Efficacy Of Noninvasive Biomarkers In Monitoring Disease Activity In Inflammatory Bowel Disease

Background and Objective:Inflammatory bowel disease (IBD) is a chronic and destructive disorder of digestive tract that has been empirically defined based on clinical, ileocolonoscopic, histopathologic, and radiologic findings. Ulcerative colitis (UC) and Crohn’s disease (CD) are frequently two major types of IBD in clinical condition. However, another clinical entity, indeterminate colitis(IC), is considered a subgroup of IBD. IC referred to those when a definitive diagnosis of UC or CD cannot be made at colonoscopy, in colonic biopsies or at colectomy in clinical and accounts 10-15% of cases of IBD. The underlying etiology of IBD remains unknown, but many studies demonstrate that immunological factors, intestinal infection, genetic background, environmental influences and psychological factors may play important roles in its etiopathogenesis. IBD is a worldwide disease, in recent decades, considerable prospective epidemiological studies have pointed out that the IBD incidence rate in traditionally high-incidence areas, such as the United States and Europe, has been relatively stable and the disease has become more prevalent in previously low incidence areas, including Asia, but the total incidence of IBD in Asia remains significantly lower than that in the West. Two recent prospective population-based studies, conducted in Guang Zhou and Wu Han, also demonstrated that IBD had a high incidence and wide geographical coverage in China. In 2013,Zeng et al conducted a prospective, population-based IBD incidence study in Guangdong Province and found Age-standardized incidence rates for IBD, UC, and CD were 3.14, 2.05, and 1.09 per 100000 persons, respectively. With the rapid economic growth and industrialization degree of China, IBD is becoming one of the common diseases of digestive system in our country.The disease course of IBD is characterized by periods of inflammatory activity alternating with periods of remission and is difficult to cure. Clinicians should choose the therapy regimen based on the comprehensive evaluation of the disease and tailor therapy according to different disease stage. They need a monitoring technique to recognize an imminent flare in endoscopic active patients for timely intensification of treatment and better disease control in endoscopic remission patients. So determination of inflammatory activity is essential for clinician choosing reasonable therapy and predicting prognosis. According to a questionnaire study, seventy-eight percent judged clinical activity to be the most relevant criterion for monitoring IBD activity,15% chose endoscopic severity. Seventy percent of gastroenterologists based their therapeutic decisions on clinical activity,24% on endoscopic severity. To date, we found that most clinicians monitoring IBD activity and guiding therapeutic decisions based on clinical activity indexes. In UC, the common used clinical activity indexes were "Truelove and Witts Activity Index" and "Rachmilewitz Clinical Colitis Activity Index" (CAI). In CD, Harvey and Bradshow Index and Best CDAI were commonly used. However, emerging data show that IBD patients with clinically quiescent disease may still have residual mucosal inflammation and remission of symptoms may not available predictors of long term favorable outcome in IBD patients. However, mucosal inflammatory changes have been proven to be strong predictors of disease related outcomes in IBD and mucosal healing (MH) become a therapeutic treatment goal in IBD. So endoscopy is currently considered the gold standard for assessing disease activity, and a number of endoscopic scoring systems exist to quantify inflammatory activity in UC and CD, for CD with the "Crohn’s Disease Endoscopic Index of Severity" (CDEIS), "simple endoscopic score for Crohn’s disease" (SES-CD), for CD-related surgery patients with the Rutgeerts score and ulcerative colitis (UC) with the Mayo score. Although endoscopy can have a direct visualisation of intestinal lesions, however, endoscopy is invasive, uncomfortable, expensive, and not well tolerated by patients. Therefore, regular monitoring of disease activity using endoscopy is not feasible. Furthermore, with some patients terrified by painful endoscopy experiences, they are reluctant to visit clinicians until the disease manifests by rectal bleeding or obstruction, which affects the prognosis. As symptom is subjective to assess disease activity and endoscopy procedure is often a burden to IBD patient. A simple, acceptable, and specific biomarker is needed to play an adjunctive or even primary role in the assessment of disease activity, enabling the most cost-effective use of medical resources.Currently, generally used laboratory markers include CRP, ESR, white blood cell count, platelets, and procalcitonin. As serum markers can be elevated in a variety of conditions, it seems likely that fecal inflammation markers would more specific for IBD. Many studies have reported FC to be a reliable surrogate marker of endoscopic activity in IBD.FC, a 36 kDa calcium and zinc binding protein, accounts 60% of the cytosolic protein in the granulocyte. Unlike’traditional’serological biological markers, FC has higher specificity for the assessment of gastrointestinal inflammatory conditions. FC also has the advantage of showing excellent stability in feces at room temperature, for up to 3 days, and remarkably resistant to proteolytic degradation. It is a surrogate marker of neutrophil turnover in the digestive tract, has become an attractive biomarker in measuring bowel inflammation for researchers and clinicians. What’s more, clinical assessment indexes correlate poorly with endoscopic activity and remission of symptoms may not indicate remission of IBD. Found a biomarker that can detect increased disease activity earlier before any clinical symptoms have occurred was an unmet clinical need. The aim of this study, therefore, was to assess the efficacy of noninvasive biomarkers, especially FC, for evaluation of disease activity in three categories of IBD patients (colonic or ileo-colonic CD patients, CD-related surgery patients and UC patients) and further to explore the accuracy of those non-invasive biomarkers in detecting active residual mucosal inflammation in clinical remission IBD patients.Materials and MethodsThis study was approved by the Ethics Committee of Nan Fang Hospital, Southern Medical University (NFEC-2014-065).In this prospective study, we consecutively recruited 136 adult outpatients and inpatients (69 CD,23 CD related surgery,44 UC) with previously confirmed diagnoses of IBD referred for colonoscopy at the Department of Gastroenterology of Nan fang Hospital. A second cohort of 25 IBS patients, defined according to the Rome-Ⅲ criteria, served as controls. The endoscopic activities of IBD patients were graded according to validated endoscopic score tools. The SES-CD was used to quantify the colonic or ileo-colonic CD patients endoscopy activity (SES-CD<3 endoscopic remission,>4 endoscopic active). The Rutgeerts score was a well-established endoscopic scoring system for assessing the neo-terminal ileum for patients having prior CD-related surgery (iO-il endoscopic remission,≥i2 endoscOpic active).The disease severity of UC was evaluated according to the Mayo score scale (Mayo score≤2 endoscopic remission,≥3 endoscopic active). For CD patients, clinical activity was assessed based on the’Crohn’s disease activity index’(CDAI:< 150 clinical remission,>150 clinical active). For UC patients, clinical activity was assessed according to the ’clinical colitis activity index’(CAI), according to Rachmilewitz (CAI≤4 clinical remission,≥5 clinical active). Stool samples were collected from 3 to 1 day before bowel preparation. Upon fecal sample arrival in the laboratory, we sampled from 6 sites in each stool sample and mixed the sample with a stirrer, then 6-10g fecal sample were stored at-80℃ until analysis. The Buhlmann Calprotectin ELISA kit was designed for the quantitative determination of FC in stool samples. Before endoscopy examinations, patients were asked to measure CRP, ESR and PCT for IBD patients. At the same time, the patients were asked to complete a questionnaire to record their demographic data, symptoms, and current medication usage. Data were analyzed using the SPSS 15.0 software, The Mann-Whitney U-test was used to assess differences between the groups. The Kruskal-Wallis H test was used to assess differences in more than two independent groups. A Bonferroni adjustment was carried out in cases of multiple testing. Associations between endoscopic disease activity and laboratory parameters were assessed with Spearman’s correlation. The area under the ROC curve (AUC) was used to measure the ability of parameter to predict endoscopic severity.ResultsDuring the study period, six patients (3 CD,2 CD-related surgery, and 1 UC patients) underwent colonoscopy twice, thus a total of 142 endoscopies were performed in the 136 IBD patients. FC, CDAI/CAI, CRP and ESR were successfully measured in all included IBD patients.PCT was measured in 67case with CD,18case with CD-related surgery and 41case with UC patients. The median FC was significant higher [468.0 (IQR,137.1-1278.1)μg/g] in the IBD group compared with 25 IBS patients [9.9 (IQR,0-49.7)μg/g], P< 0.001. When the IBD patients were grouped into three categories-colonic or ileo-colonic CD patients, CD-related surgery patients, and UC patients-the median FC values were 695.0 (IQR,147.1-1805.0), 188.5 (IQR,72.06-559.7), and 497.9 (IQR,131.7-1198.0)μg/g, respectively. Within each IBD diagnosis group, FC was compared between patients with and without endoscopic inflammation, yielding significant differences between colonic or ileo-colonic CD patients and UC patients, all P< 0.001. In the CD-related surgery patients, FC values showed no difference based on Rutgeerts score classified endoscopic inflammation, [142.97 (IQR,62.86-4117.75) ug/g VS 229.27 (IQR, 75.85-726.65) ug/g; p= 0.52]. When compared with the three IBD patient groups without endoscopic inflammation, IBS patients also had significantly lower levels of FC (IBS vs. colonic or ileo-colonic CD patients and CD-related surgery patients, P< 0.001; IBS vs UC patients, p< 0.05). Differences in CDAI/CAI, CRP, and ESR according to endoscopy-based inflammation categories were observed in colonic or ileo-colonic CD patients and UC patients, but not CD-related surgery patients. With regard to PCT, no significant difference was detected in the three group IBD patients. In colonic or ileo-colonic CD and UC patients, FC was able to discriminate inactive from mild and moderate vs. severe endoscopic activity, however, FC failed to discriminate mild from moderate endoscopic activity. Of 56 colonic or ileo-colonic CD patients, the SES-CD correlated significantly with levels of FC (Spearman’s rank correlation coefficient, r= 0.802), CDAI (r= 0.734), CRP (r= 0.658), and ESR (r= 0.557) (all P< 0.01). All indexes have no correlation with Rutgeerts’score in CD-related surgery patients. In 45 UC patients, the Mayo score correlated more closely with FC (Spearman’s correlation coefficient, r= 0.837; P< 0.001), followed by CAI (r= 0.776; P< 0.001), ESR (r= 0.644; P< 0.001), and CRP (r= 0.634, P< 0.001). In colonic and ileo-colonic CD patients, at the threshold of 250μg/g, FC predicted endoscopic active disease with 97.1% sensitivity and 71.4% specificity. In UC, at cut-off value of 250μg/g, FC yielded a sensitivity and specificity of 87.9%, 100%, respectively. Furthermore, in CD-related surgery patients, the proposed cut-off level for endoscopic active CD of 250μg/g FC failed to discriminate between endoscopic recurrent disease and endoscopic remission. In the three subgroups of IBD clinical remission patients,41% colonic or ileo-colonic CD patients,36.8% CD-related surgery patients, and 52% of UC patients had active endoscopic inflammation. In colonic and ileo-colonic CD patients with an acknowledged cut-off value of 250μg/g, FC has 93% sensitivity/70% specificity and 86.4% sensitivity/ 100% specificity, respectively, in detecting endoscopy residual inflammation. Also, the diagnostic accuracy of FC in identifying patients with mucosal inflammation in clinical remission was reflected by an AUC of 0.91 for colonic or ileo-colonic CD and 0.96 for UC.ConclusionsFC was the most surrogate marker for the evaluation of colonic or ileo-colonic CD and UC patient endoscopic disease activity. However, FC did not correlate with the Rutgeerts scoring system indicated that FC may not for evaluation of patients having prior CD-related surgery. What’s more, a considerable number of IBD patients in clinical remission actually had active endoscopy inflammation, FC had good diagnostic accuracy in detecting residual mucosal inflammation in colonic or ileo-colonic CD and UC patients in clinical remission.