OptiCal Study -Optimizing Fecal Calprotectin Monitoring: a Clinical Study Comparing CalproLab Against PhiCal and Evaluating Its Association With the Gut Microbiome

Study Aims: To analyze stool specimens to test and validate the CalproLab assay against the predicate PhiCal in order to determine performance characteristics. And to correlate Calpro levels to the gut microbiome composition.

Fecal Calprotectin has become one of the most frequently used biomarkers in patients suffering from Inflammatory Bowel Diseases. Although its use for monitoring disease activity and therapeutic efficiency has previously been demonstrated, the test performance should further be optimized to improve clinical accuracy. The PhiCal™ test is a quantitative ELISA for measuring, in human stool, concentrations of fecal calprotectin, a neutrophilic protein that is a marker of mucosal inflammation. The PhiCal™ test can be used as an in vitro diagnostic to aid in the diagnosis of inflammatory bowel diseases (Crohn's disease and Ulcerative Colitis), and to differentiate IBD from irritable bowel syndrome. Recently, the CalproLab™ in vitro device has become available which provides a greater reporting range than PhiCal™. The OptiCal study aims to test and validate the CalproLab™ assay against the PhiCal™ assay and determine its performance characteristics.

Calprotectin is a valuable clinical marker for inflammation. Calprotectin belongs to a group of calcium- binding neutrophil-derived proteins. Calprotectin makes up 60% of the cytosolic proteins within the neutrophil. It is very resistant to bacterial degradation in the gut and is stable in stool for up to one week

at room temperature. Calprotectin is the noninvasive "test of choice" for quantifying the degree of GI inflammation and differentiating Irritable Bowel Syndrome (IBS) from Inflammatory Bowel Disease (IBD).

Inflammatory bowel disease (IBD) is considered to result from interplay between host and intestinal microbiota. Recently it was shown that the microbiota varied along a gradient of increasing intestinal inflammation (indicated by calprotectin levels), which was associated with reduced microbial richness, abundance of butyrate producers, and relative abundance of Gram-positive bacteria (especially Clostridium clusters IV and XIVa). A significant association between microbiota composition and inflammation was indicated by a set of bacterial groups predicting the calprotectin levels. So, intestinal microbiota may represent a potential biomarker for correlating the level of inflammation and therapeutic responses but this needs to be further validated.

• This study is a prospective case series during which patients with either Crohn's disease or ulcerative colitis, Celiac Disease, Irritable Bowel Syndrome and healthy controls (N=175) will be invited to participate.
• Participants will receive a home kit (with a plain white plastic cup for calprotectin sample collections) for stool sampling, which then will be returned and processed at Genova Diagnostics. All participants are required to complete a patient survey, which will be included in the collection kit.
• Participants will be recruited through the UCLA Division of Digestive Diseases.
• All patients will have confirmation of their diagnosis according to published clinical guidelines and standards of care using gold standard diagnostics (e.g. endoscopy)
• Stool specimens will be analyzed for fecal Calprotectin using both the CalproLab™ assay and the PhiCal™ assay. No blood draws or other testing will be performed.
• Patients will undergo a gut microbiome assessment utilizing the GI Effects™ 2200 Comprehensive Profile (Genova Diagnostics).