Noninvasive Detection of Bladder Cancer by Shallow-Depth Genome-Wide Bisulfite Sequencing of Urinary Cell-Free DNA for Methylation and Copy Number Profiling

Background: The current diagnosis and monitoring of bladder cancer are heavily reliant on cystoscopy, an invasive and costly procedure. Previous efforts in urine-based detection of bladder cancer focused on targeted approaches that are predicated on the tumor expressing specific aberrations. We aimed to noninvasively detect bladder cancer by the genome-wide assessment of methylomic and copy number aberrations (CNAs). We also investigated the size of tumor cell-free (cf)DNA fragments. Methods: Shallow-depth paired-end genome-wide bisulfite sequencing of urinary cfDNA was done for 46 bladder cancer patients and 39 cancer-free controls with hematuria. We assessed ( a ) proportional contribution from different tissues by methylation deconvolution, ( b ) global hypomethylation, ( c ) CNA, and ( d ) cfDNA size profile. Results: Methylomic and copy number approaches were synergistically combined to detect bladder cancer with a sensitivity of 93.5% (84.2% for low-grade nonmuscle-invasive disease) and a specificity of 95.8%. The prevalence of methylomic and CNAs reflected disease stage and tumor size. Sampling over multiple time points could assess residual disease and changes in tumor load. Muscle-invasive bladder cancer was associated with a higher proportion of long cfDNA, as well as longer cfDNA fragments originating from genomic regions enriched for tumor DNA. Conclusions: Bladder cancer can be detected noninvasively in urinary cfDNA by methylomic and copy number analysis without previous knowledge or assumptions of specific aberrations. Such analysis could be used as a liquid biopsy to aid diagnosis and for potential longitudinal monitoring of tumor load. Further understanding of the differential size and fragmentation of cfDNA could improve the detection of bladder cancer.