Our laboratory has developed cf-RRBS to assess DNA methylation in circulating free DNA, enabling liquid biopsy diagnostics in cancer. However, completed clinical studies revealed that the detection limit of cf-RRBS is 3-5% tumor fraction, which needs to be enhanced for applications in early diagnostics and minimally residual disease. Our goal is to develop the next generation high sensitivity cf-RRBS by incorporating innovation steps so that sensitivity is increased by 10 fold to 0.5%. Next we will examine our method in actionable case studies: early diagnosis of hepatocellular carcinoma (HCC) in cirrhosis patients and prognosis of first-line drug response in diffuse large cell B lymphoma (DLBCL) and ER+ breast cancer. In order to demonstrate cf-RRBS potential in biomarker discovery, we will exploit the data derived from the clinical studies to identify differentially methylated regions associated with these differential diagnosis questions. In future work, these will also be useful for monitoring minimal residual disease upon treatment of these tumor types.