Background:
Somatic molecular profiling is more important than ever, as precision treatment for ovarian cancer advances. In lieu of a tissue biopsy, tumour material for profiling can be accessed via malignant ascites, a common manifestation of ovarian cancer, however cancer cells are often sparse in this fluid and cell-free DNA (cfDNA) is not well studied.
Objectives:
We aimed to identify clinically relevant variants in cfDNA from ascites and to examine tumour evolution by comparing matched samples or historical biopsies from different points in the disease trajectory.
Methods:
Ascites-derived cfDNA (n=14 participants, aged 36-82) was sequenced with the Illumina TSO-500 assay, along with DNA from ascites-derived tumour cells, archived FFPE-tissue from surgery and cfDNA from sequential ascites samples (interval range, 13-559 days). cfDNA from one patient was additionally analysed using an Oxford Nanopore Technology R9.4.1 MinION.
Results:
Abundant cfDNA was identified in all ascites samples (up to 660 ng/mL), achieving similar alignment success and improved coverage compared to cell or FFPE-derived DNA. Somatic ovarian cancer-associated driver mutations were detected in 100% of cfDNA samples at mutation fractions of up to 79%. All clinically known variants (including 6 in BRCA1 or BRCA2) were identified in ascites cfDNA and cells, aside from in one case, where a TP53 mutation identified in FFPE tissue was not present in ascites, sampled over 9 months later, due to tumour evolution. In this case, we found tumour-derived cfDNA to contain a deletion of the entire p-arm of chromosome 17, where TP53 is located, explaining the mutation’s absence in the later samples. This was indicated by a significant decrease in Oxford Nanopore sequencing reads aligning to the chromosome 17 p-arm relative to the q-arm, adjusted for size (p=0.0015). With non-parametric analyses, we found a significant increase in tumour mutational burden between samples after multiple lines of chemotherapy, further indicating tumour evolution (p=0.042).
Conclusions:
Our work demonstrates the reliability of using cfDNA from ascites for molecular profiling, allowing a liquid biopsy of ovarian cancer when tumour tissue access is restricted. We show that equivalent information can be attained from cfDNA from ascites as from traditional biopsies. This approach provides better access to tumour tissue, allowing capture of clinically actionable mutations prior to surgery or upon recurrence, following tumour evolution.