Functional GI disorders: from animal models to drug development

Transcription

1 1 UCLA Center for Neuroviseral Sienes & Women s Health, Departments of Mediine, Physiology and Psyhiatry, David Geffen Shool of Mediine at UCLA, Los Angeles, CA, USA; 2 UCLA Center for Neuroviseral Sienes & Women s Health, Departments of Mediine, David Geffen Shool of Mediine at UCLA, Los Angeles, CA, USA; 3 UCLA Center for Neuroviseral Sienes & Women s Health, Departments of Mediine, Center for Outomes Researh and Eduation, David Geffen Shool of Mediine at UCLA, Los Angeles, CA and VA Greater Los Angeles Healthare System, Los Angeles, CA, USA; 4 UCLA Center for Neuroviseral Sienes & Women s Health, Departments of Mediine and Psyhiatry, David Geffen Shool of Mediine at UCLA, Los Angeles, CA and VA Greater Los Angeles Healthare System, Los Angeles, CA, USA Correspondene to: Professor E A Mayer, UCLA Center for Neuroviseral Sienes & Women s Health, VAGLAHS, Bldg. 115, Room 223, Wilshire Blvd, Los Angeles, CA 90073, USA; emayer@ula.edu Revised 27 August 2007 Aepted 8 September 2007 Published Online First 19 Otober 2007 Funtional GI disorders: from animal models to drug development E A Mayer, 1 S Bradesi, 2 L Chang, 2 B M R Spiegel, 3 J A Bueller, 2 B D Naliboff 4 ABSTRACT Despite onsiderable efforts by aademi researhers and by the pharmaeutial industry, the development of novel pharmaologial treatments for irritable bowel syndrome (IBS) and other funtional gastrointestinal (GI) disorders has been slow and disappointing. The traditional approah to identifying and evaluating novel drugs for these symptom-based syndromes has relied on a fairly standard algorithm using animal models, experimental mediine models and linial trials. In the urrent artile, the empirial basis for this proess is reviewed, fousing on the utility of the assessment of viseral hypersensitivity and GI transit, in both animals and humans, as well as the preditive validity of prelinial and linial models of IBS for identifying suessful treatments for IBS symptoms and IBS-related quality of life impairment. A review of published evidene suggests that abdominal pain, defeation-related symptoms (urgeny, straining) and psyhologial fators all ontribute to overall symptom severity and to health-related quality of life. Correlations between readouts obtained in prelinial and linial models and respetive symptoms are small, and the ability to predit drug effetiveness for speifi as well as for global IBS symptoms is limited. One possible drug development algorithm is proposed whih fouses on pharmaologial imaging approahes in both prelinial and linial models, with dereased emphasis on evaluating ompounds in symptom-related animal models, and more rapid sreening of promising andidate ompounds in man. Despite the tremendous efforts by aademia and industry alike during the past 15 years, the suess rate for effetive drug development for irritable bowel syndrome (IBS) and other funtional gastrointestinal (GI) disorders (FGIDs) remains unimpressive. Only two new IBS treatments that is, alosetron and tegaserod have gained initial Food and Drug Administration (FDA) approval as a new IBS treatment so far. However, both of these mediations are only available in the USA as part of a restrited aess programme, due to side effets. 1 Of the many failed ompounds that never saw the light of the peer-reviewed literature, only a small number were offiially announed as a failure, inluding the peripheral viseral k-opioid agonist fedotozine, the seletive m 3 musarini reeptor antagonist darifenain and the seletive NK 3 reeptor antagonist talnetant. Despite this disouraging news, and despite the fat that several major ompanies have deided to leave the IBS field altogether, several new and promising ompounds are in various stages of early linial development. 2 3 There have been advanes in the field inluding development of agreed upon definitions of the major syndromes (as well as a myriad of other FGIDs), 4 animal models with some fae and onstrut validity for the human syndrome 5 and identifiation of a ontinuously inreasing number of moleular targets on epithelial and immune ells and viseral afferent neurons, 6 and in the entral nervous system (CNS). 3 Furthermore, several potential biomarkers have been postulated in IBS patients (ranging from abnormal baterial flora 7 to muosal immune ativation (reviewed in Spiller et al 8 ) and abnormal pereptual and brain responses (reviewed in Mayer et al 9 ). Despite the lak of an agreed upon pathophysiology, a standard strategy for drug disovery, inluding prelinial and linial studies on GI motility and oloretal sensitivity, as well as large phase II and III effiay trials, have been followed by most of the pharmaeutial ompanies involved in this field. Why have these aomplishments not translated into more effetive treatments? Is there a fundamental flaw in the drug disovery and development strategies that have been followed during the past deade? Is it premature to embark on ostly drug development strategies for omplex symptom-based disorders like IBS or other FGIDs as long as a full understanding of the pathophysiology of these syndromes is not available and the treatment targets remain moving targets? The situation is further ompliated by rapid advanes in the sope and speed of drug disovery efforts in the pharmaeutial industry over the past deade so that the field is now faed with a rapidly growing number of andidate ompounds emerging from prelinial development without a ost-effetive strategy to sreen these ompounds effiiently for their usefulness in human patients. In the following review, we will fous on the proess of drug development in IBS, by ritially reviewing three areas in the urrent assumptions and strategies of drug development efforts in IBS: (1) What is the urrent approah to drug development in IBS, and what is the strength of the evidene supporting this approah? (2) What is the preditive validity of ommonly employed experimental mediine models for IBS symptoms. (3) What is the preditive validity of existing animal models and the moleular targets identified in these models for the human disorder? Based on this review, we are proposing possible modifiations in the existing strategies whih ould avoid the pitfalls of the past and present, and hopefully translate into more ost-effetive development 384 Gut 2008;57: doi: /gut

2 strategies for this important area of gastroenterology. THE CURRENT APPROACH TO DRUG DEVELOPMENT The urrent approah to drug development for IBS and other FGIDs is illustrated in fig 1. Syndromes are defined by their primary symptoms, in the ase of IBS in terms of hronially reurring abdominal pain or disomfort assoiated with alterations in bowel habits 4 (fig 1A). Human biologial markers inluding pereptual hypersensitivity to experimental retal or sigmoid distension (referred to throughout as viseral hypersensitivity ), and altered intestinal transit (whole gut or regional oloni transit) whih are thought to underlie speifi IBS symptoms serve as a basis for the development of animal models (noieptive reflex responses to oloretal distension, faeal pellet output or transit studies) whih in turn aim to mimi the human biomarkers or intermediate phenotypes (fig 1C). 10 Moleular targets are identified in these animal models whih are thought to mediate the harateristi features exhibited by the animal model (eg, ion hannels and reeptors on viseral afferent neurons, enterohromaffin ells, enteri neurons, entral stress iruits). Highly seletive, andidate ompounds aimed at these moleular targets are developed and optimised (fig 1E), whih in turn are tested in the respetive animal models and, if shown to be effetive and safe, are tested in human experimental mediine models (phase I and IIa) for their ability to affet Gut: first published as /gut on 26 Otober Downloaded from Figure 1 General strategy in irritable bowel syndrome (IBS) drug disovery and development. The shemati illustrates the vertial progression from the symptom omplex of IBS to target identifiation in animal ells. For eah step the urrent approah is shown in the left portion of the boxes. In the right portion of eah box, suggested modifiations to the urrent approah are shown. The urrent approah is lengthy, expensive and based on poor orrelations between the individual steps. GI, gastrointestinal. on 23 January 2019 by guest. Proteted by opyright. Gut 2008;57: doi: /gut

3 GI transit or viseral sensitivity. Either sequentially or in parallel, andidate ompounds are being evaluated in small proof of onept (POC) studies (phase IIa) and large phase II/III multientre studies, using global (as opposed to speifi symptom-related) end points, in partiular the adequate relief end point. 11 Even though this drug development approah appears to be rational at first glane, the fat that it has not proven to be more suessful or osteffetive for IBS may be related to several major flaws in its logi. Translation of human symptom-based disorder into human biomarkers (fig 1A) In ontrast to most organi diseases with identifiable lesions or biohemial abnormalities, so-alled funtional syndromes have traditionally relied on subjetive, symptom-based outome measures. In parallel to attempts to refine these symptom riteria, a major effort has been underway during the last deade to identify surrogate biomarkers for FGIDs, whih an be used as more objetive end points for drug evaluation. The ideal surrogate marker would be seen in most affeted patients (or a learly identifiable subset of patients), its demonstration would be independent of a partiular laboratory, it would exhibit high test retest reliability both within and between patients, and, most importantly, would orrelate highly with either a speifi IBS symptom or, preferably, with global IBS symptom severity or health-related quality of life (HRQoL) that is, the fators that ultimately determine if an individual beomes a healthare-seeking IBS patient. The finding of an abnormality without a high orrelation with symptom severity (eg, muosal immune ativation 12 ) may be interesting from a sientifi standpoint to understand IBS pathophysiology better or to identify subgroups of patients, but suh a finding probably does not have partiular value for prediting drug effetiveness or determining optimal drug dose. Examples of the latter inlude non-speifi alterations in GI motility, altered number of enterohromaffin ells, intraepithelial lymphoytes or plasma ortisol values. There has been reported a orrelation of abdominal pain with the number of muosal mast ells in lose proximity to nerve endings, but this finding will need to be reprodued by other laboratories before it an be onsidered a true biomarker of IBS symptoms. Potential biomarkers for whih orrelation with IBS symptoms have not been published inlude reently reported abnormalities in stool miroflora 7 and in stool proteases. 15 The assessment of a good surrogate marker in validated experimental paradigms in human subjets should have a high validity in prediting the effetiveness of a ompound in the treatment of IBS symptoms. Based on the ardinal symptoms of IBS (abdominal pain/disomfort, altered bowel habits), a series of suh experimental mediine models have been developed and used in the drug development proess during the past 10 years. However, before aepting the most ommonly used surrogate markers for abdominal pain and for altered bowel habits as meeting the requirements of prediting the effetiveness of a andidate drug on IBS, several points have to be onsidered. The best and most ommonly used subjetive end point to assess IBS drug effetiveness is global end points, inluding the adequate relief end point. 4 While the gold standard in IBS linial trials, suh global end points do not employ speifi symptom-based readouts, but rather a subjetive value judgement (eg, how an individual patient feels that her/ his symptoms have been relieved adequately), whih ould be orrelated with surrogate markers. Surprisingly, despite deades of IBS drug development, there is unertainty and few data to demonstrate how best to measure IBS severity. A preliminary report identified several preditors for patient-assessed overall severity of gastrointestinal symptoms. 16 As hypothesised, the preditors inluded multiple symptoms, suh as ratings of abdominal pain and disomfort (bloating), defeation-related symptoms (straining, urgeny) and a symptom-related anxiety ( something serious is wrong with my body ). These fators jointly aounted for only one-third of the variane of IBS severity. Therefore, there are other fators whih were not speified in this model or have not been measured. However, within this group of preditors, abdominal pain was the most powerful preditor of severity. The assoiation between abdominal pain and severity was almost three times as strong as the next losest preditor, whih was something serious is wrong with my body. A reent study identified several preditors of IBSrelated HRQoL impairment. 17 Surprisingly, alteration in bowel habits was not identified as a signifiant preditor of HRQoL, while several non-gi symptoms, suh as vital exhaustion and symptom-related anxiety, were identified. These fators jointly aounted for 39% of the variane of HRQoL. Amongst the symptom preditors of HRQoL, symptoms of vital exhaustion suh as low energy and tiring easily were the most important preditors of physial HRQoL, followed losely behind by overall symptom severity. Feeling tense was the most important preditor, followed by nervousness and hopelessness for mental HRQoL. Assuming that the global end points used in linial trials ( did you get adequate relief of your bowel symptoms ) are influened by both IBS severity and IBS-related HRQoL impairments, it is likely that a range of fators inluding measures of abdominal pain or disomfort, defeation-related symptoms and psyhologial symptoms ontributes to the way we urrently measure the effetiveness of IBS drugs. If this hypothesis is orret, suessful pharmaologial relief of one of these omponents will result in only moderate global symptom relief, while pharmaologial relief of two or more of these fators should result in a highly effetive drug. The identifiation of other preditors of severity and HRQoL impairment is likely to improve the model and in fat may lead to 386 Gut 2008;57: doi: /gut

4 new surrogate markers for drug development. An alternative or omplementary approah would be the identifiation of CNS orrelates of IBS symptom severity or of adequate relief, using the respetive brain iruits as objetive surrogate markers (see also final setion). This hypothesis is illustrated in fig 2B. In the following, we will fous on the two most ommonly used surrogate markers for whih evidene for orrelation of human biomarkers with linial IBS symptoms is available: enhaned pereptual responses to ontrolled retal or sigmoid distension (referred to from hereon as viseral hypersensitivity ) and gut transit studies, Viseral hypersensitivity as a biomarker for IBS Viseral hypersensitivity has been the most widely used and perhaps most ontroversial of the biologial markers of IBS. In fat, in humans the metri of sensitivity is almost always based on a subjetive report and, therefore, is an indiret measure of a hypothesised neurophysiologial proess. Traditionally, basi sientists have extrapolated from results obtained from viseral afferent reordings and pseudoaffetive reflex responses in animals to relevane for the treatment of enhaned viseral pereption and IBS symptoms in patients. However, it has to be kept in mind that what an be assessed in human subjets is mostly the subjetive sensitivity to experimental stimuli in a safe environment, and not measures of afferent sensitivity for stimuli diretly related to linial symptoms. 18 Given the onsiderable number of ognitive and emotional variables that influene the ultimate experiene of the sensation, it an be assumed that the relationship between viseral afferent sensitivity and pereption is highly nonlinear. The peripheral stimulus for testing viseral sensation is most often a mehanial distension using an air-filled bag and a omputer-ontrolled pump (or barostat), but may also inlude eletrial or hemial stimuli. 19 Both the type and the harateristis of the stimulus (eg, slow inreasing ramp distension vs square wave phasi distension) as well as its preditability an signifiantly impat the outome of experiments independent of the absolute amount of pressure or volume. Is viseral sensitivity a reliable marker of IBS? For over 30 years viseral hypersensitivity to balloon distension has been desribed as a harateristi of IBS. 22 Viseral hypersensitivity ould in priniple result from a number of peripheral (eg, muosal immune ativation, mast ell degranulation, altered flora) as well as from entral mehanisms. A large number of papers have ompared groups of IBS patients with ontrols using various distension proedures, and several generalisations an be made from this heterogeneous group of studies. 18 IBS patients as a group typially show lower average thresholds for pain or disomfort than healthy ontrols in response to brief (30 60 s) phasi retal distensions. About 40 60% of IBS subjets show baseline hypersensitivity when this is defined as a pain or disomfort threshold to phasi distensions lower than the 95% CI of a healthy ontrol group. 29 Bouin et al 24 studied a large lini sample of IBS patients (n = 86, 60 females, 26 males) and examined the sensitivity and speifiity of an asending phasi distension test (ie, a paradigm whih maximises hypervigilane) for separating IBS patients from healthy subjets and other GI patients with inflammatory bowel disease, funtional onstipation, et. They found an 80% speifiity and a 90.7% sensitivity at a 40 mm Hg threshold (86.3% effiieny) for separating IBS and ontrols, and similar values for omparing IBS with the ombined GI patients (86.8% effiieny at 40 mm Hg). IBS patients do not show the same hypersensitivity to slow ramp inflations ompared with ontrols, and may even be less sensitive to suh stimuli than healthy ontrols. Similarly, IBS patients seem to demonstrate somati hypersensitivity when tested with toni heat stimuli, 27 but normosensitive or hyposensitive to phasi eletrial or pressure stimuli. IBS patients, in partiular female patients, demonstrated enhaned pereptual responses to repeated distensions in the retum and sigmoid olon, suggesting that abnormal sensitisation or temporal summation of viseral stimulation may also be a marker of the disorder In summary, hypersensitivity based on barostat assessment using phasi stimuli and a subjetive readout does show speifiity for IBS, although there is onsiderable variability aross patients and between different laboratories, and overlap with non-patient samples. The role of hypervigilane in viseral hypersensitivity Clearly a signifiant omponent of pereptual hypersensitivity seen in the IBS studies desribed above is due to attentional proesses that an be desribed as hypervigilane to viseral sensations. 21 Experimental designs that use non-random presentation of stimuli (eg, those that use an asending series of distensions) tend to foster response bias in that subjets an give appropriate responses without paying lose attention to the atual sensation. IBS subjets may be partiularly prone to suh bias due to anxiety, even though it is urrently not known if this type of anxiety is a primary fator or has developed as a onsequene of hroni pain. Although non-random and random proedures are often orrelated, in general, differenes between IBS and ontrols are greater in studies using asending versus randomly presented stimuli In addition, even with randomly presented stimuli, IBS patients show signifiant Gut 2008;57: doi: /gut

5 Gut: first published as /gut on 26 Otober Downloaded from Figure 2 Progression from animal models to human biomarkers to human symptoms and health-related quality of life (QoL) impairment. (A) Current onept. Shown are ommonly used assays and measures at these three levels of investigation. Arrows denote orrelations/preditive validity from the lower to higher level. Signs next to arrows denote strength of onnetions: + good, (+) weak,? not known. (B) Proposed onept. Shown is a proposed modified version of (A), with greater emphasis on brain imaging approahes at both the prelinial and linial level. The orrelations between prelinial and linial brain imaging approahes, symptom severity and health-related QoL are urrently not known (for details, see also fig 3). IBS, irritable bowel syndrome; GI, gastrointestinal. on 23 January 2019 by guest. Proteted by opyright. 388 Gut 2008;57: doi: /gut

6 habituation of responses with repeated exposure to retal distensions, 28 and pereption of distensions inreases during even mild psyhologial stress, 41 demonstrating the influene of affetive and ognitive fators on viseral pereption beyond simple judgemental bias. This view is onsistent with brain imaging results showing greater ativation in emotional arousal regions, and habituation of suh hyperarousal with repeated stimulus appliation. 28 Correlation of viseral sensitivity with linial symptoms (IBS symptoms, symptom severity, psyhologial symptoms) Despite the large literature omparing viseral sensitivity aross patient and non-patient groups, there are very few data examining the relationship between these measures and IBS symptom severity. Of the few studies that have reported these data, the relationships appear to be small or nonexistent. Two studies, reported in abstrat form, suggest that the orrelation between pain symptoms and retal and sigmoid sensitivity range between 0.3 and 0.4 that is, only between 9% and 16% of abdominal pain symptoms are explained by retosigmoid sensitivity during experimental distension. Several more reent studies have also reported a signifiant positive relationship between a viseral sensitivity measure and abdominal pain symptoms. There are no lear data relating viseral sensitivity observed during barostat-mediated retal distension and more global IBS symptoms, and in fat several studies have shown a lak of suh a relationship There is a larger literature examining the relationship between viseral sensitivity and bowel habit; however, the results have not been onsistent. Some studies have found inreased sensitivity in diarrhoea-predominant patients ompared with those with onstipation, but other studies have found the opposite or no differenes (see Staher et al 47 ). In addition, there are no lear data on the relationship of severity of bowel habit and measures of sensitivity. A reent study suggests that the hypersensitivity seen in IBS patients is primarily seen in female patients. 35 Preditive validity of viseral sensitivity testing for FGID drug development Table 1 lists studies using viseral pereption testing proedures to evaluate the potential benefits of andidate drugs as viseral analgesis and as potential mediations for treating IBS symptoms. From these publitions, several important onlusions an be drawn with regard to this experimental mediine proedure for IBS drug development. (1) Viseral sensitivity tests learly show robust analgesi responses to m- and k-opiate analgesis in small rossover samples suh as used in typial trials These studies support the viability of distension proedures in the retum (and probably the stomah as well) to detet aute hanges in analgesia. In other words, the barostat test is a valid human experimental model for viseral pain. It should also be noted that the pereptual differenes in these studies were independent of hanges in ompliane or tone. (2) To be useful as a surrogate marker, viseral testing should be helpful in disriminating mediations that do or do not have a positive impat on either speifi or global IBS symptoms. This riterion is often referred to as the ability to reah a go/no-go deision based on the test results. Data from multiple lasses of drugs have now been ompiled and often there is little preditability between the two types of outomes, with some drugs showing positive hanges on viseral sensitivity testing by barostat, but no hange on symptoms, and vie versa (see Kuiken et al. 72 ). Some ompounds, suh as the k-opioid-preferring antagonist fedotozine, have a signifiant effet on viseral testing in some studies, and also seemed to impat IBS symptoms positively in initial small lini studies. However, further development was not pursued, presumably due to lak of effiay in well-designed phase II linial trials. Similarly, the syntheti somatostatin analogue otreotide onsistently showed viseral analgesi and antihyperalgesi properties during laboratory testing. Preliminary evidene from an 8-week ontrolled linial treatment trial 68 showed that otreotide treatment was assoiated with a redution in the pereption of barostat-indued retal distension in non-onstipated IBS patients, as well as a redution in abdominal omplaints and improved stool onsisteny. The NK3 reeptor antagonist talnetant had no effet on viseral pereption (or mehanoelasti parameters of the retum) in a large well-designed study in healthy volunteers 61 (it was never tested in IBS patients), and had no signifiant effet on IBS symptoms in two well-designed randomised ontrolled trials (RCTs). 60 On the other hand, the 5-HT 3 reeptor antagonists inluding ondansetron and alosetron have shown no diret effet on pereption, but at least for alosetron have shown positive effets on global IBS symptoms. 52 Similarly, the 5-HT 4 reeptor agonist, tegaserod, is another serotonergi ompound that has shown some effiay on global IBS symptoms but no lear hange in studies on human viseral sensitivity. 74 Sine these ompounds are likely to affet serotonin modulation of GI motility and seretion, and in the ase of alosetron possible CNS mehanisms related to anxiety, 75 it is likely that for many drugs their impat on IBS global symptoms is not via the presumed viseral analgesi mehanism but instead through their effet on entral or peripheral autonomi (inluding enteri nervous system) pathways. Antidepressants inluding triylis have not been well studied in viseral sensitivity tests, but there is little evidene that they derease the pereption of noxious retal distension per se, even though they have been found to have a small to moderate therapeuti effet on IBS symptoms Gut 2008;57: doi: /gut

7 Table 1 Drug effets on viseral sensitivity in IBS Drug Dose IBS study subjets Study design Barostat methodology Effet on sensitivity in IBS Effet on motor funtion Effet on IBS symptoms Conlusions from phase II and III trials Opiates Fentanyl 40 Two doses (HD and LD), bolus + infusion 10 IBS (6 F) DB, PC, rossover Phasi pressure HD and LD: q DisTh, q SensTh HD: q PainTh None NR* NR Fedotozine k 100 mg single dose 14 IBS (8 F) PC, rossover Phasi pressure q PainTh, q SensTh None NR Small but signifiant agonist 48 hange in pain in small trial 49 Asimodoline k 0.5 mg single dose 20 IBS (20 F), all agonist 50 hypersensitive 5-HT Alosetron 51 Either 0.25 mg bd or 4mgbd6 7 days Alosetron 53 Either 1 mg alosetron or 4 mg alosetron bd 6 4 weeks DB, PC, rossover Phasi pressure No hange in PainTh, Q sensory ratings None NR NR 22 IBS (9 F) PC, parallel group study Phasi pressure No effet on PainTh (pressure) q ompliane Effetive for global symptom relief IBS (19 F) PC, parallel group study Phasi pressure? q retal ompliane Ondansetron 54 One dose at 0.15 mg/kg 12 IBS (6 F) DB, PC, parallel group study Phasi pressure No effet on sensitivity q retal ompliane NA No effet in pilot study 55 Ondansetron mg, 3 times/day 6 IBS PC, rossover? q SensTh None Dereased number of episodes of abdominal pain Ondansetron 57 Single dose of 0.15 mg/kg Granisetron 58 Either 40 mg/kg or 160 mg/kg 5 IBS (3 F) PC, parallel group study? Phasi pressure No hange in sensitivity None NR 12 IBS (8 F) DP, PC, rossover Phasi pressure or volume?? Tegaserod 63 6 mg bid 49 IBS (49 F) DB, PC, parallel group study q DisTh, q UrgeTh None Partiipants noted onstipation Phasi sigmoid pressure No effet q sigmoid aomodation NR No symptom hange Effetive for global symptoms and onstipation 59 Talnetant and 100 mg 102 healthy ontrols (60 F) Antidepressant Amitriptyline mg/day 6 2 weeks and then 25 mg h for the following 4 weeks DB, PC, parallel group study Phasi retal pressure No effet No effet NA Not different from plaebo IBS (7 F) Parallel group study? Phasi pressure q PainTh None Symptom redution Some effetiveness in meta-analysis and in high quality linial Fluoxetine mg 6 6 weeks 40 IBS DB, PC, parallel group study Phasi pressure and volume ramp No hange in sensitivity. Q abdominal pain in hypersensitivity patients none Redued the number of patients reporting signifiant abdominal pain trial May be effetive. Positive effet on pain in small trial 65 Other CNS Gabapentin mg/day 6 5 days 43 IBS-D (no sex data) DB, PC, parallel group study Phasi pressure q PainTh, DisTh q ompliane NR NR Otreotide mg/kg (s) 10 IBS DB, PC, rossover Phasi pressure q PainTh, DisTh None NR Preliminary evidene Otreotide mg (s) 8 IBS-D DB, PC, rossover Ramp volume q tolerane q ompliane NR suggests some Otreotide 70 7 IBS (4 F), 9 IBS (9 F) DB, PC, rossover (2 studies) Phasi pressure alone and after sigmoid stimulation q DisTh, Q sensitisation from sigmoid stimulation None NR effetiveness on IBS symptoms 68 bd, twie a day; CNS, entral nervous system; DB, double-blind; DisTh, disomfort threshold; F, female; HD, high dose; 5-HT, 5-hydroxytryptamine; IBS, irritable bowel syndrome; IBS-D, diarrhoea-predominant irritable bowel syndrome; LD, low dose; NA, not appliable; NR, not reported; PainTh, pain threshold; PC, plaebo-ontrolled; SensTh, sensory threshold; s, subutaneous; UrgeTh, urgeny threshold. 390 Gut 2008;57: doi: /gut

8 How then does viseral sensitivity testing fare against the riteria for a biomarker presented above? It is lear that a majority of IBS (and probably funtional dyspepsia) patients show hypersensitivity to at least phasi distension, although a substantial minority do not show this abnormality. No speifi linial harateristi has been identified that would differentiate the hypersensitive and normosensitive patients, with the possible exeption of female sex. Muosal immune ativation, differenes in baterial flora or geneti fators may all be ontributing to this heterogeneity. The barostat tehniques do appear to be reliable aross labs, with muh onsisteny for manipulations (eg, analgesis) that should diretly impat responding. The results are reproduible in a single subjet at least for two testings, as evidened by the suess of rossover studies; however, newer data show gradual habituation over multiple testing. 28 Viseral sensitivity testing has not been shown to be reliably assoiated with aute or hroni symptom intensity. Unfortunately, there are atually few data on this relationship, but to date there is no evidene for a relationship with global symptoms, a small set of data indiating some relationship with abdominal pain, and inonsistent data from ompounds that either do or do not impat linial symptoms. Overall, the barostat-mediated distension of the human gut emerges as a potentially reliable and valid approah to test pereption of viseral sensation and hanges in viseral pereption. However, it is most important in the formulation of a trial design to understand that the subjetive (pereptual) and objetive (mehanoelasti) responses obtained in this invasive and lengthy test are not highly related to global or even speifi IBS symptoms (suh as pain) and, therefore, when used by itself, may not be suitable to make soalled go/no-go deisions in drug development. There has also been some interest in examining the lower limb (or RIII) noieptive reflex as a potential objetive marker of viseral sensitivity. 29 The RIII is dereased during slow ramp distension of either the stomah or retum in healthy ontrols, a response that has been hypothesised to result from supraspinal modulation of the reflex as part of a pain inhibiting pain system referred to as desending noxious inhibitory ontrols (DNICs). This RIII tehnique has been used in one study of Tegaserod 71 and has shown alterations in IBS (inreased RIII during a ramp retal distension). However, while of interest, it has not to date been validated suffiiently as a marker of viseral hypersensitivity in patient populations or with any other mediations. GI transit as a biomarker for the IBS symptom: altered bowel habits In general, subjetive reports of onstipation or diarrhoea have been assoiated with alterations in GI transit, and measurements of global or regional GI transit have been used extensively to evaluate andidate drugs assuming a preditive value of suh tests for drug effetiveness on IBS symptoms. However, while defeation-related symptoms (straining, urgeny) were found to be preditive of IBS severity, 16 altered bowel habits were not found to be an important fator in the impairment of HRQoL in a large survey of IBS patients. 17 It is well known that the symptom of onstipation may our with or without slowing of oloni transit. 76 In many patients omplaining of onstipation, partiularly those with normal transit onstipation, it may be more a sensory symptom of feeling onstipated, rather than a symptom of altered motility or seretion. In suh patients, using the objetive marker of gut transit would be expeted to show poor orrelation with overall IBS symptoms. Transit studies to quantitate intestinal motility Gut transit refers to the time taken for food or other material to pass through the GI trat. Transit is a linially relevant and onvenient measure of GI funtion primarily related to GI motility and seretion. 77 There are various methods of measuring GI transit, and the more ommonly used tehniques in linial studies are radio-opaque markers and sintigraphy (reviewed in Camilleri et al 78 and Metalf et al 79 ). Validation of the radioopaque marker study to evaluate gut transit was performed by omparing the three different markers used in a group of healthy individuals. 79 The mean olon transit times as measured by the three different markers orrelated fairly well (r = ). The variability in transit times within an individual was thought to reflet true day-to-day variation. The authors suggested that for linial purposes only major differenes from normal values an be aepted as a signifiant finding. The reproduibility of sintigraphy was assessed in 21 healthy individuals over a 3-week period of time. 80 Gastri emptying at 4 h was highly reproduible (oeffiient of variation was 4%) on repeat testing. The oloni measurement was less reproduible and varied by more than one geometri entre unit in 37% of subjets at 24 h and in 26% of subjets at 48 h. Furthermore, in almost 75% of subjets, the residuals for oloni transit by sintigraphy were within one geometri entre in up to 72% of subjets aross the range of mean oloni transit times by radio-opaque markers. 80 The sample sizes for sintigraphi transit measurements needed to detet linially meaningful differenes were alulated and differed depending on the end point. The transit end points, whih appeared to need the smallest number of subjets, were the perentage of gastri emptying at 4 h (n = 6 to detet a differene of 25%) and the oloni geometri entre at 48 h (n = 14 to detet a differene of 1.5 geometri entre units). A sample size of 23 was needed to detet a differene of 1.7 geometri entre units at 24 h. Gut 2008;57: doi: /gut

9 Do gut transit measurements orrelate with IBS symptoms? There is evidene of aelerated gut transit in diarrhoea-predominant IBS (IBS-D) patients ompared with normal values, although group sizes are relatively small in many of the studies. In one of the largest studies, Horikawa and olleagues ompared gut transit times in 72 IBS patients (48 IBS-D, 24 onstipation-predominant IBS (IBS-C)) and 23 healthy ontrols using radio-opaque markers. 81 Total gut transit times were signifiantly aelerated in IBS-D patients ompared with ontrols and IBS-C patients. Segmental olon transit times of the asending, transverse, desending and retosigmoid olon were signifiantly shorter in IBS-D than in the other two groups. However, there were no signifiant differenes in transit times between the IBS-C and ontrol groups. In another study, baseline oroaeal transit was shorter in six patients with IBS-D ompared with eight healthy ontrols. 82 This finding onflited with another study whih found that gastri emptying and small intestinal transit times measured by sintigraphy were not different between eight IBS-D patients and six ontrols. 83 Vasallo and olleagues demonstrated in a study of 10 IBS-D patients that overall oloni transit was aelerated in seven patients (five also had rapid emptying of the proximal olon). 84 Although gut transit measurements are not used speifially to identify IBS bowel habit subgroups, studies have shown that transit times differ between those with IBS-C and IBS-D, as was shown in Horikawa s study. 81 Breath hydrogen tests showed signifiantly shorter small intestinal transit times in IBS-D patients and prolonged transit in IBS-C patients or those with predominant pain and distension. 85 Psyhologial stress may aelerate oroaeal transit in IBS-D, but slow transit time in IBS-C. 85 A sintigraphy study reported signifiantly faster ileoaeal transit in IBS-D than in IBS-C. 86 Interestingly, the reently established Rome III sublassifiation of IBS is based on stool form (and not stool frequeny), whih shows good orrelations with intestinal transit time Stool form has been shown to differentiate IBS bowel habit subgroups best, 90 partiularly IBS with alternating bowel habits (IBS-A), 91 but to date has not been found to be a signifiant preditor of overall IBS symptom severity. Constipated patients with delayed oloni transit may respond to treatment differently from patients with normal transit. 77 A reently published study investigated if olon transit measured by radio-opaque markers in IBS patients orrelated with symptoms in the Rome II diagnosti riteria. 92 In 148 healthy ontrol subjets and 1385 onseutive IBS patients, overall oloni transit time was measured, as were transit times for three segments of the olon (right, left and retosigmoid). Fifty-four perent of IBS patients (12.3% diarrhoea, 60.4% onstipation, 27.7% alternating) and 91% of ontrols had normal overall oloni transit times (,70 h). There were no signifiant differenes in overall oloni transit times between IBS patients and healthy ontrols within eah gender group. However, a small subgroup of healthy men had slower right olon transit ompared with men with IBS, and women with IBS had shorter transit times in the left olon and retosigmoid olon ompared with healthy women. In IBS patients with normal overall oloni transit times, luster analysis revealed heterogeneity in segmental olon transit times in both ontrols and IBS patients. In IBS patients, there was no signifiant differene in linial symptoms between the four different oloni transit luster groups. However, another reent study in a muh smaller group of IBS patients (n = 28) demonstrated that oloni transit was independently assoiated with bowel urgeny. 93 Those with bowel urgeny had shorter oloni transit times, partiularly in the left and retosigmoid olon, than those without urgeny. This is not unexpeted sine the patients with bowel urgeny had IBS-D (n = 11 of 13) and IBS-A (n = 2), while over half of the patients without urgeny had IBS-C (n = 8 of 15) and the remainder had IBS-D (n = 4) and IBS-A (n = 3). In summary, GI transit times (measured using radio-opaque markers) within the normal range do not appear to orrelate well with bowel symptoms in IBS. Gas transit has been measured in IBS patients with abdominal bloating. These patients were found to have impaired reflex ontrol of gut handling of ontents, whih leads to gas retention and symptoms of bloating. However, the orrelation of gas retention with subjetive symptoms was poor. Total gut transit of gas was delayed due to impaired small bowel transit, whereas oloni transit was normal In IBS patients, intraluminal lipids impaired intestinal gas learane, whih was thought to be due to an upregulated reflex inhibition of small bowel transit, without signifiant oloni effets. How preditive are GI transit studies in the evaluation of andidate ompounds aimed at overall IBS symptoms? There are a number of studies whih have evaluated the effet of treatment interventions on GI transit in IBS. The studies whih ompared the effet of the therapeuti agent with baseline measurement or plaebo in IBS patients are shown in table 2. These IBS therapies may prolong or shorten transit times in IBS depending on their mehanism of ation and the IBS bowel habit subgroup in whih they are being evaluated. However, only a few studies reported if the hanges in transit times orrelated with IBS symptoms. Using radio-opaque markers to measure olon transit, the bulking agent alium arbophil was found to prolong olon transit time in IBS-D patients but redue olon transit time in IBS-C patients ompared with a drug-free baseline ondition. The holeystokinin (CCK)-1 antagonist dexloxiglumide slowed asending olon emptying (and aelerated gastri emptying) as measured by sintigraphy but had no effet on overall olon transit time ompared with plaebo in 36 women with IBS-C. 101 Colon transit time 392 Gut 2008;57: doi: /gut

10 Table 2 Correlations between GI transit and symptoms in IBS during ontrolled treatment interventions Drug Dose IBS study subjets Study design Transit methodology Effet on GI transit vs plaebo (or no drug) in IBS Effet on IBS symptoms Bulking agents Bran g 6 3 days 12 Bloated IBS (10 F, 2M) Calium polyarbophil 98 3 g/day 6 8 weeks 26 IBS (14 IBS-D and 12 IBS-C, 19F, 7 M) DB, PC, rossover Sintigraphy Aelerated overall transit NR Open-labelled study (drug vs baseline) Radio-opaque markers Prolonged CT time in IBS-D and redued CT time in IBS-C ompared with baseline Antispasmodi agents Pinaverium bromide mg po tid 6 2 weeks 43 IBS (38F, 5M) PC rossover Radio-opaque markers NS NR Cimetropium bromide mg tid 6 4 weeks 40 IBS (25 IBS-C, 15 IBS-D, 24 F, 16 M) Triyli antidepressants Imipraine mg po qhs and inreased to 100 po qhs 6 5 days 6 IBS-D (3 F, 3 M) Open-labelled study (drug >3 weeks vs baseline) Choleystokinin-1 antagonist Dexloxiglumide mg po tid days 36 IBS-C (all F) DB, randomised, parallel group design Serotonergi agents Alosetron (5-HT3 antagonist) mg po bid 6 8 days 13 non-onstipated IBS (9 F, 4 M) DB, PC, parallel group design Radio-opaque markers Aelerated CT in those with prolonged transit times; no effet in IBS with shorter transit times DB, PC rossover Radio-opaque markers and breath hydrogen test Alosetron (5-HT 3 antagonist) 53 1 mg bid or 4 mg bid 25 non-onstipated IBS DB, PC, 2 dose study (n = 10 in eah drug group and n = 5 in plaebo group) Alosetron (5-HT 3 antagonist) mg bid 6 6 weeks 30 IBS-D (15 F, 15M) Open-labelled study (drug vs baseline) Ondansetron (5-HT 3 antagonist) mg tid 6 4 weeks with 4 week washout between treatment periods 14 IBS-D (6 F, 8 M) DB, PC rossover study Breath hydrogen test and radio-opaque markers Latulose hydrogen breath test Inrease in oroaeal transit time NR Sintigraphy Aelerated GE and slowed asending olon emptying but no effet on overall CT Slowed proximal olon transit but no effet on oroaeal or left olon transit Sintigraphy No signifiant hange in GE, small intestinal transit time or CT time Sintigraphy Slowed oroaeal transit (but not GE) and CT ompared with baseline Trend for greater slowing in CT in F vs. M No effet on oroaeal or small intestinal transit times; no signifiant effet on CT Tegaserod (5-HT 4 agonist) mg po bid 6 1 week 24 IBS-C (all F) DB, PC, parallel group study Sintigraphy Aelerated proximal olon emptying but no effet on GE or overall CT Renzapride (5-HT 4 agonist/5-ht 3 1, 2 or 4 mg/day 6 antagonist) days Renzapride (5-HT 4 agonist/5-ht 3 Plaebo 6 4 weeks, then antagonist) 106 2mgqd6 4 weeks and inreasing to 2 mg bid 6 4 weeks 48 IBS-C (46 F, 2M) DB, PC, dose-ranging, parallel group (n = 12/group) Sintigraphy No signifiant effet on GE and small intestinal transit, but signifiant linear dose response for CT to renzapride and for 4 mg dose vs plaebo 20 IBS-C (12 F, 8 M) SB, PC, esalating dose study Radio-opaque markers Dose of 2 mg bid redued overall CT time and in aeum/ac and DC (n = 11) 5-HT 4 antagonist mg po qd 6 10 days 12 IBS-D (6 F, 6 M) DB, PC rossover study Breath hydrogen test Aelerated oroaeal transit NR CT time negatively orrelated with stool form (Bristol sale, r= 20.60) and stool frequeny (r = 20.56) but not abdominal pain (r = 20.18) NR CT time positively orrelated with omposite sore of bowel funtion (r = 0.43), but no effet on IBS symptoms NR NR NR NR NR Aeleration of CT positively orrelated with improvements in ease of passage (r = 0.54) and stool form (r = 0.59) but not stool frequeny NR Continued Gut 2008;57: doi: /gut

11 Table 2 Continued Effet on GI transit vs plaebo (or no drug) in IBS Effet on IBS symptoms Drug Dose IBS study subjets Study design Transit methodology 40 IBS-D (31 F, 9 M) DB, PC, parallel group doseranging Sintigraphy No effet on GE, small intestinal NR study transit or CT DB, PC, parallel group study Sintigraphy Slowed CT NR Other agents Clonidine (a 2 -adrenergi agonist) , 0.1 or 0.2 mg/day 6 4 weeks VSL#3 (probioti) 109 Oral dose bid weeks 48 IBS with bloating (16 IBS-C, 20 IBS-D, 12 IBS-A; NR Latulose hydrogen breath test Prolonged mouth to aeum transit time DB, randomised 2 session study 1 week apart 21 F, 27 M) 12 IBS with diarrhoea (9 F, 3 M) Otreotide 110 Single 50 mg subutaneous injetion AC, asending olon; bid, twie daily; CT, olon transit; DB, double-blind; DC, desending olon; F, female; GE, gastri emptying; IBS, irritable bowel syndrome; IBS-C, onstipation-predominant irritable bowel syndrome; IBS-D, diarrhoea-predominant irritable bowel syndrome; M, male; NR, data not reported; NS, not signifiant; PC, plaebo-ontrolled; po, orally; qd, four times a day; qhs; 4 hourly; SB, single-blind; tid, three times a day. positively orrelated with omposite sore of bowel funtion (ie, stool frequeny and form, ease of passage and inomplete evauation), but this effet on transit did not translate into effiay for IBS-C symptoms in two, well-designed phase III RCTs. 111 The omposite sore was saled so that the higher sore was assoiated with symptoms more assoiated with a diarrhoea-like pattern (ie, less formed stool, higher number of bowel movements per day and easier passage), and a lower sore was assoiated with a onstipation-like pattern. Renzapride is an investigational drug whih is a ombined 5-HT 4 agonist and 5-HT 3 antagonist, and is urrently being assessed in phase III trials for IBS-C. In a double-blind, plaebo-ontrolled, parallel group sintigraphi study in 48 patients with IBS-C (n = 12/group), there was a signifiant linear dose response for olon transit to renzapride (1, 2 or 4 mg/day) and for 4 mg dose versus plaebo (but no signifiant effet on gastri emptying and small intestinal transit). 105 Aeleration of olon transit positively orrelated with improvements in ease of passage and stool form, but not with stool frequeny. In another study, Tak et al 106 found that renzapride at a dose of 2 mg twie daily produed a statistially signifiant redution in overall oloni transit measured with radio-opaque markers ompared with plaebo. IBS symptoms showed some improvement with renzapride ompared with plaebo but did not reah statistial signifiane although sample sizes were small. The relationships between the hange in olon transit and symptoms were not reported. In summary, based on existing literature, gut transit times do not appear to be altered in the majority of IBS patients, 92 although it seems to differentiate IBS subtypes that is, GI transit is more rapid in those with IBS-D, bowel urgeny and looser stools, while slower transit is more likely to be seen in patients with IBS-C, hard stools and bloating. Sintigraphy has been shown to be reproduible in healthy individuals, but this may be diffiult to assess in IBS patients unless performed in a relatively short spae of time beause IBS patients ommonly transition between subtypes, partiularly IBS-M with mixed bowel habit, and IBS-C. 112 Based on a limited number of studies, hanges in oloni transit in response to drug treatment appear to orrelate onsistently with stool form, and to a lesser extent stool frequeny and ease of stool passage. However, as pointed out earlier, none of these symptoms has been shown to be preditive of IBS severity or HRQoL. Thus, gut transit is a good surrogate marker for stool form and, therefore, may be a useful tool to evaluate drugs whih affet bowel habit in IBS, but is not likely to be an ideal surrogate marker for overall IBS severity, abdominal pain and HRQoL. Strengths and limitations of the two most ommonly used human biomarkers for IBS symptoms Pereptual sensitivity to oloretal distension is assoiated with the presene of IBS, but is probably 394 Gut 2008;57: doi: /gut

12 only moderately orrelated with the presene and severity of abdominal pain (orrelation oeffiients between 0.3 and 0.5) and muh less with global IBS symptoms. It therefore has some preditive validity when evaluating drugs with known analgesi or antihyperalgesi effets. However, similar to GI transit measurements, the value of suh studies to predit the effetiveness of a andidate ompound to redue global IBS symptoms in RCTs, and to form the basis for so-alled go/no-go deisions in IBS drug development is limited. GI transit measures performed in IBS patients probably have the best preditive validity for speifi IBS symptoms, suh as stool form and possible ease of defeation (straining, urgeny), even though orrelation oeffiients have not been reported. It is likely that these tests are also preditive in slow transit onstipation in normalising transit and reduing the symptom of onstipation. GI transit measures are also essential in determining possible undesired side effets of ertain drugs, in partiular onstipation. However, in view of the normal overall oloni transit times seen in the majority of IBS patients without diarrhoea-predominant symptoms, 92 and the poor preditive value of bowel movements, stool form and stool frequeny for global IBS symptom severity and HRQoL, the value of suh studies to predit the effetiveness of a andidate ompound to treat global IBS symptoms as assessed in RCTs is limited. In ontrast, transit studies are presumably highly preditive for symptom relief by a ompound of suh disorders as slow transit onstipation or diarrhoea. WHAT IS THE VALIDITY OF ANIMAL MODELS AND MOLECULAR TARGETS IN THESE MODELS FOR SPECIFIC AND GLOBAL IBS SYMPTOMS? Viseral pain models As disussed in the previous setion, viseral hypersensitivity refleted by enhaned pereption of physiologial signals from the gut or by enhaned pereption of experimental viseral stimuli is ommonly onsidered to play a major role in the pathophysiology of IBS. In ontrast to the relatively straightforward modelling of the objetive measure of GI transit in animal models, there are several problems with viseral pain models. For example, it is urrently not known if this harateristi finding in humans is a refletion of peripheral sensitisation of primary afferent pathways (eg, true viseral afferent hypersensitivity), of entral sensitisation, of entral pain amplifiation or a ombination of these inter-related mehanisms. Furthermore, readouts from the most ommonly used animal models (based on pseudoaffetive reflex responses or omplex, unlearned behaviours) may also show poor orrelations with the human symptom of pain (a subjetive pain experiene whih is highly modulated by ortial influenes), despite their fae validity. There are four basi approahes to modelling pain in animals using quantifiation of (1) segmental (spinal reflexes), (2) omplex, unlearned behaviours mediated by brainstem mehanisms (inluding the viseromotor response to oloretal distension), (3) operant behavioural responses, involving learned operant behaviours 113 and, most reently, (4) brain imaging approahes in awake and unrestrained animals Even though tehnially more demanding, the major advantage of the latter two approahes is the fat that in ontrast to (1) and (2) these approahes provide information about higher order erebral proessing of noieptive information, greatly inreasing their validity as animal models of human pain. The assessment of pseudoaffetive reflex responses (and to a lesser degree of behavioural responses) to the ontrolled distension of different regions of the GI trat and other visera (oesophagus, stomah, urinary bladder, vagina/ervix and olon/retum) has beome the primary readout for the assessment of viseral pain. Sine it was developed by Ness and Gebhart 118 in 1988, the oloretal distension model of viseral pain has been extensively haraterised and has beome the standard tool for the assessment of viseral sensitivity in rodents. When applied to rats at pressures omparable with the one produing pain in humans, oloretal distension is aversive and produes a range of autonomi and behavioural pseudoaffetive reflexes suh as hanges in arterial pressure and heart rate (inreased arterial blood pressure and tahyardia in awake animals, and dereased arterial blood pressure and bradyardia in anaesthetised animals), 118 passive avoidane behaviours (immobility, bak arhing, hind leg spreading) and ontration of the abdominal musulature. This viseromotor response is the most ommonly used index of viseral pain response in rats. It is important to point out that in ontrast to the subjetive experiene of pain in humans whih involves a network of ortial regions, 9 the response is a noieptive brainstem reflex whih shows a good orrelation with the intensity of the stimulus applied to the olon (pressure or volume). 118 It an be reorded as a measure of eletromyographi signals or ounts of the number of spike bursts, but also as manometri hanges in balloon pressure. 119 It has been reently adapted to mie Eletrophysiologial reordings from primary afferent neurons or seond order spinal neurons has also been used as more diret evidene of afferent ativity. The validation of prelinial animal models for the study of viseral hyperalgesia has almost exlusively relied on suh methods measuring the pseudoaffetive response to oloretal distension in experimental models of viseral hypersensitivity. Loal treatment with inflammatory agents or irritants has been repeatedly shown to trigger aute hypersensitivity to distension of different parts of the gut, and these aute models probably have good validity for suh human disorders as aute gastroenteritis or flare of inflammatory bowel disease. Certain interventions, suh as stress in the neonatal period, and the delayed effets of stress, gut inflammation 129 or infestation Gut 2008;57: doi: /gut

13 with parasites, have demonstrated the development of hroni viseral sensitivity, way beyond the time of the interventions, thereby inreasing the fae validity of these models for a hroni disorder, suh as IBS. However, the translation of pseudoaffetive responses to noxious oloretal distension in rodents as an index of viseral hypersensitivity, and abdominal pain in humans, is ompliated by several fators: (1) stimulus intensities (more than twie as high in rodents) and balloon dimensions differ greatly between prelinial and linial appliations of the test; (2) humans and rodents differ in the entral proessing and modulation of noieptive signals from the GI trat; 9 and (3) a signifiant ontribution to the human pain response is fators related to ognitive and emotional dimensions related to the experimental situation. As mentioned earlier, brainstemmediated reflex responses are less likely to apture suh ortial inputs ompared with operant behavioural pain models Novel approahes suh as operant behavioural assays or funtional brain imaging of integrated brain responses to noieptive stimuli may be superior as animal models for viseral hypersensitivity and IBS symptoms. GI transit studies/faeal pellet output In general, the tehniques used to reord GI motility or measure transit in animals provide measurement of gastri emptying, duodenojejunal migrating motor omplex patterns and oloni motility and transit (reviewed in Canilleri et al 135 ). The methods established for evaluation of GI motor and biohemial funtion in vivo inlude luminal pressure reordings whih determine the ontratile pattern in a gut region by measuring the fore via a pressure measurement in the lumen and in liquid-filled balloons. Other tests inlude transit time studies, faeal pellet output, phmetry, and imaging suh as radiography. In a reent artile, Camilleri et al 135 reviewed the animal models that have been validated for the study of the effets of pharmaologial agents on GI motility. Stress, under different forms, an affet gastri emptying, motor patterns, and oloni motility and transit. Stressors suh as restraint, aousti stress, old stress, ombined aousti and old stress, or passive avoidane have been assoiated with delayed gastri emptying. Aute stress exposure an trigger alteration in migrating motor omplex patterns, and has been used to stimulate oloni motility, oloni transit and faeal exretion in rats. In addition, prolonged oloni distension and duodenal infusion of lipids were found to inhibit gastri emptying. Also, inhibition of oloni motility and transit an be indued by pharmaologial agents suh as a 2 -adrenergi and m-opioid reeptor agonists. In ontrast to the readouts from oloretal distension experiments, whih differ greatly between rodents and human subjets, objetive GI transit measurements translate more diretly between prelinial and linial models. Anxiety-like behaviours Extensive epidemiologial evidene has demonstrated the ommon omorbidity of IBS with anxiety disorders and to a lesser degree with depression. 136 More reent evidene has demonstrated an important role of symptom- or illnessrelated anxiety in the symptom severity in IBS, and this is illustrated in fig 2. Furthermore, reent brain imaging studies impliate alterations in ortiolimbi interations in IBS patients. Although peripheral and entral sensitisation may play a role in viseral hypersensitivity in this patient population, a signifiant omponent of the hronially enhaned pereptual response to gut stimuli may be due to altered affetive (symptom-related anxiety) and ognitive modulation (hypervigilane, atastrophising) of viseral sensation. To date, the role of the limbi system in the modulation of viseral noieption in prelinial studies has been indiretly demonstrated in animal models of stress-indued viseral hypersensitivity using neonatal maternal separation stress, neonatal pain 140 or aute/hroni stress in adult animals. In these models, enhaned stress responsiveness was assoiated with inreased anxiety-like behaviours measured as the response to openfield exposure. However, most of the animal models of enhaned viseral noieption assoiated with hemial inflammation or irritation, mehanial distension or infetion have not been haraterised for hanges in anxiety level. There are urrently a number of paradigms that are being used to measure anxiety in animals or detet the anxiolyti ation of different lasses of ompounds, inluding the measurement of exploratory behaviour in response to novelty (plus-maze, openfield, light dark transition), soial behaviours (soial interation, separation-voalisation) or the aousti startle response. 142 Even though the preditive validity of these tests for human forms of anxiety is well established, 143 little is known about the preditive validity of suh measures for IBS symptom-related anxiety or global IBS symptoms. Preditive validity of animal models for IBS symptoms As shown in table 3, for a seletive number of IBS andidate drugs, the preditive validity of prelinial transit models has been relatively good for example, similar effets of 5-HT 3 and 5-HT 4 reeptor modulators of otreotide and of m-opioid reeptor agonists were observed in both prelinial and human experimental models. For the two serotonin reeptor drugs, this preditive validity of prelinial models also applies to overall IBS symptoms, as assessed by a modest benefiial effet on a global end point. In ontrast, the preditive validity of prelinial viseral pain models has been less onsistent. For example, while robust viseroanalgesi and antihyperalgesi effets of the k-opioid agonist fedotozine were seen in several rodent models, effets in human viseral sensitivity testing were largely negative, and 396 Gut 2008;57: doi: /gut

14 Table 3 Effet of andidate ompounds and prelinial and linial read outs Prelinial Clinial Reeptor targeted Compound Motility Viseral analgesia/ antihyperalgesia Anxiety Transit Pereption Brain imaging for viseral pain IBS symptoms (phase II or III) k1-opioid Fedotozine (agonist) Inreased transit after ileus indued by laparotomy or irritation Redued viseral hypersensitivity in a model of oloni irritation Antinoieptive effet on duodenal pain reflexes in rats 148 NR NR Dereased gastri sensitivity to distension in healthy humans 149 Relieved hypersensitivity to oloretal distension in IBS patients 48 NR Relief of abdominal pain and bloating in IBS patients ompared with ontrol. Effet on transit not reported 49 m-opioid Fentanyl (agonist) Dereased GI transit Prevented the sensitisation assoiated with repetitive oloretal distensions in mie Fentanyl attenuated fear-potentiated startle in rats 153 Anxiolyti effet of entral m-opioid agonist on pain-indued anxiety 154 Slowed GI Attenuated the pereption transit 155 of phasi retal distension in IBS patients 40 NR NR 5-HT 3 Alosetron (antagonist) 5-HT 4 Tegaserod (agonist) Redution of oloni Centrally mediated viseral NR Redution of GI Inreased oloni motility 156 antihyperalgesi effet transit 102 ompliane. Lak of true viseroanalgesi effet 51 Enhaned GI motor Redution in viseral NR Aeleration of Generally no evidene for funtion 161 sensitivity GI transit 104 Changes in entral modulation of gut funtion and pain Modulation of entral viseroanalgesi effet proessing of viseral afferent information 165 Global improvement of symptoms in male and female patients with IBS-D 52 Effetive in the treatment of IBS-C symptoms 166 Somatostatin 2,3,5 Otreotide (agonist) CCK-1 Dexloxiglumide (antagonists) NK3 Talnetant (antagonist) Redution of GI transit Viseroanalgesi effet 168 time 167 Aelerated transit Dereased sensitivity to time 173 oloretal distension in rats with inflamed olon 174 Inhibited motility, redued exitatory reflex indued by streth in the olon 175 NR Redution of GI Viseroanalgesi and transit time 169 antihyperalgesi effet during retal distension NR Overall symptom improvement 68 NR + aelerates NR NR NR: Press release, Forest, 1 transit time 101 Otober 2003: therapeuti effet not onfirmed Antihyperalgesi effet 175 Anxiolyti effet NR No effet in healthy NR No effet ompared with plaebo 60 ontrols 61 CCK, holeystokinin; GI, gastrointestinal; IBS, irritable bowel syndrome; IBS-C, onstipation-predominant irritable bowel syndrome; IBS-D, diarrhoea-predominant irritable bowel syndrome; NR, data not reported. Gut 2008;57: doi: /gut

15 results of phase II linial trials were mixed. In the ase of the 5-HT 3 reeptor antagonist alosetron, viseral antihyperalgesi effets were seen in prelinial testing (presumably mediated by entral 5-HT 3 reeptors), no viseroanalgesi effets were seen in human viseral pain tests, while robust effets were seen on linial symptoms. For the 5- HT 4 reeptor agonist tegaserod, viseroanalgesi effets were seen in prelinial models, while no effets were seen in human patient viseral pain assessments. Nevertheless, tegaserod has been shown to be effetive in reduing linial symptoms. For the non-seletive somatostatin reeptor antagonist otreotide, the robust effets seen in prelinial models were repliated in human viseral pain testing, and preliminary results suggest that this may translate into relief of IBS symptoms. 68 Finally, the viseroanalgesi effet of the NK 3 reeptor antagonist talnetant seen in a small number of prelinial studies 175 did not translate into positive results in human viseral pain testing, and no effets on IBS symptoms were observed in well-designed linial trials. Even though in-depth analysis of eah ompound (eg, doses, plasma levels, et.) an yield important information regarding the positive or negative predition by the respetive prelinial model, it is lear that urrent prelinial pain models in rodents, using pseudoaffetive reflex responses as a readout for viseral pain, have generally shown inonsistent preditive validity for IBS symptoms. Emerging strategies hallenges Even though it is oneivable that differenes in dosing in prelinial, human biomarker and linial trials may be responsible in part for the poor orrelations between results obtained in these different tests of IBS drugs, we believe that the problem is more related to the shortomings of the drug development strategy. As disussed in detail above and illustrated in figs 1 and 2A, the traditional drug development strategy taken by the majority of pharmaeutial ompanies is well suited to produe effetive drugs to treat symptoms of onstipation and diarrhoea, but muh less effetive in identifying ompounds early in development with high impat on global IBS symptoms. Considering that abdominal pain is a major preditor of IBS symptom severity and presumably HRQoL impairment, this is learly a limitation of urrent strategies. Fators that have ontributed to the urrent model of drug development inlude a more omplete understanding of the enteri nervous system and its role in the regulation of GI motility and seretion (ompared with the limited understanding of entral mehanisms involved in the modulation of these funtions, and in pain modulation), and, therefore, a primary fous on peripheral drug targets in the GI trat. This has resulted in a fous on prelinial and linial models for modifying GI transit, despite the fat that transit alterations in IBS patients are small and inonsistent, and probably have only a small role in global IBS symptom generation. It has also resulted in the fous on prelinial models of peripheral sensitisation of viseral afferent pathways, using models of questionable validity for funtional GI syndromes, suh as hemial sensitisation, massive inflammation or infestation with parasites, using reflex responses (as opposed to operant behaviours or brain responses) as readouts of abdominal pain, whih do not have a good preditive validity for the omplex human experiene of pain and disomfort. Based on the model summarised in fig 2A, we would like to make the following preditions: A andidate drug will be most effetive if it affets more than one of the omponent symptoms shown in the lower half of the figure, sine eah of these symptoms ontributes at best 10% to global symptoms. In other words, a ompound that redues abdominal pain AND bloating, as well as normalises defeation-related symptoms will be more effetive than one that only affets one of these symptoms. Similarly, a ompound that in addition affets symptom-related anxiety will have the greatest impat on global symptoms and QoL improvement. This obvious onlusion implies that ompounds would be most effetive if they have demonstrated effetiveness on more than one human biomarker and more than one prelinial model. If this assumption is orret, then based on their prelinial effetiveness, drugs suh as somatostatin reeptor agonists, or antagonists for the ortiotropin-releasing fator 1 should have a greater impat on reduing IBS symptom severity than peripherally restrited prokineti agents or pure antidiarrhoeals, even though these drugs may be highly effetive in treating these individual symptoms. The 5-HT 3 reeptor antagonist alosetron, one of the few examples of a ompound that has gone through all the stages of drug development, is in line with this hypothesis: 5-HT 3 reeptor antagonists have been shown in animal models to have anxiolyti, antihyperalgesi and transitreduing properties, 75 even though only one of these properties (GI transit redution) has been demonstrated in human experimental models. However, the drug was assoiated with redued ativity in limbi brain iruits, and this redution orrelated with a redution in global IBS symptoms. 160 It remains to be determined if symptom- or illness-related anxiety, an important fator whih influenes both IBS symptom severity and IBS-related QoL impairment, an be affeted seondarily simply by normalising altered defeation (eg. by reduing urgeny or the sensation of fullness), or if this abnormality reflets a primary entral alteration whih needs to be targeted diretly. In other words, in a head to head omparison, is a ompound suh as Imodium equally effetive in reduing global IBS symptoms as alosetron (suggesting that it is primarily the antidiarrhoeal effet) or is alosetron more effetive, beause of its independent effets on motility, seretion, entral pain 398 Gut 2008;57: doi: /gut

16 Figure 3 Funtional brain imaging in drug disovery. Known agents and proedures are used to develop disease-relevant biomarkers by rossvalidation of data between normal animal and human brain funtion and between animal disease models and human disease. This proess also provides new understanding of brain system funtions and aids pharmaologial target identifiation for drug disovery. The effet of a new ompound on the previously established biomarker an then be assessed, effetive doses evaluated and regulatory evidene olleted in effiient POC studies. These POC studies in small ohorts of patients and healthy volunteers an validate our understanding of a mehanism of ation and give initial data on effiay. Reprodued with permission from Wise and Traey. 182 amplifiation and symptom-related anxiety. Similarly, is the peripherally ating hloride hannel opener lubiprostone, whih is highly effetive in treating hroni onstipation, equally effetive in treating global IBS symptoms? Animal models that mimi more than one symptom of the human syndrome and inlude operational aspets, rather than being limited to reflex responses (eg, have greatest fae validity), should have the greatest preditive validity for effetive IBS drug development. For example, several rodent models have been reported whih demonstrate anxiety-like behaviour, enhaned stress sensitivity, transit abnormalities and viseral hyperalgesia On the other hand, models that only mimi an individual symptom or human biomarker would be expeted to have the least preditive validity for global symptom severity. Based on the earlier disussions about animal models, rodent models of ertain human biomarkers (suh as GI transit, stress sensitivity and possibly anxiety) may show a better orrelation with the human biomarker than measures of abdominal pain, given the profound differenes in CNS mehanisms underlying the human pain/disomfort experiene and the rodent noieptive responses. Sine the majority of IBS symptoms are subjetive human experienes generated by dediated brain iruits onerned with the proessing of viseral homeostati afferent input to the brain, 9 diret imaging of abnormalities of the ativity and onnetivity of these iruits holds signifiant promise as a biomarker for drug development, both in humans and in animal models (for details, see figs 2B and 3). Through the use of brain imaging tehniques suh as positron emission tomography (PET) and funtional MRI (fmri), onsiderable progress has been made in the identifiation and haraterisation of speifi, yet overlapping brain iruits onerned with pain proessing, pain modulation, stress sensitivity, emotional reativity and entral autonomi regulation in humans, 9 as well as in different animal speies, inluding rodents and non-human pri mates. While ontinued researh is needed to onsolidate our understanding of these iruitries, many neural models of sensory and affetive proessing have already emerged as a result of the growing interation between ognitive neurosiene and linial researh. Using these models, markers of disease states have been defined by determining the degree to whih this iruitry is altered in disorders suh as anxiety, depression or hroni pain onditions. Suh markers have subsequently been used as targets for pharmaologial modulation. While pharmaologial brain imaging approahes have been used to find surrogate markers of drug effiay, there are other advantages to using brain imaging in drug disovery and development. For instane, neuroimaging offers the advantage of aquiring objetive measures of regional brain ativity, while traditional behavioural tehniques (ie, subjet report, reation time and auray) are heavily biased by subjetive experiene and mood states. Thus, pharmaologial imaging may require onsiderably fewer subjets (12 15 vs hundreds of subjets) to identify signifiant effets of interest. 186 Also, many ompounds have been pulled from the market due to rare adverse events appearing late in development or during postmarketing surveys, whih may be CNS mediated (an example is nausea). Therefore, it may be important Gut 2008;57: doi: /gut

17 to determine the degree to whih a partiular drug may ontribute to the CNS side effet profile before the drug is released. By haraterising the neural networks involved in the presentation of different side effets suh as nausea, depression, anxiety or suiidal ideation, one an determine if the neuroprofile of a drug inludes the modulation of these iruits, thereby inreasing the risk of suh an adverse event. In short, the ability to identify and haraterise abnormalities in terms of ativity and signalling mehanisms in the CNS (brain, brainstem and spinal ord) of well-defined IBS populations, identifying the orrelates of these abnormalities in the CNS of animal models, evaluating the drug effets and dose requirements on both human and animal targets, and the ability to perform relatively small phase IIa studies in patients to sreen ompounds before taking the most promising ompounds into full-sale linial trials are all potential benefits of this approah (for a summary, see fig 3). In light of these advantages, pharmaologial brain imaging approahes in IBS patients have been reported for alosetron, 160 amitriptyline 187 and tegaserod, 165 and analogous phase IIa studies are urrently underway to evaluate the effets of reeptor antagonists for ortiotropin-releasing fator and substane P. Preliminary reports suggest the feasibility of identifying brain iruit ativation in response to viseral stimuli in rodents SUMMARY AND CONCLUSIONS Despite the onsiderable efforts by the pharmaeutial industry, the suess of IBS drug development has been disappointing to patients, industry and involved investigators alike. As we tried to emphasise in the disussions above, part of this disappointment is related to the inomplete understanding of IBS pathophysiology, and to limitations intrinsi to the traditional drug development strategy taken. Primary fous on targets related to speifi symptoms, and reliane on limited prelinial and linial models has in general been quite suessful in the development of drugs aimed at treating onstipation and diarrhoea, but muh less so for abdominal pain or disomfort, or for global IBS symptoms. We propose that translational (bidiretional) pharmaologial brain imaging approahes in both animal models and humans (in addition to novel linial trial designs) have the potential to improve and aelerate the drug disovery and development proess, inluding the identifiation of more effetive ompounds, and the dramati shortening of the drug development proess. To validate this novel approah will require onsiderable investments both by forwardlooking pharmaeutial ompanies and by publi funding agenies. 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21 Editor s quiz: GI snapshot 175. Sanger GJ. Neurokinin NK1 and NK3 reeptors as targets for drugs to treat gastrointestinal motility disorders and pain. Br J Pharmaol 2004;141: Salome N, Stemmelin J, Cohen C, et al. Seletive blokade of NK2 or NK3 reeptors produes anxiolyti- and antidepressantlike effets in gerbils. Pharmaol Biohem Behav 2006;83: Spooren W, Riemer C, Meltzer H. Opinion: NK3 reeptor antagonists: the next generation of antipsyhotis? Nat Rev Drug Disov 2005;4: Spiegel BM, Strikland A, Chang L. Preditors of patientassessed illness severity in irritable bowel syndrome (IBS). Gastroenterology 2007;132:A Li Q. Cellular and moleular alterations in mie with defiient and redued serotonin transporters. Mol Neurobiol 2006;34: Million M, Wang L, Stenzel-Poore MP, et al. Enhaned pelvi responses to stressors in female CRF-overexpressing mie. Am J Physiol Regul Integr Comp Physiol 2007;292:R Shweinhardt P, Bountra C, Traey I. Pharmaologial fmri in the development of new analgesi ompounds. NMR Biomed 2006;19: Bleeding duodenal uler with a right upper quadrant mass CLINICAL PRESENTATION A 72-year-old woman presented with right upper quadrant abdominal pain and haematemesis. She was known to have gallstones, diabetes and hypertension. On admission, she was afebrile and haemodynamially stable. Clinial examination revealed a tender right upper quadrant egg-sized mass. Her haemoglobin was 9 g/dl and white ell ount /l. Liver funtion tests were deranged (bilirubin 31 mmol/l, alkaline phosphatase (ALP) 833 IU/l, alanine aminotransferase (ALT) 133 IU/l). She underwent an upper gastrointestinal endosopy whih demonstrated an uler at the first part of the duodenum, with a Figure 1 Upper gastrointestinal endosopi image of an uler seen in the duodenum Wise RG, Traey I. The role of fmri in drug disovery. J Magn Reson Imaging 2006;23: Lowe AS, Beeh JS, Williams SC. Small animal, whole brain fmri: innouous and noieptive forepaw stimulation. Neuroimage 2007;35: Martinez V, Wang L, Tahé Y. Proximal olon distension indues Fos expression in the brain and inhibits gastri emptying through apsaiin-sensitive pathways in onsious rats. Brain Res 2006;1086: Matthews PM, Honey GD, Bullmore ET. Appliations of fmri in translational mediine and linial pratie. Nat Rev Neurosi 2006;7: Paulus MP, Feinstein JS, Castillo G, et al. Dose-dependent derease of ativation in bilateral amygdala and insula by lorazepam during emotion proessing. Arh Gen Psyhiatry 2005;62: Morgan V, Pikens D, Gautam S, et al. Amitriptyline redues retal pain related ativation of the anterior ingulate ortex in patients with irritable bowel syndrome. Gut 2005;54: Figure 2 Arterial ontrast phase of the abdominal CT image revealing a spherial mass (white arrows). lot in the base but no ative bleeding (fig 1). In the following days, the gastrointestinal bleeding ontinued. Two subsequent endosopies performed had similar findings. An abdominal CT san revealed a thik-walled gallbladder surrounded by a lot, within whih an unexpeted 1063 m spherial mass was seen on the arterial ontrast phase (fig 2). QUESTION What was the ause of her bleeding? How should it be managed? See page 423 for answers F J Foo, 1 A M Smith, 1 M Sheridan, 2 A A Niholson 2 1 Department of Hepatobiliary Surgery, St James s University Hospital, Leeds, UK; 2 Department of Radiology, St James s University Hospital, Leeds, UK Correspondene to: Mr A M Smith, Department of Hepatobiliary Surgery, St James s University Hospital, Bekett Street, Leeds LS9 7TF, UK; andrewm.smith@ leedsth.nhs.uk Competing interests: None. Gut 2008;57:404. doi: /gut Gut Marh 2008 Vol 57 No 3