Molecular Biomarkers of Sessile Serrated Adenoma/Polyps

Abstract

Objectives: Sessile serrated adenoma/polyps (SSA/Ps) contribute up to 30% of all colon cancers. There is considerable histological overlap between SSA/Ps and hyperplastic polyps. Inadequate consensus exists among pathologists, and no molecular biomarkers exist to differentiate these lesions with high accuracy. Lack of reliable diagnosis adversely affects clinical care. We previously defined a novel 7-gene panel by RNA sequencing that differentiates SSA/Ps from hyperplastic polyps. Here, we use the 7-gene panel as a molecular approach to differentiate SSA/Ps and HPs with higher sensitivity and specificity in a large sample set from a tertiary health care center.

Methods: Reverse transcription quantitative polymerase chain reaction of the 7-gene panel was performed on 223 formalin-fixed, paraffin-embedded serrated polyp and normal colon samples. We compare the sensitivity and specificity of the 7-gene panel with the BRAF and KRAS mutation incidence in differentiating SSA/Ps and HPs. We also evaluate the clinical data of patients with SSA/Ps showing high and low expression of the gene panel.

Results: The 7-gene RNA expression panel differentiates SSA/Ps and HPs with 89.2% sensitivity and 88.4% specificity. The gene panel outperforms BRAF mutation in identification of SSA/Ps. Clinical data suggest that expression of the 7-gene panel correlates with the development of SSA/Ps in the future.

Discussion: This study describes a novel 7-gene panel that identifies SSA/Ps with improved accuracy. Our data show that RNA markers of SSA/Ps advance the distinction of serrated lesions and contribute to the study of the serrated pathway to colon cancer.

Figure 1.

RNA expression by reverse transcription quantitative polymerase chain reaction of the 7-gene panel, graphing each gene individually (FSCN1, MUC6, SEMG1, TRNP1, ZIC2, ZIC5, and CRYBA2) in sessile serrated adenomas/polyps (SSA/Ps, n = 99), in hyperplastic polyps (HPs = 78), in uninvolved colon from patients with serrated polyps (Uninvl, n = 27), and in control colon samples from patients without polyps on colonoscopy (Cntl, n = 19). Y axis shows the fold change in expression relative to the mean of HP expression. X axis shows each of the 4 sample groups. Mean ± SE bars are presented for each of the 4 groups. Mann-Whitney U-test statistics in GraphPad Prism 5.0d version software (San Diego, CA) were used to determine statistical significance between sample groups. The expression of all 7 genes was statistically significant (**P < 0.0001) between SSA/Ps and the other 3 groups (HPs, uninvolved, and control).

Figure 2.

RNA expression of the 7-gene panel based on the size and location of SSA/Ps (n = 99), graphing each gene individually (FSCN1, MUC6, SEMG1, TRNP1, ZIC2, ZIC5, and CRYBA2). Y axis shows the fold change in expression relative to the mean of HP expression. X axis shows each of the 4 groups (proximal-proximal to splenic flexure; large >/ = 10 mm). Mean ± SE bars are presented for each of the 4 groups. Mann-Whitney U-test statistics in GraphPad Prism 5.0d version software were used to determine statistical significance between sample groups. The expression of FSCN1, MUC6, SEMG1, ZIC2, and CRYBA2 was statistically significant (*P < 0.05) by location. The expression of TRNP1 and CRYBA2 was statistically significant (#P < 0.05) by size. HP, hyperplastic polyp.

Figure 3.

ROC curves of the 7-gene expression panel (FSCN1, MUC6, SEMG1, TRNP1, ZIC2, ZIC5, and CRYBA2) (a), and BRAF (b) and KRAS (c) mutation data, showing relative sensitivity and specificity in distinguishing SSA/Ps (n = 36) and HPs (n = 41). Serrated polyps fulfilling 2 or 3 of 3 criteria were considered SSA/Ps, and serrated polyps fulfilling 0 or 1 of 3 criteria were considered HPs. The 7-gene panel had high sensitivity and specificity (AUC 92.5%) in differentiating SSA/Ps and HPs, whereas BRAF and KRAS mutations did not (AUCs 57.4% and 58.4%, respectively. AUC, area under the curve; ROC, receiver operator characteristic; SSA/P, sessile serrated adenoma/polyp.

Figure 4.

Range of fold change expression of the 7-gene panel (FSCN1, MUC6, SEMG1, TRNP1, ZIC2, ZIC5, and CRYBA2) in serrated polyps fulfilling 2 or 3 of the 3 criteria (n = 87). Red and blue dots show polyps with serrated sessile polyp-type expression of a minimum of 4 of 7 genes in the top or bottom 25% for all polyps, respectively. Lines separate the 25% highest and lowest-expressing polyp samples for each gene. Y axis depicts fold change from the mean of 0 of 3 criteria polyps (n = 72), and x axis shows individual SSA/P fold change values.