Single‐cell genetic analysis of clonal dynamics in colorectal adenomas indicates CDX2 gain as a predictor of recurrence

Colorectal adenomas are common precancerous lesions with the potential for malignant transformation to colorectal adenocarcinoma. Endoscopic polypectomy provides an opportunity for cancer prevention; however, recurrence rates are high. We collected formalin‐fixed paraffin‐embedded tissue of 15 primary adenomas with recurrence, 15 adenomas without recurrence, and 14 matched pair samples (primary adenoma and the corresponding recurrent adenoma). The samples were analysed by array‐comparative genomic hybridisation (aCGH) and single‐cell multiplex interphase fluorescence in situ hybridisation (miFISH) to understand clonal evolution, to examine the dynamics of copy number alterations (CNAs) and to identify molecular markers for recurrence prediction. The miFISH probe panel consisted of 14 colorectal carcinogenesis‐relevant genes (COX2, PIK3CA, APC, CLIC1, EGFR, MYC, CCND1, CDX2, CDH1, TP53, HER2, SMAD7, SMAD4 and ZNF217), and a centromere probe (CEP10). The aCGH analysis confirmed the genetic landscape typical for colorectal tumorigenesis, that is, CNAs of chromosomes 7, 13q, 18 and 20q. Focal aberrations (≤10 Mbp) were mapped to chromosome bands 6p22.1‐p21.33 (33.3%), 7q22.1 (31.4%) and 16q21 (29.4%). MiFISH detected gains of EGFR (23.6%), CDX2 (21.8%) and ZNF217 (18.2%). Most adenomas exhibited a major clone population which was accompanied by multiple smaller clone populations. Gains of CDX2 were exclusively seen in primary adenomas with recurrence (25%) compared to primary adenomas without recurrence (0%). Generation of phylogenetic trees for matched pair samples revealed four distinct patterns of clonal dynamics. In conclusion, adenoma development and recurrence are complex genetic processes driven by multiple CNAs whose evaluations by miFISH, with emphasis on CDX2, might serve as a predictor of recurrence. What's new? Colonoscopy screening helps detecting and removing adenomas before progression to cancer but predicting recurrence remains challenging. Here the authors performed single‐cell multiplex interphase fluorescence in situ hybridisation to detect copy number changes of 14 genes implicated in colon carcinogenesis in primary and recurrent colorectal adenomas. Intestine‐specific transcription factor CDX2 was identified as a potential marker of recurrence as it was exclusively upregulated in adenomas with recurrence.