The forskolin-induced swelling assay (FIS) in patient-derived intestinal organoids (PDIOs), used to determine in vitro responsiveness to elexacaftor/tezacaftor/ivacaftor (ETI), showed variability in swelling among PDIOs obtained from people with CF (pwCF) carrying the same F508del/F508del CFTR genotype. We aimed to characterise the effect of ETI on the transcriptional activity of PDIOs-derived cells to understand the intracellular processes triggered by ETI and the differences in treatment response. Six high- and six low-responding PDIOs to ETI, derived from F508del/F508del pwCF, were incubated with or without ETI for 2 to 6 h. Gene expression was assessed using 3'-mRNA sequencing and modelled using negative binomial models. Incubation with ETI resulted in a significant upregulation of several biological processes: mostly related to chemokines and signalling, chemotaxis, and tissue development processes. No changes were observed in abundance of the CFTR transcripts or in CFTR-related gene sets and pathways. The genes and pathways associated with ETI did not overlap with those whose expression changed with time only. PDIOs with a high FIS response did not significantly differ in any interpretable gene from the FIS-low organoids. The changes in the PDIOs gene expression upon the exposure to ETI cannot explain differences in the magnitude of PDIOs FIS-measured response to ETI. In conclusion, on incubation with ETI, genes of the CFTR-related pathways do not change their transcriptional activity; instead, overexpression was observed in genes of inflammatory-like cytokine response and receptor activation pathways.
- MeSH
- aktivátory chloridových kanálů terapeutické užití farmakologie MeSH
- aminofenoly * terapeutické užití farmakologie MeSH
- benzodioxoly * terapeutické užití farmakologie MeSH
- chinolony * farmakologie terapeutické užití MeSH
- cystická fibróza * genetika farmakoterapie MeSH
- fixní kombinace léků MeSH
- indoly * farmakologie MeSH
- lidé MeSH
- organoidy * metabolismus MeSH
- protein CFTR genetika MeSH
- pyrazoly * farmakologie MeSH
- pyridiny farmakologie MeSH
- pyrrolidiny farmakologie MeSH
- pyrroly farmakologie MeSH
- stanovení celkové genové exprese metody MeSH
- střeva účinky léků MeSH
- transkriptom MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Anthelmintic drugs are widespread environmental contaminants, but their impact is still poorly understood. Although contact of parasitic nematode Haemonchus contortus with traces of anthelmintic drug albendazole (ABZ) altered the expression and activity of several UDP-glycosyl transferases (UGTs) and P-glycoproteins (Pgps, belonging to ABC-transporters), key enzymes in endogenous and xenobiotic metabolism, it is not known whether these changes will last during the life cycle and pass to the next generations. In the present study simulating the environmental-like exposure, free-living stages of H. contortus were exposed or unexposed to a sub-lethal dose of ABZ and its transformation products (ABZs) during L3 development. The L3 served for lambs' infection and obtaining of H. contortus adults and eggs, which were again exposed or unexposed to ABZs during L3 development. The expression pattern of UGTs and Pgps was analysed and compared in the first generation of L3, in the adults, and in the second generation of L3. The results showed that ABZs exposition during larvae development altered the expression of several ugt and pgp genes in L3 and adults. The intrageneration stability of ABZs-evoked changes was observed in the case of three genes, four genes maintained the intergeneration stability. Interestingly, ABZs-induced changes in the expression of some genes became apparent only in the second generation of L3. Taking together, contact of free-living stages of H. contortus with traces of ABZs in the environment evokes changes in the expression of certain UGTs and Pgps, with some of these changes being intra- and inter-generation stable.
Chronic Kidney Disease (CKD) is associated with heightened risk of thrombosis. Prescription of anticoagulants is key to manage it; however, CKD patients have shown an increased risk of bleeding under anticoagulation therapy compared to non-CKD patients. We hypothesized that the sex could modify the metabolism of indoxyl sulfate (IS), a uremic toxin and Apixaban. Our intoxication model shows that higher doses of IS and apixaban accumulate in the plasma of female mice because of expression differences in efflux transporters and cytochromes in the liver, ileum and kidneys, when compared to males. Furthermore, we found that accumulation of apixaban in females contributes to increased bleeding. Transcriptional analysis of liver samples revealed elevated Sult1a1 but reduced Abcg2 and Cyp3a11 in female mice, while in the kidneys the expression rates of Oat1 and Oat3 were respectively lower and higher than those observed in males, potentially affecting drug clearance. Whole proteomics liver analysis confirmed the previous transcriptional results at the protein level and revealed that sex had a major influence in regulating both coagulation and drug metabolism pathways. Thus, our findings underline the need for inclusive clinical and preclinical trials to accurately reflect sex-specific metabolic variations, and to consider CKD-specific changes to optimize dosing, minimize side effects, and improve patient outcomes.
- MeSH
- ABC transportér z rodiny G, člen 2 metabolismus genetika MeSH
- antikoagulancia aplikace a dávkování metabolismus MeSH
- chronická renální insuficience metabolismus farmakoterapie MeSH
- cytochrom P-450 CYP3A metabolismus genetika MeSH
- indican * metabolismus krev MeSH
- játra * metabolismus účinky léků MeSH
- krvácení metabolismus MeSH
- ledviny metabolismus účinky léků MeSH
- myši inbrední C57BL MeSH
- myši MeSH
- přenašeče organických aniontů nezávislé na sodíku metabolismus genetika MeSH
- protein 1 přenášející organické anionty metabolismus genetika MeSH
- pyrazoly * farmakologie MeSH
- pyridony * aplikace a dávkování metabolismus farmakologie MeSH
- sexuální faktory MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
This is the second in a series of four papers updating the European Cystic Fibrosis Society (ECFS) standards for the care of people with CF. This paper focuses on establishing and maintaining health. The guidance is produced using an evidence-based framework and with wide stakeholder engagement, including people from the CF community. Authors provided a narrative description of their topic and statements, which were more directive. These statements were reviewed by a Delphi exercise, achieving good levels of agreement from a wide group for all statements. This guidance reinforces the importance of a multi-disciplinary CF team, but also describes developing models of care including virtual consultations. The framework for health is reinforced, including the need for a physically active lifestyle and the strict avoidance of all recreational inhalations, including e-cigarettes. Progress with cystic fibrosis transmembrane conductance regulator (CFTR) modulator therapy is reviewed, including emerging adverse events and advice for dose reduction and interruption. This paper contains guidance that is pertinent to all people with CF regardless of age and eligibility for and access to modulator therapy.
- MeSH
- aktivátory chloridových kanálů terapeutické užití MeSH
- aminofenoly * terapeutické užití MeSH
- benzodioxoly * terapeutické užití MeSH
- chinoliny terapeutické užití MeSH
- chinolony * terapeutické užití MeSH
- cystická fibróza * farmakoterapie genetika MeSH
- dospělí MeSH
- exokrinní pankreatická insuficience farmakoterapie etiologie genetika MeSH
- fixní kombinace léků MeSH
- genotyp MeSH
- indoly * terapeutické užití MeSH
- lidé MeSH
- protein CFTR * genetika MeSH
- pyrazoly * terapeutické užití MeSH
- pyridiny terapeutické užití MeSH
- pyrrolidiny terapeutické užití MeSH
- Check Tag
- dospělí MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- dopisy MeSH
- kazuistiky MeSH
OBJECTIVES: To determine whether selected single nucleotide polymorphisms (SNPs) of genes encoding proteins responsible for the activation, transport, or metabolism of dabigatran and apixaban might be associated with a risk of gastrointestinal bleeding in a cohort of adult patients treated with these drugs. No previous study has focused specifically on the association with gastrointestinal bleeding. MATERIALS AND METHODS: Ninety-one patients treated with dabigatran or apixaban were genotyped for selected polymorphisms. The following polymorphisms were studied: ABCB1 gene rs1045642, rs4148738, rs1128503 and rs2032582; CES1 gene rs2244613, rs8192935 and rs2244614; and SULT1A1 gene rs9282861 and SULT1A2 gene rs1136703. Two groups divided by particular drugs and genotypes were compared in terms of the presence (bleeding group) or absence (nonbleeding group) of gastrointestinal bleeding. The genotype distribution was expressed via dominant and recessive models. RESULTS: In patients treated either with dabigatran or with apixaban, no evidence was found to support the association of gastrointestinal bleeding with any genotype for any of the studied SNPs. CONCLUSION: In both dabigatran- and apixaban-treated patients, no associations between the selected polymorphisms and gastrointestinal bleeding risk were found, however the results should be interpreted with caution because of the small cohort size.
- MeSH
- antitrombiny škodlivé účinky terapeutické užití MeSH
- dabigatran * škodlivé účinky MeSH
- dospělí MeSH
- farmakogenetika MeSH
- gastrointestinální krvácení * genetika chemicky indukované MeSH
- genotyp MeSH
- inhibitory faktoru Xa škodlivé účinky terapeutické užití MeSH
- jednonukleotidový polymorfismus * MeSH
- lidé středního věku MeSH
- lidé MeSH
- P-glykoproteiny MeSH
- pyrazoly * škodlivé účinky terapeutické užití MeSH
- pyridony * škodlivé účinky terapeutické užití MeSH
- senioři nad 80 let MeSH
- senioři 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
CFTR is a membrane protein that functions as an ion channel. Mutations that disrupt its biosynthesis, trafficking or function cause cystic fibrosis (CF). Here, we present a novel in vitro model system prepared using CRISPR/Cas9 genome editing with endogenously expressed WT-CFTR tagged with a HiBiT peptide. To enable the detection of CFTR in the plasma membrane of live cells, we inserted the HiBiT tag in the fourth extracellular loop of WT-CFTR. The 11-amino acid HiBiT tag binds with high affinity to a large inactive subunit (LgBiT), generating a reporter luciferase with bright luminescence. Nine homozygous clones with the HiBiT knock-in were identified from the 182 screened clones; two were genetically and functionally validated. In summary, this work describes the preparation and validation of a novel reporter cell line with the potential to be used as an ultimate building block for developing unique cellular CF models by CRISPR-mediated insertion of CF-causing mutations.
- MeSH
- buněčná membrána metabolismus MeSH
- buněčné linie MeSH
- CRISPR-Cas systémy genetika MeSH
- cystická fibróza * genetika metabolismus MeSH
- lidé MeSH
- protein CFTR * genetika metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Physiology-based pharmacokinetic modeling suggests that rifabutin can out-balance P-glycoprotein (P-gp) induction by concurrent P-gp inhibition. However, clinical or experimental evidence for this Janus-faced rifabutin effect is missing. Consequently, LS180 cells were exposed to a moderately (2 μM) and strongly (10 μM) P-gp-inducing concentration of rifampicin or rifabutin for 6 days. Cellular accumulation of the fluorescent P-gp substrate rhodamine 123 was evaluated using flow cytometry, either without (induction only) or with adding rifamycin drug to the cells during the rhodamine 123 efflux phase (induction + potential inhibition). Rhodamine 123 accumulation was decreased similarly by both drugs after 6-day exposure (2 μM: 55% residual fluorescence compared to non-induced cells, P < 0.01; 10 μM: 30% residual fluorescence compared to non-induced cells, P < 0.001), indicating P-gp induction. Rhodamine 123 influx transporters mRNA expressions were not affected, excluding off-target effects. Acute re-exposure to rifabutin, however, considerably re-increased rhodamine 123 accumulation (2 μM induction: re-increase by 55%, P < 0.01; 10 μM induction: 49% re-increase, P < 0.001), suggesting P-gp inhibition. In contrast, rifampicin only had weak effects (2 μM induction: no re-increase; 10 μM induction: 16% re-increase; P < 0.05). Molecular docking analysis eventually revealed that rifabutin has a higher binding affinity to the inhibitor binding site of P-gp than rifampicin (ΔG (kcal/mol) = -11.5 vs -5.3). Together, this study demonstrates that rifabutin can at least partly mask P-gp induction by P-gp inhibition, mediated by high affinity binding to the inhibitory site of P-gp.
- MeSH
- ABC transportéry MeSH
- cytostatické látky aplikace a dávkování terapeutické užití MeSH
- doxorubicin aplikace a dávkování terapeutické užití MeSH
- geny MDR MeSH
- inhibitory HIV-proteasy farmakologie terapeutické užití MeSH
- inhibitory proteas farmakologie terapeutické užití MeSH
- lidé MeSH
- Lopinavir aplikace a dávkování terapeutické užití MeSH
- mnohočetná léková rezistence * účinky léků MeSH
- nádory farmakoterapie MeSH
- nanočásticový lékový transportní systém MeSH
- P-glykoprotein metabolismus účinky léků MeSH
- protinádorové látky aplikace a dávkování terapeutické užití MeSH
- transkripční faktor STAT3 metabolismus MeSH
- Check Tag
- lidé MeSH
INTRODUCTION: In silico tools capable of predicting the functional consequences of genomic differences between individuals, many of which are AI-driven, have been the most effective over the past two decades for non-synonymous single nucleotide variants (nsSNVs). When appropriately selected for the purpose of the study, a high predictive performance can be expected. In this feasibility study, we investigate the distribution of nsSNVs with an allele frequency below 5%. To classify the putative functional consequence, a tier-based filtration led by AI-driven predictors and scoring system was implemented to the overall decision-making process, resulting in a list of prioritised genes. METHODS: The study has been conducted on breast cancer patients of homogeneous ethnicity. Germline rare variants have been sequenced in genes that influence pharmacokinetic parameters of anticancer drugs or molecular signalling pathways in cancer. After AI-driven functional pathogenicity classification and data mining in pharmacogenomic (PGx) databases, variants were collapsed to the gene level and ranked according to their putative deleterious role. RESULTS: In breast cancer patients, seven of the twelve genes prioritised based on the predictions were found to be associated with response to oncotherapy, histological grade, and tumour subtype. Most importantly, we showed that the group of patients with at least one rare nsSNVs in cystic fibrosis transmembrane conductance regulator (CFTR) had significantly reduced disease-free (log rank, p = 0.002) and overall survival (log rank, p = 0.006). CONCLUSION: AI-driven in silico analysis with PGx data mining provided an effective approach navigating for functional consequences across germline genetic background, which can be easily integrated into the overall decision-making process for future studies. The study revealed a statistically significant association with numerous clinicopathological parameters, including treatment response. Our study indicates that CFTR may be involved in the processes influencing the effectiveness of oncotherapy or in the malignant progression of the disease itself.
- MeSH
- dospělí MeSH
- frekvence genu MeSH
- jednonukleotidový polymorfismus MeSH
- lidé středního věku MeSH
- lidé MeSH
- nádory prsu * genetika farmakoterapie patologie MeSH
- protein CFTR * genetika MeSH
- senioři MeSH
- studie proveditelnosti * MeSH
- umělá inteligence * MeSH
- výsledek terapie MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
