Drug Transporters ABCB1 Research Articles

Brain accumulation of tivozanib is restricted by ABCB1 (P-glycoprotein) and ABCG2 (breast cancer resistance protein) in mice.

Tivozanib is a potent and selective inhibitor of VEGFR1-3, recently approved by the EMA for first-line treatment of renal cell carcinoma. We used wild-type, knockout, and transgenic mouse strains to study the effects of the drug transporters ABCB1, ABCG2, and OATP1A/1B, and of the CYP3A enzymes on the oral availability and tissue distribution of tivozanib. Tivozanib was transported by human ABCB1 and mouse Abcg2 in polarized MDCK-II cells. Upon oral administration, tivozanib showed rapid absorption and the plasma concentration-time curves showed secondary peaks in all mouse strains, suggesting enterohepatic recirculation. The brain-to-plasma ratios were significantly increased in Abcb1a/1b-/- (2.2-fold) and Abcb1a/1b;Abcg2-/- (2.6-fold) mice compared to wild-type mice, indicating a modest protective role of these transporters in the blood-brain barrier. Slco1a/1b-/- mice showed a 1.2-fold lower liver-to-plasma ratio than wild-type mice, suggesting a minor role of mOatp1a/1b in tivozanib liver distribution. Oral plasma pharmacokinetics of tivozanib was not significantly altered in these mouse strains, nor in Cyp3a knockout and CYP3A4-humanized mice. The modest effect of ABC transporters on tivozanib brain accumulation, if also true in humans, might mean that this drug is not strongly limited in its therapeutic efficacy against malignant lesions situated partly or completely behind the blood-brain barrier.

Interactions between Maraviroc and the ABCB1, ABCG2, and ABCC2 Transporters: An Important Role in Transplacental Pharmacokinetics.

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

ABCB1 rs1045642 C>T Change Is Associated with AML Cell Line Sensitivity to Calicheamicin

Introduction: Gemtuzumab ozogamicin (GO), a CD33-antibody conjugated to DNA damaging cytotoxin-calicheamcin is a re-emerging and promising drug in AML treatment. Promising results from multiple clinical trials demonstrated GO improved outcomes in both adult and pediatric patients with most pronounced effect in favorable-risk cytogenetics. In light of these results GO was reapproved for treatment of AML by FDA in Sep 2017. Given the anticipated increase in GO use post approval, it is very timely to understand mechanisms contributing to differences in patient response. Calicheamicin releases intracellularly and causes DNA strand breaks which leads to apoptosis and cell death. The anti-leukemic effects of GO are thus entirely due to calicheamicin-induced DNA damage. Given that calicheamicin is a substrate of drug transporter ABCB1 (Pgp1), genetic polymorphisms in ABCB1 can impact the intra-cellular accumulation of calicheamicin and thus clinical outcome. Our group have recently shown that ABCB1 synonymous SNP rs1045642 C>T (also well known as 3435C>T) is associated with clinical outcome in children with AML trial, now we report the results of the in vitro evaluation of impact of ABCB1 SNP on calicheamicin mediated cell cytotoxicity.Methods: To understand differences in in vitro calicheamicin toxicity mediated by ABCB1 variation, HL-60 cells with low native ABCB1 expression were transfected with expression plasmid of ABCB1*1 (rs1045642-C and also WT for other two other SNPs in ABCB1) (WT Plasmid), and rs1045642-T. Cells were harvested 48 hours' post-transfection and treated with 40nM calicheamicin for 24 hours. Posttreatment cell were stained with permeant nuclear stain (NucRed® 647 Live Invitrogen) that emits bright far-red fluorescence upon binding to DNA in live cells. One-hour after incubation with the dye, calicheamicin toxicity was assessed by checking cell viability using flow cytometry (BDTM LSR II) and Nikon fluoresce microscope for additional visualization.Results: As shown in Figure 1a, our previous data show rs1045642 a synonymous coding change in ABCB1, in patients treated with GO presence of variant allele (CT/TT) is associated with significantly improved outcome as compared to patients with CC genotype (risk of relapse CC=45±10%, CT=30±7% TT=28±10%, CC vs. CT/TT p=0.007). Consistent with these results we observe significant decrease in cell viability with transfection with ABCB1-rs1045642/3435T as compared to the ABCB1-rs1045642/3435-C (P < 0.05) expression constructs. These results indicate that higher levels of calicheamicin toxicity in ABCB1-3435T transfected cells as opposed to cells transfected with the WT plasmid (Figure 1b and c) probably due to impact of T allele on ABCB1 levels. rs1045642-T allele has been shown to be associated with lower ABCB1 expression in several studies and with higher expression in one study. Hofmeyer et al has previously inked TT genotype of rs1045642 with lower expression and activity of ABCB1 which is consistent with our observation with respect to calicheamicin. Although more work is required to establish the clear relationship between ABCB1 genotypes and calicheamicin efflux, our results suggest that presence of ABCB1- low expressing rs1045642 TT genotype might be resulting in lower-efflux and thus higher intracellular abundance of calicheamicin in leukemic cells which in turn makes cells more sensitive to calicheamicin.Acknowledgments: Funding support from R21CA155524 and Children's Oncology group [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Detection of the Mitochondrial Membrane Potential by the Cationic Dye JC-1 in L1210 Cells with Massive Overexpression of the Plasma Membrane ABCB1 Drug Transporter.

JC-1, a cationic fluorescent dye when added to living cells, is known to be localized exclusively in mitochondria, particularly in good physiological conditions characterized by sufficient mitochondrial membrane potential (ΔΨ). The accumulation of JC-1 in these organelles leads to the formation J-aggregates (with a specific red fluorescence emission maximum at 590 nm), which is in addition to the typical green fluorescence of J-monomers (emission maximum of ∼529 nm). The lack of mitochondrial ΔΨ leads to the depression of JC-1 mitochondrial accumulation and a decrease in J-aggregate formation. Therefore, the ratio between the red and green fluorescence of cells loaded with JC-1 is often used for the detection of the mitochondrial membrane potential. However, JC-1 represents a suitable substrate of the multidrug transporter P-glycoprotein (P-gp). Therefore, the depression of the JC-1 content in intracellular space and particularly in the mitochondria to a level that is inefficient for J-aggregate formation could be expected in P-gp-positive cells. In the current paper, we proved this behavior on parental P-gp-negative L1210 (S) cells and their P-gp-positive variants obtained by either selection with vincristine (R) or transfection with the human gene encoding P-gp (T). P-glycoprotein inhibitors cyclosporine A and verapamil fail to restore JC-1 loading of the R and T cells to an extent similar to that observed in S cells. In contrast, the noncompetitive high affinity P-gp inhibitor tariquidar fully restored JC-1 accumulation and the presence of the typical red fluorescence of J-aggregates. In the presence of tariquidar, measurement of the JC-1 fluorescence revealed similar levels of mitochondrial membrane potential in P-gp-negative (S) and P-gp-positive cells (R and T).

Abstract 4895: Human ATP-binding cassette proteins ABCB1 and ABCG2 confer resistance to histone deacetylase 6 inhibitor ricolinostat (ACY-1215) in cancer cell lines

Abstract Histone deacetylase 6 (HDAC6), localized primarily in the cytoplasm, is responsible for the deacetylation of non-histone substrates such as α-tubulin and heat shock protein 90 (Hsp90). HDAC6 has recently emerged as an important epigenetic therapeutic target for cancer, and much effort has been invested into developing novel inhibitors of HDAC6. Ricolinostat (ACY-1215) is the first safe and well tolerated inhibitor of HDAC6 that has progressed into human clinical trials, both as a single agent and in combination therapy. Ricolinostat is currently under evaluation in phase I and II clinical trials in patients with multiple myeloma, relapsed or refractory lymphoid malignancies, relapsed chronic lymphocytic leukemia, unresectable or metastatic breast cancer and for the treatment of gynecological cancer. The overexpression of ATP-binding cassette (ABC) drug transporters ABCB1 and ABCG2 in cancer cells is often associated with the development of multidrug resistance (MDR) phenotype and poor clinical outcome in patients with metastatic breast cancer, chronic lymphocytic leukemia or relapsed multiple myeloma. In this study, we report that ABCB1 and ABCG2 are capable of actively transporting and conferring resistance to ricolinostat. Moreover, we demonstrated that the efficacy of ricolinostat on the activity of HDAC6 and drug-induced apoptosis was significantly reduced in ABCB1- and ABCG2-overexpressing cancer cells, which can be fully restored by an competitive inhibitor of ABCB1 or ABCG2.Our finding is supported by biochemical data and a docking analysis of ricolinostat in the drug-binding pocket of ABCB1 and ABCG2. Taken together, our study indicate that the overexpression ABCB1 or ABCG2 in cancer cells may potentially play a significant role in the development of resistance to ricolinostat in cancer cells. Citation Format: Sung-Han Hsiao, Shahrooz Vahedi, Suresh V. Ambudkar, Chung-Pu Wu. Human ATP-binding cassette proteins ABCB1 and ABCG2 confer resistance to histone deacetylase 6 inhibitor ricolinostat (ACY-1215) in cancer cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4895.

Abstract LB-056: Novel molecular mechanism underlying cancer drug resistance: Involvement of p58IPKchaperone pathway

Abstract Rhabdomyosarcoma (RMS) is an aggressive and highly malignant form of pediatric cancer that develops from skeletal muscle cells. A third of these cancers become drug resistant, metastatic and fatal. Current therapeutic approaches have not resulted in significant improvement in outcome for patients with recurrent and/or metastatic disease. Here, employing two RMS-derived cell lines (RH30, a drug sensitive and RD, a drug resistant), we demonstrate a novel underlying molecular mechanism of drug resistance in these tumor subtypes. We show that in response to chemotherapeutics drugs [actinomysin D, torisol, etoposide and arsenic trioxide (ATO)], endoplasmic reticulum (ER) stress associated unfolded protein response (UPR) signaling pathway could be induced in RH30 cell only, while drug resistance RD cells largely lack this ability. The mechanism underlying UPR induction involves activation of multifunctional molecular chaperone P58IPK that besides regulating protein homeostasis is an inhibitor of the interferon-induced double-stranded RNA-activated protein kinase (PKR). We observed that the expression of p58IPK is high in drug resistance RD cells as compared to drug sensitive RH30 cells. To confirm its role in drug resistance, we ablated or overexpressed p58IPK respectively in drug resistant RD and drug sensitive RH30 cells. The p58IPK depleted RD cells became sensitive to chemotherapeutic agents while overexpressing p58IPK RH30 cells manifested drug resistance. To further demonstrate the known functional consequences of manipulation in the expression of p58IPK we found that P58IPK over-expression in RH30 cells inhibited eIF2α phosphorylation and diminished apoptosis by downregulating ATF4/CHOP/Cleaved caspase-3, while silencing of p58IPK in RD cells enhanced eIF2α phosphorylation and increased ATF4/CHOP/Cleaved caspase-3-dependent apoptosis in response to chemotherapeutic drug treatment. The underlying mechanism by which P58IPK alters drug sensitivity in RMS cells involves its chaperone function by its binding to drug transporters ABCB-1 and ABCC-1. At baseline, in RD cells with high p58IPK manifest significantly higher dye efflux as compared to RH30 with low p58IPK. This baseline drug exclusion profile correlated with the ATPase activity of these ABC transporters. However, ablation or overexpression of p58IPK respectively, in RH30/RD cells reversed their drug resistance phenotype. Thus, we have identified a novel drug resistance mechanism in RMS. These results may have translational implications in overcoming drug-resistance in deadly neoplasm. These studies are in progress in our laboratory. Citation Format: Mohammad Athar, Ritesh Srivastava. Novel molecular mechanism underlying cancer drug resistance: Involvement of p58IPKchaperone pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-056.

Chemically Induced Degradation of Anaplastic Lymphoma Kinase (ALK).

We present the development of the first small molecule degraders that can induce anaplastic lymphoma kinase (ALK) degradation, including in non-small-cell lung cancer (NSCLC), anaplastic large-cell lymphoma (ALCL), and neuroblastoma (NB) cell lines. These degraders were developed through conjugation of known pyrimidine-based ALK inhibitors, TAE684 or LDK378, and the cereblon ligand pomalidomide. We demonstrate that in some cell types degrader potency is compromised by expression of drug transporter ABCB1. In addition, proteomic profiling demonstrated that these compounds also promote the degradation of additional kinases including PTK2 (FAK), Aurora A, FER, and RPS6KA1 (RSK1).

Genotypes and haplotypes of ABCB1 contribute to TAC chemotherapy response in Malaysian triple negative breast cancer patients

Triple negative breast cancer (TNBC) which is characterized by absent expression of ER, PR and HER2 receptors, is typically associated with poor treatment response and high recurrence risk rate or metastasis. ABCB1 is a drug efflux enzyme, involved in the drug transport across the membrane. Evidence have shown that, activation (or inactivation) of ABCB1 drug transporter gene may contribute to interindividual variability in treatment response. The present study aimed to investigate the association of SNPs (1236 C>T, 2677 G>T/A and 3435 C>T) and mRNA expression of ABCB1 with chemoresistance on TNBC patients undergoing chemotherapy with Taxane, Adriamycin and Cyclophosphamide (TAC) regimen. Our results showed that, homozygous variant (TT) genotype and variant allele (T) of ABCB1 3435 C>T polymorphism were significantly associated with higher risk for development of chemoresistance. The haplotypes ABCB1 3435T–1236T–2677T and ABCB1 3435T–1236G–2677T also showed significantly higher risk for development of resistance to chemotherapy. The mRNA expression of ABCB1 was significantly down regulated in cancerous as compared to normal adjacent tissues. Interestingly, up-regulation of ABCB1 mRNA expression was observed in patients who showed resistance compared to response to the chemotherapy. Thus, this study concludes that, genotypes and haplotypes of ABCB1 may contribute to the risk of chemoresistance in triple negative breast cancer patients undergoing TAC chemotherapy and may be useful as biomarkers for predicting chemoresistance.

Role of 2677G&gt;T&gt;A Genetic Polymorphism in the ABCB1 Drug Transporter in the Interaction between Voriconazole and Cyclosporine and Cyclosporine Related Nephrotoxicity After Allogeneic Stem Cell Transplantation

Role of 2677G&gt;T&gt;A Genetic Polymorphism in the ABCB1 Drug Transporter in the Interaction between Voriconazole and Cyclosporine and Cyclosporine Related Nephrotoxicity After Allogeneic Stem Cell Transplantation

Overexpression of the ABCB1 drug transporter in acute myeloid leukemia cells is associated with downregulation of latrophilin-1.

Finding new markers with appropriate prognostic levels for the differential diagnosis of neoplastic diseases represents an important issue for biomedical research. Recently, latrophilin-1 (LPHN1) was reported to be expressed in human monocytic leukemia cell lines and in primary human acute myeloid leukemia (AML) cells. However, this expression was found to be absent in healthy leukocytes. LPHN1 was therefore considered a novel biomarker of human AML. In previous papers, we established two P-gp-positive variants (SKM-1/VCR and MOLM-13/VCR) of AML cell lines derived from parental human AML cells SKM-1 and MOLM-13 by selection with VCR. The present paper addresses the measurement of LPHN1 expression in SKM-1 and MOLM-13 cells and their P-gp-positive variants. Both parental AML lines were positive for LPHN1 expression at the mRNA and protein levels. However, the expression of LPHN1 at both the mRNA and protein levels was reduced in both P-gp-positive SKM-1/VCR and MOLM-13/VCR variants of AML cells. Interestingly, we observed an elevation of the latrophilin-3 transcript in P-gp-positive variants of AML cell lines. The combined results suggest that alterations in latrophilin expression occur in AML cells expressing P-gp.

Apoptosis Induced by the Curcumin Analogue EF-24 Is Neither Mediated by Oxidative Stress-Related Mechanisms nor Affected by Expression of Main Drug Transporters ABCB1 and ABCG2 in Human Leukemia Cells.

The synthetic curcumin analogue, 3,5-bis[(2-fluorophenyl)methylene]-4-piperidinone (EF-24), suppresses NF-κB activity and exhibits antiproliferative effects against a variety of cancer cells in vitro. Recently, it was reported that EF-24-induced apoptosis was mediated by a redox-dependent mechanism. Here, we studied the effects of N-acetylcysteine (NAC) on EF-24-induced cell death. We also addressed the question of whether the main drug transporters, ABCB1 and ABCG2, affect the cytotoxic of EF-24. We observed that EF-24 induced cell death with apoptotic hallmarks in human leukemia K562 cells. Importantly, the loss of cell viability was preceded by production of reactive oxygen species (ROS), and by a decrease of reduced glutathione (GSH). However, neither ROS production nor the decrease in GSH predominantly contributed to the EF-24-induced cell death. We found that EF-24 formed an adduct with GSH, which is likely the mechanism contributing to the decrease of GSH. Although NAC abrogated ROS production, decreased GSH and prevented cell death, its protective effect was mainly due to a rapid conversion of intra- and extra-cellular EF-24 into the EF-24-NAC adduct without cytotoxic effects. Furthermore, we found that neither overexpression of ABCB1 nor ABCG2 reduced the antiproliferative effects of EF-24. In conclusion, a redox-dependent-mediated mechanism only marginally contributes to the EF-24-induced apoptosis in K562 cells. The main mechanism of NAC protection against EF-24-induced apoptosis is conversion of cytotoxic EF-24 into the noncytotoxic EF-24-NAC adduct. Neither ABCB1 nor ABCG2 mediated resistance to EF-24.

Tyrphostin RG14620 selectively reverses ABCG2-mediated multidrug resistance in cancer cell lines

The multidrug resistance (MDR) phenotype associated with the overexpression of ATP-binding cassette (ABC) drug transporters ABCB1, ABCC1 and ABCG2 is a major obstacle in cancer chemotherapy. Numerous epidermal growth factor receptor (EGFR) inhibitors have previously been shown capable of reversing MDR in ABCG2-overexpressing cancer cells. However, most of them are not transporter-specific due to the substantial overlapping substrate specificity among the transporters. In this study, we investigated the interaction between ABCG2 and tyrphostin RG14620, an EGFR inhibitor of the tyrphostin family, in multidrug-resistant cancer cell lines. We found that at nontoxic concentrations, tyrphostin RG14620 enhances drug-induced apoptosis and restores chemosensitivity to ABCG2-overexpressing multidrug-resistant cancer cells. More importantly, tyrphostin RG14620 is selective to ABCG2 relative to ABCB1 and ABCC1. Our findings were further supported by biochemical assays demonstrating that tyrphostin RG14620 stimulates ATP hydrolysis and inhibits photoaffinity labeling of ABCG2 with IAAP, and by a docking analysis of tyrphostin RG14620 in the drug-binding pocket of this transporter. Taken together, our findings indicate that tyrphostin RG14620 is a potent and selective modulator of ABCG2 that may be useful to overcome chemoresistance in patients with drug-resistant tumors.

NEW INSIGHTS INTO THE ROLE OF ABC-TRANSPORTERS IN AMYLOID-BETA CLEARANCE FROM CELLS IN THE BRAIN

Accumulation of toxic amyloid-beta peptides in the brain is an established hallmark of Alzheimer's Disease. Several members of the ABC transporter family of proteins implicated in the clearance of amyloid-beta peptides across the blood brain barrier (BBB) endothelium include the drug transporter ABCB1 (or P-glycoprotein) and the lipid transporter ABCG4. However, despite years of research in this area, there are still fundamental gaps in our understanding of whether amyloid-beta peptides are direct substrates for these transporters, and how these transport processes are regulated in various cell types in the brain. We have recently published that ABCG4 is a substrate for an E3-ubiquitin ligase, NEDD4–1 (see Aleidi et al JBC 2015;290:24604), which has also been implicated in regulating ABCB1 activity. Our research aims to investigate whether these transporters may potentially have a combined function in this clearance pathway and how this may be regulated. ABC transporter expression was measured via PCR and Western Blotting in Be(2)-C neuronal and hCMEC/D3 BBB endothelial cells. Cellular ABCB1 activity was measured using Calcein-AM assays while purified recombinant ABCB1, reconstituted in vesicles, was used to measure ATP-consumption in the presence of non-aggregated amyloid-beta (1–40)-peptide. ABCB1 was expressed in BBB endothelial cells as expected but was also surprisingly highly expressed in neuronal cells, a cell type in which ABCB1 function has not been previously characterised. Cellular ABCB1 activity was in line with expression of the transporter, suggesting that ABCB1 is active in neurons. Preliminary data show that amyloid-beta (1–40)-peptide directly stimulates ATP consumption of purified reconstituted ABCB1. In addition, ABCG4 was expressed in neurons as previously described, but also in BBB endothelial cells, where its function is similarly unknown. In both cell types, ABCG4 was not regulated by Liver X receptor (LXR) ligands, as expected from previously published work on the regulation of its close family member, the lipid transporter ABCG1. ABCB1 has the potential to be of importance in the initial export of amyloid-beta peptides from neurons. We are currently investigating a potentially combined function of these two transporters in net amyloid-beta export from endothelial cells and neurons in the brain.

Effect of ABCB1 (3435C>T) and CYP3A5 (6986A>G) genes polymorphism on tacrolimus concentrations and dosage requirements in liver transplant patients

BackgroundTacrolimus (TAC) is an immunosuppressant used in organtransplant recipients. It is a substrate of drug transporter ABCB1 as well as of cytochrome P4503A (CYP3A). AimTo assess the influence of ABCB1 (3435 C>T) and CYP3A5 (6986 A>G) genes polymorphism of liver transplant donors and recipients on blood level and dose requirements of oral tacrolimus, to help in designing an individualized tacrolimus regimen for liver transplant recipients. Subjects and methodsForty-eight adult liver transplant recipients and their matching living donors were prospectively enrolled in this study. TAC doses and blood concentration were recorded on 1st, 2nd and 3rd days, after 1 and 2weeks, and at 1, 3 and 6months postoperatively using ultra performance liquid chromatography Tandem mass spectrometry. Genotyping of ABCB1 (3435C>T) and CYP450 3A5 (6986A>G) genes were determined by Polymerase chain reaction followed by restriction fragment length polymorphism and by TaqMan allelic discrimination assay techniques, respectively. ResultsOf the enrolled 48 recipients, CYP3A5∗3/∗3 and CYP3A5∗1/∗3 genotypes were detected in 18 (37.5%) and in 20 (41.7%) recipients respectively, while ABCB1 CT and TT genotypes were detected in16 (33.3%) and 10 (20.8%) recipients respectively. TAC daily dose was significantly increased among recipients carrying ABCB1 CC genotype compared to recipients carrying CT and TT genotypes during and after the first month postoperatively. During 1st, 2nd days and 2weeks post-transplant, a significant increase of TAC concentration / dose ratio was observed among recipients carrying CYP3A5∗3∗3 genotype than recipients carrying 1∗1∗ and 1∗3∗ genotypes, and among recipients carrying ABCB1 CT and TT genotypes compared to those carrying CC genotype on 1st, 3rd days and at 3months postoperatively. ConclusionsABCB1 and CYP3A5 genetic polymorphism is one of the factors influencing TAC pharmacokinetics, screening for these SNPs prior to liver transplantation might be helpful for individualization of tacrolimus treatment.

Hernandezine, a Bisbenzylisoquinoline Alkaloid with Selective Inhibitory Activity against Multidrug-Resistance-Linked ATP-Binding Cassette Drug Transporter ABCB1.

The overexpression of ATP-binding cassette (ABC) drug transporter ABCB1 (P-glycoprotein, MDR1) is the most studied mechanism of multidrug resistance (MDR), which remains a major obstacle in clinical cancer chemotherapy. Consequently, resensitizing MDR cancer cells by inhibiting the efflux function of ABCB1 has been considered as a potential strategy to overcome ABCB1-mediated MDR in cancer patients. However, the task of developing a suitable modulator of ABCB1 has been hindered mostly by the lack of selectivity and high intrinsic toxicity of candidate compounds. Considering the wide range of diversity and relatively nontoxic nature of natural products, developing a potential modulator of ABCB1 from natural sources is particularly valuable. Through screening of a large collection of purified bioactive natural products, hernandezine was identified as a potent and selective reversing agent for ABCB1-mediated MDR in cancer cells. Experimental data demonstrated that the bisbenzylisoquinoline alkaloid hernandezine is selective for ABCB1, effectively inhibits the transport function of ABCB1, and enhances drug-induced apoptosis in cancer cells. More importantly, hernandezine significantly resensitizes ABCB1-overexpressing cancer cells to multiple chemotherapeutic drugs at nontoxic, nanomolar concentrations. Collectively, these findings reveal that hernandezine has great potential to be further developed into a novel reversal agent for combination therapy in MDR cancer patients.

Osimertinib (AZD9291) Attenuates the Function of Multidrug Resistance-Linked ATP-Binding Cassette Transporter ABCB1 in Vitro.

The effectiveness of cancer chemotherapy is often circumvented by multidrug resistance (MDR) caused by the overexpression of ATP-binding cassette (ABC) drug transporter ABCB1 (MDR1, P-glycoprotein). Several epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) have been shown previously capable of modulating the function of ABCB1 and reversing ABCB1-mediated MDR in human cancer cells. Furthermore, some TKIs are transported by ABCB1, which results in low oral bioavailability, reduced distribution, and the development of acquired resistance to these TKIs. In this study, we investigated the interaction between ABCB1 and osimertinib, a novel selective, irreversible third-generation EGFR TKI that has recently been approved by the U.S. Food and Drug Administration. We also evaluated the potential impact of ABCB1 on the efficacy of osimertinib in cancer cells, which can present a therapeutic challenge to clinicians in the future. We revealed that although osimertinib stimulates the ATPase activity of ABCB1, overexpression of ABCB1 does not confer resistance to osimertinib. Our results suggest that it is unlikely that the overexpression of ABCB1 can be a major contributor to the development of osimertinib resistance in cancer patients. More significantly, we revealed an additional action of osimertinib that directly inhibits the function of ABCB1 without affecting the expression level of ABCB1, enhances drug-induced apoptosis, and reverses the MDR phenotype in ABCB1-overexpressing cancer cells. Considering that osimertinib is a clinically approved third-generation EGFR TKI, our findings suggest that a combination therapy with osimertinib and conventional anticancer drugs may be beneficial to patients with MDR tumors.

Role of the Drug Transporter ABCC3 in Breast Cancer Chemoresistance.

Increased expression of ABC-family of transporters is associated with chemotherapy failure. Although the drug transporters ABCG2, ABCB1 and ABCC1 have been majorly implicated in cancer drug resistance, recent studies have associated ABCC3 with multi drug resistance and poor clinical response. In this study, we have examined the expression of ABCC3 in breast cancers and studied its role in drug resistance and stemness of breast cancer cells in comparison with the more studied ABCC1. We observed that similar to ABCC1, the transcripts levels of ABCC3 was significantly high in breast cancers compared to adjacent normal tissue. Importantly, expression of both transporters was further increased in chemotherapy treated patient samples. Consistent with this, we observed that treatment of breast cancer cell lines with anti-cancer agents increased their mRNA levels of both ABCC1 and ABCC3. Further, similar to knockdown of ABCC1, knockdown of ABCC3 also significantly increased the retention of chemotherapeutic drugs in breast cancer cells and rendered them more chemo-sensitive. Interestingly, ABCC1 and ABCC3 knockdown cells also showed reduction in the expression of stemness genes, while ABCC3 knockdown additionally led to a reduction in the CD44high/CD24low breast cancer stem-like subpopulation. Consistent with this, their ability to form primary tumours was compromised. Importantly, down-modulation of ABCC3 rendered these cells increasingly susceptible to doxorubicin in xenograft mice models in vivo. Thus, our study highlights the importance of ABCC3 transporters in drug resistance to chemotherapy in the context of breast cancer. Further, these results suggest that combinatorial inhibition of these transporters together with standard chemotherapy can reduce therapy-induced resistance in breast cancer.

Human ATP-Binding Cassette Transporter ABCG2 Confers Resistance to CUDC-907, a Dual Inhibitor of Histone Deacetylase and Phosphatidylinositol 3-Kinase.

CUDC-907 is a novel, dual-acting small molecule compound designed to simultaneously inhibit the activity of histone deacetylase (HDAC) and phosphatidylinositol 3-kinase (PI3K). Treatment with CUDC-907 led to sustained inhibition of HDAC and PI3K activity, inhibition of RAF-MEK-MAPK signaling pathway, and inhibition of cancer cell growth. CUDC-907 is currently under evaluation in phase I clinical trials in patients with lymphoma or multiple myeloma, and in patients with advanced solid tumors. However, the risk of developing acquired resistance to CUDC-907 can present a significant therapeutic challenge to clinicians in the future and should be investigated. The overexpression of ATP-binding cassette (ABC) drug transporter ABCB1, ABCC1, or ABCG2 is one of the most common mechanisms of developing multidrug resistance (MDR) in cancers and a major obstacle in chemotherapy. In this study, we reveal that ABCG2 reduces the intracellular accumulation of CUDC-907 and confers significant resistance to CUDC-907, which leads to reduced activity of CUDC-907 to inhibit HDAC and PI3K in human cancer cells. Moreover, although CUDC-907 affects the transport function of ABCG2, it was not potent enough to reverse drug resistance mediated by ABCG2 or affect the expression level of ABCG2 in human cancer cells. Taken together, our findings indicate that ABCG2-mediated CUDC-907 resistance can have serious clinical implications and should be further investigated. More importantly, we demonstrate that the activity of CUDC-907 in ABCG2-overexpressing cancer cells can be restored by inhibiting the function of ABCG2, which provides support for the rationale of combining CUDC-907 with modulators of ABCG2 to improve the pharmacokinetics and efficacy of CUDC-907 in future treatment trials.

Patient-Reported Outcomes (PRO) in the Setting of Relapsed Myeloma: The Influence of Treatment Strategies and Genetic Variants Predict Quality of Life and Pain Experience

Patient-Reported Outcomes (PRO) in the Setting of Relapsed Myeloma: The Influence of Treatment Strategies and Genetic Variants Predict Quality of Life and Pain Experience

Isoreserpine Reverses Multidrug Resistance Mediated by ABCB1

One of the major obstacles to successful cancer chemotherapy is the development of multidrug resistance (MDR) that is associated with the overexpression of ATP-binding cassette (ABC) drug transporter ABCB1 (P-glycoprotein/ MDR1). Currently, the most efficient way to overcome ABCB1-mediated MDR in cancer is by direct inhibition of ABCB1 function. Many drugs with known biological activities have been discovered to inhibit the function of ABCB1 and reverse ABCB1-mediated MDR in cancers. However, clinical trial results suggested that many of these clinically active drugs should not be used as ABCB1 modulators due to direct toxicity or undesirable side effects. In this study, we demonstrated that isoreserpine, an indole alkaloid with relatively low toxicity, can significantly inhibit ABCB1-mediated efflux of calcein-AM, a known substrate of ABCB1, in a dose-dependent manner. Moreover, we showed that at non-toxic concentrations, isoreserpine potently reversed ABCB1-mediated resistance to doxorubicin and colchicine in ABCB1-overexpressing human KB-V-1 epidermal cancer cells. Collectively, our findings revealed thatby inhibiting the transport function of ABCB1, isoreserpine can restore drug sensitivity of ABCB1-overexpressing cells to conventional chemotherapeutic drugs. In conclusion, isoreserpine should be further developed into a promising reversal agent for the treatment of MDR in ABCB1-overexpressing cancers.