Current Challenges of Methylation-Based Liquid Biopsies in Cancer Diagnostics

Simple Summary This review discusses the importance of advancements in cancer diagnosis and treatment, emphasizing the need for early detection methods. It highlights the role of DNA methylation, an epigenetic change, as a promising marker for detecting cancer. This review explains how DNA methylation patterns differ between cancerous and healthy cells, influencing gene expression and contributing to genomic instability. It also delves into the challenges and methods associated with detecting DNA methylation, particularly in cell-free DNA (cfDNA) from bodily fluids, known as liquid biopsies. Different tests for single-cancer detection, such as colorectal, lung, and bladder cancer, as well as multi-cancer detection tests, are discussed along with their methodologies and clinical applications. Biological factors influencing DNA methylation, such as age, gender, and environmental influences, are also explored. The conclusion emphasizes the potential of cfDNA methylation analysis in cancer diagnostics but acknowledges the current limitations and challenges, highlighting the need for further research and advancements in methodology and understanding.Abstract In current clinical practice, effective cancer testing and screening paradigms are limited to specific types of cancer, exhibiting varying efficiency, acceptance, and adherence. Cell-free DNA (cfDNA) methylation profiling holds promise in providing information about the presence of malignity regardless of its type and location while leveraging blood-based liquid biopsies as a method to obtain analytical samples. However, technical difficulties, costs and challenges resulting from biological variations, tumor heterogeneity, and exogenous factors persist. This method exploits the mechanisms behind cfDNA release but faces issues like fragmentation, low concentrations, and high background noise. This review explores cfDNA methylation's origins, means of detection, and profiling for cancer diagnostics. The critical evaluation of currently available multi-cancer early detection methods (MCEDs) as well as tests targeting single genes, emphasizing their potential and limits to refine strategies for early cancer detection, are explained. The current methodology limitations, workflows, comparisons of clinically approved liquid biopsy-based methylation tests for cancer, their utilization in companion diagnostics as well as the biological limitations of the epigenetics approach are discussed, aiming to help healthcare providers as well as researchers to orient themselves in this increasingly complex and evolving field of diagnostics.