Molecular Detection of Colorectal Neoplasia David A. Ahlquist  Gastroenterology  Volume 138, Issue 6, Pages 2127-2139 (May 2010) DOI: 10.1053/j.gastro.2010.01.055 Copyright © 2010 AGA Institute Terms and Conditions

Figure 1 Digital quantification of low abundance DNA methylation in clinical samples by methyl-beads, emulsion, amplification, and magnetics (BEAMing) method. (A) Methyl-BEAMing laboratory flow. (B) Representative results of methyl-BEAMing obtained with flow cytometry. A number of beads representing methylated vimentin, unmethylated vimentin, a mixture of these, and virgin beads are indicated in the top right corner of each box. Virgin beads represent those in aqueous nanocompartments that did not contain a template molecule. From Li M, Chen WD, Papadopoulos N, et al. Sensitive digital quantification of DNA methylation in clinical samples. Nat Biotechnol 2009;27:858−863, reprinted with permission. Gastroenterology 2010 138, 2127-2139DOI: (10.1053/j.gastro.2010.01.055) Copyright © 2010 AGA Institute Terms and Conditions

Figure 2 Stool DNA (SDT-2) vs occult blood testing for detection of screen-relevant neoplasia (curable-stage colorectal cancer, high-grade dysplasia, and adenomas >1 cm), n = 142. Sensitivity is plotted for Hemoccult and HemoccultSensa with 1, 2, and 3 stools per patient and for SDT-2 on a single stool per patient. Marker panel for SDT-2 comprises methylated vimentin, KRAS mutations, and APC mutations. Vertical lines represent 95% confidence intervals. *P < .001 vs Hemoccult or HemoccultSensa. From Ahlquist DA, Sargent DJ, Loprinzi CL, et al. Stool DNA and occult blood testing for screen detection of colorectal neoplasia. Ann Intern Med 2008;149:441−450, W81, reprinted with permission. Gastroenterology 2010 138, 2127-2139DOI: (10.1053/j.gastro.2010.01.055) Copyright © 2010 AGA Institute Terms and Conditions

Figure 3 Blinded comparison of stool tests for detection of advanced adenomas in the screening setting. A next generation stool DNA test, digital melt curve (DMC), is compared to common fecal blood tests (Hemoccult and HemoccultSENSA (Beckman-Coulter) and a first generation single-base-extension stool DNA test from Exact Sciences. (A) Sensitivities for advanced adenomas with tissue-proven KRAS mutations (n = 27). The DMC assay detected significantly more adenomas than the other tests (P < .05 vs each other test). (B) Specificities based on stools from age- and gender-matched controls; differences were not statistically significant. From Zou H, Taylor WR, Harrington JJ, et al. High detection rates of colorectal neoplasia by stool DNA testing with a novel digital melt curve assay. Gastroenterology 2009;136:459−470, reprinted with permission. Gastroenterology 2010 138, 2127-2139DOI: (10.1053/j.gastro.2010.01.055) Copyright © 2010 AGA Institute Terms and Conditions

Figure 4 Fraction of mutant APC fragments in the plasma of patients with colorectal neoplasms [advanced adenomas (Ad) and Dukes' stage A, B, and D carcinomas]. For each mutation analyzed, DNA from normal lymphoid cells or plasma DNA from healthy donors was used as a control (Normal). The red lines represent the mean, minimum, and maximum values of the normal controls. From Diehl F, Li M, Dressman D, et al. Detection and quantification of mutations in the plasma of patients with colorectal tumors. Proc Natl Acad Sci U S A 2005;102:16368−16373, reprinted with permission. Gastroenterology 2010 138, 2127-2139DOI: (10.1053/j.gastro.2010.01.055) Copyright © 2010 AGA Institute Terms and Conditions

Figure 5 Scatter plots of urinary nucleosides (A) cytidine, (B) adenosine, (C) 8-hydroxy-2′-deoxyguanosine (D) N2,N2-dimethylguanine from colorectal patients (gray squares) and normal controls (white squares). Each variable is directly plotted as a data point. The horizontal lines correspond to the normal group mean ± 2 standard deviations. Although there is some overlap in distributions of urinary marker levels between groups in (A) and (B), marker levels in (C) and (D) discriminated tumors from normal samples. From Hsu WY, Chen WT, Lin WD, et al. Analysis of urinary nucleosides as potential tumor markers in human colorectal cancer by high performance liquid chromatography/electrospray ionization tandem mass spectrometry. Clin Chim Acta 2009;402:31−37, reprinted with permission. Gastroenterology 2010 138, 2127-2139DOI: (10.1053/j.gastro.2010.01.055) Copyright © 2010 AGA Institute Terms and Conditions

David A. Ahlquist Gastroenterology 2010 138, 2127-2139DOI: (10.1053/j.gastro.2010.01.055) Copyright © 2010 AGA Institute Terms and Conditions