Abstract
The aim of this study was to determine an optimal workflow to detect TP53 mutations in baseline and longitudinal serum cell free DNA (cfDNA) from high-grade serous ovarian carcinomas (HGSOC) patients and to define whether TP53 mutations are suitable as biomarker for disease. TP53 was investigated in tissue and archived serum from 20 HGSOC patients by a next-generation sequencing (NGS) workflow alone or combined with digital PCR (dPCR). AmpliSeq™-focused NGS panels and customized dPCR assays were used for tissue DNA and longitudinal cfDNAs, and Oncomine NGS panel with molecular barcoding was used for baseline cfDNAs. TP53 missense mutations were observed in 17 tissue specimens and in baseline cfDNA for 4/8 patients by AmpliSeq, 6/9 patients by Oncomine, and 4/6 patients by dPCR. Mutations in cfDNA were detected in 4/6 patients with residual disease and 3/4 patients with disease progression within six months, compared to 5/11 patients with no residual disease and 6/13 patients with progression after six months. Finally, mutations were detected at progression in 5/6 patients, but not during chemotherapy. NGS with molecular barcoding and dPCR were most optimal workflows to detect TP53 mutations in baseline and longitudinal serum cfDNA, respectively. TP53 mutations were undetectable in cfDNA during treatment but re-appeared at disease progression, illustrating its promise as a biomarker for disease monitoring.
Highlights
Epithelial ovarian cancer is the most lethal malignancy among gynecological cancers in the Western world, partly due to the advanced disease stage at the time of diagnosis in most patients [1].A large proportion of these patients will have a recurrence or progression within two years and die of their disease
As TP53 mutations in our archived serum might originate from tumor cells, but can arise from clonal hematopoiesis, we first defined tumor-specific TP53 mutations in tissue by next-generation sequencing (NGS) and immunohistochemistry
Non-synonymous TP53 mutations with nuclear staining in tumor tissue were evaluated in serum cell-free DNA (cfDNA)
Summary
Epithelial ovarian cancer is the most lethal malignancy among gynecological cancers in the Western world, partly due to the advanced disease stage at the time of diagnosis in most patients [1].A large proportion of these patients will have a recurrence or progression within two years and die of their disease. Higher levels of cfDNA were associated with advanced disease stage, high grade, and poorer prognosis [1,8]. It has been reported in an orthotopic mouse model that progression of disease could be monitored by measuring human cfDNA. In this model, the amount of cfDNA correlated significantly with tumor weight [9]. Increased level of cfDNA can be found in patients with benign lesions, inflammatory disease, and tissue trauma and is not a specific tumor biomarker [10]. The presence of tumor-specific genetic alterations in cfDNA could potentially offer a more specific approach
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