Current Topics in Medicinal Chemistry - Volume 10, Issue 17, 2010

ABC transporters play a crucial role in several physiological barriers such as the blood-brain barrier (BBB), bloodcerebrospinal fluid (B-CSF) barrier, and blood-testis barrier (BTB) modulating the absorption and excretion of xenobiotics across these barriers. In the brain, these transporters are localized at the luminal membrane of endothelial cells of blood capillaries where they actively modulate the permeation of xenobiotics. Transporter over-expression has been observed in several tumors causing multidrug resistance (MDR) to treatment with chemotherapeutics. Moreover, resistance to central nervous system (CNS) drugs, such as antidepressant, antiepileptic and anti-HIV medicine, may also be related to overexpression of ABC transporters. Recently, it has been reported that alterations in ABC expression and function are related to the etiology and pathogenesis of neurologic disorders, such as Alzheimer's disease (AD) and Parkinson's (PD) disease. Moreover, a recent study reported that ABC expression and function are strongly decreased during the neuro-inflammation process in Multiple Sclerosis (MS). Positron Emission Tomography (PET) is of great importance for non-invasive studies of physiological and pathological processes in the brain, contributing to a understanding of the etiology of neurological diseases. The tracers used for PET are small molecules belonging to several classes of compounds developed in the medicinal chemistry field. Several in vitro and ex vivo assays can be used to characterize their interaction mechanism with ABC transporters. ABC substrates labeled with positron emitters are tracers for monitoring the activity of these transporters whereas labeled ABC inhibitors may permit to estimate their expression. The first section of this issue is focused on the contribution of medicinal chemistry in developing potent inhibitors, substrates and modulators of various ABC transporters [1-3]. An overview of natural compounds able to modulate such transporters is also provided [4], and innovative strategies for the design of novel radiotracers are discussed [5]. A second section focuses on PET imaging to assess the function of ABC transporters in neurodegenerative diseases [6] in epilepsy [7] and in chemoresistant tumors [8]. Other manuscripts highlight how PET imaging can be used to elucidate mechanisms underlying drug disposition [9,10] and how cerebral tracer uptake can be quantified with compartment models [11]. A final contribution describes SPECT studies of ABC transporter function in oncology [12]. As guest editors, we would like to thank all authors for their comprehensive contributions and the prompt submission of their manuscripts. Moreover, we thank Prof. Allen Reitz, Editor-in Chief of Current Topics in Medicinal Chemistry, for his kind invitation and support.

In recent years, several PET tracers for monitoring the activity and expression of P-gp at the BBB have been tested. P-gp substrates such as [11C]verapamil and [11C]loperamide can be employed to visualize P-gp activity, but they display a moderate baseline uptake in the brain and formation of radiolabeled metabolites which hamper the interpretation of PET data. P-gp inhibitors such as [11C]elacridar, [11C]laniquidar and [11C]tariquidar have been tested to investigate Pgp expression and the results need further investigation. Recently, we developed MC18, MC266 and MC80, that have been characterized as an inhibitor, substrate and inducer of P-gp both by in vitro assays and in the everted gut sac method. These compounds have been radiolabelled with 11C and been evaluated in vivo. In the present review, we compare the outcome of biological in vitro assays and the corresponding in vivo PET data for the P-gp inhibitors [11C]MC18 and [11C]elacridar, the P-gp substrates [11C]MC266 and [11C]verapamil, the P-gp inducer [11C]MC80 and the P-gp modulator cyclosporin A. Since a satisfactory overlap was found comparing in vivo results and the corresponding in vitro findings, the proposed biological in vitro assays could be predictive for the in vivo PET data of novel radiotracers. PET tracers could be employed for various purposes: radiolabeled P-gp inhibitors to monitor decreased expression of P-gp at the BBB in neurodegenerative disorders such as Alzheimer's and Parkinson's disease; and radiolabeled P-gp substrates with a high baseline uptake to monitor increased expression of P-gp in epileptic foci.

Multidrug resistance (MDR) is a kind of acquired resistance of microorganisms and cancer cells to chemotherapeutic drugs that are characterized by different chemical structure and different mechanism of action. Classic MDR is due to a lower intracellular concentration of cytotoxic drugs that is associated with accelerated efflux of the chemotherapeutic drugs and is the consequence of the over expression of transporter proteins that act as extrusion pumps. Pglycoprotein (P-gp/ABCB1) is the most important and studied member of such proteins belonging to the ATP Binding Cassette (ABC) superfamily of transporters that use ATP as energy source. Inhibition of the functions of P-gp and other ABC proteins could represent a way to circumvent appearance of MDR in cancer cells and the most classical pharmacological strategy is the administration of agents able to modulate the P-gp function. On the basis of the known characteristics of the recognition site of P-gp, we have designed a new class of P-gp-mediated MDR reverters. These compounds are flexible molecules carrying a basic nitrogen atom flanked, at properly modulated distance, by two aromatic moieties; most of them possess MDR inhibitory activity on anthracycline-resistant erytroleukemia K562 cells. By applying the frozen analog approach to that series of very flexible MDR reverters, we identified a new series of N,N-bis(cyclohexanol)amine aryl esters that show very interesting MDR-reversing properties. Among them, compound 15d, that consistently shows low nanomolar potency and high efficacy in all the tests used, appears as a new pharmacological tool for P-gp studies and a promising lead for the development of potent, efficient and safe MDR reverters.

Human multidrug resistance protein (MRP/ABCC) family contains 9 members (MRP1-9) which transport a structurally diverse array of anticancer and antimicrobial drugs and several important endogenous substances including prostaglandins (PGs) and leukotrienes (LTs) with different substrate specificity. MRP1-5 can collectively confer resistance to natural product anticancer drugs and their conjugated metabolites, platinum compounds, folate antimetabolites, nucleoside and nucleotide analogs, and alkylating agents. MRP1-3 are often associated with tumor resistance which is often caused by an increased efflux and decreased intracellular accumulation of natural product anticancer drugs and other anticancer agents. Both PGE1 and PGE2 are known high-affinity substrates of MRP4, but not MRP1, MRP2, MRP3 and MRP5. LTC4 is a substrate of MRP1, MRP2, MRP3, and MRP6-8. MRP2 is also able to transport LTD4 and LTE4. Experimental studies in Abcc1-defficient mice have demonstrated a role of MRP1 in inflammation process in vivo. Abcc3- deficcient mice have normal bile salt transport, however, they have decreased blood bilirubin glucuronide levels. Abcc6- deficient mice show remarkable mineralization of the connective tissues, including skin, arterial blood vessels, and retina. Most MRP/ABCC transporters are subject to inhibition by a variety of compounds. Drug targeting of these transporters to overcome MRP/ABCC-mediated multidrug resistance may play a role in cancer and infection (e.g. HIV infection) chemotherapy. Some modulators of MRPs have shown reversing effects on MDR phenotype in preliminary clinical studies and some modulators of MRPs may modify the inflammatory process and consequently ameliorate the inflammatory symptoms. A better understanding of the interactions of these modulators with MRPs has important implications in development of novel drugs for treatment of cancer, infection and inflammation.

The multidrug resistance (MDR) proteins that belong to the ATP-binding cassette superfamily such as Pglycoprotein (P-gp) and MRP1, are present in a majority of human tumors and constitute an important cause of therapeutic failure. Selective inhibitors of the MDR-efflux proteins may improve the effectiveness of cancer chemotherapy. Their mechanism of action was believed to be a competition between resistance modifiers and drugs for the same binding site of P-gp. In our previous work we studied modulation of MDR in cancer cells expressing P-gp or MRP1 by selected carotenoids, flavonoids and extracts from medically important Chinese plants. Capsanthin and capsorubin, carotenoids isolated from paprika, were identified as potent P-gp inhibitors, while lycopene, lutein, antheraxanthin and violaxanthin induced moderate effects. Among flavonoids, effective modulators were rotenone, chrysin, phloretin and sakuranetin. Some chloroform extracts of Chinese herbs were also found to inhibit MDR efflux pumps. The effects of the modulators on Pgp activity were studied by measuring rhodamine 123 uptake in several cancer cells such as the human MDR1 genetransfected mouse lymphoma cells (L1210) and human breast cancer cells MDA-MB-231 expressing the MRP1 pump (HTB26). Additionally, the ability to alter biophysical properties of lipid bilayers by selected carotenoids was studied by differential scanning calorimetry. The antiproliferative effects as well as the MDR reversal activity of the studied compounds, applied in combination with anticancer drugs, were also discussed.

Design of inhibitors of P-glycoprotein still represents a challenging task for medicinal chemists. The polyspecificity of the transporter combined with the limited structural information renders rational drug design approaches rather ineffective. Within this article we will exemplify how recent insights into structure and mechanism of Pglycoprotein may aid in design of potent inhibitors. P>

P-glycoprotein (P-gp) at the blood-brain barrier (BBB) functions as an active efflux pump by extruding a wide range of substrates from the brain. This is important for maintaining loco-regional homeostasis and for protecting the brain against blood-borne toxic substances. Altered P-gp function seems to be involved in the pathophysiology of neurodegenerative disease and various neurological and psychiatric disorders. Positron emission tomography (PET) with the radiotracer 11C-verapamil (VPM-PET) is a validated technique allowing measurement of P-gp function at the human BBB. In this review, we highlight changes of P-gp function, as measured with VPM-PET, in aging and in the pathogenesis and progression of neurodegenerative disease, as well as their role in depressive disorders.

The issue of pharmacoresistance in epilepsy has received considerable attention in recent years, and a number of plausible hypotheses have been proposed. Of these, the so-called transporter hypothesis is the most extensively researched and documented. This hypothesis assumes that refractory epilepsy is associated with a localised over-expression of drug transporter proteins such as P-glycoprotein (Pgp) in the region of the epileptic focus, which actively extrudes antiepileptic drugs (AEDs) from their intended site of action. However, although this hypothesis has biological plausibility, there is no clinical evidence to support the assertion that AEDs are sufficiently strong substrates for transportermediated extrusion from the brain. The use of modern brain imaging techniques to determine Pgp function in patients with refractory epilepsy has started only recently, and may ultimately determine whether increased expression and function of Pgp or other efflux transporters are involved in AED resistance.

The failure of solid tumors to respond to chemotherapy is a complicated and clinically frustrating issue. The ability to predict which tumors will respond to treatment could reduce the human and monetary costs of cancer therapy by allowing pro-active selection of a chemotherapeutic to which the tumor does not express resistance. PET/CT imaging with a radiolabeled form of paclitaxel, F-18 fluoropaclitaxel (FPAC), may be able to predict the uptake of paclitaxel in solid tumors, and as a substrate of P-glycoprotein, it may also predict which tumors exhibit multidrug resistance (MDR), a phenotype in which tumors fail to respond to a wide variety of chemically unrelated chemotherapeutic agents. This article reviews the synthetic, preclinical and early human data obtained during the development phase of this promising new radiopharmaceutical.

This paper discusses the basic principles of drug/P-glycoprotein (P-gp) interaction, focusing on the methodology and design of positron emission tomography (PET) studies investigating P-gp function. The requirements of a good PET P-gp radiotracer are also evaluated. (R)-[11C]verapamil is used as an example, as this drug is the most common tracer for P-gp studies, but [11C]loperamide, [11C]desmethyl-loperamide and other compounds are also mentioned. The article also discusses the various study designs that can be used for PET drug disposition studies, such as administration of the inhibitor before or after the radiolabeled drug (tracer) and the use of bolus injections or infusions. Concepts such as the unbound partition coefficient (Kp,uu) and the volume of distribution of unbound drug in brain (Vu,brain), which are not easily measured directly with PET, can be used to describe the impact of protein binding and non-specific binding on drug distribution in brain tissue. It is concluded that new imaging probes will be required if the role of PET in studies of the interactions of drugs with efflux transporters is to expand.

Multidrug resistance-associated protein 1 (MRP1) functions as a primary active transporter utilizing energy from ATP hydrolysis. In the central nervous system (CNS), MRP1 plays an important role in limiting the permeation of xenobiotic and endogenous substrates across the blood-brain and blood-cerebrospinal fluid barriers, and across brain parenchymal cells. While MRP1 contributes to minimizing the neurotoxic effects of drugs, it may also restrict the distribution of drugs for the treatment of CNS diseases. Moreover, neurodegenerative disease may be associated with abnormal expression of efflux transporters in the brain. Noninvasive measurement of MRP1 function will therefore be useful for directly evaluating the effect of modulators on enhancing the penetration of drugs into the brain and for examining the pathophysiological role of MRP1 in the brain. Positron emission tomography (PET) is a powerful molecular imaging technique. While several PET probes have been proposed for imaging function of the efflux transporter P-glycoprotein, few reports discuss the probes for imaging MRP1 function in the brain. Ideally, brain radioactivity should consist of a single radioactive compound that is selectively transported by the efflux transporter of interest, without other efflux routes. However, most PET probes for MRP1 or P-glycoprotein are eliminated by both a transporter and simple diffusion, resulting in inaccurate measurement of pump function. This review addresses a new strategy to avoid this problem, and suggests the design of a PET probe based on this strategy, particularly for MRP1 imaging. Several published reports on imaging MRP1 function with PET are also discussed.

P-glycoprotein (P-gp) is a drug efflux transporter with broad substrate specificity localized in the blood-brain barrier and in several peripheral organs. In order to understand the role of P-gp in physiological and patho-physiological conditions, several carbon-11 labelled P-gp tracers have been developed and validated. This review provides an overview of the spectrum of radiopharmaceuticals that is available for this purpose. A short overview of the physiology of the blood-brain barrier in health and disease is also provided. Tracer kinetic modelling for quantitative analysis of P-gp function and expression is highlighted, and the advantages and disadvantages of the various tracers are discussed.

Multidrug resistance (MDR) mediated by overexpression of MDR1 (ABCB1) P-glycoprotein (Pgp) is one of the best characterized transporter-mediated barriers to successful chemotherapy in cancer patients and is also a rapidly emerging target in the progression of neurodegenerative disorders, such as Alzheimer's and Parkinson's diseases. Therefore, molecular imaging probes capable of imaging noninvasively Pgp and closely related transporter activities in tissues as well as tumors would be expected to contribute to personalized medicine. Interrogation of Pgp-mediated transport activity in vivo via noninvasive SPECT imaging could be beneficial for stratification of patient populations likely to benefit from a given therapeutic treatment, assist in the management of chemotherapy and aid the study of neurodegenerative diseases.