may be responsible for the low cord-blood/maternal-blood ratio observed in humans.

• MeSH
• ABC transportér z rodiny G, člen 2 metabolismus   MeSH
• antagonisté receptoru CCR5 farmakokinetika   terapeutické užití   MeSH
• buňky MDCK MeSH
• HIV infekce farmakoterapie   MeSH
• infekční komplikace v těhotenství farmakoterapie   MeSH
• krysa rodu rattus MeSH
• lidé MeSH
• maravirok farmakokinetika   terapeutické užití   MeSH
• maternofetální výměna látek * MeSH
• modely u zvířat MeSH
• P-glykoprotein metabolismus   MeSH
• permeabilita MeSH
• placenta metabolismus   MeSH
• placentární oběh MeSH
• plod metabolismus   MeSH
• proteiny spojené s mnohočetnou rezistencí k lékům metabolismus   MeSH
• psi MeSH
• těhotenství MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• lidé MeSH
• psi MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND: Bioavailability of tacrolimus (Tac) and cyclosporine is determined by cytochrome P450IIIA and by P-glycoprotein encoded by the CYP3A4/CYP3A5 and ABCB1 genes. Polymorphisms in these genes have been suggested to influence acute rejection and pharmacokinetics in renal transplantation. We aimed to validate these findings in a haplotype analysis. METHODS: A total of 832 renal transplant recipients were genotyped for the CYP3A4 -288A>G, CYP3A5 +6986G>A, ABCB1 +1236C>T, +2677G>T>A, and +3435C>T polymorphisms. Their association with acute rejection and with pharmacokinetic parameters was analyzed in haplotype models. RESULTS: Apart from human leukocyte antigen-DR mismatches, delayed graft function and age at renal transplantation, acute rejection was also predicted by the [ABCB1 +1236C; +2677G; +3435T] haplotype. Allograft survival was determined by donor age, age at renal transplantation, delayed graft function, cold ischemia, and history of more than two acute rejections. Homozygotes for the [CYP3A4 -288A; CYP3A5 +6986G] haplotype achieved earlier therapeutic concentrations of Tac and a higher concentration to dose ratio at week 1. ABCB1 haplotypes did not influence pharmacokinetic parameters. CONCLUSIONS: ABCB1 haplotypes modify the risk of acute rejection, suggesting that ABCB1 allelic arrangement is a stronger regulator of P-glycoprotein activity than single polymorphisms. The risk of acute rejection determined by ABCB1 is independent of pharmacokinetic parameters. CYP3A haplotypes control the bioavailability of Tac, but do not modify the risk of acute rejection.

• MeSH
• alely MeSH
• dítě MeSH
• dospělí MeSH
• financování organizované MeSH
• genotyp MeSH
• haplotypy genetika   MeSH
• homologní transplantace MeSH
• jednonukleotidový polymorfismus genetika   MeSH
• kohortové studie MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• P-glykoprotein genetika   metabolismus   MeSH
• předškolní dítě MeSH
• přežívání štěpu genetika   MeSH
• rejekce štěpu genetika   metabolismus   MeSH
• retrospektivní studie MeSH
• rizikové faktory MeSH
• senioři MeSH
• studie případů a kontrol MeSH
• transplantace ledvin MeSH
• Check Tag
• dítě MeSH
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• senioři MeSH
• ženské pohlaví MeSH

OBJECTIVE AND DESIGN: Tenofovir (TFV) is used in pregnant women as a part of combination antiretroviral treatment to prevent mother-to-child transmission of HIV infection. We aimed to detect whether TFV and/or its prodrug, tenofovir disoproxil fumarate (TDF), are substrates of ATP-binding cassette (ABC) transporters that are functionally expressed in the placenta, namely P-glycoprotein (ABCB1/MDR1), Breast Cancer Resistance Protein (ABCG2/BCRP) and Multidrug Resistance-Associated Protein 2 (ABCC2/MRP2). We employed in-vitro cell-based assays and in-situ animal model to assess possible role of the efflux transporters in transplacental pharmacokinetics of TFV and TDF. METHODS: In-vitro transport assays were performed in MDCKII cells transduced with human ABCB1, ABCG2 or ABCC2. To quantify the effect of these transporters on TFV/TDF transplacental passage, we employed the in-situ model of dually perfused rat term placenta in open and closed setup. RESULTS: In-vitro assays revealed that TDF is a dual substrate of ABCB1 and ABCG2 but not of ABCC2. In contrast, TFV transport was not influenced by any of these transporters. Applying concentration-dependent studies and selective inhibitors, we further confirmed these findings in situ on the organ level; both ABCB1 and ABCG2 limited mother-to-fetus transfer of TDF whereas TFV transplacental passage was not affected by these ABC transporters. CONCLUSION: We propose limited mother-to-fetus transport of both TFV and TDF. While placental transport of TFV is restricted passively, by physical-chemical properties of the molecule, mother-to-fetus passage of TDF is actively hindered by placental ABCB1 and ABCG2 transporters, pumping this compound from trophoblast back to maternal circulation.

• MeSH
• ABC transportéry metabolismus   MeSH
• adenin analogy a deriváty   metabolismus   farmakokinetika   MeSH
• buněčné linie MeSH
• krysa rodu rattus MeSH
• látky proti HIV metabolismus   farmakokinetika   MeSH
• lidé MeSH
• nádorové proteiny metabolismus   MeSH
• organofosfonáty metabolismus   farmakokinetika   MeSH
• P-glykoprotein metabolismus   MeSH
• potkani Wistar MeSH
• proteiny spojené s mnohočetnou rezistencí k lékům metabolismus   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• lidé MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Moderate neonatal jaundice is the most common clinical condition during newborn life. However, a combination of factors may result in acute hyperbilirubinemia, placing infants at risk of developing bilirubin encephalopathy and death by kernicterus. While most risk factors are known, the mechanisms acting to reduce susceptibility to bilirubin neurotoxicity remain unclear. The presence of modifier genes modulating the risk of developing bilirubin-induced brain damage is increasingly being recognised. The Abcb1 and Abcc1 members of the ABC family of transporters have been suggested to have an active role in exporting unconjugated bilirubin from the central nervous system into plasma. However, their role in reducing the risk of developing neurological damage and death during neonatal development is still unknown.To this end, we mated Abcb1a/b-/- and Abcc1-/- strains with Ugt1-/- mice, which develop severe neonatal hyperbilirubinemia. While about 60% of Ugt1-/- mice survived after temporary phototherapy, all Abcb1a/b-/-/Ugt1-/- mice died before postnatal day 21, showing higher cerebellar levels of unconjugated bilirubin. Interestingly, Abcc1 role appeared to be less important.In the cerebellum of Ugt1-/- mice, hyperbilirubinemia induced the expression of Car and Pxr nuclear receptors, known regulators of genes involved in the genotoxic response.We demonstrated a critical role of Abcb1 in protecting the cerebellum from bilirubin toxicity during neonatal development, the most clinically relevant phase for human babies, providing further understanding of the mechanisms regulating bilirubin neurotoxicity in vivo. Pharmacological treatments aimed to increase Abcb1 and Abcc1 expression, could represent a therapeutic option to reduce the risk of bilirubin neurotoxicity.

• MeSH
• bilirubin toxicita   MeSH
• glukuronosyltransferasa fyziologie   MeSH
• lidé MeSH
• modely nemocí na zvířatech * MeSH
• mozeček účinky léků   patologie   MeSH
• myši knockoutované MeSH
• myši MeSH
• neurotoxické syndromy etiologie   metabolismus   patologie   MeSH
• novorozená zvířata MeSH
• novorozenecká hyperbilirubinemie komplikace   metabolismus   patologie   MeSH
• P-glykoprotein genetika   metabolismus   MeSH
• proteiny spojené s mnohočetnou rezistencí k lékům genetika   metabolismus   MeSH
• viabilita buněk MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Ribociclib is a novel cyclin-dependent kinase (CDK) 4 and 6 selective inhibitor that recently gained breakthrough therapy status and global approval for advanced breast cancer treatment. ATP-binding cassette (ABC) transporters may become a site of severe drug interactions and a mechanism of multidrug resistance (MDR) development. With respect to rapid progress of ribociclib in the clinical field, we aimed to identify its interactions with ABC transporters and cytochrome P450 (CYP) isoenzymes and evaluate its potential to overcome transporter-mediated MDR using established in vitro methods. Our data showed accelerated ABCB1 inhibitor LY335979-sensitive, basolateral-to-apical transport of ribociclib across MDCKII-ABCB1 cell monolayers, which identified ribociclib as an ABCB1 substrate. The antiproliferative studies supported this finding by demonstrating significantly higher EC50 value in ABCB1-, but not ABCG2- or ABCC1-expressing MDCKII cells, than in the parent MDCKII cell line. Furthermore, we observed significant inhibitory effects of ribociclib on ABCB1 and ABCG2 transporters and CYP1A2, CYP3A4, CYP3A5, and CYP2C9 isoform activity in human CYP-expressing insect microsomes. The ribociclib-induced ABCB1 and ABCG2 inhibition further reversed daunorubicin and mitoxantrone resistance in MDCKII and human MCF-7 breast carcinoma cell lines, indicating a synergistic antiproliferative effect, without affecting ABCB1 or ABCG2 expression. In summary, our data indicate that ABCB1 affects ribociclib transport across the membranes and the high potential of ribociclib for drug-drug interactions (DDIs) through ABCB1 and ABCG2 transporters and CYP isoforms. Moreover, we demonstrate the beneficial MDR-reversing potential of ribociclib, which could be further exploited in novel anticancer treatment strategies.

• MeSH
• ABC transportér z rodiny G, člen 2 antagonisté a inhibitory   metabolismus   MeSH
• aminopyridiny farmakokinetika   MeSH
• buňky MDCK MeSH
• inhibitory cytochromu P450 farmakokinetika   MeSH
• izoenzymy antagonisté a inhibitory   metabolismus   MeSH
• lékové interakce fyziologie   MeSH
• lidé MeSH
• MFC-7 buňky MeSH
• nádorové proteiny antagonisté a inhibitory   metabolismus   MeSH
• P-glykoproteiny antagonisté a inhibitory   metabolismus   MeSH
• psi MeSH
• puriny farmakokinetika   MeSH
• substrátová specifita účinky léků   fyziologie   MeSH
• systém (enzymů) cytochromů P-450 metabolismus   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• psi MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Low curability of patients diagnosed with acute myeloid leukemia (AML) must be seen as a call for better understanding the disease's mechanisms and improving the treatment strategy. Therapeutic outcome of the crucial anthracycline-based induction therapy often can be compromised by a resistant phenotype associated with overexpression of ABCB1 transporters. Here, we evaluated clinical relevance of ABCB1 in a context of the FMS-like tyrosine kinase 3 (FLT3) inhibitor midostaurin in a set of 28 primary AML samples. ABCB1 gene expression was absolutely quantified, confirming its association with CD34 positivity, adverse cytogenetic risk, and unachieved complete remission (CR). Midostaurin, identified as an ABCB1 inhibitor, increased anthracycline accumulation in peripheral blood mononuclear cells (PBMC) of CD34+ AML patients and those not achieving CR. This effect was independent of FLT3 mutation, indicating even FLT3- AML patients might benefit from midostaurin therapy. In line with these data, midostaurin potentiated proapoptotic processes in ABCB1-overexpressing leukemic cells when combined with anthracyclines. Furthermore, we report a direct linkage of miR-9 to ABCB1 efflux activity in the PBMC and propose miR-9 as a useful prognostic marker in AML. Overall, we highlight the therapeutic value of midostaurin as more than just a FLT3 inhibitor, suggesting its maximal therapeutic outcomes might be very sensitive to proper timing and well-optimized dosage schemes based upon patient's characteristics, such as CD34 positivity and ABCB1 activity. Moreover, we suggest miR-9 as a predictive ABCB1-related biomarker that could be immensely helpful in identifying ABCB1-resistant AML phenotype to enable optimized therapeutic regimen and improved treatment outcome.

• MeSH
• akutní myeloidní leukemie * farmakoterapie   genetika   metabolismus   MeSH
• antracykliny farmakologie   MeSH
• inhibitory proteinkinas farmakologie   MeSH
• leukocyty mononukleární účinky léků   metabolismus   MeSH
• lidé MeSH
• mikro RNA * genetika   metabolismus   MeSH
• mutace MeSH
• P-glykoproteiny * genetika   metabolismus   MeSH
• staurosporin * analogy a deriváty   farmakologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH