Simple SummaryCirculating tumor DNA (ctDNA) has emerged as a potential biomarker for clear cell renal cell carcinoma (ccRCC). However, the ctDNA characteristics have not been demonstrated in small ccRCC. The aim of our pilot study is to explore the characteristics of ctDNA in pT1a ccRCC (ccRCC less than 4 cm in diameter). We included 53 patients with pT1a ccRCC, a greater number than in previous studies using next-generation sequencing. We found that the ctDNA detection rate was low in pT1a ccRCC. The relationship between ctDNA and clinicopathological features, such as tumor size, tumor grade, and patient age, was not clear. Increasing the sensitivity and removing background noise in ctDNA analysis may aid in further understanding the characteristics of ctDNA, thereby enhancing its clinical utility in pT1a ccRCC.AbstractCirculating tumor DNA (ctDNA) is a promising biomarker for clear cell renal cell carcinoma (ccRCC); however, its characteristics in small renal masses of ccRCC remain unclear. In this pilot study, we explored the characteristics of ctDNA in pT1a ccRCC. Plasma samples were collected preoperatively from 53 patients with pT1a ccRCC. The ctDNA of pT1a ccRCC was profiled using next-generation sequencing and compared with that of higher-stage ccRCC. The association of ctDNA in pT1a ccRCC with clinicopathological features was investigated. The positive relationship of mutations between ctDNA and matched tissues was evaluated. In pT1a ccRCC, the ctDNA detection rate, cell-free DNA concentration, and median variant allele frequency were 20.8%, 5.8 ng/mL, and 0.38%, respectively, which were significantly lower than those in metastatic ccRCC. The ctDNA gene proportions in pT1a samples differed from those in metastatic ccRCC samples. The relationships between ctDNA and tumor size, tumor grade, and patient age were not elucidated. The positive concordance between ctDNA and matched tissues was poor for pT1a ccRCC. Strategies are needed to increase sensitivity while eliminating noise caused by clonal hematopoiesis to increase the clinical utility of ctDNA analysis in small renal masses of ccRCC.