STUDY DESIGN

At an initial screening visit, subjects underwent a complete medical history, routine laboratory evaluation, faecal occult blood testing, and pregnancy testing (serum β-HCG levels) for female subjects. Subjects were asked to abstain completely from alcoholic beverages, spicy foods, and smoking for seven days before and during the study and were instructed to discontinue any prior prescription and non-prescription medications (including over the counter preparations) containing any NSAIDs and/or antisecretory or cytoprotective drug therapies.

Approximately one week after screening, eligible subjects underwent a baseline evaluation of intestinal permeability (see below). A total of 39 subjects were randomised to receive one of four treatment regimens for seven days: placebo; rofecoxib 25 mg once daily in the morning; rofecoxib 50 mg once daily in the morning; or indomethacin 50 mg (two 25 mg tablets) three times daily. Each subject received all four seven day treatment regimens in a predefined balanced sequence and each treatment regimen was separated by a washout period of 7–10 days. Study medication was dispensed in individually marked boxes (three boxes per treatment period) containing a seven day supply of study drugs. During the study, subjects completed diaries to record compliance with study medication. After the initial dose of study medication, subjects were reminded by electronic pager to take subsequent doses throughout the entire treatment period. Subjects who discontinued prior to completion of the protocol were replaced with subjects assigned to the same treatment order to preserve statistical balance.

MEASUREMENT OF INTESTINAL PERMEABILITY

Intestinal permeability was determined by measuring urinary recovery of ⁵¹CrEDTA (a marker of intercellular permeation) and l-rhamnose (a marker of non-carrier mediated transcellular permeation).8 The urinary excretion ratio of⁵¹CrEDTA/l-rhamnose was chosen as the primary index of intestinal permeability because the differential excretion ratio of these agents is largely unaffected by premucosal and postmucosal factors.25 We chose ⁵¹CrEDTA rather than lactulose as a paracellular marker in this study to maximise the sensitivity of the small intestinal permeability test. Previous studies have shown that differential urinary excretion of⁵¹CrEDTA/l-rhamnose is more sensitive than lactulose/l-rhamnose (when administered as an iso-osmolar test solution) in patients with coeliac disease26 or taking NSAIDs.8

Intestinal permeability was measured before treatment in each of the four treatment periods and on day 7 at the conclusion of each treatment period. For each permeability measurement, fasting subjects drank a test solution (100 ml) containing ⁵¹CrEDTA (30 μCi, Amersham International, Amersham, UK), l-rhamnose (1.0 g), and d-xylose (0.5 g) one hour after consuming the study medication (except at baseline when no study medication was taken). The radiation dose received from a 30 mCi dose of ⁵¹CrEDTA taken orally is less than 0.04 millisieverts (effective dose equivalent). d-Xylose (0.5 g) was included in the test solution as a marker of intestinal absorption, as some,27 ,28 but not all,12 studies have suggested that NSAIDs impair absorption. Urine samples were subsequently collected over the next five hours, during which subjects continued fasting for the first two hours. Urine was collected in a bottle containing 1 ml (10% w/v) of sodium ethylmercurithiosalicylate (thiomersal, Sigma Chemical Co., St Louis, Missouri, USA) as a preservative. Urine volume was measured on completion of the test, and a 5 ml sample was removed for measurement of radioactivity using an LKB 1281 gamma counter. Urinary concentrations of l-rhamnose and d-xylose were determined using a modified thin layer chromatographic technique which involved scanning densitometry with an arabinose internal standard to overcome errors of application, as described previously.29 The percentage of the⁵¹CrEDTA and l-rhamnose dose was determined in each urine sample, from which the ratio of the percentage excreted⁵¹CrEDTA/ l-rhamnose was calculated.

TOLERABILITY ASSESSMENTS

Adverse experiences were monitored throughout the study and were defined as any unfavourable or unintended change in structure, function, or chemistry of the body, or a worsening of a pre-existing condition. Investigators evaluated all clinical adverse experiences in terms of intensity (mild, moderate, or severe), duration, seriousness, outcome, and relation to test drugs.

STATISTICAL ANALYSIS

Data obtained from a previously published study8indicated that the ⁵¹CrEDTA/ l-rhamnose ratio is log normally distributed. Therefore, between treatment comparisons were made by computing the geometric mean ratio of⁵¹CrEDTA/l-rhamnose values. Based on pre- and post-treatment data from this study,8 a limit for comparability to placebo was prespecified; rofecoxib would be considered equivalent to placebo if the between treatment ratio of the⁵¹CrEDTA/l-rhamnose values was <1.37. The sample size was chosen to provide 95% power to prove that the upper limit of a 95% two sided confidence interval was below this value. The primary statistical analysis was based on a per protocol assessment of data. However, if a subject completed periods 1 and 2 and then discontinued the study, the subject was still included in the analysis, even though the subject failed to complete the entire study.

Analysis of covariance suitable for a four period crossover design (with treatment, period, subject, carryover, and log baseline as covariate factors) was used to analyse differences in the log of⁵¹CrEDTA/l-rhamnose ratio on day 7 compared with baseline values and to assess the statistical significance of differences in the log of ⁵¹CrEDTA/l-rhamnose ratio among the four study groups.

STUDY POPULATION AT ENTRY

A total of 39 subjects were enrolled in the study. Mean age was 28 years (range 18–39 years) and 54% were male. Subjects were assigned to one of four treatment sequence groups and received, in a predefined balanced sequence (for first order carryover effect), the four study drugs (placebo, rofecoxib 25 mg, rofecoxib 50 mg, and indomethacin 150 mg). The treatment sequence groups were well matched with respect to baseline characteristics, including age, sex, race, caffeine use, secondary diagnoses, prior medication use, and baseline intestinal permeability. All subjects had a negative faecal occult blood test. Nine subjects violated the study protocol by consuming alcohol and were therefore excluded from the per protocol analysis (alcohol may affect intestinal permeability). In addition, isolated visits from five other subjects were also excluded because of protocol violations; all decisions regarding protocol violations were made before unblinding the treatment code. Three subjects discontinued prior to study completion of all four treatment groups; one subject developed an adverse experience during treatment. At the time of discontinuation, one subject was taking placebo, and the other two subjects discontinued during a washout period following rofecoxib 50 mg. A comparison of the effects of the four study drugs on intestinal permeability (per protocol) was made on the following numbers of patients: 29 in the placebo group, 29 in the rofecoxib 25 mg group, 27 in the rofecoxib 50 mg group, and 26 in the indomethacin 150 mg group.

EFFECT ON INTESTINAL PERMEABILITY

Pretreatment differential urinary ratios of⁵¹CrEDTA/l-rhamnose were comparable across the four treatment regimens, with mean values of 0.036, 0.036, 0.035, and 0.038 for placebo, rofecoxib 25 mg, rofecoxib 50 mg, and indomethacin 150 mg, respectively (fig 1). Following seven days of administration of indomethacin, the permeability of the intestine increased significantly, as indicated by an increase in the ratio of⁵¹CrEDTA/l-rhamnose from baseline values of 0.038 to 0.055 on day 7 (p<0.05). In contrast, the permeability of the intestine following seven days of administration of placebo, rofecoxib 25 mg, or rofecoxib 50 mg was not significantly different from baseline values (fig 1). As illustrated in fig 2, day 7 to baseline ratios for⁵¹CrEDTA/l-rhamnose averaged 0.97, 0.80, 0.98, and 1.53 for placebo, rofecoxib 25 mg, rofecoxib 50 mg, and indomethacin 150 mg groups, respectively. Analysis of intent to treat data, which included all patients enrolled in the study, revealed similar findings to those seen with the per protocol analysis.

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Figure 1

Five hour geometric mean urinary excretion ratios for⁵¹CrEDTA/l-rhamnose at baseline and on day 7 for subjects who received placebo, rofecoxib 25 mg, rofecoxib 50 mg, and indomethacin 150 mg (per protocol analysis). Indomethacin 150 mg significantly increased the ⁵¹CrEDTA/l-rhamnose ratio compared with baseline values, whereas placebo, rofecoxib 25 mg, and rofecoxib 50 mg did not. *p<0.05 versus baseline values

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Figure 2

Day 7 to baseline geometric mean ratios (with 95% confidence intervals) for five hour urinary excretion of⁵¹CrEDTA/l-rhamnose in subjects who received placebo, rofecoxib 25 mg, rofecoxib 50 mg, and indomethacin 150 mg (per protocol analysis).

Comparison of day 7 to baseline ratios for⁵¹CrEDTA/l-rhamnnose revealed no significant differences between placebo and either the rofecoxib 25 mg or rofecoxib 50 mg groups (table 1); the upper limit on the 95% confidence interval was <1.37, the prespecified definition of clinical equivalence. In contrast, day 7 to baseline ratio for ⁵¹CrEDTA/l-rhamnnose was significantly higher in subjects who received indomethacin compared with either the placebo or rofecoxib groups (p⩽0.001).

Absorption of d-xylose was not significantly different between the four treatment groups (p=0.382). Day 7 to baseline mean ratios for urinary excretion of d-xylose were 0.95, 1.05, 0.99, and 0.87 for placebo, rofecoxib 25 mg, rofecoxib 50 mg, and indomethacin 150 mg groups, respectively. None of these ratios was significantly different from 1.0 (data not shown).

SAFETY

No serious clinical or laboratory adverse experiences occurred during the study. Non-serious adverse effects were reported more often in subjects receiving indomethacin 150 mg (83.3%) than in subjects who received placebo, rofecoxib 25 mg, or rofecoxib 50 mg (53.8%, 64.1%, and 60.5%, respectively). Overall, the incidence of clinical adverse experiences was similar across treatment groups except for dizziness and headache which were reported more often in indomethacin treated subjects than in placebo or rofecoxib treated subjects. Analysis of the occurrence of predefined NSAID-type gastrointestinal symptoms (acid reflux, dyspepsia, epigastric discomfort, heartburn, nausea, and vomiting) showed that overall, gastrointestinal symptoms occurred most frequently with indomethacin treatment (30.6%), had a similar frequency in the rofecoxib 50 mg and placebo groups (18.4% and 12.8%, respectively), and were least frequent in the rofecoxib 25 mg group (5.1%). One subject discontinued the study due to an adverse event which occurred during treatment with placebo (angioedema). Two subjects discontinued because of adverse events during washout periods (one due to pyelonephritis, one due to upper respiratory infection with fever, abdominal pain, and diarrhoea). None of the reported adverse experiences necessitated unblinding of treatment.