HD Chromoendoscopy Coupled with DNA Mass Spectrometry Profiling Identifies Somatic Mutations in Microdissected Human Proximal Aberrant Crypt Foci

Abstract

Despite increased implementation of screening colonoscopy, interval cancers in the proximal colon remain a major public health concern. This fact underscores the limitations of current screening paradigms and the need for developing advanced endoscopic techniques. The density of aberrant crypt foci (ACF), the earliest identifiable mucosal abnormality, may serve as a surrogate marker for colon cancer risk, but has rarely been studied in the proximal colon. To this end, high-definition (HD) chromoendoscopy was conducted to define the relevance of ACF in the proximal colon. In addition, due to limited ACF size, the development of a combinatorial approach was required to maximize data acquisition obtained from individual biopsy samples. Proximal and distal ACF samples were characterized for a total of 105 mutations across 22 known tumor suppressor and proto-oncogenes using high-throughput Sequenom MASSarray analysis. From this profiling, a discrete number of somatic mutations were identified, including APCR876* and FLT3I836M, as well as a deletion within the EGFR gene. Combined, these data highlight the significance of ACF within the context of colon cancer pathogenesis, particularly in the proximal colon. Implications The identification of cancer-related mutations in commonly overlooked mucosal lesions underscores the preventive benefit of implementing advanced endoscopic screening to larger patient populations, particularly in the proximal colon.

A Phase IIa Randomized, Double-Blind Trial of Erlotinib in Inhibiting Epidermal Growth Factor Receptor Signaling in Aberrant Crypt Foci of the Colorectum

“…Previously, a significant percentage of distal colon and rectum ACF were shown to have significant rates of KRAS and BRAF mutations (20-23) that can drive ACF growth. In colorectal and non-small cell lung adenocarcinomas, KRAS (33, 34), BRAF (35, 36) and EGFR kinase domain mutations (37) have been previously associated with anti-EGFR targeted therapy chemoresistance, which is thought to arise from both pre-existing mutations and induction of mutations from EGFR-inhibitor exposure.…”

“…Furthermore, many of the molecular derangements described in colon cancers are also found in ACF, including KRAS , APC , and CTNNB1 mutations and growth-promoting alterations in cell cycle–controlling genes(22). In the distal colon and rectum, approximately 32-63% and 30-37% of ACFs respectively have KRAS and BRAF mutations(20-23), which can drive downstream RAS/RAF/ERK pathway activation. Along with less common EGFR mutations, KRAS and BRAF mutations are thought to drive thegrowth of almost all ACFs, and are typically mutually exclusive in individual ACFs (23).…”

“…In the distal colon and rectum, approximately 32-63% and 30-37% of ACFs respectively have KRAS and BRAF mutations(20-23), which can drive downstream RAS/RAF/ERK pathway activation. Along with less common EGFR mutations, KRAS and BRAF mutations are thought to drive thegrowth of almost all ACFs, and are typically mutually exclusive in individual ACFs (23). …”

Distinct Transcriptional Changes and Epithelial–Stromal Interactions Are Altered in Early-Stage Colon Cancer Development

“…HD-chromoendoscopy was performed in the distal 20-cm of the colorectum and throughout the entire proximal colon using 0.1% indigo-carmine dye-spray for contrast enhancement. The identification and histologic evaluation of ACF has been described previously (8,9). In addition, each subject had a histologically confirmed corresponding normal biopsy specimen removed from the same side of the colon, generally within 2-cm of the ACF biopsy.…”

“…In addition, each subject had a histologically confirmed corresponding normal biopsy specimen removed from the same side of the colon, generally within 2-cm of the ACF biopsy. Mutational spectra of ACF were determined using DNA-MS analysis (Sequenom) (8). Only ACF with confirmed, non-overlapping somatic mutations to either KRAS , BRAF or APC were selected for further analysis.…”