Mo1973 Immunohistochemical Markers Differentiating Colonic Sessile Serrated Adenomas/Polyps (SSA/Ps) From Hyperplastic Polyps

Abstract

Differentiating SSA/Ps from hyperplastic polyps (HPs), with little or no risk for progression to cancer, can be a major challenge using conventional endoscopic and histological examinations. SSA/Ps are frequently right sided, flat and have a mucus cap. The goal of this study was identifying potential immunohistochemical (IHC) markers that help to differentiate SSA/ Ps from HPs. Methods: We compared RNA sequencing transcriptional signatures of SSA/Ps (n=15) and control (n=20) colonic mucosa from patients undergoing screening colonoscopy. Reads from the analysis were aligned to the GRCh37/Hg19 human reference genome and filtered for high quality alignments (http://novocraft.com). Hierarchical clustering of log2 ratios comparing SSA/Ps and controls were performed and mined for candidate IHC targets. IHC analysis was performed on formalin-fixed paraffin-embedded sections of 12 SSA/Ps, 10 HPs, 5 adenomas, 5 colon cancers and 5 control colon specimens to assess staining for selected antigens. Expression of MUC5AC and TFF1 protein were quantified with ImageJ software. Results: Bioinformatics analysis identified 1,294 (≥1.5-fold change) differentially expressed annotated genes (false discovery rate (FDR) <0.05) in SSA/Ps as compared to normal controls. The dataset was mined for the most highly expressed genes and genes with possible biological functions related to SSA/Ps. A total of 249 genes were overexpressed ≥5fold in SSA/Ps compared to controls. SSA/Ps markedly overexpressed MUC5AC (582-fold), a component of the gastric mucus layer, and a secretory protein that may stabilize mucin layers, trefoil factor 1 (TFF1, 79-fold). Neither protein is typically expressed in normal colonic mucosa. SSA/Ps exhibited strong co-localized staining for both proteins (Fig. 1 and 2). Intense MUC5AC staining was primarily seen in the serrated crypts, whereas expression in adjacent non-serrated mucosa was identical to controls. Conversely, HPs showed only weak patchy staining of MUC5AC and variable TFF1 staining that was infrequently colocalized. Normal colon had no detectable MUC5AC staining and minimal TFF1 staining. Conventional adenomas showed trace MUC5AC and TFF1 staining. Conclusions: RNA-seq and IHC analysis showed markedly increased expression of two mucin-associated genes/ proteins in SSA/Ps, MUC5AC and TFF1, which are not typically expressed in normal colonic mucosa or adenomas. There was also intense co-localization of MUC5AC and TFF1 in SSA/ Ps but not in HPs. These results identify intense co-localized expression of MUC5AC and TFF1 as a possible diagnostic marker that would aid differentiating SSA/Ps from HPs. Further studies will be needed with larger numbers of SSA/Ps. Since MUC5AC has been linked to signaling related to bacteria, these findings suggest that luminal factors may play a role in the formation or progression of SSA/Ps.