Using DNA Methylation for Advancements in Early Cancer Detection

This blog post was written by Hologic Diagenode, a leading provider of epigenomics with more than 20 years of expertise in the field. Their services are available on the Scientist.com marketplace.

Epigenomics is rapidly gaining recognition as a crucial approach in the field of cancer research. Although an individual’s gene set remains constant, the expression of these genes can vary significantly during cancer development. Therefore, analyzing epigenomic methylation profiles yields more comprehensive insights than traditional genomic sequencing alone. By tracking these profiles over time, researchers can detect changes in gene expression related to carcinogenesis, treatment responses or relapse as they occur.

Liquid biopsies, which involve the collection of blood and other bodily fluids containing tiny fragments of cell-free DNA (cfDNA), are increasingly favored for their non-invasive nature. Paired with sensitive techniques capable of analyzing minimal DNA input, liquid biopsies can be performed regularly to capture dynamic changes in cellular states. These biopsies are particularly promising for the early detection of cancer, potentially identifying the disease before conventional methods reveal malignancies, thus increasing the chance of survival. With assays that are both sensitive and specific, changes in cancer progression can be monitored during treatment, thereby informing therapeutic decisions.

In collaboration with Universal Diagnostics (UDx), we have contributed our two decades of epigenomic knowledge to aid in the development of an early-detection workflow for colorectal cancer (CRC) that utilizes liquid biopsies. Our expertise facilitated the creation, validation and implementation of an epigenomic protocol based on DNA methylation, tailored for low-sample input scenarios.

During the discovery phase, UDx conducted genome-wide methylation analyses on samples from both healthy individuals and cancer patients. This process led to the identification of DNA methylation sites with the potential to serve as biomarkers.

A novel, tailored validation approach for these biomarkers was conceived, employing targeted methyl-seq technology (Figure 1). Specifically, a bead-based cfDNA extraction method was employed to optimize yield and ensure reproducibility from 4 ml plasma samples. Given that the input material was scarce and extensively fragmented, methyl-seq library preparation was conducted using an enzymatic conversion approach rather than bisulfite treatment to minimize loss and prevent further degradation of the cfDNA. Furthermore, to address the challenges posed by the limited initial cfDNA input, libraries were pooled before undergo probe capture and sequencing, thereby enhancing the robustness of the workflow.

The approach proved to be highly reproducible for analysing both hypomethylated and hypermethylated CpG sites within the biomarker signature (Figure 2).

Given the scarce presence of cfDNA in plasma, numerous parameters were evaluated to ensure the reliability of the results. Notably, spike-ins were incorporated into samples to determine conversion efficiency based on the DNA low input range. Our findings indicated that the targeted methyl-seq workflow was exceptionally consistent and maintained accuracy at the conversion level despite the low DNA input used (Figure 3).

Hologic Diagenode has partnered with UDx to further the development of a tailored workflow for their blood-based methyl-seq assay. The finalized workflow boasts reproducibility even at low DNA input with high specificity and sensitivity, making it suitable for early-detection colorectal cancer screening.

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