Fragment Length of Circulating Tumor DNA

During cell death, DNA that is not contained within a membrane (i.e., cell-free DNA) enters the circulation. Detecting cell-free DNA originating from solid tumors (i.e., circulating tumor DNA, ctDNA), particularly solid tumors that have not metastasized, has proven challenging due to the relatively abundant background of normally occurring cell-free DNA derived from healthy cells. Our study defines the subtle but distinct differences in fragment length between normal cell-free DNA and ctDNA from a variety of solid tumors. Specifically, ctDNA was overall consistently shorter than the fragment length of normal cell-free DNA. Subsequently, we showed that a size- selection for shorter cell-free DNA fragments increased the proportion of ctDNA within a sample. These results provide compelling evidence that development of techniques to isolate a subset of cell-free DNA consistent with the ctDNA fragment lengths described in our study may substantially improve detection of non-metastatic solid tumors. As such, our findings may have a direct impact on the clinical utility of ctDNA for the non-invasive detection and diagnosis of solid tumors (i.e., the “liquid biopsy”), monitoring tumor recurrence, and evaluating tumor response to therapy.