Linaclotide Improves Abdominal Pain and Bowel Habits in a Phase IIb Study of Patients With Irritable Bowel Syndrome With Constipation

GASTROENTEROLOGY 2010;139:1877 1886 CLINICAL Linaclotide Improves Abdominal Pain and Bowel Habits in a Phase IIb Study of Patients With Irritable Bowel Syndrome With Constipation JEFFREY M. JOHNSTON,* CAROLINE B. KURTZ,* JAMES E. MacDOUGALL,* BERNARD J. LAVINS,* MARK G. CURRIE,* DONALD A. FITCH,* CHRIS O DEA,* MOLLIE BAIRD,* and ANTHONY J. LEMBO *Ironwood Pharmaceuticals, Inc, Cambridge, Massachusetts; and Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts BACKGROUND & AIMS: Linaclotide, a minimally absorbed, 14-amino acid peptide agonist of guanylate cyclase-c, has shown benefit in a proof-of-concept study for the treatment of patients with irritable bowel syndrome (IBS) with constipation (IBS-C). We assessed the efficacy and safety of linaclotide at a daily dose range of 75 600 g in IBS-C. METHODS: We performed a randomized, double-blind, multicenter, placebo-controlled study of 420 patients with IBS-C given oral linaclotide at doses of 75, 150, 300, or 600 g or placebo once daily for 12 weeks. End points included change from baseline in daily bowel habits, daily abdominal symptoms, and weekly global assessments, in addition to responder criteria. RE- SULTS: All doses of linaclotide significantly improved bowel habits, including frequency of spontaneous bowel movements and complete spontaneous bowel movements (primary end point), severity of straining, and stool consistency. Abdominal pain was significantly reduced from baseline, compared with placebo; mean changes in abdominal pain (assessed on a 5-point scale) from baseline were 0.71, 0.71, 0.90, and 0.86 for linaclotide doses of 75, 150, 300, and 600 g, respectively, compared with 0.49 for placebo. Likewise, most doses of linaclotide significantly improved other abdominal symptoms, including discomfort and bloating, and global measures of IBS-C compared with placebo. Effects were observed within the first week and were sustained throughout 12 weeks of treatment. Except for diarrhea, the incidence of adverse events was similar between placebo and linaclotide groups. CONCLUSIONS: Linaclotide, across a wide range of doses, significantly improved symptoms of IBS-C, including abdominal pain and bowel symptoms. Diarrhea was the only dose-dependent adverse event and was usually of mild or moderate severity. Keywords: Functional Bowel; Clinical Trial; Colon; Complete Spontaneous Bowel Movement. Irritable bowel syndrome (IBS) is reported in approximately 10% 15% of the population in North America. 1 IBS is a chronic functional gastrointestinal disorder characterized by abdominal pain and discomfort associated with altered bowel habits. 2 Patients often experience additional symptoms such as bloating, sensation of incomplete evacuation, and straining. The symptoms of IBS not only adversely affect a person s health-related quality of life (QOL) 3 but also place a significant financial burden on society because of reduced work productivity and an over 50% increase in the use of health-related resources. 4 Among the IBS subtypes, which are categorized based on predominant bowel symptom (ie, constipation, diarrhea, or mixed), IBS with constipation (IBS-C) accounts for approximately one-third of all IBS cases and occurs more commonly in women than men. To date in the United States, only tegaserod and lubiprostone have been approved for the treatment of IBS-C. 5 Tegaserod, a 5-HT 4 partial agonist, was removed from the market in 2007 because of concerns regarding increased cardiovascular adverse events (AEs) in patients receiving the medication. Other therapies that are used but not approved by the US Food and Drug Administration to treat IBS-C, such as laxatives, are primarily directed toward the patient s constipation symptoms. Given the unmet medical need and limited options to manage the multiple symptoms of IBS-C, alternative therapies are needed. Linaclotide (MD-1100), a novel 14-amino acid peptide that is minimally absorbed, has been shown to reduce measures of intestinal pain 6 and increase fluid secretion and transit in animal models. 7 The actions of linaclotide are mediated through its binding to the guanylate cyclase-c (GC-C) receptor on the luminal surface of intes- Abbreviations used in this paper: AE, adverse event; ANCOVA, analysis of covariance; BM, bowel movements; cgmp, cyclic guanosine monophosphate; ECG, electrocardiogram; IBS, irritable bowel syndrome; IBS-C, irritable bowel syndrome with constipation; IBS-SSS, irritable bowel syndrome symptom severity scale; CSBM, complete spontaneous bowel movement; GC-C, guanylate cyclase-c; IVRS, interactive voice response system; QOL, quality of life; SAE, serious adverse events; SBM, spontaneous bowel movement. 2010 by the AGA Institute 0016-5085/$36.00 doi:10.1053/j.gastro.2010.08.041

1878 JOHNSTON ET AL GASTROENTEROLOGY Vol. 139, No. 6 tinal enterocytes. 8 Activating GC-C receptors increases intracellular cyclic guanosine monophosphate (cgmp), triggering a signal transduction cascade that results in the activation of the cystic fibrosis transmembrane conductance regulator. 9 This activation causes secretion of chloride and bicarbonate into the intestinal lumen resulting in an increase in fluid secretion and acceleration of colonic transit. 9 Linaclotide also has been shown to ameliorate visceral hypersensitivity 10 through a mechanism believed to be dependent on GC-C/cGMP. 6 Evidence indicates that cgmp fluxes out of the epithelial cells into the serosal space to act on the submucosal afferent pain fibers to suppress the nerve firing rate. 11 In healthy human volunteers, linaclotide was generally well tolerated at single oral doses of up to 3000 g and multiple oral doses (7 days of once daily treatment) of up to 1000 g. 12 Linaclotide was undetectable in the serum of healthy volunteers at therapeutic doses (doses of up to 1000 g). In women with IBS-C, 100 and 1000 g once daily of linaclotide for 5 days improved stool frequency, stool consistency, straining, and time to first bowel movement. Colonic transit was increased with the 1000 g once daily dose of linaclotide. 13 The objectives of this study were to assess the efficacy, safety, and dose response of linaclotide administered at 75, 150, 300, and 600 g once daily for 12 weeks in patients with IBS-C. Patients and Methods Study Design A randomized, double-blind, parallel-group, placebo-controlled, phase IIb study was conducted at 92 clinical centers in the United States and Canada between March 2007 (first patient randomized) and February 2008 (last patient completed). The study was performed in accordance with the Declaration of Helsinki and US21 Code of Federal Regulations and registered on clinicaltrials.gov (NCT00460811). Written informed consent was obtained from each patient prior to participation in the study. The study protocol and informed consent were approved by an institutional review board from each center participating in the study. After giving informed consent, patients entered an initial screening period of up to 28 days. During the screening period, routine blood tests (ie, hematology, chemistry, urinalysis, and pregnancy test if appropriate) were obtained, and laxatives and other ineligible medications (eg, anticholinergic agents, opioids) were discontinued for at least 3 days prior to patient entry into the pretreatment phase of the study. Patients meeting the inclusion and exclusion criteria then entered a 2-week pretreatment baseline period and used an interactive voice response system (IVRS) to provide their daily assessment of bowel symptoms and IBS symptoms. Patients eligible for the 12-week treatment period were randomized to receive 75, 150, 300, or 600 g gelatin capsules or placebo orally once daily before the first meal of the day. After completing the treatment period, patients immediately entered into a 2-week posttreatment period. Patients continued to make daily calls to the IVRS throughout the treatment and posttreatment periods. Study visits occurred during the screening period, pretreatment baseline period (beginning at day 14), treatment period (days 1, 15, 29, 57, and 85), and posttreatment period (2 weeks after completion of treatment period or day 99). All personnel involved in the design and implementation of the study remained blinded to study medication until study conclusion. Study Participants Patients were men and women 18 years of age or older who met the Rome II criteria for IBS 14 and who reported having fewer than 3 spontaneous bowel movements (SBMs) per week and 1 or more of the following symptoms for at least 12 weeks in the 12 months preceding entry into the study: (1) straining during 25% of bowel movements (BMs); (2) lumpy or hard stools during 25% of BMs; or (3) sensation of incomplete evacuation during 25% of BMs. In addition, during the 2-week pretreatment baseline period, patients were required to report a mean score of 2.0 for the daily assessment of nonmenstrual abdominal pain or abdominal discomfort (ie, at least mild on a 5-point scale ranging from 1 none to 5 very severe), as well as a mean of 3 complete SBMs (CSBMs) and 6 SBMs per week. Rescue and Concomitant Medications Rescue medications were allowed for severe constipation (ie, at least 72 hours after the patient s previous BM) including oral bisacodyl up to 15 mg daily, bisacodyl suppository, or sodium phosphate enema. Rescue medication was allowed during the pretreatment baseline period, provided that there were no more than 2 uses and that none was used during the 3 days before the first dose of study medication. Patients were instructed to contact the investigator prior to taking a rescue medication for severe constipation, and all rescue medication usage was recorded during the patient s daily call to the IVRS. Patients were prohibited from taking prescription or over-the-counter IBS and constipation medications (except in rescue medication situations described above). Patients on a stable, continuous regimen of fiber therapy for at least 30 days prior to the pretreatment baseline period were allowed to continue, provided that they continued at a constant dose throughout the study. Antidepressants were allowed provided that the patient had been on a stable dose for at least 30 days before entry into the study and that no dose adjustments were planned during the study. Assessments All efficacy assessments were recorded by phone using an IVRS. Daily assessments included time of taking

December 2010 LINACLOTIDE IN IBS-C 1879 the study medication, use and timing of rescue medications, frequency and timing of BMs as well as the following characteristics of each BM: (1) sensation of complete bowel emptying (yes/no); (2) stool consistency using the 7-point Bristol Stool Form Scale 15 ; and (3) degree of straining using a 5-point ordinal scale (1 not at all, 2 a little bit, 3 a moderate amount, 4 a great deal, 5 an extreme amount). A BM was determined to be a SBM if no laxative, enema, or suppository was taken in the preceding 24 hours and a CSBM if the patient indicated that the SBM was associated with a sensation of complete bowel emptying. Symptoms of abdominal pain, abdominal discomfort, and bloating were measured daily using a 5-point ordinal severity scale (1 none, 2 mild, 3 moderate, 4 severe, 5 very severe). In addition to daily symptom assessments, the following weekly assessments were obtained: (1) adequate relief of IBS symptoms (yes/no); (2) global relief of IBS using a 7-point balanced scale (1 completely relieved, 2 considerably relieved, 3 somewhat relieved, 4 unchanged, 5 somewhat worse, 6 considerably worse, 7 as bad as I can imagine); (3) IBS symptom severity; and (4) constipation severity (both using the 5-point ordinal severity scale: 1 none, 2 mild, 3 moderate, 4 severe, 5 very severe). The following assessments were measured at the beginning and end of treatment: (1) IBS symptom severity scale (IBS-SSS; 0 500 based on 5 equally weighted questions that measure severity of abdominal pain, frequency of abdominal pain, severity of abdominal distention, dissatisfaction with bowel habits, and interference with QOL) 16 and (2) IBS-QOL (34 items each rated on a 5-point Likert scale assessing the degree to which IBS interferes with patient s QOL). 17 Finally, the following were assessed only at the end of treatment: (1) overall satisfaction with the study medication s ability to relieve IBS (1 not at all satisfied, 2 a little satisfied, 3 moderately satisfied, 4 quite satisfied, 5 very satisfied) and (2) likelihood that the patient would continue taking study medication (1 not at all, 2 a little likely, 3 moderately likely, 4 quite likely, 5 very likely). The investigator recorded all patient-reported AEs and serious AEs (SAEs) and assessed their relationship to study treatment as well as severity and outcome. Other safety evaluations included physical examinations; electrocardiogram (ECG) recordings; measurement of vital signs; and standard laboratory tests including hematology, clinical chemistry, urinalysis, and pregnancy test. Table 1. Summary of Baseline Patient Characteristics: ITT Population All (N 419) Placebo (n 85) 75 g (n 79) 150 g (n 82) Linaclotide 300 g (n 84) 600 g (n 89) Demographic data Age, y, mean (range) 44.4 (18 72) 44.3 (21 65) 42.3 (18 69) 45.6 (24 70) 46.0 (21 72) 43.7 (19 69) Sex: female, n (%) 386 (92) 78 (92) 74 (94) 78 (95) 77 (92) 79 (89) Race, n (%) African American 71 (17) 16 (19) 8 (10) 18 (22) 12 (14) 17 (19) White 335 (80) 67 (79) 68 (86) 62 (76) 69 (82) 69 (78) BMI, mean (range) 27.5 (14 58) 27.0 (16 45) 27.3 (20 46) 28.2 (19 58) 26.9 (14 56) 27.8 (17 47) Pretreatment baseline, mean (SD) CSBMs/wk 0.3 (0.6) 0.3 (0.5) 0.4 (0.7) 0.2 (0.5) 0.2 (0.5) 0.3 (0.6) SBMs/wk 3.0 (2.2) 3.1 (1.8) 3.2 (2.1) 2.5 (1.7) 3.2 (2.8) 3.1 (2.6) Stool consistency a 2.3 (0.9) 2.3 (0.9) 2.3 (0.8) 2.2 (0.9) 2.4 (0.9) 2.4 (1.0) Straining b 3.5 (0.8) 3.4 (0.8) 3.4 (0.8) 3.5 (0.9) 3.4 (0.7) 3.6 (0.7) Abdominal pain c 3.0 (0.7) 3.0 (0.7) 3.1 (0.8) 3.1 (0.8) 2.9 (0.7) 3.1 (0.7) Abdominal discomfort c 3.2 (0.6) 3.2 (0.6) 3.3 (0.7) 3.3 (0.8) 3.1 (0.5) 3.3 (0.6) Bloating c 3.4 (0.7) 3.3 (0.7) 3.5 (0.8) 3.6 (0.8) 3.3 (0.6) 3.5 (0.7) IBS severity c 3.6 (0.7) 3.6 (0.7) 3.6 (0.7) 3.7 (0.8) 3.6 (0.5) 3.7 (0.6) Constipation severity c 3.7 (0.7) 3.6 (0.7) 3.7 (0.7) 3.8 (0.8) 3.5 (0.7) 3.8 (0.7) IBS-SSS d 357 (83) 355 (83) 360 (78) 361 (91) 346 (78) 364 (85) IBS-QOL e 55.6 (22.7) 53.6 (22.1) 59.5 (23.5) 53.3 (26.2) 58.4 (19.0) 53.5 (22.3) SBM, spontaneous bowel movement; CSBM, complete SBM; SD, standard deviation; BMI, body mass index; IBS, irritable bowel syndrome; IBS-QOL, IBS quality of life; IBS-SSS, IBS symptom severity scale; ITT, intent-to-treat; QOL, quality of life; wk, week. a Stool consistency was assessed using the 7-point Bristol Stool Forms Scale. b Straining was assessed on a 5-point ordinal scale (1 not at all, 2 a little bit, 3 a moderate amount, 4 a great deal, 5 an extreme amount). c Abdominal pain, abdominal discomfort, bloating, IBS severity, and constipation severity were assessed using a 5-point ordinal severity scale (1 none, 2 mild, 3 moderate, 4 severe, 5 very severe). d IBS-SSS: IBS symptom severity scale: range 0 500 ( 175, mild; 175 300, moderate; 300, severe). e IBS-QOL: range 0 100 (lower score indicates worse QOL).

1880 JOHNSTON ET AL GASTROENTEROLOGY Vol. 139, No. 6 Statistical Analysis Patients were randomized in approximately equal proportions to 1 of the 5 treatment groups using central randomization balanced within each site using a block size of 5. A computer-generated randomization schedule was generated before the study by a statistician not otherwise associated with the trial. The sponsor, all randomized patients, and study center personnel were blinded to treatment group allocation. The planned sample size was based on the results of a previous phase IIa linaclotide study 13 in which the change from baseline in the mean CSBM frequency was 1.2 for placebo and 3.55 for linaclotide with a common standard deviation estimate of 3.90. Under these assumptions, with the intention of providing approximately 95% power at a 5% significance level, and allowing for a 12.5% dropout rate, enrollment of at least 80 patients per treatment arm was required. The primary efficacy end point was the change from the 2-week pretreatment baseline in normalized weekly CSBM rate (number of CSBMs per week) during the 12-week treatment period. CSBMs were also analyzed using a 75% responder definition with a CSBM responder defined as a patient who, for 75% of the treatment weeks, had a weekly CSBM rate 3 and an increase 1 relative to baseline (SBM responders were similarly defined). An adequate relief responder was defined as a patient who reported adequate relief for at least 75% of the treatment period weeks. A global relief responder was defined as a patient who responded somewhat relieved, considerably relieved, or completely relieved for all 12 weeks or considerably relieved or completely relieved for at least 6 of the 12 weeks. Change from baseline end points was analyzed using an analysis of covariance (ANCOVA) model, with fixed effect terms for treatment group and geographical region and the corresponding baseline value as a covariate. Because of the potential of the data not meeting parametric normality assumptions, the percent of abdominal painfree days and percent of days with mild or no abdominal pain end points were rank transformed prior to the ANCOVA. 18 The mean changes from baseline are the least squares means from the ANCOVA model. Responder end points were analyzed using a Cochran Mantel Haenszel test controlling for geographical region. Missing data were not imputed nor was a last observation carried forward approach utilized for continuous end points. For responder end points, patients were considered to be nonresponders for the weeks in which their data were missing. US geographic region (Northeast, Southeast, Midwest, Southwest, and West) was used as a factor in Table 2. Bowel Function Efficacy End Points Bowel function Placebo (n 85) Linaclotide 75 g (n 79) 150 g (n 82) 300 g (n 84) 600 g (n 89) Complete spontaneous BMs Mean number CSBM per wk 1.47 3.55 2.79 3.93 3.10 Change from baseline a,b 1.01 2.90 c 2.49 d 3.61 c 2.68 c CSBM 24 hours first dose, (%) b 12.9 36.7 c 28.0 e 32.1 d 27.0 e CSBM 75% responder, (%) f 11.8 25.3 e 19.5 32.1 d 23.6 e Spontaneous BMs Mean number SBM per wk 4.55 7.69 6.67 7.98 8.59 Change from baseline a,b 1.68 4.62 c 4.36 c 4.97 c 5.64 c SBM 24 hrs first dose, (%) b 45.9 74.7 c 64.6 e 77.4 c 71.9 c SBM 75% responder, (%) f 29.4 54.4 d 39.0 65.5 c 52.8 d Stool consistency Mean BSFS score 2.90 4.27 4.13 4.67 4.62 Change from baseline a,b 0.56 1.91 c 1.80 c 2.28 c 2.20 c % SBMs without hard or lumpy stools 60.2 78.9 c 76.8 c 85.3 c 83.4 c (BSFS 3) b Straining Mean straining score 2.8 2.2 2.3 2.0 2.1 Change from baseline a,b 0.71 1.23 c 1.20 c 1.48 c 1.35 c % SBM without significant straining (score 3) b 42.9 64.7 c 62.5 c 72.0 c 65.6 c BM, bowel movement; SBM, spontaneous bowel movement; CSBM, complete SBM; BSFS, Bristol Stool Form Scale. a Changes from baseline are the least-squares means from the analysis of covariance model. b P values were based on a comparison of each linaclotide group vs the placebo group using the analysis of covariance model. c P.001, unadjusted comparisons. d P.01, unadjusted comparisons. e P.05, unadjusted comparisons. f P values were based on a comparison of each linaclotide group vs the placebo group using the Cochran Mantel Haenszel test. Overall 75% responder: patients who reported an SBM or CSBM rate 3 and an increase 1 from their pretreatment weekly rate for at least 9 out of 12 treatment weeks.

December 2010 LINACLOTIDE IN IBS-C 1881 the analyses as opposed to trial center because of the potential for low enrollment at the majority of trial centers. All P values are based on 2-sided tests. In this phase IIb dose range-finding study, the reported P values were not adjusted for multiple comparisons. A logistic regression was performed to assess the relationship between baseline severity as measured by IBS-SSS (on a 0 500 point scale) and adequate relief, controlling for treatment. Results Patient Disposition, Demographics, and Baseline Characteristics Nine hundred ninety-six (996) patients signed consent forms, of whom 254 were screen failures and 322 were pretreatment failures. Four hundred twenty patients were randomized to 1 of the 5 treatment arms of the study and received study medication (safety population, N 420). All but 1 of these patients had at least 1 assessment of the primary efficacy end point (intent-totreat population, n 419). A total of 83 (19.8%) patients did not complete the study: 25 (6.0%) because of AEs, 4 (1.0%) because of noncompliance, 34 (8.1%) who withdrew consent, 1 (0.2%) at the investigator s request, and 19 (4.5%) who were lost to follow-up (Supplementary Figure 1). The treatment groups were generally well balanced with respect to demographics and baseline IBS-C characteristics (Table 1). Bowel Function The primary end point, mean change from pretreatment baseline to the 12-week treatment period in CSBM rate (number of CSBMs per week), was 2.90, 2.49, 3.61, and 2.68 for linaclotide doses of 75, 150, 300, and 600 g, respectively, compared with 1.01 for placebo (P.01 for each of the linaclotide doses) (Table 2). Weekly CSBM rates are shown in Figure 1A. The mean changes from baseline in SBM rate were 4.62, 4.36, 4.97, and 5.64 for linaclotide doses of 75, 150, 300, and 600 g, respectively, compared with 1.68 for placebo (P.001 for each of the linaclotide doses) (Table 2). Significant differences between each of the linaclotide doses and placebo were evident within 24 hours of starting treatment (Table 2). Compared with placebo, the percentage of patients who were CSBM and SBM 75% responders was significantly greater for all doses of linaclotide except the 150 g dose group (Table 2). Stool consistency and straining were also significantly improved with all doses of linaclotide compared with placebo (P.001 for all doses) (Table 2). Additionally, linaclotide-treated patients reported a greater percentage of SBMs with a straining score of 3 (ie, less than moderate straining, P.001 for all doses) as well as a greater percentage of SBMs with a stool consistency score of 3 (ie, not hard or lumpy, P.001 for all doses) compared with patients receiving placebo (Table 2). Figure 1. Weekly mean change from baseline in CSBM rate (A) and abdominal pain (B) for linaclotide 75, 150, 300, and 600 g and placebo. Abdominal Symptoms Mean decreases in abdominal pain scores from the 2-week pretreatment baseline to the 12-week treatment period were statistically significantly greater for all doses of linaclotide, with mean changes of 0.71, 0.71, 0.90, and 0.86 for linaclotide doses of 75, 150, 300, and 600 g, respectively, compared with 0.49 for placebo (P.05 for all doses; Figure 2A). Absolute mean changes from baseline to the 12-week treatment period in percentage of days with an abdominal pain score 2 (ie, mild or no pain) were 32.4%, 31.1%, 38.1%, and 38.7% for linaclotide doses of 75, 150, 300, and 600 g, respectively, compared with 22.7% for placebo (P.01 for 300 and 600 g doses and P.05 for the 75- g dose). Improvement in abdominal pain with linaclotide occurred within the first week of treatment and was sustained throughout the 12 weeks of treatment (Figure 1B). Upon cessation of dosing in linaclotide-treated patients during the posttreatment period, abdominal pain scores increased, approaching levels of pain found in the placebo group. Patients who reported severe or very severe abdominal pain (ie, abdominal pain score of 4 or 5) for at least

1882 JOHNSTON ET AL GASTROENTEROLOGY Vol. 139, No. 6 Figure 2. Mean abdominal pain (A), abdominal pain in severe patients (B), abdominal discomfort (C), and bloating (D) ratings during the 2-week pretreatment baseline and 12-week treatment periods for linaclotide 75, 150, 300, and 600 g and placebo. 50% of the days during the baseline period (107 [25.5%] of the 419 intent-to-treat patients) demonstrated the greatest reduction in abdominal pain during the treatment period compared with placebo (Figure 2B). In these patients, absolute mean changes from baseline in the percentage of days during the treatment period with a pain score 2 (ie, mild or no pain) were 35.1%, 34.5%, 52.9%, and 52.4% for linaclotide doses of 75, 150, 300, and 600 g, respectively, compared with 10.3% for placebo (P.01 for 300 and 600 g doses and P.05 for the 75 and 150 g doses). The mean reductions in abdominal discomfort and bloating from baseline to the 12-week treatment period were statistically significantly greater than placebo for all doses except 75 g (abdominal discomfort) and 150 g (bloating) (Figure 2C and D). As with abdominal pain, improvements in abdominal discomfort and bloating with linaclotide were present during first week of dosing and were generally sustained throughout the 12 weeks of treatment (results not shown). Global Measures of IBS Symptoms A greater percentage of patients in each of the linaclotide dose groups were adequate relief responders (ie, reported adequate relief for at least 9 of the 12 treatment weeks) than in the placebo group (38.0%, 32.9%, 51.2%, and 42.7% for linaclotide 75, 150, 300, and 600 g, respectively, compared with 22.4% for placebo; P.01 for 300 and 600 g and P.05 for 75 g). Each of the linaclotide doses significantly improved the mean change from baseline in global relief of IBS, IBS severity, and constipation severity (Table 3). For each of the linaclotide doses except the 75- g dose, a significantly greater percentage of patients were global relief responders compared with placebo (Figure 3). The mean change from baseline as well as the percentage of patients with a 50% reduction in the IBS-SSS score compared with placebo was statistically significantly higher for the 75-, 300-, and 600- g dose groups compared with placebo (Table 3). Among the individual component questions of the IBS-SSS, the severity and frequency of abdominal pain, dissatisfaction with bowel habits, and the impact of IBS on life in general showed statistically significant improvement in the linaclotide 300- and 600- g dose groups (Table 3). Whereas the differences from placebo at the end of the 12-week treatment period were not statistically significant for any dose, IBS-QOL scores showed clinically meaningful improvement (ie, 14 points) 19 in over one-third of patients in all treatment groups (37% of linaclotide-treated patients had a change from baseline in the overall IBS-QOL score of 14). Patient-reported satisfaction with the study medication s ability to relieve IBS was significantly greater for all doses of linaclotide except 75 g compared with placebo. The likelihood to continue on study medication score was statistically significantly greater for each of the linaclotide doses compared with placebo. Safety Table 4 shows the most common AEs (occurring in at least 3% of all linaclotide patients). Diarrhea, the most common and only dose-dependent AE, was reported by 11.4%, 12.2%, 16.5%, and 18.0% of patients in the 75, 150, 300, and 600 g linaclotide dose groups, respectively, compared with 1.2% in the placebo group. Most of these diarrhea AEs were graded by the investigator as mild or moderate in intensity; 9 were graded as severe (2 in the 75 g group, 2 in the 150 g group, 1 in

December 2010 LINACLOTIDE IN IBS-C 1883 Table 3. Global Outcome Efficacy End Points Linaclotide Placebo 75 g 150 g 300 g 600 g Assessments of relief No. 85 79 82 84 89 Adequate relief responder, (%) a 22.4 38.0 b 32.9 51.2 c 42.7 d Global relief responder, (%) a 29.4 44.3 43.9 b 58.3 c 53.9 d Assessments of severity IBS severity No. 79 72 74 75 83 Mean e 3.03 2.65 b 2.75 b 2.35 c 2.54 c Constipation severity No. 82 77 80 81 89 Mean e 2.95 2.55 c 2.63 d 2.15 c 2.43 c IBS-SSS No. 85 79 82 84 86 Mean (0 500) e 274 226 d 248 204 c 210 c Reduction 50% (%) a 20.0 38.0 b 25.6 35.7 b 34.8 b Subscales Severity of abd pain (0 100) e 49.2 33.9 d 40.6 30.4 c 29.9 c Days with abd pain ( 10) e 47.7 36.2 b 41.8 36.0 b 35.3 b Severity of abd distention (0 100) e 53.2 49.9 53.2 45.0 46.7 Dissatisfaction with BMs (0 100) e 71.9 62.1 b 65.3 51.6 c 55.9 d Interference with your life (0 100) e 49.5 44.4 47.9 36.6 d 41.0 b Quality of life No. 85 79 82 84 87 IBS-QOL (mean) e 68.1 71.5 67.5 72.4 67.6 IBS-QOL responder (%) a 36.5 34.2 41.5 36.9 36.0 Satisfaction with medication No. 54 61 65 61 64 Treatment satisfaction (mean) f 2.78 3.21 3.28 b 3.74 c 3.34 b Treatment continuation (mean) f 2.94 3.54 b 3.48 b 3.95 c 3.70 d abd, abdominal; IBS, irritable bowel syndrome; IBS-SSS, IBS symptom severity scale; IBS-QOL, IBS quality of life. a P values were based on a comparison of each linaclotide group vs the placebo group using the Cochran Mantel Haenszel test. Adequate relief responder: a patient who reported adequate relief for at least 9 of 12 of the treatment period weeks. Global relief responder: a patient who responded somewhat relieved, considerably relieved, or completely relieved for all 12 weeks or considerably relieved or completely relieved for at least 6 of the 12 weeks of the treatment period. IBS-QOL responder: a patient with a clinically meaningful change (ie, 14 points). b P.05, unadjusted comparisons. c P.001, unadjusted comparisons. d P.01, unadjusted comparisons. e P values were based on the change from baseline of each linaclotide group vs the placebo group using the analysis of covariance model. f P values were based on a comparison between each linaclotide group vs the placebo group using the analysis of covariance model. Satisfaction with the study medication s ability to relieve IBS was assessed using a 5-point scale. Likelihood to continue on study medication was assessed using a 5-point scale. the 300- g group, and 4 in the 600- g group). Among the patients who discontinued treatment because of AEs, discontinuation was because of diarrhea in 0 of 2 placebo patients, 2 of 4 patients in the 75 g group, 4 of 6 patients in the 150 g group, 1 of 3 patients in the 300 g group, and 6 of 10 patients in the 600 g group. In linaclotide patients reporting diarrhea, the median number of days from the first dose of study drug to the initial onset of a diarrhea AE was 4 days. No dehydration, electrolyte changes, or other adverse clinical sequelae from diarrhea were reported. One patient in the 300 g linaclotide group experienced an SAE during the treatment period. This patient was hospitalized for fecal impaction resulting in a temporary withholding of the study medication. The patient recovered and completed the study. The event was reported by the investigator to be unrelated to the study medication. No clinically significant differences in ECG, laboratory values, vital signs, or physical examination abnormalities were found between the linaclotide and placebo groups. Discussion The results of this large, 12-week, placebo-controlled trial show that across a wide range of doses linaclotide statistically significantly improved multiple symptoms of IBS-C, including abdominal pain, bloating, and constipation. Linaclotide also resulted in statistically significant improvements in several global measure-

1884 JOHNSTON ET AL GASTROENTEROLOGY Vol. 139, No. 6 Figure 3. Percentage of patients who were (A) adequate relief responders during the 12-week treatment periods and (B) global relief responders during the 12-week treatment period for linaclotide 75, 150, 300, and 600 g and placebo. ments. Linaclotide s effects occurred within the first week of therapy and were sustained for the entire 3-month duration of this study. Thus, linaclotide resulted in sustained effects in the treatment of a common condition for which few therapies are currently available. Abdominal pain is an important feature of IBS and is the most important factor of overall illness severity, health care utilization, and health-related QOL. 20 In this study, linaclotide reduced abdominal pain as demonstrated by the decrease in pain score (significant for all doses) and the percent of days with a pain score of none or mild (ie, 2) (significant for the 75, 300, and 600 g doses). In addition, the effect of linaclotide on pain was particularly apparent in patients with severe abdominal pain, defined as patients reporting severe or very severe abdominal pain at least 50% of the days during the 2-week baseline period, with a mean increase in the percent of days with mild or no pain ranging from 34.5% to 52.9% for the linaclotide dose groups compared with 10.3% for placebo. Treatment for this subgroup of patients is particularly challenging because they typically have more severe IBS, utilize more health care resources, experience greater impairment in their daily activities, and require more intensive treatment. 20 Improvement in the primary end point, change from baseline to the 12-week treatment period in CSBM rate, was statistically significantly greater in all linaclotide dose groups compared with the placebo group. Because fewer than 3 CSBMs per week is considered constipated, requiring that patients have at least 3 CSBMs per week and improvement over baseline of at least 1 CSBM per week to be considered a responder reflects normalization of bowel function. The responder rate using this definition was 2- to 3-fold greater for linaclotide patients than for placebo patients, representing an approximate 10-fold improvement over baseline (eg, 0.3 to 3.0 CSBMs per week). Because CSBM integrates SBM with the qualitative symptom of complete emp- Table 4. Summary of Adverse Events Experienced by 3% of Linaclotide Patients: Safety Population MedDRA preferred term Placebo (n 85) All (n 335) 75 g (n 79) Linaclotide, n (%) a 150 g (n 82) 300 g (n 85) 600 g (n 89) Diarrhea 1 (1.2) 49 (14.6) 9 (11.4) 10 (12.2) 14 (16.5) 16 (18.0) Abdominal pain 3 (3.5) 18 (5.4) 4 (5.1) 3 (3.7) 4 (4.7) 7 (7.9) Urinary tract infection 2 (2.4) 14 (4.2) 7 (8.9) 1 (1.2) 5 (5.9) 1 (1.1) Nausea 5 (5.9) 13 (3.9) 1 (1.3) 8 (9.8) 1 (1.2) 3 (3.4) Nasopharyngitis 5 (5.9) 11 (3.3) 3 (3.8) 6 (7.3) 1 (1.2) 1 (1.1) Headache 6 (7.1) 11 (3.3) 1 (1.3) 3 (3.7) 5 (5.9) 2 (2.2) Upper respiratory tract infection 3 (3.5) 11 (3.3) 0 (0.0) 2 (2.4) 4 (4.7) 5 (5.6) NOTE. AEs are presented by MedDRA preferred term. A patient with multiple occurrences of the same AE was counted only once for that preferred term. AEs were those reported at any time during the study or within 24 hours after the study medication was stopped. AE, adverse events; MedDRA, Medical Dictionary for Regulatory Activities. a n (%) Number (percentage) of patients in each dose group who have 1 AE matching the specified preferred term.

December 2010 LINACLOTIDE IN IBS-C 1885 tying of the bowel, it is a more rigorous and clinically meaningful end point, as some patients have frequent small SBMs but do not feel that they have completely emptied their bowels. 21 The Rome II, 14 rather than Rome III, 2 diagnostic criteria were used in this study for several reasons. The Rome III criteria were relatively new with limited clinical trial experience at the time that this study was being developed. In addition, a phase II pilot IBS study 13 with linaclotide used the Rome II criteria, and, therefore, the Rome II criteria were preferred to maintain consistency throughout the linaclotide development program. Based on the Bowel Habits Questionnaire administered at the screening visit for this study, most patients (90%) meeting the Rome II criteria for IBS also met Rome III criteria. Given that the primary end point of this study was the change from baseline in CSBM frequency, this study used a modification of the Rome II criteria for subtyping IBS-C. 2 Specifically, this study limited entry to IBS patients who reported 3 BMs per week and at least 1 other constipation symptom (ie, lumpy or hard stools, incomplete evacuation, or straining) for at least 25% of BMs. Manual maneuvers to facilitate defecation and sensation of anorectal obstruction were not included as constipation symptoms to fulfill IBS-C subtype criteria for entry into the study in an attempt to avoid enrolling patients with dyssynergic defecation. 2 These results for abdominal pain and constipation are supported by the effect of linaclotide on abdominal discomfort and bloating, as well as global assessments of IBS, including adequate relief of IBS, global relief of IBS, IBS severity, constipation severity, IBS-SSS, satisfaction with study medication, and likelihood of continuing study medication. 5 This study confirmed 2 previous reports demonstrating an inverse relationship between baseline disease severity (determined by IBS-SSS) and adequate relief of symptoms. 22,23 The IBS-QOL showed clinically meaningful improvement in all linaclotide treatment groups, although the change from baseline at the end of treatment for the linaclotide groups was not significantly different from the placebo group for the overall IBS-QOL score or any of its 8 subscales. However, the 300- and 600- g linaclotide dose groups experienced statistically significant improvement compared with placebo for the QOL question included in the IBS-SSS, which asks patients how their IBS symptoms affect or interfere with their life in general. Taken together, these results, along with the improvement in abdominal symptoms (pain, discomfort, and bloating) and bowel symptoms (CSBM, SBM, consistency, and straining), support the robust effect of linaclotide for the treatment of IBS-C. AEs were generally related to the GI tract, particularly diarrhea. Most events of diarrhea were mild to moderate in severity, and all cases resolved spontaneously without medical intervention. The incidence of other AEs, excluding diarrhea, were similar to placebo and were not dose related. One SAE (fecal impaction) occurred in a patient receiving 300 g of linaclotide and resulted in temporary withholding of study medication; this event, which resolved with treatment, was reported to be unrelated to the study medication by the investigator. Finally, there were no clinically significant changes in vital signs, serum electrolytes, or ECGs observed. The safety results of this study are consistent with those of previous trials with linaclotide in patients with IBS-C. 13,24 The effects observed in this study are believed to be a consequence of linaclotide s activation of GC-C receptors in the intestinal lumen leading to the intracellular production of cgmp. In animal models, this activation has been shown to result in the opening of anion channels with a significant increase in fluid secretion into the intestinal lumen and an associated acceleration of intestinal transit. Importantly, linaclotide s activation of GC-C receptors has also been shown to have potent antinociceptive effects in a range of rodent models of visceral hypersensitivity. 6,25,26 Although all linaclotide doses were associated with a statistically significant improvement compared with placebo for most end points, the higher doses of linaclotide (ie, 300 and 600 g) were generally more effective across most parameters. Because the 300 and 600 g doses provided comparable efficacy and the higher dose was associated with an increase in GI side effects, linaclotide 300 g per day was selected for continued evaluation in phase III trials. In summary, the results of this placebo-controlled, dose-range-finding, phase IIb study support further development of linaclotide for treatment of adults with IBS-C. Supplementary Material Note: To access the supplementary material accompanying this article, visit the online version of Gastroenterology at www.gastrojournal.org, and at doi: 10.1053/j.gastro.2010. References 1. Hungin AP, Chang L, Locke GR, et al. Irritable bowel syndrome in the United States: prevalence, symptom patterns and impact. Aliment Pharmacol Ther 2005;21:1365 1375. 2. Longstreth GF, Thompson WG, Chey WD, et al. Functional bowel disorders. Gastroenterology 2006;130:1480 1491. 3. Drossman DA, Morris CB, Schneck S, et al. International survey of patients with IBS: symptom features and their severity, health status, treatments, and risk taking to achieve clinical benefit. J Clin Gastroenterol 2009;43:541 50. 4. Pare P, Gray J, Lam S, et al. Health-related quality of life, work productivity, and health care resource utilization of subjects with irritable bowel syndrome: baseline results from LOGIC (Longitudinal Outcomes Study of Gastrointestinal Symptoms in Canada), a naturalistic study. Clin Ther 2006;28:1726 1735. 5. Brandt LJ, Chey WD, Foxx-Orenstein A, et al. An evidence-based systematic review on the management of irritable bowel syndrome. Am J Gastroenterol 2009;104:S1 S34.

1886 JOHNSTON ET AL GASTROENTEROLOGY Vol. 139, No. 6 6. Eutamene H, Gilet M, Beaufrand C, et al. Effect of oral cyclic GMP on two distinct visceral pain models. Neurogastroenterol Motil 2008;22:75. 7. Eutamene H, Bradesi S, Larauche M, et al. Guanylate cyclase C-mediated antinociceptive effects of linaclotide in rodent models of visceral pain. Neurogastroenterol Motil 2010;22:312 e84. 8. Currie MG, Fok KF, Kato J, et al. Guanylin: an endogenous activator of intestinal guanylate cyclase. Proc Natl Acad Sci U S A 1992;89:947 951. 9. Forte LR Jr. Uroguanylin and guanylin peptides: pharmacology and experimental therapeutics. Pharmacol Ther 2004;104:137 162. 10. Bueno L, Beaufraud C, Mahajan-Miklos S. Antinocicetive actions of MD-1100, a novel therapeutic agent for C-IBS, in animal models of visceral pain. Am J Gastroenterol 2004;99(Suppl 2): A283. 11. Ustinova E, Bryant A, Tammi R, et al. Oral cyclic guanosine monophosphate (cgmp) desensitizes colonic afferents in a animal model of experimental pain. Am J Gastroenterol 2008;486. 12. Kurtz CB, Fitch D, Busby RW. Effects of multidose administration of MD-1100 on safety, tolerability, exposure, and pharmacoldynamics in healthy subjects. Gastroenterology 2006;130(Suppl 2):A26. 13. Andresen V, Camilleri M, Busciglio IA, et al. Effect of 5 days linaclotide on transit and bowel function in females with constipation-predominant irritable bowel syndrome. Gastroenterology 2007;133:761 768. 14. Thompson WG, Longstreth GF, Drossman DA, et al. Functional bowel disorders and functional abdominal pain. Gut 1999; 45(Suppl 2):II43 47. 15. Heaton KW, O Donnell LJ. An office guide to whole-gut transit time. Patients recollection of their stool form. J Clin Gastroenterol 1994;19:28 30. 16. Francis CY, Morris J, Whorwell PJ. The irritable bowel severity scoring system: a simple method of monitoring irritable bowel syndrome and its progress. Aliment Pharmacol Ther 1997;11: 395 402. 17. Patrick DL, Drossman DA, Frederick IO, et al. Quality of life in persons with irritable bowel syndrome: development and validation of a new measure. Dig Dis Sci 1998;43:400 411. 18. Conover WJ. Practical nonparametric statistics. Hoboken, NJ: Wiley; 1999. 19. Drossman D, Morris CB, Hu Y, et al. Characterization of health related quality of life (HRQOL) for patients with functional bowel disorder (FBD) and its response to treatment. Am J Gastroenterol 2007;102:1442 1453. 20. Spiegel B, Strickland A, Naliboff BD, et al. Predictors of patientassessed illness severity in irritable bowel syndrome. Am J Gastroenterol 2008;103:2536 2543. 21. Camilleri M, Kerstens R, Rykx A, et al. A placebo-controlled trial of prucalopride for severe chronic constipation. N Engl J Med 2008;358:2344 2354. 22. Passos MC, Lembo AJ, Conboy LA, et al. Adequate relief in a treatment trial with IBS patients: a prospective assessment. Am J Gastroenterol 2009;104:912 919. 23. Whitehead WE, Palsson OS, Levy RL, et al. Reports of satisfactory relief by IBS patients receiving usual medical care are confounded by baseline symptom severity and do not accurately reflect symptom improvement. Am J Gastroenterol 2006;101: 1057 1065. 24. Johnston JM, Kurtz CB, Drossman DA, et al. Pilot study on the effect of linaclotide in patients with chronic constipation. Am J Gastroenterol 2009;104:125 132. 25. Bryant AP, Busby RW, Bartolini WP, et al. Linaclotide is a potent and selective guanylate cyclase C agonist that elicits pharmacological effects locally in the gastrointestinal tract. Life Sciences 2010;86:760 765. 26. Busby RW, Bryant AP, Bartolini WP, et al. Linaclotide, through activation of guanylate cyclase C, acts locally in the gastrointestinal tract to elicit enhanced intestinal secretion and transit. European Journal of Pharmaology 2010 Sep 20. [Epub ahead of print]. Received February 12, 2010. Accepted August 19, 2010. Reprint requests Address requests for reprints to: Jeffrey M. Johnston, MD, 301 Binney Street, Cambridge, Massachusetts 02141. e-mail: jjohnston@ ironwoodpharma.com Acknowledgments The authors thank the investigators for their participation in this study and Nicole LaVallee, PhD, who independently reviewed the data set and confirmed the analyses. Clinicaltrials.gov ID: NCT00460811. The statistical analysis of the entire data sets pertaining to efficacy (specifically primary and major secondary efficacy end points) and safety (specifically, serious adverse events as defined in federal guidelines) have been independently confirmed by Nicole LaVallee, PhD, a biostatistician who is not employed by the corporate entity; and the corresponding author had full access to all of the data and takes full responsibility for the veracity of the data and analysis. Dr. LaVallee was compensated by Ironwood Pharmaceutical for statistical review of the data. Conflicts of interest The authors disclose the following: Jeffrey Johnston, Caroline Kurtz, James MacDougall, B. J. Lavins, Mark Currie, Donald Fitch, Chris O Dea, and Mollie Baird are employees of Ironwood Pharmaceuticals. Anthony Lembo is a paid consultant to Ironwood Pharmaceuticals. Funding Supported by Ironwood Pharmaceuticals.

December 2010 LINACLOTIDE IN IBS-C 1886.e1 Supplementary Patients and Methods Exclusion Criteria Patients were excluded from the study if they reported loose (mushy) or watery stools in the absence of laxatives for 25% of bowel movements (BM) during the 12 weeks preceding the study. Likewise, patients were excluded if, during the 2-week pretreatment baseline period, they reported more than 1 loose, mushy stool (Bristol Stool Form Scale [BSFS] score of 6) or any watery, liquid stool (BSFS score of 7) without laxative use during the previous 24 hours. Patients with a history of pelvic floor dysfunction, the need to use manual maneuvers to achieve a BM, surgery of the colon at any time, abdominal surgery within 60 days prior to entry into the study, or laxative abuse were excluded from the study. Patients with other medical conditions (eg, neurologic disorders, metabolic disorders, or other significant disease) or pretreatment laboratory or electrocardiogram findings determined by the investigator to impair their ability to participate in the study were also excluded. Use of prohibited medications (eg, narcotics, prokinetics), any surgery within the past 30 days, pregnancy, breast-feeding, or use of an investigational drug within 30 days prior to the start of the study were also disqualifying factors. Patients over the age of 50 years were required to have colonoscopy screening according to the American Gastroenterological Association guidelines. 1 Likewise, patients of any age with alarm symptoms (eg, lower gastrointestinal bleeding, iron-deficiency anemia, unexplained clinically significant weight loss, systemic signs of infection or colitis, or a family history of celiac disease or inflammatory bowel disease) were required to complete a colonoscopy with negative findings. Women of childbearing age were required to have a negative serum pregnancy test. Sexually active women were required to have used oral or implanted contraceptives or a double barrier method for at least 3 months prior to entering the study and to continue using such contraceptive methods throughout the study. Patients were asked to refrain from making any major lifestyle changes (eg, starting a new diet or changing their exercise pattern) during the study. Rescue and Concomitant Medications Rescue medications were allowed for severe constipation (ie, at least 72 hours after the patient s previous BM) including oral bisacodyl up to 15 mg daily, bisacodyl suppository, or sodium phosphate enema. Rescue medication was allowed during the pretreatment baseline period, provided that there were no more than 2 uses and that none was used during the 3 days before the first dose of study medication. Patients were instructed to contact the investigator prior to taking a rescue medication for severe constipation, and all rescue medication usage was recorded during the patient s daily call to the interactive voice response system. Patients were prohibited from taking prescription or over-the-counter inflammatory bowel syndrome (IBS) and constipation medications (except in rescue medication situations described above). Patients on a stable, continuous regimen of fiber therapy for at least 30 days prior to the pretreatment baseline period were allowed to continue, provided that they continued at a constant dose throughout the study. Antidepressants were allowed provided that the patient had been on a stable dose for at least 30 days before entry into the study and that no dose adjustments were planned during the study. Results Rescue Medication Rescue medication use in this study was low, averaging between 1.0% and 2.5% of days over the 12-week study treatment period; the change from baseline was lower for all linaclotide groups compared with placebo and reached statistical significance for the 600- g dose group (P.0361). In addition, there was no significant change in rescue medication use during the posttreatment period compared with the pretreatment or treatment periods. Posttreatment During the 2-week posttreatment period, mean values for abdominal pain, bloating, complete spontaneous bowel movement and spontaneous bowel movement rates, stool consistency, straining, and global relief of IBS symptoms returned toward pretreatment levels. However, there was no worsening of IBS with constipation symptoms relative to baseline (ie, no rebound effect). Baseline Severity and Adequate Relief Responders There was a statistically significant inverse relationship between baseline severity as determined by the IBS-symptom severity scale (IBS-SSS) and outcome as assessed by the percentage of patients who were adequate relief responders (P.0236). When results on the 0- to 500-point IBS-SSS scale were categorized as mild (0 174), moderate (175 300), and severe (301 500), the percentages of patients who reported adequate relief for at least 9 out of 12 weeks of the treatment period stratified by baseline IBS-SSS were 64%, 42%, and 35%, respectively; this inverse relationship was maintained when patients who reported baseline adequate relief were excluded from the analysis (11% of the intent-to-treat population). References 1. Winawer S, Fletcher R, Rex D, et al. Colorectal cancer screening and surveillance: clinical guidelines and rationale-update based on new evidence. Gastroenterology 2003;124:544 560.