Liquid biopsies, in particular the profiling of circulating tumor DNA (ctDNA), has become a principle centre of interest in precision oncology and promises to reshape cancer care in the near-future. Despite spectacular progress in this field, however, remarkably little is known about several aspects of ctDNA molecules, including (i) the mechanisms underlying its generation and release into body fluids, (ii) various epigenetic features, and (iii) the plethora of physiological/biological factors that modulate both the genetic and epigenetic characteristics of ctDNA in vivo and in collected biospecimens. Yet, it is becoming increasingly clear that a deeper understanding of these aspects is crucial for the successful development and wide-spread implementation of ctDNA assays into routine clinical diagnostics. On one hand, this knowledge is critical for understanding fluctuations in the baseline values of ctDNA molecules, which informs the development of assays with increased diagnostic sensitivity and specificity. On the other hand, systematic mapping of ctDNA biology will lead to the discovery of increasingly specific biomarkers in relation to various cancer indications.