Implications of Epigenetic Drift in Colorectal Neoplasia

Abstract

Many normal tissues undergo age-related drift in DNA methylation, providing a quantitative measure of tissue age. Here, we identify and validate 781 CpG islands (CGI) that undergo significant methylomic drift in 232 normal colorectal tissues and show that these CGI continue to drift in neoplasia while retaining significant correlations across samples. However, compared with normal colon, this drift advanced (∼3-4-fold) faster in neoplasia, consistent with increased cell proliferation during neoplastic progression. The observed drift patterns were broadly consistent with modeled adenoma-to-carcinoma sojourn time distributions from colorectal cancer incidence data. These results support the hypothesis that, beginning with the founder premalignant cell, cancer precursors frequently sojourn for decades before turning into cancer, implying that the founder cell typically arises early in life. At least 77% to 89% of the observed drift variance in distal and rectal tumors was explained by stochastic variability associated with neoplastic progression, whereas only 55% of the variance was explained for proximal tumors. However, gene-CGI pairs in the proximal colon that underwent drift were significantly and primarily negatively correlated with cancer gene expression, suggesting that methylomic drift participates in the clonal evolution of colorectal cancer. Methylomic drift advanced in colorectal neoplasia, consistent with extended sojourn time distributions, which accounts for a significant fraction of epigenetic heterogeneity in colorectal cancer. Importantly, these estimated long-duration premalignant sojourn times suggest that early dietary and lifestyle interventions may be more effective than later changes in reducing colorectal cancer incidence. SIGNIFICANCE: These findings present age-related methylomic drift in colorectal neoplasia as evidence that premalignant cells can persist for decades before becoming cancerous.See related commentary by Sapienza, p. 437.

Fig. 1:

DNA methylation drift measured in a cancer tissue sample provides a measure of the sojourn time between initiation of the founder premalignant cell and the cancer that arises along this lineage. Premalignant clones may grow gradually for decades prior to generating an observable adenoma or cancer.

Fig. 2.

Estimated CpG drift rates of 182,498 CpG probes vs the (log2) ratio of methylation variance in tumor samples relative to the corresponding variance in normal tissue samples from the Luo study [11]. Variances and drift rates were computed using M-values. The drift rates were estimated using linear regression of methylation vs patient age (in years). CpGs in dark grey undergo significant methylomic drift (q-value <10⁻⁴), CpGs in medium grey are considered static, i.e., do not show significant linear trends with patient age. The subset of CpGs marked in light grey serves as a control group for the analysis of gene expression and methylomic drift (see Results).

Fig. 3.

Drift rate distributions in SMS for 781 CGI with a minimum of 5 identified drift-CpGs by sex (solid curves) versus analogous distributions at the probe-level comprising 12,700 CpGs (dashed curves).

Fig. 4.

Boxplots of the drift rate distributions for the same CGI as in Fig.(2), but validated in samples from the GICR study for left (distal) and right (proximal) colon samples. For each group the individual drift rate estimates are shown as data points. Due to small sample sizes for males and females in right colon, drift rates were determined for both sexes combined in right colon.

Fig. 5.

A) Expected (mean) premalignant sojourn times (in years) for males by age of cancer diagnosis and anatomical site with 95% confidence bands, based on the model fits described in Meza et al. [10] for UK males. B) Sojourn time-dependent drift curves fitted to normal/stromal cell content corrected TCGA (solid symbols) and Luo (empty symbols) methylomic CGI-level drift in tumors by sex and anatomical site. Regression model described in Material and methods. C, D) same as A and B, respectively, but for females.

Fig. 6.

Methylation heat map of 300 static CGI (top rows) and 781 drift-related CGI (bottom rows) for 68 normal samples and 141 TCGA colon cancer samples (left colon only). Sample groups (normal, cancers) are shown ordered by their mean island level methylation.