RESUMO
5-fluorouracil (5-FU) is an inexpensive treatment for colon cancer; however, its efficacy is limited by chemoresistance. This study investigates the combination therapy approach of 5-FU with Sitagliptin (Sita), a diabetic drug with potential cancer-modulating effects. The combination was evaluated in vitro and in silico, focusing on the effects of Sita and 5-FU on colon cancer cells. The results showed that the addition of Sita significantly decreased the IC50 of 5-FU compared to 5-Fu monotherapy. The study also found that Sita and 5-FU interact synergistically, with a combination index below 1. Sita successfully lowered the 5-FU dosage reduction index, decreasing the expression of MDR1 mRNA and p-AKT and NFκB2 subunits p100/p52 protein. Molecular docking analyses confirmed Sita's antagonistic action on MDR1 and thymidylate synthase proteins. The study concludes that sitagliptin can target MDR1, increase apoptosis, and significantly reduce the expression of p-AKT and NFκB2 cell-survival proteins. These effects sensitize colon cancer cells to 5-FU. Repurposing sitagliptin may enhance the anticancer effects of 5-FU at lower dosages.
Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP , Neoplasias do Colo , Sinergismo Farmacológico , Fluoruracila , Proteínas Proto-Oncogênicas c-akt , Fosfato de Sitagliptina , Humanos , Fosfato de Sitagliptina/farmacologia , Fluoruracila/farmacologia , Neoplasias do Colo/metabolismo , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/patologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Regulação para Baixo/efeitos dos fármacos , Linhagem Celular Tumoral , Simulação de Acoplamento MolecularRESUMO
Pancreatic malignancy is the fourth cause of cancer-related death in Western countries and is predicted to become the second leading cause of cancer-related mortality by 2030. The standard therapies (FOLFIRINOX and gemcitabine with nab-paclitaxel) are not resolutive because this type of cancer is also characterized by a high chemoresistance, due in part to the activity of the ATP Binding Cassette (ABC) pumps accounting for the reduction in the intracellular concentration of the drugs. In this work, we analyze the occurrence of single-nucleotide polymorphisms (SNPs) in the MDR-1 gene, in different pancreatic cancer cell lines, and in tissues from pancreatic cancer patients by DNA sequencing, as well as the expression levels of MDR-1 mRNA and protein, by qRT-PCR and Western Blot analysis. We found that gemcitabine-resistant cells, in conjunction with homozygosis of analyzed SNPs, showed high MDR-1 basal levels with further increases after gemcitabine treatment. Nevertheless, we did not observe in the human PDAC samples a correlation between the level of MDR-1 mRNA and protein expression and SNPs. Preliminary, we conclude that in our small cohort, these SNPs cannot be used as molecular markers for predicting the levels of MDR-1 mRNA/protein levels and drug responses in patients with PDAC.
Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP , Desoxicitidina , Resistencia a Medicamentos Antineoplásicos , Gencitabina , Neoplasias Pancreáticas , Polimorfismo de Nucleotídeo Único , Humanos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/metabolismo , Resistencia a Medicamentos Antineoplásicos/genética , Linhagem Celular Tumoral , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacologia , Desoxicitidina/uso terapêutico , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Masculino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Feminino , Pessoa de Meia-Idade , Idoso , RNA Mensageiro/genética , RNA Mensageiro/metabolismoRESUMO
Prostate cancer (PCa) is the second leading cause of cancer-related death in American men. PCa that relapses after hormonal therapies, referred to as castration resistant PCa (CRPC), often presents with metastases (mCRPC) that are the major cause of mortality. The few available therapies for mCRPC patients include taxanes docetaxel (DTX) and cabazitaxel (CBZ). However, development of resistance limits their clinical use. Mechanistically, resistance arises through upregulation of multidrug resistance (MDR) proteins such as MDR1/ABCB1, making ABCB1 an attractive therapeutic target. Yet, ABCB1 inhibitors failed to be clinically useful due to low specificity and toxicity issues. To study taxanes resistance, we produced CBZ resistant C4-2B cells (RC4-2B) and documented resistance to both CBZ and DTX in cell culture and in 3D prostaspheres settings. RNAseq identified increased expression of ABCB1 in RC4-2B, that was confirmed by immunoblotting and immunofluorescent analysis. ABCB1-specific inhibitor elacridar reversed CBZ and DTX resistance in RC4-2B cells, confirming ABCB1-mediated resistance mechanism. In a cell-based screen using a curated library of cytotoxic drugs, we found that DNA damaging compounds Camptothecin (CPT) and Cytarabine (Ara-C) overcame resistance as seen by similar cytotoxicity in parental C4-2B and resistant RC4-2B. Further, these compounds were cytotoxic to multiple PC cells resistant to taxanes with high ABCB1 expression and, therefore, can be used to conquer the acquired resistance to taxanes in PCa. Finally, inhibition of cyclin-dependent kinases 4/6 (CDK4/6) with small molecule inhibitors (CDK4/6i) potentiated cytotoxic effect of CPT or Ara-C in both parental and resistant cells. Overall, our findings indicate that DNA damaging agents CPT and Ara-C alone or in combination with CDK4/6i can be suggested as a new treatment regimen in CRPC patients, including those that are resistant to taxanes.
Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP , Docetaxel , Resistência a Múltiplos Medicamentos , Resistencia a Medicamentos Antineoplásicos , Neoplasias de Próstata Resistentes à Castração , Taxoides , Humanos , Masculino , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/patologia , Neoplasias de Próstata Resistentes à Castração/metabolismo , Neoplasias de Próstata Resistentes à Castração/genética , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Linhagem Celular Tumoral , Docetaxel/farmacologia , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Taxoides/farmacologia , Taxoides/uso terapêutico , Antineoplásicos/farmacologiaRESUMO
BACKGROUND: Pharmacotherapy for brain diseases is severely compromised by the blood-brain barrier (BBB). ABCB1 and ABCG2 are drug transporters that restrict drug entry into the brain and their inhibition can be used as a strategy to boost drug delivery and pharmacotherapy for brain diseases. METHODS: We employed elacridar and tariquidar in mice to explore the conditions for effective inhibition at the BBB. Abcg2;Abcb1a/b knockout (KO), Abcb1a/b KO, Abcg2 KO and wild-type (WT) mice received a 3 h i.p. infusion of a cocktail of 8 typical substrate drugs in combination with elacridar or tariquidar at a range of doses. Abcg2;Abcb1a/b KO mice were used as the reference for complete inhibition, while single KO mice were used to assess the potency to inhibit the remaining transporter. Brain and plasma drug levels were measured by LC-MS/MS. RESULTS: Complete inhibition of ABCB1 at the BBB is achieved when the elacridar plasma level reaches 1200 nM, whereas tariquidar requires at least 4000 nM. Inhibition of ABCG2 is more difficult. Elacridar inhibits ABCG2-mediated efflux of weak but not strong ABCG2 substrates. Strikingly, tariquidar does not enhance the brain uptake of any ABCG2-subtrate drug. Similarly, elacridar, but not tariquidar, was able to inhibit its own brain efflux in ABCG2-proficient mice. The plasma protein binding of elacridar and tariquidar was very high but similar in mouse and human plasma, facilitating the translation of mouse data to humans. CONCLUSIONS: This work shows that elacridar is an effective pharmacokinetic-enhancer for the brain delivery of ABCB1 and weaker ABCG2 substrate drugs when a plasma concentration of 1200 nM is exceeded.
Assuntos
Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Acridinas , Barreira Hematoencefálica , Encéfalo , Camundongos Knockout , Tetra-Hidroisoquinolinas , Animais , Tetra-Hidroisoquinolinas/farmacologia , Tetra-Hidroisoquinolinas/administração & dosagem , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/antagonistas & inibidores , Encéfalo/metabolismo , Encéfalo/efeitos dos fármacos , Camundongos , Acridinas/farmacologia , Acridinas/administração & dosagem , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/efeitos dos fármacos , Quinolinas/farmacologia , Quinolinas/farmacocinética , Quinolinas/administração & dosagem , Sistemas de Liberação de Medicamentos/métodos , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/antagonistas & inibidores , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Masculino , Camundongos Endogâmicos C57BLRESUMO
BACKGROUND: Helicobacter pylori eradication therapy based on antimicrobial susceptibility in Vietnamese children currently get low efficiency. There are causes of treatment failure, among host genetic factors namely MDR1 C3435T and CYP2C19 affect the absorption and metabolism of proton pump inhibitors - a crucial component of eradication therapy. The study aimed to investigate the effect of MDR1 C3435T and CYP2C19 genetic polymorphisms on the cure rate. METHODS: 207 pediatric patients with gastritis and peptic ulcer infecting Helicobacter pylori completed the eradication therapy based on antimicrobial susceptibility with proton pump inhibitor esomeprazole. Eradication efficacy was assessed after at least 4 weeks by the urease breath test. MDR1 C3435T genetic polymorphism and CYP2C19 genotype were determined using a sequencing method based on Sanger's principle. RESULTS: Among 207 children recruited in this study, the ratio of CYP2C19 EM, IM, and PM phenotypes was 40.1%, 46.4%, and 16.9%, respectively. The patient with MDR1 3435 C/C polymorphism accounted for 43.0%, MDR1 3435 C/T was 40.1%, and MDR1 3435T/T was 16.9%. The cure rate of Helicobacter pylori infection in patients with CYP2C19 EM genotype was 78.3%; 83.3% of those with the IM genotype, and PM genotype was 96,4% (p = 0.07). Successful eradication rates for Helicobacter pylori were 85.4%, 86.7%, and 68.6% in patients with the MDR1 3435 C/C, C/T, and T/T, respectively (p = 0.02). Multiple logistic regression analysis found that MDR1 C3435T genetic polymorphisms of patients were significant independent risk factors for treatment failure, and CYP2C19 genotype did not affect Helicobacter pylori eradication. CONCLUSIONS: The Helicobacter pylori eradication rates by regimens based on antibiotic susceptibility and esomeprazole were not significantly different between the CYP2C19 phenotypes. The MDR1 C3435T polymorphism is one of the factors impacting Helicobacter pylori eradication results in children.
Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP , Citocromo P-450 CYP2C19 , Gastrite , Infecções por Helicobacter , Helicobacter pylori , Úlcera Péptica , Inibidores da Bomba de Prótons , Humanos , Citocromo P-450 CYP2C19/genética , Infecções por Helicobacter/tratamento farmacológico , Infecções por Helicobacter/genética , Helicobacter pylori/efeitos dos fármacos , Criança , Masculino , Feminino , Vietnã , Gastrite/tratamento farmacológico , Gastrite/microbiologia , Gastrite/genética , Úlcera Péptica/tratamento farmacológico , Úlcera Péptica/genética , Úlcera Péptica/microbiologia , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Inibidores da Bomba de Prótons/uso terapêutico , Adolescente , Pré-Escolar , Genótipo , Polimorfismo Genético , Resultado do Tratamento , Esomeprazol/uso terapêutico , Antibacterianos/uso terapêuticoRESUMO
BACKGROUND: Cardiac dysfunction, including arrhythmias, may be one of the main clinical manifestations of Becker muscular dystrophy (BMD). Amiodarone is widely used to treat arrhythmia. However, multi-systemic toxicity caused by amiodarone, especially hepatotoxicity, should not be neglected. Here, we introduce a novel case of multi-systemic amiodarone toxicity involving the liver, renal and coagulation in BDM patient with ABCB4 gene mutation. CASE PRESENTATION: We present a case of a 16-year-old boy admitted with heart failure and atrial fibrillation (AF). He was diagnosed with Becker muscular dystrophy (BMD) and gene testing showed comorbid mutations in gene DMD, ABCB4 and DSC2. Amiodarone was prescribed to control the paroxysmal atrial fibrillation intravenously. However, his liver enzyme levels were sharply elevated, along with cardiac shock, renal failure and coagulation disorders. After bedside continuous renal replacement therapy, the patient's liver function and clinical status rehabilitated. CONCLUSIONS: ABCB4 gene mutation might be involved in amiodarone-induced hepatotoxicity. Studies in a cohort might help to prove this hypothesis in the future.
Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP , Amiodarona , Antiarrítmicos , Insuficiência Cardíaca , Distrofia Muscular de Duchenne , Mutação , Humanos , Amiodarona/efeitos adversos , Amiodarona/administração & dosagem , Masculino , Adolescente , Insuficiência Cardíaca/induzido quimicamente , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Antiarrítmicos/efeitos adversos , Antiarrítmicos/uso terapêutico , Antiarrítmicos/administração & dosagem , Distrofia Muscular de Duchenne/tratamento farmacológico , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/complicações , Doença Hepática Induzida por Substâncias e Drogas/genética , Doença Hepática Induzida por Substâncias e Drogas/etiologia , Fibrilação Atrial/tratamento farmacológicoRESUMO
Introduction: Pharmacogenetic markers, such as the ATP Binding Cassette (ABCB1) and cytochrome P450 (CYP) 3A5 enzymes, play a crucial role in personalized medicine by influencing drug efficacy and toxicity based on individuals' or populations' genetic variations.This study aims to investigate the genetic polymorphisms of CYP3A5 (rs776746) and ABCB1 (rs1045642) in the West Algerian population and compare the genotypes and allelic distributions with those of various ethnic groups. Methods: The study involved 472 unrelated healthy subjects from the Western Algerian population. DNA genotyping was performed using TaqMan allelic discrimination assay. The variants in our population were compared to those in other ethnic groups available in the 1000 Genomes Project. Genotype and allele frequencies were calculated using the chi-square test and the Hardy-Weinberg equilibrium (HWE). Results: The minor allele frequencies were found to be 0.21 for CYP3A5 6986A and 0.34 for ABCB1 3435T. These frequencies were similar to those observed in North African populations, while notable differences were observed in comparison to certain Caucasian and African populations. Conclusion: The difference in the allelic and genotypic distribution of these polymorphisms emphasize the need for dose adjustments in drugs metabolized by CYP3A5 and transported by ABCB1 to optimize treatments outcomes.
Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP , Citocromo P-450 CYP3A , Frequência do Gene , Genótipo , Polimorfismo de Nucleotídeo Único , Humanos , Citocromo P-450 CYP3A/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Argélia , Masculino , Feminino , Adulto , Farmacogenética , Pessoa de Meia-Idade , População Negra/genética , Alelos , Adulto JovemRESUMO
Previous research highlighted the involvement of the cannabinoid CB1 receptor in regulating the physiology of hepatocytes and hepatic stellate cells. The inhibition of the CB1 receptor via peripherally restricted CB1 receptor inverse agonist JD5037 has shown promise in inhibiting liver fibrosis in mice treated with CCl4. However, its efficacy in phospholipid transporter-deficiency-induced liver fibrosis remains uncertain. In this study, we investigated the effectiveness of JD5037 in Mdr2-/- mice. Mdr2 (Abcb4) is a mouse ortholog of the human MDR3 (ABCB4) gene encoding for the canalicular phospholipid transporter. Genetic disruption of the Mdr2 gene in mice causes a complete absence of phosphatidylcholine from bile, leading to liver injury and fibrosis. Mdr2-/- mice develop spontaneous fibrosis during growth. JD5037 was orally administered to the mice for four weeks starting at eight weeks of age. Liver fibrosis, bile acid levels, inflammation, and injury were assessed. Additionally, JD5037 was administered to three-week-old mice to evaluate its preventive effects on fibrosis development. Our findings corroborate previous observations regarding global CB1 receptor inverse agonists. Four weeks of JD5037 treatment in eight-week-old Mdr2-/- mice with established fibrosis led to reduced body weight gains. However, contrary to expectations, JD5037 significantly exacerbated liver injury, evidenced by elevated serum ALT and ALP levels and exacerbated liver histology. Notably, JD5037-treated Mdr2-/- mice exhibited significantly heightened serum bile acid levels. Furthermore, JD5037 treatment intensified liver fibrosis, increased fibrogenic gene expression, stimulated ductular reaction, and upregulated hepatic proinflammatory cytokines. Importantly, JD5037 failed to prevent liver fibrosis formation in three-week-old Mdr2-/- mice. In summary, our study reveals the exacerbating effect of JD5037 on liver fibrosis in genetically MDR2-deficient mice. These findings underscore the need for caution in the use of peripherally restricted CB1R inverse agonists for liver fibrosis treatment, particularly in cases of dysfunctional hepatic phospholipid transporter.
Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP , Membro 4 da Subfamília B de Transportadores de Cassetes de Ligação de ATP , Cirrose Hepática , Receptor CB1 de Canabinoide , Animais , Camundongos , Receptor CB1 de Canabinoide/genética , Receptor CB1 de Canabinoide/metabolismo , Receptor CB1 de Canabinoide/agonistas , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/deficiência , Cirrose Hepática/patologia , Cirrose Hepática/metabolismo , Cirrose Hepática/induzido quimicamente , Cirrose Hepática/tratamento farmacológico , Cirrose Hepática/genética , Fígado/efeitos dos fármacos , Fígado/patologia , Fígado/metabolismo , Masculino , Camundongos Knockout , Ácidos e Sais Biliares/metabolismo , Agonismo Inverso de Drogas , Camundongos Endogâmicos C57BLRESUMO
P-glycoprotein (P-gp) mediated multidrug resistance (MDR) is the leading cause of chemotherapy failure since it causes the efflux of chemotherapeutic drugs from the cancer cells. Solasodine, a steroidal alkaloid and oxaspiro compound, present in the Solanaceae family showed significant cytotoxic effects on various cancer cells. However, the effect of solasodine on reversing P-gp mediated drug resistance is still unknown. Primarily in this study, the integrative network pharmacology analysis found 71 common targets between solasodine and cancer MDR, among them NF-κB was found as a potential target. The results of immunofluorescence analysis showed that solasodine significantly inhibits NF-κB-p65 nuclear translocation which caused downregulated P-gp expression in KBChR-8-5 cells. Further, solasodine binds to the active sites of the TMD region of P-gp and inhibits P-gp transport activity. Moreover, solasodine significantly promotes doxorubicin intracellular accumulation in the drug resistant cells. Solasodine reduced the fold resistance and synergistically sensitized doxorubicin's therapeutic effects in KBChR-8-5 cells. Additionally, the solasodine and doxorubicin combination treatment increased the apoptotic cell populations and G2/M phase cell cycle arrest in KBChR-8-5 cells. The MDR tumor bearing xenograft mice showed tumor-suppressing characteristics and P-gp downregulation during the combination treatment of solasodine and doxorubicin. These results indicate that solasodine targets NF-κB signaling to downregulate P-gp overexpression, inhibit P-gp transport activity, and enhance chemosensitization in MDR cancer cells. Considering its multifaceted impact, solasodine represents a potent natural fourth-generation P-gp modulator for reversing MDR in cancer.
Assuntos
Apoptose , Doxorrubicina , Resistência a Múltiplos Medicamentos , Resistencia a Medicamentos Antineoplásicos , Camundongos Nus , NF-kappa B , Transdução de Sinais , Alcaloides de Solanáceas , Humanos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Animais , Alcaloides de Solanáceas/farmacologia , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , NF-kappa B/metabolismo , Camundongos , Doxorrubicina/farmacologia , Linhagem Celular Tumoral , Apoptose/efeitos dos fármacos , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Camundongos Endogâmicos BALB C , Ensaios Antitumorais Modelo de Xenoenxerto , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Neoplasias/patologia , Proliferação de Células/efeitos dos fármacos , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/genéticaRESUMO
Objective: To investigate the pathogenic mechanism and clinical characteristics of the novel splicing variant of ATP-binding cassette subfamily B member 4 (ABCB4) and provide a basis for subsequent genetic diagnosis. Methods: The clinical data of a 5-year-old child with cholestatic liver disease admitted to the Beijing Children's Hospital of Capital Medical University was retrospectively analyzed. The pathogenic variations were detected by whole exome sequencing and verified by Sanger sequencing, and bioinformatics was used to predict the pathogenicity of the mutation sites. Possible pathogenic variations were verified in vitro by Minigene assay. The clinical outcome was followed after discharge from hospital. Results: The 5-year-old boy had developed cholestasis at the age of 11 months. His physical examination showed obvious enlargement of the liver and spleen. Cholestatic cirrhosis was diagnosed by liver function tests, abdominal ultrasonography, liver biopsy and pathology. The results of genetic analysis showed that the patient was a complex heterozygote of the ABCB4 gene, with a pathogenic mutation c.2860G>A and a novel mutation c.2065-8T>G, derived from the mother and father respectively. The conservative prediction of the c.2065-8T>G site showed that this region was highly conserved and may affect splicing. Minigene assay results confirmed that the c.2065-8T>G mutation resulted in a 7 bp retention of intron 16 in the mature mRNA. In the absence of nonsense-mediated mRNA decay, the amino acid frameshift forms a truncated protein, which is represented by p.Glu689ValfsTer19. The patient was diagnosed as progressive familial intrahepatic cholestasis type 3 (PFIC3) and treated with ursodeoxycholic acid (UDCA). His clinical symptoms improved during 18 months of follow-up. Conclusions: The c.2065-8T>G variant is confirmed to affect the splicing process and exhibits complex heterozygosity with c.2860G>A, which is identified as the cause of the disease. PFIC3 children with this variant showed cholestatic liver disease as the main manifestation with a slow progression and was sensitive to treatment with UDCA.
Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP , Colestase Intra-Hepática , Mutação , Fenótipo , Humanos , Masculino , Colestase Intra-Hepática/genética , Colestase Intra-Hepática/diagnóstico , Pré-Escolar , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/deficiência , Sequenciamento do Exoma , Genótipo , Heterozigoto , Estudos Retrospectivos , Ácido Ursodesoxicólico/uso terapêutico , Fígado/patologiaRESUMO
BACKGROUND: The burden of chronic kidney disease (CKD) and kidney failure in Ghana is on the ascendency, with the prevalence of CKD estimated at 13.3%. Patients with CKD who progress to kidney failure require life sustaining kidney replacement therapy (KRT) which is almost exclusively available in Ghana as haemodialysis. Kidney transplantation is considered the best KRT option for patients with irreversible kidney failure due to its relative cost efficiency as well as its superiority in terms of survival and quality of life. However, because transplants may trigger an immune response with potential organ rejection, immunosuppressants such as tacrolimus dosing are required. OBJECTIVE: This study sought to determine single nucleotide polymorphisms in CYP3A5, CYP3A4 and MDR1 genes that affect the pharmacokinetics of Tacrolimus in a population of Ghanaian patients with kidney failure. METHOD: This cross-sectional study comprised of 82 kidney failure patients undergoing maintenance haemodialysis at the Renal and Dialysis unit of Korle-Bu Teaching Hospital (KBTH). Clinical and demographic data were collected and genomic DNA isolated. Samples were genotyped for specific SNPs using Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP). RESULTS: Participants, 58/82 (70.73%) harbored the wildtype CYP3A5*1/*1 AA genotype, 20/82 (24.39%) carried the heterozygous CYP3A5*1/*3 AG genotype, and 4/82 (4.88%) had the homozygous mutant CYP3A5*3/*3 GG genotype. Also, 6/82 (7.32%) carried the wildtype AA genotype, 11/82 (13.41%) had the heterozygous AG genotype, and 65/82 (79.27%) harbored the homozygous mutant GG genotype of CYP3A4*1B (-290 A>G). For MDR1_Ex21 (2677 G>T), 81/82 (98.78%) carried the wildtype GG genotype, while 1/82 (1.22%) had the heterozygous GT genotype. For MDR1_Ex26 (3435 C>T), 63/82 (76.83%) had the wildtype CC genotype, while 18/82 (21.95%) carried the heterozygous CT genotype, and 1/82 (1.22%) harbored the mutant TT genotype. CONCLUSION: SNPs in CYP3A4, CYP3A5, and MDR1 genes in a population of Ghanaian kidney failure patients were described. The varying SNPs of the featured genes suggest the need to consider the genetic status of Ghanaians kidney failure patients prior to transplantation and tacrolimus therapy.
Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP , Citocromo P-450 CYP3A , Imunossupressores , Polimorfismo de Nucleotídeo Único , Tacrolimo , Centros de Atenção Terciária , Humanos , Polimorfismo de Nucleotídeo Único/genética , Gana/epidemiologia , Feminino , Masculino , Citocromo P-450 CYP3A/genética , Tacrolimo/farmacocinética , Tacrolimo/administração & dosagem , Pessoa de Meia-Idade , Adulto , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Imunossupressores/administração & dosagem , Estudos Transversais , Insuficiência Renal/genética , Idoso , Diálise RenalRESUMO
Bictegravir, a key second-generation integrase strand transfer inhibitor in the treatment of HIV, is subject to active efflux transport mediated by ABCB1 (P-glycoprotein). Several coding variants of ABCB1 have been described and associated with variable effects on substrate drugs pharmacokinetics. Here, we investigated the effect of the four most common coding ABCB1 single nucleotide polymorphisms (i.e., c.1199G > A, c.1236C > T, c.2677G > T and c.3435C > T) on the intracellular accumulation of bictegravir. Using a previously validated HEK293 recombinant cell line model, we found decreased bictegravir intracellular concentrations in cell lines overexpressing ABCB1 as compared to control cell lines, in line with the known role of ABCB1 in bictegravir transport. However, we were unable to demonstrate any significant difference in bictegravir intracellular accumulation when comparing HEK293 cells overexpressing the wild type (1236C-2677G-3435C, 1199G) or the variant (1236C-2677G-3435T, 1236T-2677T-3435T or 1199A) proteins. These findings suggest that the ABCB1 c.1199G > A and c.1236C > T-c.2677G > T-c.3435C > T variants have no or at least limited impact on the active transport of bictegravir by ABCB1.
Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP , Piperazinas , Polimorfismo de Nucleotídeo Único , Humanos , Células HEK293 , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Piperazinas/metabolismo , Compostos Heterocíclicos com 3 Anéis/metabolismo , Amidas/metabolismo , Piridonas/metabolismo , Compostos Heterocíclicos de 4 ou mais Anéis/metabolismoRESUMO
Bisphenol A (BPA) has been a substantial additive in plastics until the reports on its adverse effects have led to its restrictions and replacement. Monitoring studies document the increasing occurrence of bisphenol analogs, however, data on their effects and risks is still insufficient. Based on the indications that BPA might contribute to ovarian cancer pathogenesis, we examined effects of the analogs AF (BPAF), S (BPS) and F (BPF) (10-9-10-4 M) on the Caov-3 epithelial cancer cells, including the impact on cell viability, proliferation, oxidative stress, and production and expression of several factors and genes related to ovarian cancer. At environmentally relevant doses, bisphenols did not exert significant effects. At the highest concentration, BPAF caused varied alterations, including decreased cell viability and proliferation, caspase activation, down-regulation of PCNA and BIRC5, elevation of IL8, VEGFA, MYC, PTGS2 and ABCB1 expressions. Only BPA (10-4 M) increased IL-6, IL-8 and VEGFA output by the Caov-3 cells. Each bisphenol induced generation of reactive oxygen species and decreased superoxide dismutase activity at the highest concentration. Although the effects were observed only in the supraphysiological doses, the results indicate that certain bisphenol analogs might affect several ovarian cancer cell characteristics and merit further investigation.
Assuntos
Compostos Benzidrílicos , Proliferação de Células , Sobrevivência Celular , Neoplasias Ovarianas , Fenóis , Humanos , Fenóis/toxicidade , Compostos Benzidrílicos/toxicidade , Proliferação de Células/efeitos dos fármacos , Feminino , Sobrevivência Celular/efeitos dos fármacos , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia , Linhagem Celular Tumoral , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo , Sulfonas/toxicidade , Estresse Oxidativo/efeitos dos fármacos , Antígeno Nuclear de Célula em Proliferação/metabolismo , Antígeno Nuclear de Célula em Proliferação/genética , Survivina/genética , Survivina/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Carcinoma Epitelial do Ovário/genética , Carcinoma Epitelial do Ovário/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Interleucina-8/metabolismo , Interleucina-8/genética , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Proteínas Inibidoras de Apoptose/genética , Proteínas Inibidoras de Apoptose/metabolismo , FluorocarbonosRESUMO
BACKGROUND: and Purpose: Chemotherapy-induced peripheral neuropathy (CIPN) constitutes a significant health problem due to the increasing prevalence and lack of therapies for treatment and prevention. While pivotal for routine cancer treatment, paclitaxel and vincristine frequently cause CIPN and impact the quality of life among cancer patients and survivors. Here, we investigate molecular mechanisms and drug transport in CIPN. EXPERIMENTAL APPROACH: Human sensory neurons were derived from induced pluripotent stem cells (iPSC-SNs), which were characterized using flow cytometry and immunolabeling. These iPSC-SNs were exposed to different concentrations of the two microtubule-targeting agents, paclitaxel and vincristine, with and without pre-exposure to inhibitors and inducers of efflux transporters. Neuronal networks were quantified via fluorescent staining against sensory neuron markers. Transcriptional effects of the chemotherapeutics were examined using quantitative polymerase chain reactions (qPCR). KEY RESULTS: Paclitaxel exposure resulted in axonal retraction and thickening, while vincristine caused fragmentation and abolishment of axons. Both agents increased the mRNA expression of the pain receptor, transient receptor potential vanilloid (TRPV1), and highly induced neuronal damage, as measured by activating transcription factor 3 (ATF3) mRNA. iPSC-SNs express the efflux transporters, P-glycoprotein (P-gp, encoded by ABCB1) and multidrug resistance-associated protein 1 (MPR1, encoded by ABCC1). Modulation of efflux transporters indicate that P-gp and MRP1 play a role in modulating neuronal accumulation and neurotoxicity in preliminary experiments. CONCLUSION: and Implications: iPSC-SNs are a valuable and robust model to study the role of efflux transporters and other mechanistic targets in CIPN. Efflux transporters may play a role in CIPN pathogenesis as they regulate the disposition of chemotherapy to the peripheral nervous system, and they may present potential therapeutic targets for CIPN.
Assuntos
Células-Tronco Pluripotentes Induzidas , Proteínas Associadas à Resistência a Múltiplos Medicamentos , Paclitaxel , Doenças do Sistema Nervoso Periférico , Células Receptoras Sensoriais , Vincristina , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/metabolismo , Humanos , Paclitaxel/toxicidade , Doenças do Sistema Nervoso Periférico/induzido quimicamente , Doenças do Sistema Nervoso Periférico/metabolismo , Células Receptoras Sensoriais/efeitos dos fármacos , Células Receptoras Sensoriais/metabolismo , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Proteínas Associadas à Resistência a Múltiplos Medicamentos/genética , Antineoplásicos/efeitos adversos , Antineoplásicos/toxicidade , Canais de Cátion TRPV/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Células CultivadasRESUMO
The overexpression of P-glycoprotein (P-gp/ABCB1) is a leading cause of multidrug resistance (MDR). Hence, it is crucial to discover effective pharmaceuticals that counteract ABCB1-mediated multidrug resistance. FRAX486 is a p21-activated kinase (PAK) inhibitor. The objective of this study was to investigate whether FRAX486 can reverse ABCB1-mediated multidrug resistance, while also exploring its mechanism of action. The CCK8 assay demonstrated that FRAX486 significantly reversed ABCB1-mediated multidrug resistance. Furthermore, western blotting and immunofluorescence experiments revealed that FRAX486 had no impact on expression level and intracellular localization of ABCB1. Notably, FRAX486 was found to enhance intracellular drug accumulation and reduce efflux, resulting in the reversal of multidrug resistance. Docking analysis also indicated a strong affinity between FRAX486 and ABCB1. This study highlights the ability of FRAX486 to reverse ABCB1-mediated multidrug resistance and provides valuable insights for its clinical application.
Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP , Neoplasias da Mama , Resistência a Múltiplos Medicamentos , Resistencia a Medicamentos Antineoplásicos , Humanos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/antagonistas & inibidores , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Feminino , Quinases Ativadas por p21/antagonistas & inibidores , Quinases Ativadas por p21/metabolismo , Linhagem Celular Tumoral , Western BlottingRESUMO
ABCB4 is located at the canalicular membrane of hepatocytes and is responsible for the secretion of phosphatidylcholine into bile. Genetic variations of this transporter are correlated with rare cholestatic liver diseases, the most severe being progressive familial intrahepatic cholestasis type 3 (PFIC3). PFIC3 patients most often require liver transplantation. In this context of unmet medical need, we developed a high-content screening approach to identify small molecules able to correct ABCB4 molecular defects. Intracellularly-retained variants of ABCB4 were expressed in cell models and their maturation, cellular localization and function were analyzed after treatment with the molecules identified by high-content screening. In total, six hits were identified by high-content screening. Three of them were able to correct the maturation and canalicular localization of two distinct intracellularly-retained ABCB4 variants; one molecule was able to significantly restore the function of two ABCB4 variants. In addition, in silico molecular docking calculations suggest that the identified hits may interact with wild type ABCB4 residues involved in ATP binding/hydrolysis. Our results pave the way for their optimization in order to provide new drug candidates as potential alternative to liver transplantation for patients with severe forms of ABCB4-related diseases, including PFIC3.
Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP , Simulação de Acoplamento Molecular , Humanos , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/química , Subfamília B de Transportador de Cassetes de Ligação de ATP/deficiência , Colestase Intra-Hepática/genética , Colestase Intra-Hepática/metabolismo , Transporte Proteico , Ensaios de Triagem em Larga Escala/métodos , Células HEK293RESUMO
Liver fibrosis is characterized by chronic inflammatory responses and progressive fibrous scar formation. Macrophages play a central role in the pathogenesis of hepatic fibrosis by reconstructing the immune microenvironment. Picroside II (PIC II), extracted from Picrorhizae Rhizoma, has demonstrated therapeutic potential for various liver damage. However, the mechanisms by which macrophage polarization initiates immune cascades and contributes to the development of liver fibrosis, and whether this process can be influenced by PIC II, remain unclear. In the current study, RNA sequencing and multiple molecular approaches were utilized to explore the underlying mechanisms of PIC II against liver fibrosis in multidrug-resistance protein 2 knockout (Mdr2-/-) mice. Our findings indicate that PIC II activates M1-polarized macrophages to recruit natural killer cells (NK cells), potentially via the CXCL16-CXCR6 axis. Additionally, PIC II promotes the apoptosis of activated hepatic stellate cells (aHSCs) and enhances the cytotoxic effects of NK cells, while also reducing the formation of neutrophil extracellular traps (NETs). Notably, the anti-hepatic fibrosis effects associated with PIC II were largely reversed by macrophage depletion in Mdr2-/- mice. Collectively, our research suggests that PIC II is a potential candidate for halting the progression of liver fibrosis.
Assuntos
Apoptose , Cinamatos , Células Estreladas do Fígado , Glucosídeos Iridoides , Cirrose Hepática , Macrófagos , Animais , Masculino , Camundongos , Apoptose/efeitos dos fármacos , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Membro 4 da Subfamília B de Transportadores de Cassetes de Ligação de ATP/genética , Cinamatos/farmacologia , Células Estreladas do Fígado/efeitos dos fármacos , Glucosídeos Iridoides/farmacologia , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/imunologia , Cirrose Hepática/tratamento farmacológico , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
Mitoxantrone resistant variant of SW620 line was developed, characterized and subsequently used as a model system to determine oncostatin M ability to modulate MDR phenomenon. The selection regimen allowed for overexpression of ABCG2 and ABCB1 both at the RNA and protein level, which was further confirmed by functional assays. Oncostatin M supplementation resulted in partial reversal of MDR phenotype by decreasing overexpression of ABCG2 demonstrating for the first time the ability of this cytokine for selective down-regulation of one of MDR proteins.
Assuntos
Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Resistencia a Medicamentos Antineoplásicos , Mitoxantrona , Proteínas de Neoplasias , Oncostatina M , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/genética , Mitoxantrona/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Humanos , Proteínas de Neoplasias/metabolismo , Proteínas de Neoplasias/genética , Oncostatina M/metabolismo , Linhagem Celular Tumoral , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/genéticaRESUMO
BACKGROUND AND OBJECTIVES: Clozapine is the medication of choice for treatment-resistant schizophrenia. However, it has a complex metabolism and unexplained interindividual variability. The current work aims to develop a pharmacokinetic model of clozapine and norclozapine in non-smokers and assess the impact of demographic and genetic predictors. METHODS: Healthy volunteers were recruited in a population pharmacokinetic study. Blood samples were collected at 30 min and 1, 2, 3, 5 and 8 h following a single flat dose of clozapine (12.5 mg). The clozapine and norclozapine concentrations were measured via high-performance liquid chromatography-ultraviolet method. A semi-physiological pharmacokinetic model of clozapine and norclozapine was developed using nonlinear mixed-effects modeling. Clinical and genetic predictors were evaluated, including CYP1A2 (rs762551) and ABCB1 (rs2032582), using restriction fragment length polymorphism. RESULTS: A total of 270 samples were collected from 33 participants. The data were best described using a two-compartment model for clozapine and a two-compartment model for norclozapine with first-order absorption and elimination and pre-systemic metabolism. The estimated (relative standard error) clearance of clozapine and norclozapine were 27 L h-1 (31.5 %) and 19.6 L h-1 (30%), respectively. Clozapine clearance was lower in sub-Saharan Africans (n = 4) and higher in Caucasians (n = 9) than Asians (n = 20). Participants with CYP1A2 (rs762551) (n = 18) and ABCB1 (rs2032582) (n = 12) mutant alleles had lower clozapine clearance in the univariate analysis. CONCLUSIONS: This is the first study to develop a semi-physiological pharmacokinetic model of clozapine and norclozapine accounting for the pre-systemic metabolism. Asians required lower doses of clozapine as compared with Caucasians, while clozapine pharmacokinetics in sub-Saharan Africans should be further investigated in larger trials.
Assuntos
Antipsicóticos , Clozapina , Citocromo P-450 CYP1A2 , Voluntários Saudáveis , Modelos Biológicos , Clozapina/farmacocinética , Clozapina/análogos & derivados , Clozapina/administração & dosagem , Clozapina/sangue , Humanos , Adulto , Masculino , Antipsicóticos/farmacocinética , Antipsicóticos/administração & dosagem , Feminino , Citocromo P-450 CYP1A2/genética , Citocromo P-450 CYP1A2/metabolismo , Adulto Jovem , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Pessoa de Meia-IdadeRESUMO
The leaves of Araucaria cunninghamii are known to be nonedible and toxic. Previous studies have identified biflavones in various Araucaria species. This study aimed to investigate the in vitro cytotoxicity of the isolated compounds from Araucaria cunninghamii after metabolomics and network pharmacological analysis. Methanol extract of Araucaria cunninghamii leaves was subjected to bioassay-guided fractionation. The active fraction was analyzed using LC-HRMS, through strategic database mining, by comparing the data to the Dictionary of Natural Products to identify 12 biflavones, along with abietic acid, beta-sitosterol, and phthalate. Eight compounds were screened for network pharmacology study, where in silico ADME analysis, prediction of gene targets, compound-gene-pathway network and hierarchical network analysis, protein-protein interaction, KEGG pathway, and Gene Ontology analyses were done, that showed PI3KR1, EGFR, GSK3B, and ABCB1 as the common targets for all the compounds that may act in the gastric cancer pathway. Simultaneously, four biflavones were isolated via chromatography and identified through NMR as dimeric apigenin with varying methoxy substitutions. Cytotoxicity study against the AGS cell line for gastric cancer showed that AC1 biflavone (IC50 90.58 µM) exhibits the highest cytotoxicity and monomeric apigenin (IC50 174.5 µM) the lowest. Besides, the biflavones were docked to the previously identified targets to analyze their binding affinities, and all the ligands were found to bind with energy ≤-7 Kcal/mol.