Combining circulating tumor cell and circulating cell-free DNA analyses broadens clinical applicability of liquid biopsy in high grade serous tubo-ovarian carcinoma.

Liquid biopsy is a promising strategy for detecting and monitoring neoplastic diseases, with circulating tumor cells (CTCs) and circulating tumor DNA (ctDNA) being the most common objects of investigation. Studies are mainly focusing on these biomarkers separately, and simultaneous detection has never been attempted in high grade serous tubo-ovarian carcinoma (HGSOC). Here, we assess whether tandem CTC/ctDNA analysis improves the efficiency of detecting HGSOC via peripheral blood liquid biopsy. For CTC identification, gene expression assays and TP53 next-generation sequencing (NGS) were tested using healthy donor samples spiked with known cancer cell numbers. Both approaches detected as few as five spiked cancer cells, showing high analytical sensitivity and specificity. Validation in HGSOC patients and healthy controls revealed better performance of TP53 NGS, as it correctly identified the disease in 47% liquid biopsies, compared to 13% sensitivity obtained by gene expression assay. TP53 NGS was also applied for ctDNA detection, where analytical validity was ensured by calculating 0.31% as the optimal variant allele frequency threshold for mutation calling. Clinical validation demonstrated that ctDNA approach, with sensitivity of 70%, outperformed CTC-based methods. Combining ctDNA/CTC analysis improved disease detection rate in two HGSOC cohorts, achieving, respectively, 73.3% and 93.3% sensitivity, which would translate to an absolute gain of additional 13 patients per 100 cases being detected if combined approach is applied compared to ctDNA method alone. Interestingly, we found private CTC variants, and shared ctDNA/CTC mutations undetected in solid biopsy, highlighting the ability of dual-analyte approach to capture tumor heterogeneity and allow mutation cross-validation, which is particularly useful in contexts when solid biopsy is not available. Our study reveals the complementary value of simultaneous ctDNA/CTC analysis in HGSOC, advancing the translational potential of liquid biopsy integration for management of this disease, especially regarding early detection, chemotherapy response evaluation and relapse detection.