A new study introduces a sensitive and cost-effective method for MCED and diagnostic using multimodal cfTAPS, enhancing the potential of liquid biopsies. In this post I’ll take a dive into the new paper, lay out a short overview of DNAme-mod methods development, and the possible implications for CRC screening and Exact’s ColoGuard in particular.

In the quest for early cancer detection, liquid biopsies have emerged as a promising tool. These minimally invasive tests analyze circulating tumor DNA (ctDNA) in the bloodstream to identify cancer presence. This paper from Anna Schuh’s team in Oxford: “Multimodal cell-free DNA whole-genome TAPS is sensitive for early cancer detection” presents an innovative approach, which enhances the utility, sensitivity and cost-effectiveness of these tests.

Understanding the New Method

Traditional methods of ctDNA analysis often face challenges in sensitivity and cost, limiting their widespread application. The improvements to TAPS (TET-assisted pyridine borane sequencing), offers a solution by providing a more sensitive detection method at a reduced cost. This advancement could make early cancer detection more accessible and reliable.

In the paper, the authors (Dimitrios Vavoulis and Anna Schuh) present a novel approach to analysis of circulating tumour DNA (ctDNA) epigenomes using TAPS: a less destructive method than the more commonly used bisulfite, which also enables simultaneous methylome and genome analysis in cancer patients.

Key Findings

The study demonstrates that the multimodal TAPS approach can detect ctDNA with higher sensitivity compared to existing methods. This increased sensitivity is crucial for identifying cancers at an early stage when treatment options are more effective. Additionally, the cost-effectiveness of this method could facilitate broader implementation in clinical settings, potentially leading to earlier diagnoses and improved patient outcomes.

The authors employed 80x TAPS WGS to analyze cfDNA from cancer patient blood samples. This combination allowed for a comprehensive assessment of genetic and epigenetic alterations associated with various cancers i.e. SNV, CNV, fragmentome and DNA methylation. By integrating multiple data types, the method enhances the accuracy of cancer detection. Most importantly to other DNA methylation methods, this allowed the team to develop TAPS-aware analytical tools that can distinguish the TAPS mC>T from oncogenic C>T variants.

DNA methylation methods: a brief summary

Traditional bisulfite sequencing has been the gold standard for detecting 5-methylcytosine (5mC); however, it has limitations, including DNA degradation and inability to distinguish between 5mC and 5-hydroxymethylcytosine (5hmC). To overcome these challenges, alternative methods have been developed, such as TET-assisted pyridine borane sequencing (TAPS), oxidative bisulfite sequencing (oxBS-seq), and enzymatic methyl sequencing (EM-seq).

Here’s a very quick overview of those methods:

1. Oxidative Bisulfite Sequencing (oxBS-seq): Developed by Booth et al. in 2012, oxBS-seq differentiates between 5mC and 5hmC by selectively oxidizing 5hmC to 5-formylcytosine (5fC) before bisulfite treatment. This allows for the distinction between 5mC and 5hmC at single-base resolution. However, the method still relies on bisulfite treatment, which can cause DNA degradation and sequencing bias.
2. TET-assisted pyridine borane sequencing (TAPS): Introduced in 2019, TAPS offers a bisulfite-free method for detecting 5mC and 5hmC at base resolution. This technique utilizes mild chemical reactions to convert 5mC and 5hmC into thymine and cytosine, respectively, allowing for direct sequencing without the harsh conditions associated with bisulfite treatment. As a result, TAPS preserves DNA integrity, reduces sequencing bias, and provides a more accurate representation of the methylome. Available from Exact Sciences and Watchmaker Genomics (see Genomeweb).
3. Enzymatic Methyl Sequencing (EM-seq): Introduced in 2021, EM-seq employs a series of enzymatic reactions to detect 5mC and 5hmC without bisulfite treatment. TET2 and T4-BGT enzymes protect 5mC and 5hmC from deamination, while APOBEC3A deaminates unmodified cytosines. This approach preserves DNA integrity and provides single-base resolution of methylation patterns. However, the method requires multiple enzymatic steps, which may complicate the workflow. v2 now available from NEB.
4. Duet multimodal EvoC: Introduced in 2023, EvoC copys DNA strands and enzymatically converts bases to allow for coupled decoding across the original and copied strands. This captures both genetic and epigenetic information at single-base resolution. This streamlined approach simplifies analysis and provides insights into both genetic and epigenetic features. Available from Biomodal.

Compared to these methods, cfTAPS offers a simpler workflow by avoiding both bisulfite treatment and multiple enzymatic steps (I’d be interested in seeing a more detailed assessment of the complexity of the various protocols). Its mild chemical conversion preserves DNA integrity and provides accurate detection of 5mC and 5hmC at base resolution.

Implications and Future Directions

The development of this multimodal cfTAPS approach represents a significant step forward in the field of liquid biopsies. Its increased sensitivity and cost-effectiveness could lead to more widespread use in clinical practice, ultimately improving early cancer detection rates. Future research may focus on validating this method across larger and more diverse patient populations, as well as exploring its applicability to different cancer types. But, of course, the other DNA methylation methods available are going to be chomping at the heels of TAPS, Exact and Watchmaker!

Potential Applications in Colorectal Cancer Detection

The link with Exact Sciences, who obtained the TAPS IP through its acquisition of Base Genomics in 2020 suggests that cfTAPS is going to be coming to colorectal cancer early detection in the long-term. DNA methylation plays a significant role in colorectal cancer (CRC) development. Hypermethylation of promoter regions in tumor suppressor genes can lead to gene silencing, contributing to carcinogenesis. Detecting these methylation changes in cfDNA offers a promising avenue for early CRC detection.

Exact Sciences’ Cologuard® is a non-invasive, stool-based colorectal cancer (CRC) screening test designed for average-risk individuals aged 45 and older. The test analyzes stool samples for specific DNA markers and the presence of blood (hemoglobin) to detect potential cancerous or precancerous conditions. Cologuard Plus™, the next-generation version of the test, was FDA approved in 2024 based on data in the “Next-Generation Multitarget Stool DNA Test for Colorectal Cancer Screening” published in NEJM. In this paper the team evaluated the enhanced test’s efficacy and demonstrated that the next-generation multitarget stool DNA test showed 93.9% sensitivity for colorectal cancer and 43.4% sensitivity for advanced precancerous lesions.

Perhaps importantly for cfTAPS, Cologuard Plus incorporates several methylated DNA markers (LASS4, LRRC4, PPP2R5C and ZDHHC1, and Exact could leverage TAPS technology to enhance their CRC screening tools and detect CRC-specific methylation patterns with higher sensitivity and specificity. Interstingly, the Exact Sciences BLUE-C study “also included blood collection for later evaluation of a blood-based screening test being developed by Exact Sciences” so cfTAPS and other things could be on the cards.

In conclusion, TAPS represents a significant advancement in DNA methylation detection, offering a bisulfite-free, accurate, and efficient method for epigenetic analysis. Its application in cfDNA analysis holds great potential for early cancer detection, particularly in colorectal cancer, and could lead to the development of more effective diagnostic tools.