Introduction: Colorectal cancer (CRC) cells are shed into stool, providing a potential means for the early detection. Previously we demonstrated an increased amount of human DNA in stool from CRC patients. Since human DNA may also be derived from non-tumor cells we tested this parameter in a large group of patients.

Methods: From 221 patients stool was prospectively collected before colonoscopy; from 209 clinical information could be retrieved: 137 non-adenoma/non-carcinoma, 42 adenoma and 30 carcinoma patients. The non-tumor group was separated into diverticulosis, inflammation, haemorrhoids, hyperplastic polyps, blood in feces and patients without these findings. Stool sampling was performed in two ways: a) samples that have been taken by the patients themselves and resuspended directly in a preservation solution and b) samples collected from dry stool delivered to the laboratory within 24 hours. DNA was extracted from the stool samples and the relative amount of human DNA (compared to total DNA, RH-DNA) was determined using a real-time quantitative PCR for the human β-globin gene. A threshold of 7.1 ng/μg was used based on previous work.

Results: Out of the total of 222 stool samples from 221 persons, 5 stools (2.3%) could not be analysed because of no, or not detectable human β-globin amplification levels in both isolates. The RH-DNA, of samples provided by the patient and taken in the laboratory from the same stool was similar (correlation coefficient of 0.82, range 0.2-300 ng/μg total DNA). The mean RH-DNA of all samples isolated was 32 ± 132 ng/μg. The yield of total DNA per isolation for dry stool and resuspended stool varied widely, depending on the patient and the sampling by the patient (40 ng-36 μg for dry stool and 116 ng-37 μg for resuspended stool, respectively). The fraction of human DNA (RH-DNA) still varied widely for both sampling methods (0.08 -310 ng/μg for dry stool and 0.02-580 ng/μg for resuspended stool, respectively). This broad range may be due to the condition of the patient as we know from the colonoscopic examination.

In carcinoma and adenoma patients RH-DNA was increased in 21 out of 30 patients (sensitivity 70%) and 10 out of 42 patients (sensitivity 24%), respectively.

The overall sensitivity for carcinoma (70%) was much higher for left-sided carcinoma than for right-sided carcinoma, 77% and 17% respectively. The specificity of this assay for all patients in the non-adenoma/non-carcinoma group is 72%. When the patients with reported blood in their feces as clinical symptom are left out the specificity rises to 77%. The specificity rises even further to 83% when endoscopic inflammation is excluded too.

Conclusion: the sensitivity and specificity of RH-DNA in relation to colonoscopy (clinical) results is on the low side to be used for screening. We are currently investigating the use of DNA methylation markers to increase the sensitivity and specificity of CRC detection in fecal samples.

98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA