Pharmacodynamics and gene expression analysis
PD, Sigmoid colon, pre‐dose, IBD
BLOOD, SIGMOID COLON
Tissue biopsies from the sigmoid colon were obtained before start of treatment (up to Day −3) and at the end of treatment (Day 7). For operational reasons, biopsies were taken 2−4.5 h post‐dose on Day 7. Stool samples were obtained at screening, before start of treatment, throughout the study, and at the 14‐day follow‐up visit. Levels of calprotectin were determined in stool samples. Sigmoid colon tissue was processed for RNA and DNA extraction as well as for single‐cell analysis, subsequent transcriptomic analysis, and identification of PD biomarkers. Sigmoid colon tissue samples were also processed for histology (hematoxylin and eosin staining, immunohistochemistry, fluorescence in situ hybridization) in order to characterize the cell types present (i.e., neutrophils and inflammatory monocytes). Histological analysis was performed on sigmoid colon tissue samples (from baseline and Day 7) using the disease‐specific histological indices Geboes scoreA predefined IL‐1 gene signature comprising eight genes (i.e., CXCL1, CXCL2, CXCL3, CXCL5, CXCL6, CCL2, IL6, TNFAIP6) relevant in the context of IBD was assessed in RNA extracted from the sigmoid colon tissue. The signature had been determined by combining gene expression data that met the following criteria: (1) overlapping expression profiles (upregulation by IL‐1β stimulation) in multiple cell types,Blood samples were drawn pre‐dose and at various timepoints and placed in TruCulture whole blood collection and culture tubes that were pre‐filled with lipopolysaccharide (LPS) (final working concentration of 100 ng/mL LPS after addition of 1 mL of blood to the LPS‐containing Truculture tubes). Following 24 h incubation, TruCulture supernatants were collected, stored, and levels of secreted IL‐1β were subsequently measured in order to understand how these responded to NLRP3 inhibition by selnoflast. The levels of IL‐1β were also quantified in plasma K