ATP结合盒转运蛋白介导的肿瘤多药耐药及其逆转剂

肿瘤多药耐药（MDR）是导致肿瘤化疗失败的主要原因之一，是多种复杂机制：共同作用的结果。其中，肿瘤细胞膜上ATP结合盒（ABC）转运蛋白的表达或功能异常是肿瘤细胞产生MDR的重要机制之一。现以P-糖蛋白、多药耐药相关蛋白、乳腺癌耐药蛋白等为例，综述ABC转运蛋白的结构功能、与肿瘤的关系以及MDR逆转剂的研发进展。     

microRNAs与ABC转运蛋白介导的肿瘤多药耐药研究进展

目的回顾ABC转运蛋白与肿瘤耐药的研究现状,探讨miRNA在逆转肿瘤耐药过程中的作用机制。方法应用PubMed和CNKI期刊全文数据库检索系统,检索2010-01-01-2014-05-20的相关文献,以"ABC转运蛋白、miRNA和多药耐药"为关键词。纳入标准:1)ABC转运蛋白的表达水平与肿瘤耐药;2)miRNA对ABC转运蛋白表达水平的调控;3)miRNA对肿瘤细胞药物敏感性的影响,根据纳入标准符合分析的文献34篇。结果大多数癌症患者使用一种化疗药物治疗后,肿瘤细胞可能因为种种原因,不仅对该药产生耐药,而且对多种结构不同和作用机制完全不同的其他药物也产生交叉耐药。研究表明,在多药耐药导致肿瘤化疗失败的众多原因中,ABC转运蛋白过表达是导致肿瘤多药耐药的主要原因之一。在耐药肿瘤细胞当中高表达的ABC转运蛋白主要有乳腺癌耐药蛋白ABCG2,多药耐药相关蛋白ABCC1,P-糖蛋白ABCB1,这些蛋白采用ATP水解的能量将细胞内药物泵出细胞外,从而降低细胞内药物的浓度,使细胞产生耐药性。miRNA能与ABC转运蛋白mRNA的3′UTR结合,使mRNA降解或抑制其翻译,导致目标蛋白的表达受到抑制,从而增加肿瘤细胞的药物敏感性,逆转由ABC转运蛋白过表达引起的肿瘤多药耐药。结论 miRNA可以逆转由ABC转运蛋白家族高表达所引起的肿瘤耐药,这为肿瘤多药耐药的研究提供了新的思路。     

ATP结合盒转运蛋白介导的肿瘤多药耐药及其逆转剂

肿瘤多药耐药(MDR)是导致肿瘤化疗失败的主要原因之一,是多种复杂机制共同作用的结果。其中,肿瘤细胞膜上ATP结合盒(ABC)转运蛋白的表达或功能异常是肿瘤细胞产生MDR的重要机制之一。现以P-糖蛋白、多药耐药相关蛋白、乳腺癌耐药蛋白等为例,综述ABC转运蛋白的结构功能、与肿瘤的关系以及MDR逆转剂的研发进展。     

Anticancer drugs and ABC transporters

Anticancer drugs interact directly with their molecular targets in cancer cells for effective cancer chemotherapy. The direct interaction between drug and cancer cell depends on the pharmacokinetics, which consists of absorption, distribution, metabolism, and excretion phases. In the excretion phase, ATP-binding cassette (ABC) transporters are the most important proteins in cell membranes. The ABC transporters export drugs out of cells by ATP-dependent energy, leading to drug resistance with reduced concentrations of intracellular drugs. In addition, the transporters sequestrate intracellular drugs into membrane vesicles in cytoplasm, also resulting in drug resistance. On the other hand, they are also involved in drug absorption. To date, 48 ABC genes have been isolated and classified into the seven groups of ABCA to ABCG. Among them, P-glycoprotein/ABCB 1, MRP 1/ ABCC 1, MRP 2/ABCC 2, MRP 3/ABCC 3, and BCRP/ABCG 2 strongly confer anticancer drug resistance, and they have different substrate drugs. Interestingly, recent molecular-targeted drugs, such as imatinib and gefitinib, were very recently found to be substrates for P-glycoprotein and/or BCRP. Additionally, polymorphism of ABC genes affects pharmacokinetics, drug effectiveness, and adverse events. Thus, ABC transporters are clinically important molecules, and much information is needed in the clinic.     

Role of ABC transporters in cancer chemotherapy

Multidrug resistance(MDR) in cancer cells can significantly attenuate the response to chemotherapy and increase the likelihood of mortality.The major mechanism involved in conferring MDR is the overexpression of ATP-binding cassette(ABC) transporters,which can increase efflux of drugs from cancer cells,thereby decreasing intracellular drug concentration.Modulators of ABC transporters have the potential to augment the efficacy of anticancer drugs.This editorial highlights some major findings related to ABC transporters and current strategies to overcome MDR.     

Methylation of miR-129-5p CpG island modulates multi-drug resistance in gastric cancer by targeting ABC transporters

Recent studies have reported that hyper-methylation in the promoter region of miRNAs could silence the expression of tumor suppressive miRNAs and might play significant roles in the process of tumor development. However, the potential mechanisms regarding how methylation of miRNA CpG Island could regulate cancer cell chemo-resistance have not yet been studied. Using microarray and BSP (Bisulfate Sequencing PCR) assays, we found that compared with the parent SGC7901/VCR cells, expression of miR-129-5p was restored in SGC7901/VCR gastric cancer multi-drug resistant cell line treated by de-methylation reagent (5-AZA-dC). Using gain or loss of function assays, we found the over-expressed miR-129-5p reduced the chemo-resistance of SGC7901/VCR and SGC7901/ADR cells, while down-regulation of miR-129-5p had an opposite effect. Furthermore, three members of multi-drug resistance (MDR) related ABC transporters (ABCB1, ABCC5 and ABCG1) were found to be direct targets of miR-129-5p using bioinformatics analysis and report gene assays. The present study indicated that hyper-methylation of miR-129-5p CpG island might play important roles in the development of gastric cancer chemo-resistance by targeting MDR related ABC transporters and might be used as a potential therapeutic target in preventing the chemo-resistance of gastric cancer.     

The role of ABC transporters in clinical practice

Drug resistance remains one of the primary causes of suboptimal outcomes in cancer therapy. ATP-binding cassette (ABC) transporters are a family of transporter proteins that contribute to drug resistance via ATP-dependent drug efflux pumps. P-glycoprotein (P-gp), encoded by the MDR1 gene, is an ABC transporter normally involved in the excretion of toxins from cells. It also confers resistance to certain chemotherapeutic agents. P-gp is overexpressed at baseline in chemotherapy-resistant tumors, such as colon and kidney cancers, and is upregulated after disease progression following chemotherapy in malignancies such as leukemia and breast cancer. Other transporter proteins mediating drug resistance include those in the multidrug-resistance-associated protein (MRP) family, notably MRP1, and ABCG2. These transporters are also involved in normal physiologic functions. The expressions of MRP family members and ABCG2 have not been well worked out in cancer. Increased drug accumulation and drug resistance reversal with P-gp inhibitors have been well documented in vitro, but only suggested in clinical trials. Limitations in the design of early resistance reversal trials contributed to disappointing results. Despite this, three randomized trials have shown statistically significant benefits with the use of a P-gp inhibitor in combination with chemotherapy. Improved diagnostic techniques aimed at the selection of patients with tumors that express P-gp should result in more successful outcomes. Further optimism is warranted with the advent of potent, nontoxic inhibitors and new treatment strategies, including the combination of new targeted therapies with therapies aimed at the prevention of drug resistance.     

Multidrug resistance mediated by P-glycoproteins.

Overexpression of P-glycoprotein genes is a well-established cause of one form of multidrug resistance. P-glycoproteins are plasma membrane proteins containing two ATP-binding sites and twelve putative transmembrane segments. P-glycoproteins are thought to act as ATP-dependent drug efflux pumps, actively extruding a range of structurally different, hydrophobic drugs from the cell. This simple model can account for the properties of multidrug resistant cells, even those that seem to require more complex explanations. The structure and function of P-glycoprotein genes has been studied in mammals and in several lower eukaryotes. These studies are helping to delineate the range of drugs that can be transported by P-glycoproteins; the genetic mechanisms that can lead to elevated cellular P-glycoproteins levels; and the evolution of the versatile and prolific P-glycoprotein gene family. The physiological function of the human P-glycoproteins encoded by the MDR1 and MDR3 (or MDR2) genes remains a matter of speculation.     

ABCG2介导的肿瘤多药耐药

ABCG2是ATP结合盒(ABC)转运超家族中的一员,其过表达被认为是限制多种化疗药物在细胞内聚集的重要机制之一.ABCG2底物范围广泛,包括多种抗肿瘤药物和环境致癌物等,与肿瘤的多药耐药(MDR)和肿瘤发展有关.以ABCG2为靶点逆转MDR受到广泛关注.     

ABC transporters and drug resistance in leukemia: was P-gp nothing but the first head of the Hydra?

More than 30 years ago it was discovered that permeability glycoprotein (P-gp) can cause drug resistance. Over the following decades numerous studies showed that high expression of P-gp is associated with poor prognosis in acute myeloid leukemia in adults and that it causes multidrug resistance via ATP-dependent drug efflux. It was hoped that an inhibition of P-gp could sensitize resistant leukemic cells to chemotherapy and thus improve treatment results. Today we know that the family of ATP-binding cassette transporters (ABC transporters) comprises 48 different proteins. Some of them seem to be able to cause drug resistance as well as P-gp. This review focuses on emerging data on the clinical relevance of other ABC transporters, such as BCRP, MRP3, and ABCA3. When Heracles fought the ancient Hydra, he had to fight all the heads at ones but only one head was vital for the beast. Can we block all the relevant ABC transporters at once? Is there one transporter that is more important than the others?     

Multidrug resistance transporters and modulation

Multidrug resistance (MDR), whereby tumor cells simultaneously possess intrinsic or acquired cross-resistance to diverse chemotherapeutic agents, hampers the effective treatment of cancer. Molecular investigations in MDR resulted in the isolation and characterization of genes coding for several proteins associated with MDR, including P-glycoprotein (P-gp), the multidrug resistance associated protein (MRP1), the lung resistance protein (LRP), and, more recently, the breast cancer resistance protein (BCRP). These transmembrane proteins cause MDR either by decreasing the total intracellular retention of drugs or redistributing intracellular accumulation of drugs away from target organelles. These proteins are expressed at varying degrees in different neoplasms, including the AIDS-associated non-Hodgkin lymphoma and Kaposi sarcoma and are generally associated with poor prognosis. Several MDR-reversing agents are in various stages of clinical development. First-generation modulators such as verapamil, quinidine, and cyclosporin required high doses of drugs to reverse MDR and were associated with unacceptable toxicities. Second- and third-generation MDR inhibitors include PSC 833, GF120918, VX-710, and LY335979, among others. Limitations to the use of these modulators include multiple and redundant cellular mechanisms of resistance, alterations in pharmacokinetics of cytotoxic agents, and clinical toxicities. Studies to validate the role of MDR reversal in the treatment of various malignancies are underway. A potential use of these agents may be to enhance intestinal drug absorption and increase drug penetration to biologically important protective barriers, such as the blood-brain, blood-cerebrospinal fluid, and the maternal-fetal barriers. The use of MDR modulators with drugs such as the antiviral protease inhibitors and cytotoxics may enhance drug accumulation in sanctuary sites that are traditionally impenetrable to these agents.     

Uptake of meta-iodobenzylguanidine in neuroendocrine tumours is mediated by vesicular monoamine transporters

The radio-iodinated noradrenaline analogue meta-iodobenzylguanidine (MIBG) can be used for scintigraphy and radiation therapy of neuroendocrine (NE). The aim of the present study was to study the importance of vesicular monoamine transporters (VMATs) for the uptake of (123)I-MIBG in NE tumours. In nude mice, bearing the human transplantable midgut carcinoid GOT1, all organs and xenografted tumours accumulated (123)I after i.v. injection of (123)I-MIBG. A high concentration of (123)I was maintained in GOT1 tumours and adrenals, which expressed VMATs, but rapidly decreased in all other tissues. In the VMAT-expressing NE tumour cell lines GOT1 and BON and in VMAT-expressing primary NE tumour cell cultures (carcinoids, n=4 and pheochromocytomas, n=4), reserpine significantly reduced the uptake of (123)I-MIBG. The membrane pump inhibitor clomipramine had no effect on the uptake of (123)I-MIBG in GOT1 and BON cells, but inhibited the uptake in one out of four primary carcinoid cell cultures and three out of four primary pheochromocytoma cell cultures. In conclusion, VMATs and secretory granules are of importance for the uptake and retention of (123)I-MIBG in NE tumours. Information about the type and degree of expression of VMATs in NE tumours may be helpful in future to select patients suitable for radiation therapy with radio-iodinated MIBG.     

Drug transporters as targets for cancer chemotherapy

Transporter proteins play an important role in taking up nutrients into and effluxing xenobiotics out of cells to sustain cell survival. Transporters that affect drug absorption, distribution and excretion are the so-called drug transporters. In the last decade, a number of studies revealed interactions between drug transporters and clinically important anticancer agents. Utilizing the knowledge of transporter functions offers us the possibility of delivering a drug to the target tissues, avoiding distribution to other tissues and improving oral bioavailability. Many transporters have been reported to be differentially up-regulated in cancer cells compared to normal tissues, suggesting that the differential expression of transporters in cancer cells may provide good targets for enhancing drug delivery as well as diagnostic markers for cancer therapy. This review will focus on the role of drug transporters in the adaptation and growth of tumors and in their potential usefulness as therapeutic targets in cancer.     

Drug resistance

An overview of the current mechanisms identified as associated with the expression of multidrug resistance is provided. The clinical relevance of multidrug resistance associated with the overexpression of the three major drug resistance-associated proteins namely, P-glycoprotein, MRP and/or LRP, is discussed. Alternate forms of multidrug resistance involving the reduced or altered expression of either or both of the topoisomerases is next considered, followed by a short section relating to resistance to the platinum complexes and the contribution of altered DNA repair. For those involved in new drug discovery programmes for cancer chemotherapy, the identification and study of these new major intracellular targets makes this an exciting era.     

Delta-Aminolevulinic acid transport in murine mammary adenocarcinoma cells is mediated by beta transporters

Delta-aminolevulinic acid, the precursor of porphyrin biosynthesis has been used to induce the endogenous synthesis of the photosensitiser protoporphyrin IX for photodynamic therapy in the treatment of various tumours. The aim of this work was to characterise the delta-aminolevulinic acid transport system in the murine mammary adenocarcinoma cell line LM3 using (14)C-delta-aminolevulinic acid, to finally improve delta-aminolevulinic acid incorporation in mammalian cells. Our results showed that delta-aminolevulinic acid is incorporated into these cells by two different mechanisms, passive diffusion which is important at the beginning of the incubation, and active transport. Specificity assays suggested that the transporter involved in delta-aminolevulinic acid incorporation is a BETA transporter, probably GAT-2.     

Expression and induction by dexamethasone of ABC transporters and nuclear receptors in a human T-lymphocyte cell line

Abstract

The efficacy of drugs acting within lymphocytes, like antiretroviral drugs in the treatment of HIV infection, depends on their intracellular concentrations modulated by efflux proteins like ABCB1 (P-glycoprotein). In lymphocytes, two glucocorticoids, prednisone and prednisolone, have been shown to induce ABCB1 activity. Yet, no data exist regarding dexamethasone (DEX). We report the modulation of ABC transporters and nuclear receptors’ expression by DEX in a commonly used model of human lymphocytes. CCRF-CEM cells were exposed to DEX (100 nM, 2 μM) for 24 to 72 hours. ABCB1 activity was measured using DiOC₆ efflux in flow cytometry. Gene expression levels were quantified by qRT-PCR. ABCB1 activity and mRNA expression increased with DEX concentrations and incubation times. DEX (1 μM, 24 h) increased significantly ABCB1 and GR mRNA expression levels by around 8- and 3·5-fold, respectively (P<10^?6). ABCB1 induction by DEX in CCRF-CEM cells suggests a potential risk of interaction in lymphocytes when associating DEX to ABCB1 substrates in antiretroviral multitherapies in vivo.     

Role of copper transporters in platinum resistance

Platinum (Pt)-based antitumor agents are effective in the treatment of many solid malignancies. However, their efficacy is limited by toxicity and drug resistance. Reduced intracellular Pt accumulation has been consistently shown to correlate with resistance in tumors. Proteins involved in copper homeostasis have been identified as Pt transporters. In particular, copper transporter receptor 1 (CTR1), the major copper influx transporter, has been shown to play a significant role in Pt resistance. Clinical studies demonstrated that expression of CTR1 correlated with intratumoral Pt concentration and outcomes following Pt-based therapy. Other CTRs such as CTR2, ATP7A and ATP7B, may also play a role in Pt resistance. Recent clinical studies attempting to modulate CTR1 to overcome Pt resistance may provide novel strategies. This review discusses the role of CTR1 as a potential predictive biomarker of Pt sensitivity and a therapeutic target for overcoming Pt resistance.