Pharmacology Should Be at the Centre of All Preclinical and Clinical Studies on New.pdf Psychoactive

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published online 27 March 2014J PsychopharmacolA Richard Green and David J Nutt

substances (recreational drugs)Pharmacology should be at the centre of all preclinical and clinical studies on new psychoactive

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Introduction A good understanding of the actions of any drug, both desired and adverse, requires appropriate translation. That is, the availability of knowledge that allows the move from basic pharmacological science to an understanding of the effects of the drug in humans. This understanding also requires reverse translation, which is taking clinically acquired knowledge of the drug and applying it to preclinical studies to further elucidate mechanisms of action, both those involved in the therapeutic actions of the compound and those underlying its adverse or toxic effects (Green and Aronson, 2012). This is the basic underlying principle for all phar-macology research. While such an approach is fairly straightfor-ward in the development and understanding of therapeutic drugs it is almost impossible in the case of recreational drugs for a vari-ety of reasons, both scientific and ethical. However, failure to adhere to the more basic rules of good translational pharmacology continues to undermine the quality of some research and confound interpretation of the data obtained, despite earlier publi-cations outlining weaknesses that can be avoided (Gabrielsson and Green, 2009). The fact that the United Nations Office on Drugs and Crime (UNODC) recent World Drug Report (2013) states that there has been an alarming increase in the use of new psychoactive substances (NPS) makes it crucial that better research is conducted on recreational drugs in order that we can better understand their pharmacology and toxicology.

This commentary makes suggestions as to how such improve-ments can be made to better investigate recreational drugs in future. To do so it employs examples from previous studies and in particular it uses investigations on 3,4-methylenedioxymeth-amphetamine (MDMA) and 4-methylmethcathinone (mephed-rone). MDMA is arguably the most investigated recreational drug in the last 25 years (Green et al., 2003) and mephedrone, although appearing recently, has been extensively studied over the last three years both preclinically (Green et al., 2014) and also clinically in recreational users (Dargan et al., 2010, 2011; Schifano et al., 2011; Wood and Dargan, 2012; Zawilska and Wojcieszak, 2013).

Pharmacology should be at the centre of all preclinical and clinical studies on new psychoactive substances (recreational drugs)

A Richard Green1 and David J Nutt2

AbstractDespite the publication of a substantial body of preclinical and clinical information on recent recreational drugs such as 3,4-methylenedioxymethamphetamine (MDMA, ecstasy) and cathinone compounds such as mephedrone there remains a disturbing lack of consensus as to how dangerous these compounds are to the health of the individual and to society in general. This perspective proposes that use of good pharmacological practice should be mandatory in all preclinical and clinical studies. Its use will assist both translation and reverse translation of information produced in animals and clinical subjects. We propose several basic rules to be followed in all future studies. Preclinical studies should employ pharmacokinetic-pharmacodynamic integration thereby exposing animals to known or calculable drug concentrations. This will provide results relevant to pharmacology rather than toxicology and, crucially, data relevant to human drug use. Full experimental detail should be routinely provided, to allow comparison with other similar work. In clinical studies evidence should be provided that the drug under investigation has been ingested by the subjects being examined, and details given of all other drugs being ingested. Drug-drug interactions are an unavoidable confound but studies of a size that allows reliable statistical evaluation and preferably allows sub-group analysis, particularly by using meta-analysis, should help with this problem. This may require greater collaboration between investigative groups, as routinely occurs during pharmaceutical clinical trials. Other proposals include greater integration of preclinical and clinical scientists in both preclinical and clinical studies and changes in the law regarding Good Manufacturing Process (GMP) sourcing of drug for human studies.

KeywordsEcstasy, 3,4-methylenedioxymethamphetamine, mephedrone, pharmacokinetics, integrative pharmacology, quantitative pharmacology, recreational drugs, new psychoactive substances

1 School of Life Sciences, Queens Medical Centre, University of Nottingham, Nottingham, UK

2 Division of Neurosciences and Mental Health, Imperial College London, London, UK

Corresponding author:A Richard Green, School of Life Sciences, Queens Medical Centre, University of Nottingham, Nottingham NG7 2UH, UK. Email: [email protected]

528593 JOP0010.1177/0269881114528593Journal of PsychopharmacologyGreen and Nuttresearch-article2014

Perspective

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2 Journal of Psychopharmacology

Quantitative pharmacologyAn understanding of both the acute and long term effects of any drug requires knowledge of both its pharmacodynamics and phar-macokinetics. Pharmacodynamics is the study of the time course of biological effects of the drug and the relationship of the drug exposure to its effects, both wanted and adverse (basically what the drug does to the body) while pharmacokinetics investigates the time course of the drug and its metabolism in the body (basi-cally what the body does to the drug). Ideally these two scientific techniques should be pursued together, what is known as pharma-cokinetic-pharmacodynamic integration (PKPD) or quantitative pharmacology (Gabrielsson and Green, 2009). Quantitative phar-macology is now the standard approach in the pharmaceutical industry during drug discovery and development, and its use is mandatory as far as the regulatory bodies are concerned. Consequently no new chemical entity is now developed without a clear understanding of the relationship between exposure, which is not merely dose but the concentration of active drug in the body, and effect. Exposure is influenced by absorption, protein binding, metabolism and excretion. Along with these preclinical studies, investigations are made of the toxicity and possible interaction of the drug with other pharmaceutical agents.

Similarly, clinical investigation of any novel compound has to involve knowledge of the pharmacokinetics of the drug, which can be very different in humans and laboratory animals, with dose-ranging studies to determine both the minimum effective dose and those producing adverse effects. Investigation must also be made into possible interactions of the compound with other drugs. Even when all these studies have been performed and the drug is marketed, further information on possible adverse effects and drug interactions continues to be obtained with the so-called yellow card scheme, which is now an online reporting system used by physicians to report adverse effects (https://yellowcard.mhra.gov.uk).

Obviously, virtually none of these approaches have been applied in the preclinical studies of recreational drugs. Recently a plea was made for the routine use of integrative pharmacology in all preclinical pharmacology research since its use adds substan-tial value to every study and ignoring it can lead to misleading results (Gabrielsson and Green, 2009). Its use allows better trans-lation of preclinical data to clinical investigations and also better reverse-translation where clinical information can initiate further necessary preclinical studies. The suggestion that quantitative pharmacology be used in future experimental studies on MDMA was made several years ago (De la Torre and Farr, 2004; De la Torre et al. 2009; Green et al. 2009). The subsequent detailed study on the pharmacokinetics of MDMA in rats (Baumann et al., 2009) helped highlight the weaknesses of many prior preclinical investigations based on this drug in terms of their translational value. In particular it provided evidence that the pharmacokinetics of MDMA in rats and primates is fundamentally different from the pharmacokinetics of the drug in humans, as was recently reviewed (Green et al., 2012). Since the plasma half-life of the drug in rats is only one-tenth that in humans the acute adverse events in rats may be minimal compared to those in humans, and this includes both body temperature and endocrine changes. Conversely, the rapid metabolism of the drug in rats to form neurotoxic metabo-lites may result in more severe long-term brain effects in that spe-cies than those that may occur in humans.

Furthermore, the plot of dose versus plasma drug concentra-tion has a steeper slope in humans than in rats. Doubling the dose from 1 to 2 mg/kg yields a four-fold increase in plasma concen-tration, because MDMA not only is metabolised by the cytochrome P450 enzyme CYP2D6 but also inhibits it (Tucker et al., 1994).This results in the non-linear kinetics with rapid enzyme inhibition by the parent drug. In contrast there is an approximately linear relationship between dose and plasma con-centration of the drug in rats (Green et al., 2012). Consequently these differences preclude any simple projections of dosing between rats and humans and demand proper evaluation of drug exposure.

Similar detailed clinical pharmacokinetic information is una-vailable with the new recreational drugs such as mephedrone but, ideally, it should be obtained before firm conclusions are made on the translation of preclinical studies to human users. However obtaining such data may prove to be near impossible in the UK given the current recommendation that no research drug can be given to humans unless it has been prepared to Good Manufacturing Process (GMP) standards, the cost of which, being in the order of 70,000, proves impossible for most poten-tial investigators to meet (Nutt et al., 2013). The GMP require-ment is an unintended result of the poorly thought-through European clinical trials directive that has been unnecessarily applied to experimental human studies, in addition to its intended target of clinical trials of new therapeutic agents.

Subject selection, experimental conditions and reporting

Preclinical studies

Although rats are the predominant species in preclinical studies, various points must be considered. First is the fact that different strains of rats can display significant differences in both their acute and long-term responses, as has been observed in studies on both MDMA and mephedrone. For example, Wright et al. (2012) found that mephedrone induced an acute hypothermic response in Wistar rats, but that even the highest dose examined (10 mg/kg) produced little body temperature change in Sprague-Dawley rats even though both strains responded to the drug with a similar hyperactivity response. There is also substantial evi-dence that the Dark Agouti strain is much more sensitive than other strains to the neurotoxic effects of MDMA in that long term substantial loss of 5-hydroxytryptamine (5-HT) in the brain can be induced in Dark Agouti rats using doses having no effect in other strains (OShea et al. 1998). Therefore, investigations employing a single strain may be misleading in terms of the gen-erality of the effect observed, and dose-ranging studies in more than one strain are to be encouraged.

Use of a single species can also give a misleading impression of the effect the drug might have in other species including, pre-sumably, humans. MDMA has very different pharmacological and toxicological effects in mice compared to all other species that have been examined. Mice show a smaller hyperthermic effect following a single dose of MDMA than that seen in rats (Docherty and Green, 2010) and, crucially, the drug induces neu-rotoxic damage to dopamine neurones in the mouse brain rather than 5-HT neurones as observed in rat, monkey and guinea-pig


Green and Nutt 3

(Green et al., 2003). Whether this difference is due to pharma-cokinetic and metabolic differences in the way the drug is han-dled in mice versus other species is unknown.

An additional reason for examining several species comes from the fact that the plasma protein-binding of a drug can vary enormously between species (see Gabrielsson and Green, 2009) and this can lead to a near 20-fold difference in the unbound, and therefore active, drug concentration (Gabrielsson et al., 2010). Therefore, in the absence of clear information on the kinetics of the drug in man and laboratory animals, it is wise not to rely on information from only one species in projecting to possible effects in humans.

Almost all investigators use only young male animals in their studies. This may simplify the study in removing complications of the oestrus cycle, but scarcely reflects the reality of recreational drug users and there are several reports that male and female sub-jects have different psychological and physiological sensitivities to the effects of MDMA (Allott and Redman, 2007; Verheyden et al., 2002). However, the use of young animals is probably justi-fied given the demographics of recreational drug use.

The housing conditions in which the animals are investigated must also be considered. Recreational drugs such as MDMA and mephedrone are usually taken in dance club conditions. This means crowded, often warm, room conditions. It therefore makes sense for animals to be similarly housed during experiments. This is not always possible when specific behavioural studies are being undertaken and the possible implications of this on transla-tion should be acknowledged. For example, the rectal tempera-ture of singly housed rats generally decreases following a dose of MDMA (for example see Shortall et al., 2013a), but increases in grouped conditions and increases further when the ambient tem-perature is raised from around 21C to 30C (Docherty and Green, 2010; Green et al., 2005). This mimics the situation with human drug users where the drug can produce a dangerously raised body temperature in crowded hot dance club conditions, but induces a modest body temperature increase when given in controlled clin-ical situations (Farr et al., 2007). Interestingly, mephedrone administration to rats produces hypothermia in both singly housed and grouped animals and even placing them in a hot room (30C) merely abolishes the hypothermic response rather than inducing hyperthermia (Miller et al., 2013; Shortall et al., 2013a). This probably reflects the situation in recreational users as cold fingers have been reported by mephedrone users, but hyperther-mia, while it can occur, does not seem to be a generally reported adverse event (Schifano et al., 2011; Winstock et al., 2011).

A further point is that full reporting of the experimental details should always be available in any published article. Investigators often fail to give full details of methodology, often because they feel the method is so well known that some information is self evident and because journals are sometimes very restrictive in space allowed for methods. Nevertheless, such information is vital to evaluate the quality and reproducibility of the results. Often, even simple information is missing such as the blinding of the experimenter to the drug administration and the randomisa-tion of animal subjects (Sena et al., 2007). Statistical quality is also a key item and this and other matters relating to proper reporting of animal experiments have been outlined elsewhere (McGrath et al., 2010).

There is also a related problem regarding reproducibility, because negative preclinical studies (usually those not confirming

an earlier publication) are generally not reported and this leads to publication bias (Sena et al., 2010). This problem involves the investigator (generally scientists do not like publishing negative data), the referees (who see little point in publishing negative results) and journals that do not wish to use up space with an unexciting publication. However reproducibility can only be assessed when it is known whether other investigators have failed to confirm the findings of others. It is notable that it generally requires much more justification to publish a study failing to con-firm the findings of others than is required to publish the original work. It has been suggested elsewhere that journals should have an online repository for negative results (Green et al., 2011). This idea has gained the support of the British Pharmacological Society and the American Society for Pharmacology and Experimental Therapeutics for their joint journal publication (British Pharmacological Society, 2013).

Even when papers are published that fail to confirm the results of others, theoretical explanations are often suggested for the dis-crepant results. These can include strain differences or experi-mental conditions. If appropriate pharmacokinetic measures had been made and full methodological details provided, the reasons for the differing results might be more easily understood.

A recent review on preclinical studies on stroke noted how often there were a disproportionate number of studies replicating the same experimental conditions (similar dose range, experi-mental models and outcome measures). It was proposed that the use of proper collaborative studies with steering committee man-agement (as occurs in clinical trials) to co-ordinate several labo-ratories and thereby obtain full information on the experimental drug and prevent only 'easier' studies being performed would be an effective way to obtain appropriate information for subse-quent analysis of drug effects (Bath et al., 2009a). Such an approach would be valuable in psychopharmacology research by allowing studies of pharmacological, neurochemical, experimen-tal psychological and toxicological effects of the recreational drugs over appropriate dose ranges. No one laboratory could be expected to have such a wide range of skills and data obtained would also be more amenable to meta-analysis.

Clinical studies

The problems of subject recruitment are several and have been detailed by others (Cole and Sumnall, 2003; Curran, 2000). For single drug administration to a subject, double-blind administra-tion is possible; with certain ethical constraints if an illicit drug is being given. However, most studies examine the consequences of recreational drug use and the recruitment of subjects then brings about several methodological problems, as reviewed in detail by Cole and Sumnall (2003). These problems can bias the results and, while they do not need to be discussed again here, they must be considered, stated or eliminated in any publication. There is also the problem of pre-existing morbid differences which can alter interpretation since the psychological trait or pathological abnormality being reported could be the cause of the drug tak-ing rather than the consequence.

What is invariably a problem is the use of multiple drugs (polydrug use) by subjects, either together or over the course of a lifetime: this is a major confound in all studies and is discussed later.



A major concern when looking at studies that have been reported is cohort size. It is clear from perusal of meta-analysis studies that many studies that were re-analysed had investigated rather few subjects. The analysis of Kalechstein et al. (2007) on MDMA and neurocognition cites papers where the total cohort was only 20 subjects and the control and user groups were not even balanced for males and females. Nulsen et al. (2012), in a meta-analysis of short-term and working memory, similarly lists studies with low group numbers. One suspects that rather few of these studies undertook power analyses or other very simple cal-culations before starting. These analyses are widely used in human clinical trials and regulatory authorities see their use as mandatory in order to determine if an appropriate number of sub-jects is being used in order to detect a biologically important effect (if there is one). Only in that way can the experimenters be sure of avoiding a type 1 or type 2 error.

Taylor et al. (2011: 1576) observed that the more recent the study on MDMA the smaller was the effect size and they con-cluded that: perhaps the most parsimonious explanation for the observed association between year of publication and effect size is simply that study quality has increased over time, with a result-ing reduction in bias.

Doses and frequency of ingestion

Preclinical studies

There should be few problems for the preclinical scientist in this section. Most drugs are obtained from regulated chemical or pharmaceutical companies and have unequivocal evidence for purity. However, if the drugs have been supplied from confis-cated recreational supplies (by the police for example), then the purity is unknown and must be ascertained and reported, and any other adulterants removed. Whether the drug is a racemic mix-ture or a single enantiomer must also be stated since the enanti-omers of MDMA do not have the same pharmacology and pharmacokinetic parameters (Fallon et al., 1999). It should be clearly stated as to whether the stated dose is quoted as base or salt (for example, the HCl salt contains approximately 15% less MDMA than the base). While most pharmacologists tend to use and quote doses in terms of mg/kg body weight they should be aware of the limitations of this information in terms of exposure, particularly when relating the relevance of the dose to other spe-cies, as has been detailed earlier. A straight conversion based on body weight, or the use of allometric scaling can lead to very inaccurate information (Green et al., 2012) and plasma concen-trations convey more valuable information.

What is also important is the route of administration. Drugs are often given intraperitoneally, because this is technically eas-ier than oral or intravenous administration, and no attempt is made to measure absorption and exposure by measuring the plasma drug concentration. But intraperitoneal injection is not the way that recreational drugs are taken by humans, and the route employed can produce enormous differences in both peak exposure and area under the plasma concentration/time curve as has been shown by Baumann et al. (2009) who reported that administering MDMA to rats intraperitoneally, rather than orally, increased the Cmax by approximately four-fold and also increased Tmax four-fold and the AUC three-fold. Recreational drugs should ideally be given to animals by the same route as is used by human

users where possible, unless the route chosen better mimics the kinetics of the drug in humans.

Frequency of dosing can be problematic, but again can be tai-lored to reflect human use if knowledge on the pharmacokinetics of the drug in humans is available or can be inferred. Recently both MDMA and mephedrone have been given to animals in a manner designed to try and reflect human recreational use; for example by giving the drug on two consecutive days, followed by a five day gap before the next two day administration in a manner intended to reflect weekend use (Shortall et al., 2013b). The recent surge of binge dosing whereby human users take several doses in one session, an approach sometimes used with MDMA but particularly prevalent in mephedrone users (Schiffano et al., 2012), has also been mimicked in animal studies (Sanchez et al., 2004; Shortall et al., 2013b). Again this approach is only valuable if it has been shown that the drug pharmacokinetic parameters are comparable in the human and experimental animal.

Clinical studies

It is notable, and disturbing, that initial clinical reports on novel recreational drugs describing the physiological and psychologi-cal consequences of ingesting the drug often disclosed that the subjects under investigation 'believed' that they had taken it. However, blood samples were not taken to confirm that exposure to the specific named drug had occurred. This certainly happened recently with mephedrone (Dargan et al., 2010; James et al., 2011; Regan et al., 2010; Wood et al., 2011) and a recent study found that a significant proportion of reported 'mephedrone fatal-ities' were likely due to the other drugs that had been subse-quently identified at post-mortem (Schifano et al., 2012). We acknowledge the problems here in that analytical methodologies may either have not been developed or perhaps have not been accessible, and that it is important to make available to the medi-cal world the clinical problems probably associated with new recreational drugs. Some circumspection in the title and text such as 'probable mephedrone' or 'suspected mephedrone' would be preferable to the bald claim that the effects were due to mephed-rone. In the case of MDMA caution could be shown by calling the ingested drug ecstasy given the wide evidence for adultera-tion of supposed MDMA tablets by other substances (see later). This option is not possible with the many new drugs such as the cathinone derivatives and the term 'bath salts' (Joksovic et al., 2012) which is the name under which several different cathi-nones have been sold is also of no value to a pharmacologist without clarification as to the chemical formula of the substance under investigation.

Good clinical pharmacology requires accurate knowledge of the doses administered and frequency of administration. These data are never available with any precision in any study examin-ing dose effects in recreational users. Of course one has some indication as to possible dose from the number of tablets con-sumed and frequency of drug ingestion, although the dose con-tained in illicitly obtained tablets is highly variable (Vogels et al., 2009; Morefield et al., 2011). Furthermore, information on dosing and frequency is generally obtained from the user whose recall is likely to be limited, particularly if the drug being investigated has an effect on memory as has been claimed for MDMA (Parrott, 2013). Some subjects may decide to obfuscate for various reasons


Green and Nutt 5

(boasting at doses taken and therefore exaggerating or embar-rassed at doses taken and therefore under-reporting). Crucially the information can never take into account the problem of drug tablet adulteration (see later). Perhaps the most that can be stated is that users are light, medium or heavy users, together with the authors definition of these terms. The ecstasy tablet that most recreational users buy and ingest is not necessarily MDMA. Indeed in many cases, clearly, it is not.

Acute or long term effects?

Preclinical studies

What appears to be sometimes confused is whether the study is examining the consequences of acute or sub-acute dosing or long-term pathology. As a general rule with MDMA, high or repeated doses to laboratory animals can cause neurotoxic dam-age to 5-HT neurones in the forebrain. Therefore studies giving high or repeated doses and examining pharmacological effects in the next few days may be looking at the effects of long-term neu-rotoxic damage that is occurring or occurred, rather than any acute reversible action of the drug with pharmacological (rather than toxicological) relevance. Equally it should be confirmed that the drug has cleared the body if long-term effects are to be stud-ied. Knowledge of appropriate dosing and exposure should pre-vent this error and aid interpretation.

Clinical studies

Much the same comments can be made here as in the preclinical section above but with the added proviso that in the case of MDMA the 2.5 times greater period to reach peak plasma con-centration, coupled with the 10-times longer plasma half life in humans versus rats (Green et al., 2012) means that clearance of the drug will be much longer in humans. Therefore, when binge dosing has occurred, it can be several days before the drug has left the body. One is again faced with the problem of relying on subjects memory and reliability to ascertain when the last expo-sure to the drug occurred unless blood or hair samples are taken, and with hair samples this will only reflect use of the drug some days earlier.

Co-administered drugs

Preclinical studies

Mohamed and colleagues (2011) reviewed the drugs often co-administered by MDMA users. The list was extensive and included ethanol, cannabis, cocaine, nicotine, caffeine, lysergic acid diethylamide (LSD), benzodiazepines, barbiturates, opiates, ketamine, gamma-hydroxybutyrate (GHB) and psilocybin. They noted that all these compounds altered acute MDMA-induced behavioural or physiological changes in laboratory animals. In addition some drugs such as other amphetamine derivatives enhanced long-term neurotoxicity. Drug-drug interactions can result from two distinct mechanisms. One is pharmacokinetic, such as the fact that alcohol, for example, enhances plasma MDMA concentration (Hamida et al., 2009). The other is pharma-codynamic, as in the recent study that found that while mephed-rone was not itself neurotoxic it did enhance neurotoxicity induced

by methamphetamine, amphetamine and MDMA (Angoa-Prez et al., 2013). Furthermore the chemically related cathinones mephedrone and 3,4-methylenedioxypyrovalerone (MDPV), because of the different actions of the two drugs at the dopamine nerve ending, might be expected initially to release dopamine and subsequently prevent its reuptake via the dopamine uptake site (Cameron et al., 2013). Consequently, when taken together, the combination could have serious adverse effects on brain function. Our preclinical knowledge of both these types of interaction remains scant and studies are difficult to conduct as they require large investigations, using several doses, to determine not only the kinetics of the drug combinations but also whether any effect is merely additive or potentiating.

Clinical studies

Co-administered drugs can be ingested unknowingly or know-ingly. The list of drugs detailed in the previous section is that of drugs knowingly taken with MDMA by recreational users. However unknowing ingestion is a further major complication for all clinical studies since it results from tablet adulteration and is therefore impossible to factor in when reviewing the action of the active drug. One investigation has identified no less than 14 substances other than MDMA in ecstasy tablets which users pre-sumably believed contained only MDMA (Vogels et al., 2009). It is notable that fewer than 50% of ecstasy tablets confiscated in The Netherlands in 2009 contained MDMA, compared to 90% in previous years (Brunt et al., 2011). In many of these tablets MDMA was replaced by other compounds and, in 2009, mephe-drone was found to be the most prevalent new designer drug to be misleadingly sold as MDMA/ecstasy. Since the psychoactive effects of mephedrone are similar, one can presume that any of the users of these tablets recruited for a survey of the short or long-term effects of MDMA may well have reported that they were taking MDMA. Many other adulterants identified by Vogels et al. (2009) were also psychoactive and included compounds structurally related to MDMA such as 3,4-methylenedioxyethyl-amphetamine (MDEA) and 2-methylamino-1-(3,4-methylenedi-oxyphenyl)butane (MBDB) and there is little pharmacological or toxicological information on either compound. Related ampheta-mine-type compounds may well pose a particular problem as detailed in the preclinical section above.

Caffeine is a regular adulterant and again it cannot be assumed this has no interacting pharmacological action since this has been demonstrated to occur in preclinical studies on MDMA (Vanattou-Safoudine et al., 2012) and mephedrone (Shortall et al., 2013c).

With regard to knowing co-administration, ingestion of other drugs with ecstasy or other recreational drugs is endemic. Ten years ago, Cole and Sumnall (2003) detailed publications report-ing that drugs co-administered with ecstasy included other amphetamines, cannabis, LSD, cocaine, benzodiazepines, and ketamine and alcohol. Curran (2000) found that only 1.5% of sub-jects in her study had not also taken another psychotropic drug. Newer compounds such as mephedrone can now be factored in since the majority of mephedrone users (87% of 1506 survey par-ticipants with a mean age of 26 years) admitted previous illicit use of MDMA (Carhart-Harris et al., 2011; Moore et al., 2013).

In a recent review Parrott (2013: 1468) claimed that in studies on long-term neurotoxic effects of MDMA in the human brain the confounding problem of co-administered drugs has been



minimised by the fact that neurotoxicity causation by other rec-reational drugs is also unlikely on theoretical grounds, since most do not have strong serotonergic effects. However, this proposi-tion is wrong since it totally ignores the fact that when two or more compounds are taken at the same time, the effects of one of the compounds can be altered by the other because of pharmaco-dynamic or pharmacokinetic reasons. This is known as a drug-drug interaction and there are many examples in clinical pharmacology. The anticoagulant effect of warfarin is altered by a substantial number of other drugs, none of which has an effect on coagulation. Their interactions involved altering the absorp-tion, plasma protein binding and metabolism of warfarin or other more indirect actions (see Grahame-Smith and Aronson, 1992). It is reasonable to suppose therefore that some drugs may interact with MDMA and result in neurotoxicity. Consequently it is cer-tainly possible that ecstasy causes functional impairment of psychological functions or even neurotoxicity, but it is impossi-ble to ascribe the problem to MDMA itself, mainly because trans-lation cannot be made from preclinical studies using pure MDMA to studies in recreational users taking ecstasy.

MDMA is just like any other drug in that it obeys general pharmacological principles including dose-response effects (though one should never forget idiosyncratic responses can occur). It is interesting therefore that at least some meta-analyses of the effects of MDMA found no association between dosing and impairment of memory function (Laws and Kokkalis, 2007; Murphy et al. 2012; Verbaten, 2010) which makes it likely that it is a drug-drug interaction that is responsible for many of the pathologies that have been reported.

The observation in another meta-analysis that ecstasy users also consumed significantly more amphetamine, cocaine, alco-hol, nicotine and LSD, but less alcohol than polydrug controls (Verbaten, 2010), emphasises the fact that it is going to be impos-sible to remove some confounds in clinical studies. What will assist interpretation are very detailed drug histories which, together with large studies (and meta-analysis), will allow effec-tive sub-group analysis that might indicate which drug combina-tions are of particular concern. A meta-analysis on depressive symptomatology and MDMA use noted that 'in general, drug histories were poorly reported' (Sumnall and Cole, 2005: 84), so better reporting is certainly going to help when meta-analysis is performed.

What could be valuable would be the establishment of a 'yel-low card' type of system for recreational drugs by a multi-country group such as the Psychonaut Web Mapping Group to obtain information on all adverse effects of drugs experienced by users. This might help in better identifying drug combinations and their adverse effects. Whether such a system would be of value in allowing the licensing of NPS in identifying lower risk sub-stances as proposed in New Zealand is unclear.

Translation and reverse translationCan we improve the situation on the translatable value of both preclinical and clinical studies on NPS with the new recreational drugs now flooding the market? This can be answered with a qualified yes. In the case of MDMA, many major publications were appearing by the late 1980s to mid 1990s. However no good clinical pharmacokinetic study appeared until the report of Yang et al. (1996). A comprehensive preclinical study in rats was even

more recent (Baumann et al., 2009). Only then could the clear mismatch between the kinetics of the drug in humans and ani-mals be appreciated (Green et al., 2012). In contrast, there are now several good preclinical studies on mephedrone pharma-cokinetics in rats (Aarde et al., 2013; Martinez-Clemente et al., 2013; Miller et al., 2013) and some data are available on plasma concentrations in humans (Maskell et al., 2011; Schifano et al., 2012). This should allow better translation by preclinical scien-tists, provided the integrative pharmacology approach outlined above is followed. However, further detailed clinical studies on the pharmacokinetics of this drug in the UK at least will be prob-lematic as mephedrone can only be administered to humans if it has been synthesised and prepared to GMP standards as noted earlier. The same problems of drug supply and costing will also occur for anyone who wishes to examine the physiological or psychological effects of the drug in controlled laboratory condi-tions. Consequently preclinical scientists hoping to examine whether reports on the effects (both desired and adverse) reported by subjects taking the drug recreationally can be reproduced and examined in laboratory animals are therefore faced with a prob-lem in reverse translation. That is, they have to rely on clinical data collected from published observations on persons reporting adverse effects to clinicians, or data collected from users report-ing effects via questionnaires or interview. The weakness of this is evident.

Proposals and conclusionsAs detailed in this article there are many weaknesses in current research on recreational drugs and this raises the question as to how matters can be improved. We suggest the following:

1. A general understanding of the importance of pharma-cokinetic-pharmacodynamic integration in preclinical studies in providing meaningful information that will allow translation and reverse translation.

2. Better, more detailed, reporting of both preclinical and clinical studies, including negative studies.

3. Better collaboration between preclinical laboratories, thereby performing integrated studies examining diverse functional consequences of the drug.

4. Much closer interaction between preclinical and clinical scientists. This used to happen in psychopharmacology (Sjoerdsma, 2008), but interaction between preclinical and clinical pharmacologists is currently weak (Green and Aronson, 2012). Psychopharmacological studies should also encompass clinical psychologists. Clinical pharmacologists are generally more aware of the prob-lems of pharmacokinetics and could provide input on this as well as detailing what aspects of recreational drug effects are general and also clinically important and thus assist reverse translation.

5. Evidence that the drug under investigation has actually been ingested by the human recreational users that are the subject of any report, together with drug exposure information where possible.

6. Better information on the pharmacokinetics and func-tional effects of the drug in human subjects in controlled laboratory conditions. Unfortunately there are few labs with the income, skills and licenses to do this (see Nutt


Green and Nutt 7

et al., 2013). This problem could in part be ameliorated by exempting human experimental medicine studies in Europe from the current GMP production requirements, as is the case in the USA.

7. Much greater use of meta-analysis. Narrative reviews of individual studies have value but may not be comprehen-sive and include small studies and even case reports. Importantly they may fail to weigh study quality when analysing the data on which conclusions are then drawn. Meta-analysis can also be used in preclinical studies and has even been used for individual animal meta-analysis in stroke research (Bath et al., 2009b).

We now have much preclinical and clinical information on recent recreational drugs. What is disturbing is the lack of consensus as to how dangerous these compounds are to the health of the indi-vidual and society in general. Greater use of good pharmacologi-cal practice should assist in answering this point.

Conflict of interestThe authors declare that there are no conflicts of interest.

FundingThis research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors.

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