Cell-free DNA (cfDNA) methylation profiling has emerged as a promising non-invasive approach for early cancer detection, yet breast cancer remains particularly difficult to identify via liquid biopsy. Targeted methylation-based approaches have shown strong performance across cancer types, but breast cancer-specific signal remains elusive. Integrating whole-genome cfDNA methylation with matched tumor multi-omics data offers an opportunity to identify more sensitive and specific epigenetic biomarkers for early detection.
Plasma cfDNA was obtained from breast cancer patients through the Helsinki Biobank, matched to tumor samples available in the iCAN Digital Precision Cancer Medicine Flagship. Two complementary whole-genome (WG) cfDNA methylation sequencing approaches were applied. WG EM-seq was performed on 52 plasma samples from 26 individuals, each with a pre-diagnostic (mean 21 months prior; range 6–47 months) and a post-diagnostic timepoint (<2 months after diagnosis). WG TAPS+ is being applied to an expanded cohort of 181 samples from unique individuals with single timepoints post-diagnosis. Sequencing reads were processed using DRAGEN and nf-core/methylseq and integrated with matched tumor multi-omics data on the iCAN Discovery Platform.
cfDNA concentrations in the EM-seq cohort ranged from 36 to 1,170 ng/mL (mean 198 ng/mL) at a median coverage depth of 6.5× and a 95% alignment rate. Initial exploratory analysis of genome-wide methylation profiles revealed preliminary clustering patterns distinguishing breast cancer samples from external healthy controls at both pre- and post-diagnostic timepoints, suggesting a detectable epigenetic signal before diagnosis. Copy number profiles called using ichorCNA were largely copy-number neutral across the cohort, with a single case exhibiting detectable alterations.

Co-authors: Rodos Rodosthenous, Maare Arffman, Ican, Sirpa Leppä, Andrea Ganna, Esa Pitkänen