Gut, 1992, 33, 753-758 753
Effects ofacetorphan, an enkephalinase inhibitor, on experimental and acute diarrhoea
Ph Baumer, E Danquechin Dorval, J Bertrand, J M Vetel, J C Schwartz, J M Lecomte
Abstract and reduce the discomfort. Their beneficial Acetorphan is an orally active inhibitor of effect, however, has become a matter of contro- enkephalinase (EC 3.4.24.11) with antidiar- versy in view of their limited efficacy and poten- rhoeal activity in rodents apparently through tial side effects. These drugs disrupt forward protection of endogenous enkephalins and a propulsive motility, increase capacitance of the purely antisecretory mechanism. Its antidiar- gut and delay passage of fluid through the rhoeal activity in man was assessed in an intestine allowing, in turn, more time for net experimental model of cathartic induced absorption of water and electrolytes to occur.68 secretory diarrhoea as well as in acute diar- The fact that their antidiarrhoeal activity is rhoea. of presumed infectious origin. In six always associated with an antimotility effect is healthy volunteers receiving castor oil and probably responsible for some of the major pretreated with acetorphan or placebo in a gastrointestinal side effects - that is, pooling of crossover controlled trial, the drug signific- fluid in the distended bowel lumen and enhance- antly decreased the number and weight of ment of bacterial colonisation.8"'3 This has led to stools passed during 24 hours. About 200 the idea that antidiarrhoeal drugs with a purely outpatients with severe acute diarrhoea (more mucosal antisecretory activity, should lead to a than five stools per day) were included in a therapeutic improvement.3 Two main strategies randomised double blind study of acetorphan were used to reach this aim: design weak a2- against placebo. The significant antidiarrhoeal adrenergic receptor blocks devoid of cardiovas- activity of acetorphan was established using a cular effects'45 and enkephalin like pentapep- variety of criteria: (i) the duration of both tides'"'8 but, so far, the resulting investigational diarrhoea and treatment were diminished; (ii) drugs have not gained therapeutic application. no acetorphan treated patient withdrew from Another strategy was recently suggested the study whereas five dropped out because of by the observations that inhibition of enkepha- worsening in the placebo group; (iii) the fre- linase (membrane metalloendopeptidase, EC quency ofsymptoms associated with diarrhoea 3.4.24.11) delays the inactivation of endogenous - for example, abdominal pain or distension, enkephalins in the brain which, in turn, results nausea and anorexia - remaining after two in a series of naloxone reversible biological weeks was nearly halved; (iv) using visual responses.'9 21 The peptidase is present all along analogue scales acetorphan treatment was the gastrointestinal tract2223 and its inhibition found more effective than placebo by both delays the inactivation of exogenous enkephalins investigators and patients. There was statistic- by the guinea pig ileum.2>26 Moreover, enkepha- ally no significant difference between acetor- linase inhibitors exert potent, naloxone revers- phan and placebo in respect of side effects, ible antidiarrhoeal activity in the rodent castor Laboratoire Bioprojet, particularly constipation, which often accom- oil induced model of diarrhoea.2728 Interestingly, Marnes la Coquette, panies the antidiarrhoeal activity of mu opioid this action was not associated with any detectable France this difference is attribut- antitransit effect,27 30 which clearly differentiates Ph Baumer receptor agonists J M Lecomte able to the lack of antipropulsive activity of this class of compounds from mu opiate receptor acetorphan in man. The efficacy and tolerance agonists.3'-36 Centre Hospitalier of acetorphan suggest that enkephalinase In the present study we have assessed the Universitaire Trousseau, Tours, France inhibition may represent a novel therapeutic antidiarrhoeal activity of the enkephalinase E Danquechin Dorval approach for the symptomatic management of inhibitor acetorphan in man in two randomised J Bertrand acute secretory diarrhoea without impairing placebo controlled double blind trials. The first trial on six healthy volunteers Centre Hospitalier intestinal transit. was a crossover Regional, Le Mans, receiving castor oil, a validated model of experi- France mental 'secretory' diarrhoea in man,'6 37 whereas J M Vetel It is generally admitted that acute infectious the second one was a parallel study on two groups Unite de Neurobiologie et diarrhoea is accompanied with an abnormality of ofabout 100 patients each attending their general Pharmacologie, Centre intestinal water and electrolyte transport. ' As a practitioners for severe acute diarrhoea of pre- Paul Broca de l'Institut with replacement of sumed infectious origin. National de la Sante et de consequence rehydration la Recherche Medicale, electrolytes are accepted as the most important Paris, France therapeutic measures. Antimicrobial agents are J C Schwartz also of value in diarrhoea caused by identified Methods Correspondence to: intestinal protozoa but are essentially ineffective Dr J C Schwartz, Unite de Neurobiologie et in the treatment of viral enteric infections which STUDY I: CASTOR OIL INDUCED DIARRHOEA Pharmacologie de l'Institut constitute most cases of acute diarrhoea.245 In National de la Sante et de la Recherche Medicale, Centre addition, mu opiate receptor agonists such as SUBJECTS Paul Broca, 2ter rue d'Alesia, opium derived preparations, diphenoxylate or Six healthy volunteers, three men, three women, 75014 Paris, France. are used to shorten the dura- aged 21 (3) years, with no reported digestive Accepted for publication loperamide widely 16 September 1991 tion of the disease, relieve a variety of symptoms abnormality, particularly no intestinal transit 754 Baumer, Danquechin Dorval, Bertrand, Vetel, Schwartz, Lecomte
disorder, and not receiving any drug treatment, TREATMENT were selected. Their mean (SEM) body weights Identical capsules containing 100 mg either were 61-0 (3.7) kg (68 (4) kg for men, 54 (3) kg accetorphan or lactose (placebo) were supplied. for women). Written informed consent was Each patient was assigned a sequential study obtained and the study was approved by the local number and corresponding bottle containing 60 Ethical Committee. capsules. Treatment had been previously randomised using a random number table with groups of four. STUDY DESIGN Patients were given two capsules at the start of The study was a double blind, randomised, the study and were instructed to take one capsule placebo controlled crossover study ofthe preven- after each unformed bowel movement. Drug tion by acetorphan ofdiarrhoea elicited by a fixed administration was continued until recovery, dose of castor oil. The study comprised two defined as the disappearance of any unformed sessions, performed at one week intervals, stool, or for a maximum of 10 days. The remain- during which the subjects received placebo and ing capsules were returned to the investigators acetorphan capsules in a randomised order. and were counted to determine the actual Neither the subjects nor the clinicians were number of capsules ingested. No other new aware of the nature of treatment. treatment was allowed during the trial, except In the morning ofeach session, generally after for paracetamol. one spontaneous stool, each subject absorbed five capsules and, 45 minutes later, 30 g castor oil (French Pharmacopoeia). The time and weight STUDY DESIGN of each stool was then determined during the The patients were seen twice by clinicians not following 24 hours. informed of the nature oftreatments: for the first The capsules for the placebo session contained time at inclusion (visit 1) and 10-14 days later 200 mg lactose and were undiscernible from (visit 2). For each patient the clinicians had a case those containing acetorphan for which a dosage report form to complete at visits 1 and 2. It close to 10 mg/kg of body weight was selected. included a specific questionnaire with a report of The actual acetorphan dosage received by the clinical examinations. The clinicians remained subjects was 11 1 (4.2) mg/kg, a dosage eliciting blind all over the study. more than 50% inhibitions of plasma enkepha- Patients were requested to keep a diary in linase for at least six hours. which the following information was reported: All side effects were recorded during the 24 time, number and characters of bowel move- hour experimental period. ments, number of capsules taken, possible adverse affects, and their global evaluation of the treatment visual scales 0= STATISTICAL ANALYSIS using analogic (from The evaluation criteria were the cumulative no efficacy, to 100=excellent efficacy). Patients weights of stools, the total number of stools were asked to bring back the diary and remaining during the experimental 24 hour period and the capsules at visit 2 and both were checked by the delay between administration of the cathartic clinicians. and the first stool passage. Data were expressed The duration of diarrhoea was defined as the as means (SEM) and analysed by analysis of time from the first dose of the trial drug (visit 1) variance and Student's t test for matched series, to resolution of diarrhoea, that is the disappear- using the PCSM program (Deltasoft). Differ- ance of unformed stools. ences were considered significant at a 5% prob- The tolerance was assessed using the spon- level. taneously described symptoms and the visual ability analogue scales (from 0 to 100- that is, from bad to excellent tolerance). For each adverse effect, STUDY II: ACUTE DIARRHOEA the duration and the severity grade (mild, The study was a double blind, randomised, moderate or severe) were recorded. placebo controlled parallel study.
STATISTICAL ANALYSIS PATIENTS Data were analysed before disclosure of the Adult outpatients, aged at least 18 years, living randomisation. Qualitative data were compared in the Parisian district, were eligible if they using X) and Fischer tests. Quantitative data attended their physician for acute diarrhoea of were compared using two tailed Student's t test. presumed infectious origin having started less Duration of diarrhoea was analysed by the than five days before. Patients not included were actuarial method and the log rank test. Differ- those with chronic or iatrogenic diarrhoea, with ences were considered significant at a 5% prob- dysenteric syndrome characterised by fecal ability level. Values were expressed as means blood loss and those who had been given any new (SEM). drug treatment within one week of admission. No attempt at identifying the bacterial pathogen was made as fecal cultures performed in similar Results cases generally lead to negative results in a large majority of cases (over 95% in ref 38). Patients STUDY I: CASTOR OIL INDUCED DIARRHOEA were asked to participate after a full explanation Administration of castor oil during the placebo of the aims and methods of the study, and all session was followed by bowel movements gave informed consent. resulting in the passage of 3-7 (0.3) stools over Effects ofacetorphan, an enkephalinase inhibitor, on experimental andacute diarrhoea 755
TABLE I Effects ofacetorphan on castor oil induced diarrhoea Placebo Acetorphan Difference Total stool weight (g/24 h) 672 (76) 426 (83)* -(37 (9))% Number of stools (/24 h) 3-7 (0.3) 1-8 (0O2)t -(49 (6))% First stool delay (h) 4-8 (1.1) 7.0 (09)NS +(113 (59))% *p<0.01; t=0002; NS: not-significant (p=0 142).
the next 24 hours, the first one occurring after 0) 4-8 (1 1) hours and the total stool weight being 672 I and - (76) g (Table Fig 1). .) Among the six subjects, two had taken acetor- 0) phan during the first session and four during the -5 second session of the trial. The effect of the 0 treatment on total stool weights was independent 0 of the order of treatment (p=0188) and there Q) was no interaction between order and treatment (p=0 540). There was a significant reduction E (-37%) of stool weight under the action of C- acetorphan (p=0.009), all six subjects con- sidered individually having a lower stool weight during the acetorphan session than during the placebo session (Fig 2). In addition the mean number of stools during the trial was reduced by 50% (p<0-002) during the acetorphan session as compared with the placebo session. There was also a tendency toward an extended delay Placebo Acetorphan (+ 113%) between administration ofthe cathartic Figure 2: Effect ofacetorphan on castor oil induced diarrhoea and passage of the first stool but the difference in six healthy volunteers: individual stool weights (gl24 h). failed to reach significance (Table I). All subjects reported a feeling of nausea and discomfort during the two sessions which can, patients, 96 received acetorphan and 98 placebo. therefore, be attributed to the intake of castor There were no significant differences in the oil but no other side effect was reported. various characteristics of the groups on admis- sion (Table II).
STUDY II: ACUTE DIARRHOEA DURATION OF DIARRHOEA PATIENTS Five patients withdrew before the study's com- Initially, 199 patients from the Parisian area were pletion because ofworsening or lack ofimprove- admitted into the study. They were included by ment of diarrhoea; all five patients had received 28 investigators during a six month winter placebo. At visit 2, 30 patients had still unformed period. There was no exclusion because of stools during the preceding 24 hours: seven in erroneous admission. Five patients were the acetorphan group (7.4%) and 23 (23.5%) in excluded: two did not take the treatment and the placebo group (p<0002). These 35 patients three were not followed up. Filling in of diaries could not be included in the analysis of the was found correct for the 194 patients who duration of diarrhoea when expressed as mean. completed the study; particularly the number of The mean duration of diarrhoea was, however, capsules taken according to the diary was con- shorter in the acetorphan group, 3.4 (0 1) days, sistent with number deduced from capsules than in the placebo group, 4-4 (0.2) days (p= returned at visit 2. Among the 194 analysed TABLE II Characteristics ofthe two groups ofpatients with acute diarrhoea at inclusion Patients Acetorphan Placebo ,) Included (n) 96 102 Excluded/lost from view (n) 0/1 2/2 -C Analysed (n) 95 98 Age (yr)* 40.2 (1-7) 38-6(1 5) (range) (19-89) (18-84) .' Men (% of patients) 42 52 Weight(kg)* 65-1(1-1) 65.8(1-2) Figure 1: Effect of 0 Duration ofdiarrhoea before inclusion acetorphan on castor oil U) (days)* 1-7 (0-1) 1.6 (0-1) induced diarrhoea in six 0) Stools during the preceding 24 h (n)* 5.3 (0.2) 4-9 (0.2) healthy volunteers: mean Anal burning (%) 50 48 cumulative stool weights. (U Spontaneous abdominal pain (%) 88 90 The same subjects received Nausea (%) 76t 62 E Anorexia 82 77 or in two ._ (%) acetorphan placebo Pain on abdominal palpation (%) 85 85 separate sessions. Abdominal distension (%) 75 75 Comparison between the two Fever (%) 33 39 sessions by analysis of variance and Student's t test 4 8 12 16 20 24 *Values given as means (SEM). There was no significant for matched series: difference between treatment groups in any of the characteristics p Figure 3: Actuarial curves of Because, in these animals, there was a com- diarrhoea in 193 patients plete inhibition of the antidiarrhoeal effect of with acute diarrhoea treated with acetorphan or placebo. 0) enkephalinase inhibitors by naloxone, an opiate Comparison between the two cc 80 receptor blocker, it appears likely that the effect treatment groups by log rank of these agents results from protection of endo- test: p opioid receptor agonists and enkephalinase in- antimotility effects of acetorphan in rodents2728 hibitors, both devoid of antimotility effects, and healthy volunteers52 was confirmed in the were recently found active on this model. In fact present trial by the fact that the occurrence of a similar mechanism may mediate the final action constipation did not significantly differ in the two of both classes of compounds as delta receptor groups. Constipation is consistently induced by agonists exert antisecretory effects triggered mu receptor agonists.5354 In agreement, in a either directly on the intestinal mucosa or double blind randomised trial against loperamide indirectly via receptors43 of the submucosal in acute diarrhoea, acetorphan was found at least plexus.""47 Acetorphan was shown to reduce, in a as effective as the mu receptor agonist as an anti- naloxone reversible manner, the net fluxes of diarrhoeal agent but to induce significantly less water, Na+ and K+ in the jejunum of cholera reactive constipation (Roge, Baumer, Berard, toxin treated dogs4849 as well as to block inflam- Schwartz and Lecomte, submitted). From these matory fluid secretion in the cat gall bladder.50 differences, it can be reasonably expected that The second trial established the activity of side effects associated with antimotility drugs - acetorphan in acute diarrhoea, as judged from a for example, facilitation of bacterial colonisa- large variety of criteria. For instance, all five tion, invasion by Shigella, extension of the patients who spontaneously withdrew from the period of excretion of bacterial pathogens and study before completion had received placebo. precipitation ofileus and bowel dilatation ('toxic At the end of the trial - that is, after 13-14 days, megacolon')9' should not occur with enkepha- a significantly higher proportion of patients linase inhibitors. receiving placebo still had diarrhoea than when treated with acetorphan (23% against 7%, 1 Powell DW. Muscle or mucosa: the site of action of antidiar- rheal opiates? Gastroenterology 1981; 80: 406-8. p 25 Aoki K, Kajiwara M, Oka T. The role of bestatin-sensitive water and electrolytes by hydroxy fatty acids. Gastro- amino-peptidase, angiotensin converting enzyme and enterology 1973; 64: 81-8. thiorphan-sensitive 'enkephalinase' in the potency of 41 Stewart JJ, Gaginella TS, Bass P. Actions of ricinoleic acid and enkephalins in the guinea-pig ileum. JpnJ7 Pharmacol 1984; structurally related fatty acids on the gastrointestinal tract. I. 36: 59-65. Effects on smooth muscle contractility in vitro. ] Pharmacol 26 Van Amsterdam JGC, Van Bunren KJH, Krichaart MJ, Exp Ther 1975; 195: 347-54. Zuiderveld OP, Tims RPJH. Effect of inhibitors of 42 Gaginella TS, Stewart JJ, Olsen WA, Bass P. Actions of enkephalin degradation in the isolated guinea-pig ileum. ricinoleic acid and structurally relatd fatty acids on the Life Sci 1988; 43: 1529-36. gastrointestinal tract. II. Effects on water and electrolvte 27 Marcais-Collado H, Uchida G, Costentin J, Schwartz JC, absorption in vitro. J Pharmacol Exp Ther 1975; 195: 355- Lecomte JM. Naloxone-reversible antidiarrheal effects of 61. enkephalinase inhibitors. Eur]f Pharmacol 1987; 144: 125- 43 Mihara S, North RA. Opioids increase potassium conductance 32. in guinea pig submucous neurones by activating delta- 28 Lecomte JM, Costentin J, Vlaiculescu A, Chaillet P, Marcais- receptors. Br]7 Pharmacol 1986; 88: 315-22. Collado H, Llorens-Cortes C, et al. Pharmacological proper- 44 Dobbins JW, Racusen L, Binder HJ. Effect of D-alanine ties of acetorphan, a parenterally active 'enkephalinase' methionine enkephalinamide on ion transport in rabbit inhibitor.J Pharmacol Exp Ther 1986; 237: 937-44. ileum. ] Clin Invest 1980; 66: 19-28. 29 Chipkin RE, Berger JG, Billard W, Iorio LC, Chapman R, 45 Kachur JF, Miller RJ. Characterization of the opiate receptor Barnett A. Pharmacology of SCH 34826, an orally active in the guinea pig ileal mucosa. EurJ Pharmacol 1982; 81: enkephalinase inhibitor analgesic. J Pharmacol Exp Ther 177-83. 1988; 245: 829-37. 46 Kachur JF, Miller RJ, Field M. Control of guinea pig 30 Bueno L, Fioramonti J. Enkephalins, other endogenous intestinal electrolyte secretion by a delta-opiate receptor. opioids and colonic motility in dog and man. Gastroenterol Proc Natl Acad Sci USA 1982; 77: 2753-6. Clin Biol 1987; 11: 69B-76B. 47 Hautefeuille M, Brantl V, Dumontier AM, Desieux JF. In 31 Basilisco G, Bozzani A, Cambon G, Recchia M, Quatrini M, vitro effects of b-casomorphins on ion transport in rabbit Conte D, et al. Effect of loperamide and naloxone on mouth- ileum. Am]f Phvsiol 1986; 250 (Gastrointest Liver Phvsiol to-caecum transit time evaluated by lactulose hydrogen 13): G692-7. breath test. Gut 1985; 26: 700-3. 48 Primi MP. Contr6le central neuropeptidergique et hormonal 32 Basilisco G, Camboni G, Bozzani A, Paravicini M, Bianchi PA. des secr6tions intestinales chez le chien. Institut National Oral naloxone antagonizes loperamide-induced delay of Polytechnique de Toulouse, France, 1986. These de orocaecal transit. Dig Dis Sci 1987; 32: 829-32. Doctorat. 33 Kachel G, Ruppin H, Hagel J, Barina W, Meinhard T, 49 Primi MP, Bueno L, Baumer Ph, Berard H, Schwartz JC, Domschke W. Human intestinal motor activity and Lecomte JM. Action anti-secretoire intestinale d'un transport: effects of a synthetic opiate. Gastroenterology inhibiteur de l'enkephalinase, l'acetorphan: demonstration 1986; 90: 85-93. in vivo chez le chien. Gastroenterol Clin Biol 1991; 15: A73. 34 Staniforth DH. Effects of drugs on orocaecal transit time 50 Jivegard L, Pollard H, Moreau J, Schwartz JC, Thune A, assessed by the lactalose/breath hydrogen method. Eur J Svanvik J. Naloxone-reversible inhibition of gall bladder Clin Pharmacol 1987; 33: 55-8. mucosal fluid secretion in experimental cholecvstitis in the 35 Pressman JH, Hofmann AF, Witztum KF, Gertler SL, cat by acetorphan, an enkephalinase inhibitor. Clin Sci 1989; Steinbach JH, Stokes K, et al. Limitations of indirect 77: 49-54. methods of estimating small bowel transit in man. Dig Dis 51 Knisely JS, Beardsley PM, Aceto MD, Balster RL, Harris LS. Sci 1987; 32: 689-99. Assessment of the abuse potential of acetorphan, an 36 O'Brien JD, Thompson DG, McIntyre A, Burnham WR, enkephalinase inhibitor. DrugAlcohol Depend 1989; 23: 143- Walker E. Effect of codeine and loperamide on upper 51. intestinal transit and absorption in normal subjects and 52 Bergmann JF, Chaussade S, Couturier D, Baumer Ph, patients with posturogotomy diarrhoea. Gut 1988; 29: 312- Schwartz JC, Lecomte JM. Effects of ocetorphan, an 8. antidiarrheal enkephalinase inhibitor, on orocaecol and 37 Rubenstein E, Hofmann LM, Gourzis JT, Nolan RB. Com- colonic transit times in healthy volunteers. Aliment Phar- parison of a commercial bowel evacuant and castor oil with macol Therap (in press). enemas in barium enema radiology. Curr Ther Res 1986; 39: 53 Schuermans V, Lommel RV, Dom J, Brugmans J. 858-65. Loperamide (R18553), a novel type of antidiarrheal agent. 38 Gorbach SL. Infectious diarrhea. In: Sleisenger MH, Placebo-controlled comparison of the constipating activity Fordtran JS, eds. Gastrointestinal diseases. 4th ed. and safety of loperamide, diphenoxylate and codeine in Philadelphia: Saunders WB, 1989: 1191-232. normal volunteers. Arzneimittelforschung 1974; 24: 1653-7. 39 Ammon HV, Thomas PJ, Phillips S. Effects of oleic and 54 Johnson PC, Ericsson CD, Dupont HL, Moreau DR, Bitsura ricinoleic acids on net jejunal water and electrolyte move- JAM, Wood LV. Comparison of loperamide with bismuth ment. J Clin Invest 1974; 53: 374-9. subsalicvlate for the treatment of acute travelers diarrhea. 40 Bright-Asare P, Binder HJ. Stimulation of colonic secretion of JAMA i986; 255: 757-60.