ABC drug transporters: hereditary polymorphisms and pharmacological impact in MDR1, MRP1 and MRP2

Transport by ATP-dependent efflux pumps, such as P-glycoprotein (PGP) and multi-drug resistance related proteins (MRPs), influences bioavailability and disposition of drugs. These efflux pumps serve as defence mechanisms and determine bioavailability and CNS concentrations of many drugs. However, despite the fact that substantial data have been accumulated on the structure, function and pharmacological role of ABC transporters and even though modification of PGP function is an important mechanism of drug interactions and adverse effects in humans, there is a striking lack of data on variability of the underlying genes. This review focuses on the human drug transporter proteins PGP (MDR1) and the multi-drug resistance proteins MRP1 and MRP2. An overview is provided of pharmacologically relevant genetic, structural and functional data as well as on hereditary polymorphisms, their phenotypical consequences and pharmacological implications.     

The functions and structure of ABC transporters: implications for the design of new inhibitors of Pgp and MRP1 to control multidrug resistance (MDR)

Multidrug resistance (MDR) is a kind of acquired resistance of microorganisms and cancer cells to chemotherapic drugs that are characterized by different chemical structure and different mechanism of action. Classic MDR is the consequence of the over-expression of a variety of proteins that extrude the chemotherapic from the cell, lowering its concentration below the effective one. The ABC (ATP Binding Cassette) is a ubiquitous and important family of such transporter proteins. Members of this super family are present in mammals as well as in prokaryotic organisms and use ATP as the energy source to activate the extrusion process. P-glycoprotein (Pgp) and Multidrug Resistance Proteins (MRP1 and sister proteins) are the most important and widely studied members of ABC super family. Our knowledge about the structures and functions of transporter proteins has definitely improved in recent years, following the resolution of the structure of bacterial pumps which opened the way to the building of homology models for the more complex Pgp and MRP. It can be anticipated that these results will have a strong impact on the design of more potent and safer MDR reverters. A huge number of small molecules, many of natural origin, are able to reverse multidrug resistance by inhibiting the functions of Pgp, MRP1 and sister proteins and their action has been considered a possible way to reverse MDR. However, while a few compounds have reached clinical trials, none of them has, so far, been cleared for therapeutic use. Two main reasons are at the base of this difficulty: i) MDR is a complex phenomenon that may arise from several different biochemical mechanisms, with the consequence that inhibition of transporter proteins may be insufficient to reverse it; ii) the physiological role of Pgp and sister proteins requires more potent modulators with proper selectivity and pharmacokinetic in order to avoid unwanted side effects. This paper first reviews the most recent discoveries on the structures and functions of the ABC super family, in particular Pgp and MRP. Then, the medicinal chemistry of MDR reverters, in light of these findings, is discussed and the molecules that are presently in development are reviewed.     

ABC transporters and drug resistance in parasitic protozoa

Parasitic protozoa are responsible for a wide spectrum of diseases in humans and domestic animals. The main line of defence available against these organisms is chemotherapy. However, the application of chemotherapeutic drugs has resulted in the development of resistance mechanisms, which limit the number of antiprotozoal drugs that are effective in the treatment and control of parasitic diseases. Knowledge about the resistance mechanisms involved may allow the development of new drugs that minimise or circumvent drug resistance or may identify new targets for drug development. This review focuses on the role of protozoal ATP-binding cassette (ABC) transporters in drug resistance. These membrane proteins mediate the ATP-dependent transport of a wide variety of chemotherapeutic drugs away from their targets inside the parasites. The genome sequence of Plasmodium falciparum and Plasmodium yoelii has recently been completed, and the sequencing of other parasitic genomes are now underway. As a result, many new membrane transporters belonging to the ABC superfamily are being discovered. We review the ABC transporters in major parasitic protozoa, including Plasmodium, Leishmania, Trypanosoma and Entamoeba species. Transporters with an established role in drug resistance have been emphasised, but newly discovered transporters with a significant amino acid sequence identity to established ABC drug transporters have also been included.     

The role of ABC transporters in drug resistance, metabolism and toxicity

ATP Binding Cassette (ABC) transporters form a special family of membrane proteins, characterized by homologous ATP-binding, and large, multispanning transmembrane domains. Several members of this family are primary active transporters, which significantly modulate the absorption, metabolism, cellular effectivity and toxicity of pharmacological agents. This review provides a general overview of the human ABC transporters, their expression, localization and basic mechanism of action. Then we shortly deal with the human ABC transporters as targets of therapeutic interventions in medicine, including cancer drug resistance, lipid and other metabolic disorders, and even gene therapy applications. We place a special emphasis on the three major groups of ABC transporters involved in cancer multidrug resistance (MDR). These are the classical P-glycoprotein (MDR1, ABCB1), the multidrug resistance associated proteins (MRPs, in the ABCC subfamily), and the ABCG2 protein, an ABC half-transporter. All these proteins catalyze an ATP-dependent active transport of chemically unrelated compounds, including anticancer drugs. MDR1 (P-glycoprotein) and ABCG2 preferentially extrude large hydrophobic, positively charged molecules, while the members of the MRP family can extrude both hydrophobic uncharged molecules and water-soluble anionic compounds. Based on the physiological expression and role of these transporters, we provide examples for their role in Absorption-Distribution-Metabolism-Excretion (ADME) and toxicology, and describe several basic assays which can be applied for screening drug interactions with ABC transporters in the course of drug research and development.     

Role of ABC transporters in veterinary drug research and parasite resistance

A considerable body of research has been carried out in order to throw light on the pharmacological and toxicological impact of ATP-binding cassette (ABC) drug efflux transporters such as P-glycoprotein and Breast Cancer Resistance Protein (BCRP/ABCG2/MXR). Most studies focus on their role in rendering cancer cells resistant to anticancer drugs. Drug transporters are expressed in many tissues and they are strongly involved in the oral bioavailability, and the hepatobiliary, direct intestinal and renal excretion of many drugs. In veterinary therapy, some anti parasitic drugs and/or their metabolites, such as ivermectin, moxidectin, albendazole sulfoxide, which are widely used, have been shown to be actively transported by efflux pumps. This interaction plays an important role in drug disposition since its inhibition has been shown to increase the drug bioavailability in some domestic species. Moreover, some authors have reported that parasite resistance to anthelmintic drugs may be mediated by parasite P-glycoprotein efflux. In addition, the importance of milk residues for human nutrition has aroused increasing concern about the inadvertent transfer of drugs and other substances into mammary milk of domestic animals, potentially posing a health risk to consumers. Recently, the important role of BCRP in the secretion of its substrates in milk has been demonstrated.     

ABC transporters in Leishmania and their role in drug resistance.

ABC transporters have been found in several parasitic protozoa including Leishmania. At least two Leishmania ABC transporters are involved in drug resistance. One is PgpA, which is involved in resistance to arsenic and antimony-containing compounds. Antimonials are the drug of choice against Leishmania infections. Transfection and biochemical studies suggest that PgpA recognizes metals conjugated to thiols. The second ABC transporter is closely related to mammalian P-glycoproteins and confers resistance to anticancer drugs by a mechanism that remains to be elucidated. Additional ABC transporters are likely to be present in Leishmania and these are discussed in relation to the phenomenon of antimony resistance.     

ABC transporters and their role in nucleoside and nucleotide drug resistance

ATP-binding cassette (ABC) transporters confer drug resistance against a wide range of chemotherapeutic agents, including nucleoside and nucleotide based drugs. While nucleoside based drugs have been used for many years in the treatment of solid and hematological malignancies as well as viral and autoimmune diseases, the potential contribution of ABC transporters has only recently been recognized. This neglect is likely because activation of nucleoside derivatives require an initial carrier-mediated uptake step followed by phosphorylation by nucleoside kinases, and defects in uptake or kinase activation were considered the primary mechanisms of nucleoside drug resistance. However, recent studies demonstrate that members of the ABCC transporter subfamily reduce the intracellular concentration of monophosphorylated nucleoside drugs. In addition to the ABCC subfamily members, ABCG2 has been shown to transport nucleoside drugs and nucleoside-monophosphate derivatives of clinically relevant nucleoside drugs such as cytarabine, cladribine, and clofarabine to name a few. This review will discuss ABC transporters and how they interact with other processes affecting the efficacy of nucleoside based drugs.     

ATP结合盒转运蛋白介导的MDR逆转剂的实验研究现况

多药耐药性(mu ltidrug resistance,MDR)是指人肿瘤细胞对结构各异的化疗药物产生交叉耐药,是肿瘤化疗失败的主要原因之一。为了增强肿瘤细胞对化疗药物的敏感性,积极寻求有效逆转MDR的药物已成为研究重点。体外被证实具有逆转耐药活性的化合物很多,但由于体内要达到体外逆转试验的有效浓度所需剂量过大,毒副作用大,因此限制了临床应用。ATP结合盒转运蛋白家族(ATP-b ind ing cassette,ABC)介导的药物外排机制是目前MDR的主要机制,ABC转运蛋白家族中与多药耐药性相关的转运蛋有P-糖蛋白(P-gp)、多药耐药相关蛋白(MRP)和乳腺癌耐药蛋白(BCRP)。本文对ABC转运蛋白介导的MDR逆转剂的研究进展做一综述。     

Modulation of the activity of ABC transporters (P-glycoprotein,MRP2, BCRP) by flavonoids and drug response

The present article aims to review the up-to-date information on the most recent studies of the interaction of flavonoids with ABC transporters, in particular the drug pharmacokinetic consequences of such a relationship. In addition, the modulation of the expression of the ABC transporters by flavonoids is also illustrated. Flavonoids are a large group of plant polyphenols present extensively in our daily diets and herbal products. High intake of isoflavones has been associated with a variety of beneficial effects on several common diseases. These polyphenols interact with ABC drug transporters involved in drug resistance and drug absorption, distribution and excretion. A number of studies have demonstrated inhibition of drug transporters by flavonoids. This flavonoid-ABC-transporter interaction could be beneficial for poorly absorbed drugs but could also result in severe drug intoxication, especially drugs with a narrow therapeutic window. On the other hand, flavonoids are themselves substrates of ABC transporters. These proteins can affect the oral availability and tissue distribution of these compounds, modifying their beneficial effects. The challenge is to find a suitable way to predict harmful drug–flavonoid interactions mediated by these transporters. © 2009 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 99:598–617, 2010     

转运体在药物经肝脏清除过程中的作用

肝脏在药物的体内清除过程中具有重要作用,它不仅是药物代谢的主要场所,还控制着药物及其代谢物的胆汁排泄过程。转运体是控制细胞内外物质传输的一类功能性膜蛋白,其在肝脏有广泛表达,并能对药物进入肝细胞以及排泄至胆汁的过程进行调控,因而,对于肝脏清除过程具有重要作用。本文从肝脏中重要转运体的分布、功能以及底物选择性出发,对其在药物的肝脏清除中的作用、由其引起的药物药物相互作用以及重要转运体的基因多态性研究进行了综述。     

Fungal ATP-binding cassette (ABC) transporters in drug resistance & detoxification

Pleiotropic drug resistance (PDR) is a well-described phenomenon occurring in fungi. PDR shares several similarities with processes in bacteria and higher eukaryotes. In mammalian cells, multidrug resistance (MDR) develops from an initial single drug resistance, eventually leading to a broad cross-resistance to many structurally and functionally unrelated compounds. Notably, a number of membrane-embedded energy-consuming ATP-binding cassette (ABC) transporters have been implicated in the development of PDR/MDR phenotypes. The yeast Saccharomyces cerevisiae genome harbors some 30 genes encoding ABC proteins, several of which mediate PDR. Therefore, yeast served as an important model organism to study the functions of evolutionary conserved ABC genes, including those mediating clinical antifungal resistance in fungal pathogens. Moreover, yeast cells lacking endogenous ABC pumps are hypersensitive to many antifungal drugs, making them suitable for functional studies and cloning of ABC transporters from fungal pathogens such as Candida albicans. This review discusses drug resistance phenomena mediated by ABC transporters in the model system S. cerevisiae and certain fungal pathogens.     

Polymorphisms of MRP1 (ABCC1) and related ATP-dependent drug transporters

Genetic variations in drug metabolizing enzymes and targets are established determinants of adverse drug reactions and interactions, but less is known about the role of genetic polymorphisms in membrane transport proteins. MRP1 (ABCC1) is one of 13 polytopic membrane proteins that comprise the 'C' subfamily of the ATP-binding cassette (ABC) superfamily of transport proteins. MRP1 and related ABCC family members, including MRP2, 3, 4 and 5 (ABCC2, 3, 4 and 5), each have a distinctive pattern of tissue expression and substrate specificity. Together, these five transporters play important roles in the disposition and elimination of drugs and other organic anions, and in maintenance of blood-tissue barriers, as confirmed by enhanced chemosensitivity of respective knockout mice. Moreover, Mrp2 (Abcc2) deficient animals display mild conjugated hyperbilirubinemia, corresponding to a human condition known as Dubin-Johnson syndrome (DJS). Naturally occurring mutations in MRP/ABCC-related drug transporters have been reported, some of which are non-synonymous single nucleotide polymorphisms. The consequences of the resulting amino acid changes can sometimes be predicted from in vitro site-directed mutagenesis studies or from knowledge of mutations of analogous (conserved) residues in ABCC proteins that cause DJS, Pseudoxanthoma elasticum (ABCC6), cystic fibrosis (CFTR/ABCC7) or persistent hyperinsulinemic hypoglycemia of infancy (SUR1/ABCC8). Continual updating of databases of sequence variants and haplotype analysis, together with in vitro biochemical validation assays and pharmacological studies in knockout animals, should make it possible to determine how genetic variation in the MRP-related transporters contributes to the range of responses to drugs and chemicals observed in different human populations.     

Reversing agents for ATP-binding cassette (ABC) transporters: application in modulating multidrug resistance (MDR)

One of the main reasons for the failure in cancer chemotherapy is the existence of multidrug resistance (MDR) mechanisms. One form of MDR phenotype is contributed by a group of plasma membrane proteins that belong to a large superfamily of proteins called the ATP-binding cassette (ABC) transporters. There has been intense search for compounds, which can act at reversing MDR phenotype exhibited by ABC transporters such as P-glycoprotein (P-gp), multidrug-resistance protein (MRP) and breast cancer resistance protein (BCRP). Reversing agents can be designed to target MDR-associated ABC transporters at three levels - the protein, mRNA or DNA level. This review aims at describing, over-viewing and discussing currently known MDR reversing agents, which have been shown to act at either of the three levels against ABC transporters. Other potential agents and strategies, which can be used to reverse the MDR phenotype, are also discussed.     

MRP1 and its role in anticancer drug resistance

The phenomenon of multidrug resistance (MDR) in cancer is associated with the overexpression of the ATP-binding cassette (ABC) transporter proteins, including multidrug resistance-associated protein 1 (MRP1) and P-glycoprotein. MRP1 plays an active role in protecting cells by its ability to efflux a vast array of drugs to sub-lethal levels. There has been much effort in elucidating the mechanisms of action, structure and substrates and substrate binding sites of MRP1 in the last decade. In this review, we detail our current understanding of MRP1, its clinical relevance and highlight the current environment in the search for MRP1 inhibitors. We also look at the capacity for the rapid intercellular transfer of MRP1 phenotype from spontaneously shed membrane vesicles known as microparticles and discuss the clinical and therapeutic significance of this in the context of cancer MDR.     

The role of organic ion transporters in drug disposition: an update

Transporters for organic anions and organic cations in kidney, liver, intestine, brain, and placenta play essential roles in drug disposition. The cloning and characterization of these transporters have significantly advanced our understanding of the molecular and cellular mechanisms of the drug disposition process. This review aims at updating the recent knowledge of general properties, structure-function relationships, and regulation mechanisms of the organic anion transporters (OATs) and the organic cation transporters (OCTs). Such information will be essential for the design and development of new drugs to maximize therapeutic efficacy and minimize drug-induced toxicity as well as unwanted drug-drug interactions.     

Single nucleotide polymorphisms in multidrug resistance associated protein 2 (MRP2/ABCC2): its impact on drug disposition

Multidrug resistance associated protein 2 (MRP2/ABCC2), expressed on the bile canalicular membrane, plays an important role in the biliary excretion of various kinds of substrates. In addition, MRP2 is also expressed on the apical membrane of epithelial cells such as enterocytes. It is possible that the inter-individual difference in the function of MRP2 affects the drug disposition. In the present article, we will summarize the physiological and pharmacological role of MRP2, particularly focusing on the factors affecting its transport function such as single nucleotide polymorphisms and/or the induction/down regulation of this transporter. Mutations found in patients suffering from the Dubin-Johnson syndrome, along with the amino acid residues which are involved in supporting the transport activity of MRP2, are also summarized.     

Sequencing and tissue distribution of the canine MRP2 gene compared with MRP1 and MDR1

The effects of xenobiotic drugs and toxic compounds depend largely on their kinetic properties, which can be influenced by transmembrane drug transporters like MDR1/P-glycoprotein and the drug-conjugate transporters multidrug resistance protein (MRP) 1 and 2. As the dog is a preferential species used in the pharmacological and toxicological evaluation of new drugs, we sequenced the canine MRP2 cDNA and investigated its expression in various canine tissues compared with the related transporters MRP1 and P-glycoprotein. The tissue distribution pattern of these ABC-transporters differs partially from the distribution described in humans. So we found relatively high renal and low hepatic canine MRP2 expression levels, relatively high hepatic canine MRP1 expression levels, and quite high levels of MRP1 and P-glycoprotein in the dog brain. The knowledge of the tissue distribution pattern of these transporters will aid to interpret pharmacokinetic and toxicokinetic data gained from dog studies and to extrapolate them to humans.     

The role of multidrug resistance proteins MRP1, MRP2 and MRP3 in cellular folate homeostasis

Previously, we reported that the multidrug resistance proteins MRP1, MRP2 and MRP3 confer resistance to therapeutic antifolates by mediating their cellular extrusion. We now determined whether MRPs also play a role in controlling cellular homeostasis of natural folates. In MRP1, MRP2 and MRP3-transfected 2008 human ovarian carcinoma cells total cellular folate content was 32-38% lower than in 2008 cells (105+/-14pmolfolate/mgprotein) when grown in medium containing 2.3 microM folic acid (FA). Under these conditions cellular growth rates were not compromised. However, when cells were challenged under folate-depleted conditions with a short exposure (4 hr) to FA or leucovorin, MRP1 and MRP3 overexpressing cells were impaired in their growth. In contrast to wild-type cells, MRP1 transfected cells retained only 60% of the maximum growth when exposed to 500 nM leucovorin or 500 microM FA. For 2008/MRP1 and 2008/MRP3 cells FA growth stimulation capacity was dramatically decreased when, during a 4 hr exposure, metabolism into rapidly polyglutamatable and retainable dihydrofolate was blocked by the dihydrofolate reductase inhibitor trimetrexate. To retain growth under such conditions MRP1 overexpressing cells required much higher concentrations of FA (EC(50) > 500 microM) compared to 2008 cells (EC(50): 12 microM). These results suggest that down- and up-regulation of MRP1 (and MRP3) expression can influence cellular folate homeostasis, in particular when cellular retention by polyglutamylation of folates is attenuated.