Current Drug Metabolism - Volume 12, Issue 8, 2011

In designing new drugs to reach their biological target, it should be consider the synergistic role of the major systems that regulate drug permeability: P-glycoprotein (P-gp) and CYP450 metabolic enzymes. P-gp, belonging to the ABC transports family, is an efflux pump that extrudes out of the cellular barriers drugs and xenobiotics that back into the blood stream. CYP450 enzymes are a family of monooxygenases that catalyze the biotransformation of a wide variety of endogenous and exogenous compounds to give specific metabolites of the parent drug. These two mechanisms, operating in a coordinated manner, limit intracellular accumulation of xenobiotics and regulate the bioavailability, tissue distributions, and pharmacodynamic effects of drugs. Drug distribution into the Central Nervous System (CNS) is modulated by the blood-brain barrier (BBB), where, the function and expression of CYP450 enzymes and P-gp have a critical pharmacokinetic and pharmacodynamic roles. Recently, in isolated human brain microvessels, CYP1B1 and CYP2U1, have been identified as the main CYP450 isoforms present at the BBB and other studies revealed that P-gp is more present. Recent studies related to the expression, regulation and function of CYP450 enzymes and P-gp in rodent and human BBB, have been reported in the paper of Decleves X. et al. In this paper is focused also a possible interplay between some CYP450 enzymes with some ABC transporters occurring in the BBB, which makes BBB a key element determining brain concentrations of centrally acting drugs. Changes in ABC transporter expression and function are thought to be implicated in various diseases, such as cancer, epilepsy, Alzheimer's and Parkinson's diseases. The overexpression of CYP450 metabolic enzymes and P-gp at BBB level, causes drug resistance in neurological diseases such as epilepsy. In the paper of Ghosh C. et al., the role of P-gp and CYP450 enzymes in brain pathologies and the pathophysiological evolution of the drug resistant phenotype were discussed. Moreover, the experimental approaches leading to significant studies on the role of CYP450 enzymes and P-gp in the drug resistant human epileptic brain was reported. P-gp overexpression , induced by well-known chemotherapeutics agents is a major cause of multidrug resistance (MDR) of chemotherapeutic treatment in tumour cells and tissues. It is known that there is an overlap in substrate and inhibitor specificity for P-gp and CYP3A4 enzymes, so that chemotherapeutic agents also undergo metabolic transformations by various CYP450 isoforms. In the paper of Azzariti et al., data regarding the ability of some chemotherapeutics in modulating both P-gp and CYP450 enzymes are reported; in particular it is clarified whether there is a relationship between the simultaneous modulation of CYP450 and P-gp and the onset of drug resistance in tumours. A major strategy to revert MDR is the co-administration of a chemotherapeutic agent with a P-gp inhibitor. A P-gp inhibitor can concomitantly modulate CYP450 enzymes modifying pharmacokinetic properties of the administered anticancer drugs and its metabolism, causing unfavourable side effects. The paper of Darby R. et al., discusses some of the P-gp inhibitors designed and employed to date and the possible future strategies that could be implemented to achieve its design. The availability of a non-invasive imaging method which allows for measuring P-gp function or expression in vivo facilitates the identification of those patients that would benefit from treatment with P-gp modulating agents. In the article of Mairinger S. et al., the currently available P-gp radiotracers for positron emission tomography (PET) and single-photon emission computed tomography (SPECT) are described, focusing the strengths and limitations of individual probes and their potential clinical applications. This review highlights the synergistic role of P-gp and CYP450 enzymes in drug distribution, so that the simultaneously study of P-gp transport and CYP450 metabolism in drug design, for MDR and/or for neurodegenerative disorders, is fundamental. The paper of Inglese C. et al., suggests the use of everted gut sac assay, an ex vivo method that, firstly, gives simultaneous informations on absorption and metabolism; secondly, the results obtained by this assay display an high prediction degree for in vivo experiments. Mudra D. et al., describes several in vitro and in situ models to study metabolism and permeability, including transfected cell lines, isolated tissues and perfused organs. Moreover, in this paper, computational models including physiologically-based pharmacokinetic models that demonstrate the synergistic interplay between P-gp and CYP450 enzymes are reported.

P-glycoprotein (P-gp) is involved in MDR and in neurodegenerative disorders such as Parkinson disease (PD), Alzheimer disease (AD) and epilepsy. Cytochrome P450 enzymes (CYP450s) catalyze the metabolism of a wide variety of endogenous and exogenous compounds including xenobiotics, drugs, environmental toxins, steroids, and fatty acids. P-gp substrates, inhibitors and inducers should be designed and developed studying interacting mechanism with both P-gp an CYP450 enzymes before they could be employed in MDR and/or in neurodegenerative disorders. Here, the ex vivo rat everted gut sac assay has been proposed as an immediate approach to simultaneously study metabolism and transport of drugs. Elacridar, verapamil and cyclosporine A (CsA), P-gp inhibitor, substrate and modulator respectively, have been tested to validate this ex vivo approach. The new model have been used yet to develop our ligands MC18, MC266 and MC80, both as potential drugs for MDR and radiotracers for diagnosis of neurodegenerative disorders. Herein, a comparative evaluation of transport and metabolic results, by using in vitro, ex vivo and in vivo assays, is reported.

The relationship between CYP450 and P-gp occurs at different levels. It is known that certain substrates of P-gp undergo metabolic transformations by various CYP450 isoforms; in addition some of them demonstrated to be activators of both P-gp and CYP450. The majority of such compounds are well-known chemotherapeutics, therefore the purpose of this review is to clarify whether there is a relationship between the simultaneous modulation of CYP450 and P-gp and the onset of drug resistance in tumors treatment. Here, we discuss the biological aspects of the topic in relation to the various tissues distribution of CYP450 and P-gp, the recent findings regarding the ability of some chemotherapeutics in modulating both P-gp and CYP450, whether this modulation is ultimately responsible for the onset of drug resistance in cancer treatment and the promising role of gene polymorphisms in determining the interindividual variability in drug responses in clinical practice.

The multidrug resistant phenotype of cancer cells can often result from the over-production of a number of ATP binding cassette (ABC) transporters, including P-glycoprotein (P-gp). These multidrug efflux transporters expel administered anti-cancer drugs from the cancer cell, preventing sufficient intracellular drug accumulation and ultimately, drug efficacy. The co-administration of compounds that can impede the efflux of chemotherapeutic agents by these ABC transporters can concomitantly modulate various cytochrome P450 (CYP450) enzymes, consequently impacting upon anti-cancer drug metabolism. This can further result in unfavourable drug-drug interactions and altered pharmacokinetic properties of the administered anti-cancer drugs with knock-on adverse cytotoxic side effects. This review will discuss some of the P-gp inhibitors designed and employed to date, as well as expressing our views of the shortcomings of their design strategy. We present a medicinal chemist's wish list for the paradigmatic P-gp inhibitor molecule and examine the possible future strategies that could be implemented to achieve its design.

The recent identification of drug-metabolizing enzymes cytochrome P450 (CYP) in the human blood-brain barrier (BBB) raises the question of whether these enzymes act in concert with ATP-binding cassette (ABC) transporters to limit the brain distributions of drugs. We recently demonstrated several CYP genes in freshly isolated human brain microvessels; the main isoforms expressed were CYP1B1 and CYP2U1. Many studies using different experimental approaches have revealed that P-glycoprotein (P-gp, ABCB1), breast cancer resistance protein (BCRP, ABCG2) and the multidrug resistance-associated protein 4 (MRP4, ABCC4) are the main ABC transporters in the human BBB. The first part of this review covers recent studies on the expression, regulation and function of CYP450 and ABC transporters in the rodent and human BBBs. The second part focuses on the possible interplay between some CYPs and certain ABC transporters at the BBB, which makes it a determining element of brain drug concentrations and thus of the effects of centrally acting drugs.

Drug penetration into the central nervous system (CNS) is controlled by the blood-brain barrier (BBB). Even though a number of strategies to circumvent the BBB and to improve drug access have been developed, drug resistance in CNS diseases remains an unmet clinical problem. We here review the mechanisms by which a healthy or pathological BBB influences drug distribution in the brain, with emphasis on the role of P450 metabolic enzymes and multi-drug transporter (MDT) proteins. In addition to the classic hepatic and gut biotransformation pathways, CNS expression of P450 enzymes may bear pharmacokinetic and pharmacodynamic significance exerting a metabolic activity and transforming parent drugs into specific products. We propose these mechanisms to play a major role in CNS drug resistant pathologies including refractory forms of epilepsy. Changes in the cerebrovascular hemodynamic conditions can affect expression of P450 enzymes and MDT proteins. This should be taken into account when developing in vitro experimental approaches to reproduce the physiological or pathological properties of the BBB. Finally, a link between P450 and MDT expression in the diseased brain and cell survival is discussed.

The bioavailability, fraction of dose that reaches systemic circulation, of orally administered drugs is often limited by both physical barriers of the intestine (e.g., unstirred-water and mucosal layers, epithelial tight junctions) as well as biochemical barriers such as cytochromes P450 (CYP) and P-glycoprotein (P-gp). Highly expressed in intestine and liver, CYP and P-gp can limit the systemicavailability of parent-drug by metabolism and efflux, respectively, by means of similarly large and flexible active sites that accommodate a variety of structurally-diverse, lipophilic molecules over a wide-range of molecular weights. Consequently, many molecules that are substrates for CYP3A4 also demonstrate affinity for P-gp and numerous studies have reported that for these dual-substrates, CYP3A4 and P-gp afford an interplay that affects bioavailability and clearance in a manner that is non-linear. Several in vitro and in situ models of metabolism and permeability, including transfected cell lines, isolated tissues and perfused organs as well as computational models including physiologically-based pharmacokinetic models of such co-expressing systems have demonstrated this phenomenon of CYP3A/Pgp interplay. Furthermore, recent availability of ligand bound X-ray co-crystal structures of the CYP3A4 and P-gp binding sites coupled with computational docking techniques and other validated in silico models, provide medicinal chemists with tools to inform structuraldesign modifications that can modify the interaction with one or both proteins. This article provides a review of relevant in silico, in vitro, ex vivo and in situ models that allow for investigation of the extent to which clearance or bioavailability can be affected by CYP/P-gp interplay.

Adenosine triphosphate-binding cassette (ABC) transporters, such as P-glycoprotein (Pgp, ABCB1), breast cancer resistance protein (BCRP, ABCG2) and multidrug resistance-associated proteins (MRPs) are expressed in high concentrations at various physiological barriers (e.g. blood-brain barrier, blood-testis barrier, blood-tumor barrier), where they impede the tissue accumulation of various drugs by active efflux transport. Changes in ABC transporter expression and function are thought to be implicated in various diseases, such as cancer, epilepsy, Alzheimer's and Parkinson's disease. The availability of a non-invasive imaging method which allows for measuring ABC transporter function or expression in vivo would be of great clinical use in that it could facilitate the identification of those patients that would benefit from treatment with ABC transporter modulating drugs. To date three different kinds of imaging probes have been described to measure ABC transporters in vivo: i) radiolabelled transporter substrates ii) radiolabelled transporter inhibitors and iii) radiolabelled prodrugs which are enzymatically converted into transporter substrates in the organ of interest (e.g. brain). The design of new imaging probes to visualize efflux transporters is inter alia complicated by the overlapping substrate recognition pattern of different ABC transporter types. The present article will describe currently available ABC transporter radiotracers for positron emission tomography (PET) and single-photon emission computed tomography (SPECT) and critically discuss strengths and limitations of individual probes and their potential clinical applications.

The effectiveness of many anticancer agents is dependent on their disposition to the intracellular space of cancerous tissue. Accumulation of anticancer drugs at their sites of action can be altered by both uptake and efflux transport proteins, however the majority of research on the disposition of anticancer drugs has focused on drug efflux transporters and their ability to confer multidrug resistance. Here we review the roles of uptake transporters of the SLC22A and SLCO families in the context of cancer therapy. The many first-line anticancer drugs that are substrates of organic cation transporters (OCTs) organic cation/carnitine transporters (OCTNs) and organic anion- transporting polypeptides (OATPs) are summarized. In addition, where data is available a comparison of the localization of drug uptake transporters in healthy and cancerous tissues is provided. Expression of drug uptake transporters increases the sensitivity of cancer cell lines to anticancer substrates. Furthermore, early observational studies have suggested a causal link between drug uptake transporter expression and positive outcome in some cancers. Quantification of drug transporters by mass spectrometry will provide an essential technique for generation of expression data during future observational clinical studies. Screening of drug uptake transporter expression in primary tumors may help differentiate between susceptible and resistant cancers prior to therapy.