CHICAGO — Research on an investigational DNA methylation test for colorectal cancer demonstrated that the only clinical variable that influenced test results was age, according to results presented at the AACR Annual Meeting 2012, held here March 31 - April 4.
"There was a progressive increase in background methylation levels that varied widely between methylation markers tested as a patient aged," said David Ahlquist, M.D., professor of medicine and a consultant in gastroenterology and hepatology at the Mayo Clinic in Rochester, Minn. "For example, median background methylation levels of the TFPI2 gene increased 49 percent in patients from age 50 to age 80, whereas levels for the BMP3 gene increased by only 0.2 percent across this age range."
His group at the Mayo Clinic, in collaboration with Exact Sciences, developed the multimarker molecular stool test, which is highly sensitive to the critical cancer screening targets of early-stage cancers and precancerous adenomas, he said.
"This test, if broadly applied, should have a very important impact on reducing both the mortality and incidence of colorectal cancer," Ahlquist said.
The researchers examined common patient variables, including age, sex, race, alcohol consumption, tobacco use, body mass and medication use in 500 patients undergoing screening colonoscopy or polyp follow-up. Patients had a normal colonoscopy in the last three years.
With the exception of age, none of the variables influenced test results, nor did family history of colorectal cancer or polyps or personal history of polyps. These results mean that "patients don't have to change their lifestyle to have this test," Ahlquist said. "That was important from a patient-friendly standpoint for a test like this and could benefit compliance."
Researchers have selected the two markers least affected by age for further test development and validation based on these study results.
"If we can minimize the false positives, that will reduce the cost of the whole screening program by avoiding unnecessary colonoscopies," Ahlquist said.
The screening test is currently undergoing FDA validation in a multicenter study in the United States and Canada, which is expected to be completed in the fall. The Mayo Clinic and Ahlquist have a financial interest in the technology referenced in this announcement.
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Presenter: David Ahlquist, M.D.
Abstract Number: 3572
Title: Methylated gene marker levels in stool: effects of demographic, drug, and body mass and other patient characteristics
Author Block: David A. Ahlquist1, William R. Taylor1, Tracy C. Yab1, Mary E. Devens1, Douglas W. Mahoney1, Lisa A. Boardman1, Steven N. Thibodeau1, Hongzhi Zou2, Domanico Michael2, Barry M. Berger2, Graham P. Lidgard2. 1Mayo Clinic, Rochester, MN; 2EXACT Sciences, Madison, WI
Abstract: Aberrantly methylated genes represent attractive and broadly informative markers for colorectal cancer (CRC). As demographics, numerous ingestants, environmental exposures, and obesity variably influence both gene methylation rates and CRC incidence, it is important to understand the effects of such covariates on the clinical performance of CRC screening tests that incorporate assay of methylated gene markers. Based on a recent multicenter study (Ahlquist et al. Gastroenterology 2012 (in press)), stool assay of carefully selected methylated gene marker candidates (NDRG4, BMP3, vimentin, TFPI2), alone or in combination, yield high detection rates of both CRC and large adenomas.
Aim: To assess the impact of demographic, exposure, body mass, and other patient variables on stool levels of the above 4 methylated gene markers.
Methods: We studied buffered stools collected within 3 years of a normal colonoscopy from 500 patients undergoing average-risk screening or polyp surveillance (median age 64 (range 44-85); 53% women). Upon receipt, stools were promptly homogenized, aliquoted, and frozen at -80°C. On supernatants from thawed/spun aliquots, target genes were purified by hybrid capture, bisulfite treated, and assayed using the analytically-sensitive QuARTS method (quantitative allele-specific real-time target and signal amplification), as described (above citation). The reference gene β-actin was assayed along with methylation of NDRG4, BMP3, vimentin, TFPI2. Log-converted data were normalized to allow comparison of effects between markers, and effect size was expressed as % change relative to standard deviation for each marker (standardized relative change (SRΔ)).
Results: The only patient characteristic that significantly influenced all methylated marker levels in stool was age (p<0.0001 for each marker); SRΔ was greatest with TFPI2 at +91.3 (p<0.001 vs other markers), least with BMP3 at +29.7 (p<0.001 vs others), and intermediate with vimentin at +46.0 and NDRG4 at +45.1. In contrast to methylation markers, levels of β-actin did not change across age. The other demographic variables of sex, race, and geographic residence had no effect on methylation markers. Marker levels were also not affected by exposure variables (smoking, alcohol consumption, or analgesic use), by family history of CRC or polyps, or by personal history of polyps. Finally, neither overweight status (n=176) nor obesity (n=148) had an effect on marker levels.
Conclusions: Age significantly affects stool levels of tumor-associated methylated gene markers in colonoscopy-normal patients, and the degree of this age effect differs across markers. In contrast, other key patient variables had no effect on stool marker levels.
Comment: Optimal use of these methylation markers for stool screening of CRC may require age-adjustment of normal cutoff levels to maximize test specificity.