Stool DNA testing for the detection of pancreatic cancer

Abstract

BACKGROUND Pancreatic cancer (PanC) presents at late stage with high mortality. Effective early detection methods are needed. Aberrantly methylated genes are unexplored as markers for noninvasive detection by stool testing. We aimed to select discriminant methylated genes and to assess accuracy of these and mutant KRAS in stool to detect PanC. METHODS Nine target genes were assayed by real-time methylation-specific PCR (MSP) in bisulfite-treated DNA from microdissected frozen specimens of 24 PanC cases and 30 normal colon controls. Archived stools from 58 PanC cases and 65 controls matched on sex, age, and smoking were analyzed. Target genes from fecal supernatants were enriched by hybrid capture, bisulfite-treated, and assayed by MSP. KRAS mutations were assayed using the QuARTS technique. RESULTS Areas under the receiver operating characteristics curves (AUCs) for tissue BMP3, NDRG4, EYA4, UCHL1, MDFI, Vimentin, CNTNAP2, SFRP2 and TFPI2 were 0.90, 0.79, 0.78, 0.78, 0.77, 0.77, 0.69, 0.67, and 0.66, respectively. The top 4 markers and mutant KRAS were evaluated in stool. BMP3 was the most discriminant methylation marker in stool. At 90% specificity: methylated BMP3 alone detected 51% of PanCs, mutant KRAS detected 50%, and combination detected 67%. AUCs for methylated BMP3, mutant KRAS, and combination in stool were 0.73, 0.75, and 0.85, respectively. CONCLUSIONS This study demonstrates that stool assay of a methylated gene marker can detect PanC. Among candidate methylated markers discriminant in tissue, BMP3 alone performed well in stool. Combining methylated BMP3 and mutant KRAS increased stool detection over either marker alone.

The Triple-Code Model for Pancreatic Cancer

“…Besides its mechanistic importance and its relevance for cancer treatment, DNA methylation has been studied as a marker for the early detection of cancer. Cancer-specific DNA methylation patterns can be measured in DNA from detached tumor cells that are released to the blood, in pancreatic juice, or feces [73–76]. Unfortunately, in spite of its promise, there is not yet a clinically applicable assay for this purpose.…”

New DNA Methylation Markers for Pancreatic Cancer: Discovery, Tissue Validation, and Pilot Testing in Pancreatic Juice

“…We and others have detected both genetic and epigenetic markers in pancreatic juice(10, 11) and stool(12–15) from patients with pancreatic cancer and precursor lesions. A limitation with mutation markers relates to the unwieldy process of their detection; typically, numerous mutations across several genes must be assayed separately to achieve high sensitivity.…”

“…We have observed that methylation markers discriminant in primary tumor tissues often fail when assayed in an intended medium, such as stool. (15) Ideal candidate markers for pancreatic cancer screening would be universally present in pancreatic neoplasms, be absent in normal gastrointestinal mucosa, and have high signal strength.…”

Evaluation of Patients with an Apparent False Positive Stool DNA Test: The Role of Repeat Stool DNA Testing

“…However, it is known that colonoscopy is an imperfect “gold standard”, with a quoted miss rate for advanced adenomas of up to 11% (3) and 2.6%–9.0% of cancers developing within 3 years of colonoscopy (interval cancers) (4). While neoplasms in the upper gastrointestinal tract or pancreas (5) are known to produce genetic markers that may survive passage into the stool and be detectable in stool assays, a recently published cohort study that followed more than 1000 patients with a false positive sDNA test found only 8 subsequent cancers (3 lung, 3 pancreas, 1 colon, 1 biliary) at an average 4 year follow up, which was not greater than expected in the general population (6). A similarly low estimate of extracolonic sources of sDNA test positivity was found in a case-control study (2 positive aerodigestive cancers for each 10,000 patients screened) (7).…”