Clinical significance of the mutational landscape and fragmentation of circulating tumor DNA in renal cell carcinoma

Reliable biomarkers for renal cell carcinoma (RCC) have yet to be determined. Circulating tumor DNA (ctDNA) is an emerging resource to detect and monitor molecular characteristics of various tumors. The present study aims to clarify the clinical utility of ctDNA for RCC. Fifty-three patients histologically diagnosed with clear cell RCC were enrolled. Targeted sequencing was carried out using plasma cell-free DNA (cfDNA) and tumor DNA. We applied droplet digital PCR (ddPCR) to validate detected mutations. cfDNA fragment size was also evaluated using a microfluidics-based platform and sequencing. Proportion of cfDNA fragments was defined as the ratio of small (50-166 bp) to large (167-250 bp) cfDNA fragments. Association of mutant allele frequency of ctDNA with clinical course was analyzed. Prognostic potential was evaluated using log-rank test. A total of 38 mutations across 16 (30%) patients were identified from cfDNA, including mutations in TP53 (n = 6) and VHL (n = 5), and median mutant allele frequency of ctDNA was 10%. We designed specific ddPCR probes for 11 mutations and detected the same mutations in both cfDNA and tumor DNA. Positive ctDNA was significantly associated with a higher proportion of cfDNA fragments (P = .033), indicating RCC patients with ctDNA had shorter fragment sizes of cfDNA. Interestingly, the changes of mutant allele frequency in ctDNA concurrently correlated with clinical course. Positive ctDNA and fragmentation of cfDNA were significantly associated with poor cancer-specific survival (P < .001, P = .011). In conclusion, our study shows the clinical utility of ctDNA status and cfDNA fragment size as biomarkers for prognosis and disease monitoring in RCC.