The Xenobiotic Transporter Mdr1 Enforces T Cell Homeostasis in the Presence of Intestinal Bile Acids.

CD4+ T cells are tightly regulated by microbiota in the intestine, but whether intestinal T cells interface with host-derived metabolites is less clear. Here, we show that CD4+ T effector (Teff) cells upregulated the xenobiotic transporter, Mdr1, in the ileum to maintain homeostasis in the presence of bile acids. Whereas wild-type Teff cells upregulated Mdr1 in the ileum, those lacking Mdr1 displayed mucosal dysfunction and induced Crohn's disease-like ileitis following transfer into Rag1-/- hosts. Mdr1 mitigated oxidative stress and enforced homeostasis in Teff cells exposed to conjugated bile acids (CBAs), a class of liver-derived emulsifying agents that actively circulate through the ileal mucosa. Blocking ileal CBA reabsorption in transferred Rag1-/- mice restored Mdr1-deficient Teff cell homeostasis and attenuated ileitis. Further, a subset of ileal Crohn's disease patients displayed MDR1 loss of function. Together, these results suggest that coordinated interaction between mucosal Teff cells and CBAs in the ileum regulate intestinal immune homeostasis.

Myosin Vb Traffics P-Glycoprotein to the Apical Membrane of Intestinal Epithelial Cells.

BACKGROUND & AIMS: The xenobiotic efflux pump P-glycoprotein is highly expressed on the apical membrane of the gastrointestinal tract, where it regulates the levels of intracellular substrates. P-glycoprotein is altered in disease, but the mechanisms that regulate the levels of P-glycoprotein are still being explored. The molecular motor myosin Vb (Myo5b) traffics diverse cargo to the apical membrane of intestinal epithelial cells. We hypothesized that Myo5b was responsible for the delivery of P-glycoprotein to the apical membrane of enterocytes. METHODS: We used multiple murine models that lack functional Myo5b or the myosin binding partner Rab11a to analyze P-glycoprotein localization. Pig and human tissue were analyzed to determine P-glycoprotein localization in the setting of MYO5B mutations. Intestinal organoids were used to examine P-glycoprotein trafficking and to assay P-glycoprotein function when MYO5 is inhibited. RESULTS: In mice lacking Myo5b or the binding partner Rab11a, P-glycoprotein was improperly trafficked and had decreased presence in the brush border of enterocytes. Immunostaining of a pig model lacking functional Myo5b and human biopsies from a patient with an inactivating mutation in Myo5b also showed altered localization of intestinal P-glycoprotein. Human intestinal organoids expressing the motorless MYO5B tail domain had colocalization with P-glycoprotein, confirming that P-glycoprotein was trafficked by MYO5B in human enterocytes. Inhibition of MYO5 in human intestinal cell lines and organoids resulted in decreased P-glycoprotein capacity. Additionally, inhibition of MYO5 in human colon cancer cells diminished P-glycoprotein activity and increased cell death in response to a chemotherapeutic drug. CONCLUSIONS: Collectively, these data demonstrate that Myo5b is necessary for the apical delivery of P-glycoprotein.

Beyond endocrine resistance: estrogen receptor (ESR1) activating mutations mediate chemotherapy resistance through the JNK/c-Jun MDR1 pathway in breast cancer.

PURPOSE: All patients with metastatic breast cancer (MBC) expressing estrogen receptor-α (ESR1) will eventually develop resistance to endocrine therapies. In up to 40% of patients, this resistance is caused by activating mutations in the ligand-binding domain (LBD) of ESR1. Accumulating clinical evidence indicate adverse outcomes for these patients, beyond that expected by resistance to endocrine therapy. Here we aimed to study the role of ESR1 mutations in conferring chemoresistance in BC cells. METHODS: MCF-7 cells harboring Y537S and D538G ESR1 mutations (mut-ER) were employed to study the response to chemotherapy drugs, paclitaxel and doxorubicin, using viability and apoptotic assay in vitro, and tumor growth in vivo. JNK/c-Jun/MDR1 pathway was studied using qRT-PCR, western-blot, gene-reporter and ChIP assays. MDR1 expression was analyzed in clinical samples using IHC. RESULTS:  Cell harboring ESR1 mutations displayed relative chemoresistance compared to WT-ER, evidenced by higher viability and reduced apoptosis as well as resistance to paclitaxel in vivo. To elucidate the underlying mechanism, MDR1 expression was examined and elevated levels were observed in mut-ER cells, and in clinical BC samples. MDR1 is regulated by the c-Jun pathway, and we showed high correlation between these two genes in BC using TCGA databases. Accordingly, we detected higher JNK/c-Jun expression and activity in ESR1-mutated cells, as well as increased occupancy of c-Jun in MDR1 promoter. Importantly, JNK inhibition decreased MDR1 expression and restored sensitivity to chemotherapy. CONCLUSIONS: Taken together, these data indicate that ESR1 mutations confer chemoresistance through activation of the JNK/MDR1 axis. These finding suggest a novel treatment option for BC tumors expressing ESR1 mutations.

EGR1 mediates MDR1 transcriptional activity regulating gemcitabine resistance in pancreatic cancer.

BACKGROUND: Gemcitabine is a cornerstone drug for the treatment of all stages of pancreatic cancer and can prolong the survival of patients with pancreatic cancer, but resistance to gemcitabine in pancreatic cancer patients hinders its efficacy. The overexpression of Early growth response 1(EGR1) in pancreatic ductal adenocarcinoma as a mechanism of gemcitabine chemoresistance in pancreatic cancer has not been explored. The major mechanisms of gemcitabine chemoresistance are related to drug uptake, metabolism, and action. One of the common causes of tumor multidrug resistance (MDR) to chemotherapy in cancer cells is that transporter proteins increase intracellular drug efflux and decrease drug concentrations by inducing anti-apoptotic mechanisms. It has been reported that gemcitabine binds to MDR1 with high affinity. The purpose of this research was to investigate the potential mechanisms by which EGR1 associates with MDR1 to regulate gemcitabine resistance in pancreatic cancer cells. METHODS: The following in vitro and in vivo techniques were used in this research to explore the potential mechanisms by which EGR1 binds to MDR1 to regulate gemcitabine resistance in pancreatic cancer cells. Cell culture; in vitro and in vivo study of EGR1 function by loss of function analysis. Binding of EGR1 to the MDR1 promoter was detected using the ChIP assay. qRT-PCR, Western blot assays to detect protein and mRNA expression; use of Annexin V apoptosis detection assay to test apoptosis; CCK8, Edu assay to test cell proliferation viability. The animal model of pancreatic cancer subcutaneous allograft was constructed and the tumours were stained with hematoxylin eosin and Ki-67 expression was detected using immunohistochemistry. FINDINGS: We revealed that EGR1 expression was increased in different pancreatic cancer cell lines compared to normal pancreatic ductal epithelial cells. Moreover, gemcitabine treatment induced upregulation of EGR1 expression in a dose- and time-dependent manner. EGR1 is significantly enriched in the MDR1 promoter sequence.Upon knockdown of EGR1, cell proliferation was impaired in CFPAC-1 and PANC-1 cell lines, apoptosis was enhanced and MDR1 expression was decreased, thereby partially reversing gemcitabine chemoresistance. In animal experiments, knockdown of EGR1 enhanced the inhibitory effect of gemcitabine on tumor growth compared with the sh-NC group. CONCLUSIONS: Our study suggests that EGR1 may be involved in the regulation of MDR1 to enhance gemcitabine resistance in pancreatic cancer cells. EGR1 could be a novel therapeutic target to overcome gemcitabine resistance in pancreatic cancer.

Assessing the therapeutic efficacy of artemether-lumefantrine for uncomplicated malaria in Lagos, Nigeria: a comprehensive study on treatment response and resistance markers.

BACKGROUND: The burden of malaria persists in sub-Saharan Africa and the emergence of artemisinin resistance has introduced complexity to control efforts. Monitoring the efficacy of artemisinin-based treatment for malaria is crucial to address this challenge. This study assessed treatment efficacy of artemether-lumefantrine (AL) and genetic diversity of Plasmodium falciparum isolates in a Nigerian population. METHODS: Participants presenting with clinical symptoms of uncomplicated malaria at a health centre in Lagos, Nigeria, were screened for P. falciparum. Enrolled participants were treated with AL and monitored through scheduled check-up visits, clinical and laboratory examinations for 28 days. Parasite clearance and genetic diversity were assessed through polymerase chain reaction (PCR) analysis of merozoite surface proteins (msp1 and msp2). The prevalence of drug resistance mutations was assessed by P. falciparum multidrug resistance gene 1 (mdr1) genotyping followed by P. falciparum ubiquitin-specific protease 1 (ubp1) gene sequencing. RESULTS: The PCR-uncorrected treatment outcome revealed 94.4% adequate clinical and parasitological response (ACPR) and 5.6% late parasitological failure (LPF) rates. After PCR correction, no suspected LPF case was detected and ACPR 67/67 (100%) was achieved in all the individuals. Moreover, a high prevalence of wild-type alleles for mdr1 N86Y (93.7%), and mdr1 D1246Y (87.5%) was observed. Genetic diversity analysis revealed predominant K1 allelic family for msp1 (90.2%) and FC27 for msp2 (64.4%). Estimated multiplicity of infection (MOI) was 1.7, with the highest MOI observed in the 5-15 years age group. ubp1 sequence analysis identified one nonsynonymous E1528D polymorphism at a low frequency (1.6%). CONCLUSION: The study demonstrated sustained efficacy of AL for treating uncomplicated P. falciparum malaria. Genetic diversity analysis revealed various allelic types, suggesting occurrences of polyclonal infections. Nonetheless, the detection of a significant ubp1 polymorphism could have future implications for the epidemiology of anti-malarial drug resistance in the population.

Valproate modulates the activity of multidrug resistance efflux pumps, as a chemoresistance factor in gastric cancer cells.

BACKGROUND: Drug resistance is one of the most critical problems in gastric cancer therapy. This study was performed to investigate the valproic acid effects on the proliferation of sensitive and resistant cell lines of human gastric cancer, and to explore the mechanism of the agent on multi drug resistance and apoptosis genes. METHODS: The cytotoxicity effect of valproic acid on the EPG85.257 and EPG85.257RDB cells was assessed by the MTT assay, and the IC50 concentration was evaluated. Apoptosis, genotoxicity, and drug resistance pump activity were evaluated using comet assay, Real-time PCR, and flow cytometry, respectively. Cell proliferation was assayed using a scratch test. RESULTS: Dose-dependent toxicity was recorded after treatment of cells with valproic acid. Valproic acid represented a significant growth inhibition on EPG85.257 cells with IC50 values of 5.84 µM and 4.78 µM after 48 h and 72 h treatment, respectively. In contrast, the drug-resistant counterpart represented 8.7 µM and 7.02 µM IC50 values after the same treatment time. Valproic acid induced PTEN, Bcl2, P53, Bax, P21, and caspase3 expression in EPG85.257 cells, whereas p21, p53, PTEN, and ABCB1 were overexpressed in EPG5.257RDB. Valproic acid hindered cell migration in both cell lines (P < 0.01). Valproate genotoxicity was significantly higher in the parent cells than in their resistant EPG85.257RDB counterparts. Valproate led to a 62% reduction in the daunorubicin efflux of the MDR1 pump activity. CONCLUSIONS: Valproate can affect drug resistance in gastric cancer via a unique mechanism independent of MDR1 expression.

Effect of MDR1 C3435T and CYP2C19 genetic polymorphisms on the outcome of Helicobacter pylori eradication treatment in children with gastritis and peptic ulcer, Vietnam.

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.

Synergy of histone acetyltransferase inhibitor (HATi) with quercetin inhibits biofilm formation in Candida tropicalis.

Histone acetyltransferase inhibitors (HATi) are mechanism-based inhibitors that show promise in the treatment of several illnesses, including diabetes, hyperlipidemia, cancer, and Alzheimer's disease. The work emphasizes the significance of HATi as a possible treatment strategy against Candida species biofilms. Here, in this study, we found that combining a HATi, anacardic acid (AA), and quercetin, a known flavonoid, significantly prevented biofilm formation by C. tropicalis. We further show that C. tropicalis exhibited a considerable downregulation of drug-resistance gene expression (CDR1 and MDR1) when co-administrated. Additionally, in silico studies revealed that the AA interacts strongly with a histone acetyltransferase, Rtt109, which may account for the observed biofilm inhibitory effect. In conclusion, the study illustrates how HATi may be used to potentiate the inhibitory action of phytoactives or antifungals against drug-resistant yeast infections.

4-{3-[(Pyridin-4-ylmethyl)amino]-[1,2,4]triazolo[4,3-b][1,2,4]triazin-6-yl}phenol: An improved anticancer agent in hepatocellular carcinoma and a selective MDR1/MRP modulator.

Hepatocellular carcinoma is the most common type of primary liver cancer. However, multidrug resistance (MDR) is a major obstacle to the effective chemotherapy of cancer cells. This report documents the rational design, synthesis, and biological evaluation of a novel series of triazolotriazines substituted with CH2NH-linked pyridine for use as dual c-Met/MDR inhibitors. Compound 12g with IC50 of 3.06 µM on HepG2 cells showed more potency than crizotinib (IC50 = 5.15 µM) in the MTT assay. In addition, 12g inhibited c-Met kinase at a low micromolar level (IC50 = 0.052 µM). 12g significantly inhibited P-gp and MRP1/2 efflux pumps in both cancerous HepG2 and BxPC3 cells starting from the lower concentrations of 3 and 0.3 µM, respectively. 12g did not inhibit MDR1 and MRP1/2 in noncancerous H69 cholangiocytes up to the concentration of 30 and 60 µM, respectively. Current results highlighted that cancerous cells were more susceptible to the effect of 12g than normal cells, in which the inhibition occurred only at the highest concentrations, suggesting a further interest in 12g as a selective anticancer agent. Overall, 12g, as a dual c-Met and P-gp/MRP inhibitor, is a promising lead compound for developing a new generation of anticancer agents.

Application of eprinomectin-containing parasiticides at label doses causes neurological toxicosis in cats homozygous for ABCB11930_1931del TC.

The feline MDR1 mutation (ABCB11930_1931delTC) has been associated with neurological toxicosis after topical application of eprinomectin products labeled for feline use. Information was collected from veterinarians who submitted samples for ABCB11930_1931delTC genotyping. In most cases, the submission form indicated an adverse event involving eprinomectin, in other cases submitting veterinarians were contacted to determine whether the patient had experienced an adverse drug event involving eprinomectin. If so, additional information was obtained to determine whether the case met inclusion criteria. 14 cases were highly consistent with eprinomectin toxicosis. Eight cats were homozygous for ABCB11930_1931del TC (3 died; 5 recovered). Six cats were homozygous wildtype (2 died; 4 recovered). The observed ABCB11930_1931delTC frequency (57%) was higher than the expected frequency (≤1%) in the feline population (Fisher Exact test, p < 0.01). Among wildtype cats, four were concurrently treated with potential competitive inhibitors of P-glycoprotein. Results indicate that topical eprinomectin products, should be avoided in cats homozygous for ABCB11930_1931delTC. This is a serious, preventable adverse event occurring in an identifiable subpopulation treated with FDA-approved products in accordance with label directions. Acquired P-glycoprotein deficiency resulting from drug interactions may enhance susceptibility to eprinomectin-induced neurological toxicosis in any cat, regardless of ABCB1 genotype.

Cholesterol Conjugates of Small Interfering RNA: Linkers and Patterns of Modification.

Cholesterol siRNA conjugates attract attention because they allow the delivery of siRNA into cells without the use of transfection agents. In this study, we compared the efficacy and duration of silencing induced by cholesterol conjugates of selectively and totally modified siRNAs and their heteroduplexes of the same sequence and explored the impact of linker length between the 3' end of the sense strand of siRNA and cholesterol on the silencing activity of "light" and "heavy" modified siRNAs. All 3'-cholesterol conjugates were equally active under transfection, but the conjugate with a C3 linker was less active than those with longer linkers (C8 and C15) in a carrier-free mode. At the same time, they were significantly inferior in activity to the 5'-cholesterol conjugate. Shortening the sense strand carrying cholesterol by two nucleotides from the 3'-end did not have a significant effect on the activity of the conjugate. Replacing the antisense strand or both strands with fully modified ones had a significant effect on silencing as well as improving the duration in transfection-mediated and carrier-free modes. A significant 78% suppression of MDR1 gene expression in KB-8-5 xenograft tumors developed in mice promises an advantage from the use of fully modified siRNA cholesterol conjugates in combination chemotherapy.

Infection and disruption of placental multidrug resistance (MDR) transporters: Implications for fetal drug exposure.

P-glycoprotein (P-gp, encoded by the ABCB1 gene) and breast cancer resistance protein (BCRP/ABCG2) are efflux multidrug resistance (MDR) transporters localized at the syncytiotrophoblast barrier of the placenta and protect the conceptus from drug and toxin exposure throughout pregnancy. Infection is an important modulator of MDR expression and function. This review comprehensively examines the effect of infection on the MDR transporters, P-gp and BCRP in the placenta. Infection PAMPs such as bacterial lipopolysaccharide (LPS) and viral polyinosinic-polycytidylic acid (poly I:C) and single-stranded (ss)RNA, as well as infection with Zika virus (ZIKV), Plasmodium berghei ANKA (modeling malaria in pregnancy - MiP) and polymicrobial infection of intrauterine tissues (chorioamnionitis) all modulate placental P-gp and BCRP at the levels of mRNA, protein and or function; with specific responses varying according to gestational age, trophoblast type and species (human vs. mice). Furthermore, we describe the expression and localization profile of Toll-like receptor (TLR) proteins of the innate immune system at the maternal-fetal interface, aiming to better understand how infective agents modulate placental MDR. We also highlight important gaps in the field and propose future research directions. We conclude that alterations in placental MDR expression and function induced by infective agents may not only alter the intrauterine biodistribution of important MDR substrates such as drugs, toxins, hormones, cytokines, chemokines and waste metabolites, but also impact normal placentation and adversely affect pregnancy outcome and maternal/neonatal health.

Molecular epidemiology of potential candidate markers for chloroquine resistance in imported Plasmodium vivax malaria cases in Iran.

BACKGROUND: The spread of Plasmodium vivax strains resistant to chloroquine (CQ) has posed a challenge to control strategies aimed at eliminating malaria. Molecular analysis of candidate resistance markers is very important for monitoring the P. vivax resistance to CQ in different endemic regions. In the present study, the multidrug resistance 1 (pvmdr1) gene, a possible marker for CQ resistance in P. vivax, was evaluated by molecular methods. METHODS: A simple PCR-RFLP method was developed for mutation analysis in pvmdr1 gene. A number of 120 blood spots were obtained from patients with P. vivax mono-infection in 2021. All of the samples were collected from Pakistani patients who travelled to Iran. RESULTS: None of the samples had any mutation at codon 976 of pvmdr1, while the 1076 mutation was detected in 96.2% of the examined isolates. Only two pvmdr1 haplotypes were identified, including the single mutant (Y976/1076L) as the most prevalent haplotype (with 96.2% frequency) and the wild type (Y976/F1076; with 3.8% frequency). CONCLUSIONS: In this study, the major CQ resistance-mediating mutation and multiple mutant haplotypes of the pvmdr1 gene was not detected. However, continuous monitoring of drug resistance markers and close supervision of the efficacy of CQ is essential to detect the potential emergence of CQ-resistant P. vivax isolates in Iran. This data is important for performing future epidemiological surveillance to monitor CQ resistance in this endemic area and the bordering regions.

Ligand-displaying-exosomes using RNA nanotechnology for targeted delivery of multi-specific drugs for liver cancer regression.

Liver cancer such as hepatocellular carcinoma (HCC) poorly responds to chemotherapeutics as there are no effective means to deliver the drugs to liver cancer. Here we report GalNAc decorated exosomes as cargo for targeted delivery of Paclitaxel (PTX) and miR122 to liver tumors as an effective means to inhibit the HCC. Exosomes (Exos) are nanosized extracellular vesicles that deliver a payload to cancer cells effectively. GalNAc provides Exos targeting ability by binding to the asialoglycoprotein-receptor (ASGP-R) overexpressed on the liver cancer cell surface. A 4-way junction (4WJ) RNA nanoparticle was constructed to harbor 24 copies of hydrophobic PTX and 1 copy of miR122. The 4WJ RNA-PTX complex was loaded into the Exos, and its surface was decorated with GalNAc using RNA nanotechnology to obtain specific targeting. The multi-specific Exos selectively bind and efficiently delivered the payload into the liver cancer cells and exhibited the highest cancer cell inhibition due to the multi-specific effect of miR122, PTX, GalNAc, and Exos. The same was reflected in mice xenograft studies, the liver cancer was efficiently inhibited after systemic injection of the multi-specific Exos. The required effective dose of chemical drugs carried by Exos was significantly reduced, indicating high efficiency and low toxicity. The multi-specific strategy demonstrates that Exos can serve as a natural cargo vehicle for the targeted delivery of anticancer therapeutics to treat difficult-to-treat cancers.

The potential implication of MDR1 and NAC1 genetic polymorphisms on resistance to antiepileptic drugs among a Jordanian epileptic population: a cross-sectional study.

BACKGROUND: Resistance to antiepileptic drugs (AEDs) remains one of the main challenges to neurologists. Polymorphisms of drug efflux transporters such as multidrug resistance (MDR1) gene and target sites such as the nucleus accumbens-associated 1 (NAC1) gene have been suggested to influence the responsiveness to treatment. AIM: Evaluation of the association of MDR1 and NAC1 polymorphisms with AEDs resistance among Jordanian epileptic patients. SUBJECTS AND METHODS: 86 Jordanian epileptics were included in the study. DNA was extracted and genotyping was conducted by polymerase chain reaction followed by sequencing. Nine single nucleotide polymorphisms (SNPs) on the MDR1 gene and six SNPs on the NAC1 gene were investigated. RESULTS: MDR1 and NAC1 polymorphisms don't seem to influence the resistance to AEDs at the genotype or allele level. However, a strong association was found between MDR1 rs2032588 (OR = 5; 95%CI = [1.3-18.8], p = 0.01) and AEDs resistance among males at the allele level. Also, data revealed an association between MDR1 rs1128503 and AEDs resistance among females at the allele level. CONCLUSION: The data suggest that MDR1 and NAC1 polymorphisms do not influence the AEDs resistance among Jordanian epileptics. However, there is a gender-dependent association between MDR1 polymorphisms and resistance to AEDs at two SNPs (rs2032588 and rs1128503).

Canine and feline P-glycoprotein deficiency: What we know and where we need to go.

In 2001 the molecular genetic basis of so-called "ivermectin sensitivity" in herding breed dogs was determined to be a P-glycoprotein deficiency caused by a genetic variant of the MDR1 (ABCB1) gene often called "the MDR1 mutation." We have learned a great deal about P-glycoprotein's role in drug disposition since that discovery, namely that P-glycoprotein transports many more drugs than just macrocyclic lactones that P-glycoprotein mediated drug transport is present in more places than just the blood brain barrier, that some cats have a genetic variant of MDR1 that results in P-glycoprotein deficiency, that P-glycoprotein dysfunction can occur as a result of drug-drug interactions in any dog or cat, and that the concept of P-glycoprotein "inhibitors" versus P-glycoprotein substrates is somewhat arbitrary and artificial. This paper will review these discoveries and discuss how they impact drug selection and dosing in dogs and cats with genetically mediated P-glycoprotein deficiency or P-glycoprotein dysfunction resulting from drug-drug interactions.

Assessment of verdinexor as a canine P-glycoprotein substrate.

The P-glycoprotein (P-gp) substrate status of antineoplastic drugs intended for veterinary patients is an important characteristic to define for two reasons. First, neoplastic cells expressing P-gp can actively efflux drugs that are P-gp substrates curtailing their efficacy. Second, antineoplastic drugs tend to have a narrow therapeutic index. Antineoplastic drugs that are P-gp substrates can cause severe adverse reactions in animals with P-gp dysfunction such as dogs with ABCB1-1Δ and cats with ABCB11930_1931del TC. Animals with P-gp dysfunction experience greater overall exposure to P-gp substrate drugs due to mechanisms such as increased intestinal absorption, decreased biliary clearance and greater central nervous system penetration compared with animals with normal P-gp function. Accordingly, knowing the P-gp substrate status of antineoplastic drugs is an important safety consideration prior to use in canine or feline cancer patients. This study used a cell line overexpressing canine P-gp to assess the P-gp substrate status of verdinexor. Based on both a cytotoxicity assay and a competitive flow cytometry assay verdinexor is not a substrate for canine P-gp.

MDR1 Inhibition Reverses Doxorubicin-Resistance in Six Doxorubicin-Resistant Canine Prostate and Bladder Cancer Cell Lines.

Acquired chemoresistance during chemotherapy, often accompanied by cross- and multi-resistance, limits therapeutic outcomes and leads to recurrence. In order to create in vitro model systems to understand acquired doxorubicin-resistance, we generated doxorubicin-resistant sublines of canine prostate adenocarcinoma and urothelial cell carcinoma cell lines. Chemoresistance to doxorubicin, cross-resistance to carboplatin, and the reversibility of the acquired resistance by the specific MDR1-inhibitor tariquidar were quantified in metabolic assays. Resistance mechanisms were characterized by expression of the efflux transporters MDR1 and RALBP1, as well as the molecular target of doxorubicin, TOP2A, with qPCR and Western blotting. Six out of nine cell lines established stable resistance to 2 µM doxorubicin. Drug efflux via massive MDR1 overexpression was identified as common, driving resistance mechanism in all sublines. MDR1 inhibition with tariquidar extensively reduced or reversed the acquired, and also partly the parental resistance. Three cell lines developed additional, non-MDR1-dependent resistance. RALBP1 was upregulated in one resistant subline at the protein level, while TOP2A expression was not altered. Combination therapies aiming to inhibit MDR1 activity can now be screened for synergistic effects using our resistant sublines. Nevertheless, detailed resistance mechanisms and maintained molecular target expression in the resistant sublines are still to be examined.

Molecular and Clinical Links between Drug-Induced Cholestasis and Familial Intrahepatic Cholestasis.

Idiosyncratic Drug-Induced Liver Injury (iDILI) represents an actual health challenge, accounting for more than 40% of hepatitis cases in adults over 50 years and more than 50% of acute fulminant hepatic failure cases. In addition, approximately 30% of iDILI are cholestatic (drug-induced cholestasis (DIC)). The liver's metabolism and clearance of lipophilic drugs depend on their emission into the bile. Therefore, many medications cause cholestasis through their interaction with hepatic transporters. The main canalicular efflux transport proteins include: 1. the bile salt export pump (BSEP) protein (ABCB11); 2. the multidrug resistance protein-2 (MRP2, ABCC2) regulating the bile salts' independent flow by excretion of glutathione; 3. the multidrug resistance-1 protein (MDR1, ABCB1) that transports organic cations; 4. the multidrug resistance-3 protein (MDR3, ABCB4). Two of the most known proteins involved in bile acids' (BAs) metabolism and transport are BSEP and MDR3. BSEP inhibition by drugs leads to reduced BAs' secretion and their retention within hepatocytes, exiting in cholestasis, while mutations in the ABCB4 gene expose the biliary epithelium to the injurious detergent actions of BAs, thus increasing susceptibility to DIC. Herein, we review the leading molecular pathways behind the DIC, the links with the other clinical forms of familial intrahepatic cholestasis, and, finally, the main cholestasis-inducing drugs.

Effects of Apigenin on Pharmacokinetics of Dasatinib and Probable Interaction Mechanism.

Dasatinib (DAS), a narrow-therapeutic index drug, Bcr-Abl, and Src family kinases multitarget inhibitor have been approved for chronic myelogenous leukemia (CML) and Ph-positive acute lymphocytic leukemia (Ph+ ALL). Apigenin (APG) has a long history of human usage in food, herbs, health supplements, and traditional medicine, and it poses low risk of damage. The concomitant use of APG containing herbs/foods and traditional medicine may alter the pharmacokinetics of DAS, that probably lead to possible herb-drug interactions. The pharmacokinetic interaction of APG pretreatment with DAS in rat plasma following single and co-oral dosing was successfully deliberated using the UPLC-MS/MS method. The in vivo pharmacokinetics and protein expression of CYP3A2, Pgp-MDR1, and BCPR/ABCG2 demonstrate that APG pretreatment has potential to drastically changed the DAS pharmacokinetics where escalation in the Cmax, AUC(0-t), AUMC(0-inf_obs), T1/2, Tmax, and MRT and reduction in Kel, Vd, and Cl significantly in rats pretreated with APG 40 mg/kg, thus escalating systemic bioavailability and increasing the rate of absorption via modulation of CYP3A2, Pgp-MDR1, and BCPR/ABCG2 protein expression. Therefore, the concomitant consumption of APG containing food or traditional herb with DAS may cause serious life-threatening drug interactions and more systematic clinical study on herb-drug interactions is required, as well as adequate regulation in herbal safety and efficacy.