PhD defence S.R. (Silvia) Vitale

Dissertation in short:

The purpose of this thesis was to improve the prognostic and predictive value of circulating biomarkers (cell free DNA (cfDNA) and extracellular vesicles) by the optimization of pre-analytical and post-analytical conditions as well as the choice of the suitable methods to investigate rare mutations in biofluids of patients with various types of tumors. In this context, I provided evidence that the QiaSymphony and Maxwell AX1115 automated platforms enabled fast, accurate and high throughput cfDNA isolation suitable for subsequent molecular analyses of cell free tumor DNA (ctDNA). I also demonstrated that a developed NGS workflow, used alone and combined with dPCR, enabled detection of TP53 mutations at baseline and in longitudinal serum-derived cfDNA from high-grade serous ovarian carcinomas samples. Moreover, I provided a workflow to isolate and characterize EV-RNA from cancer patients. Similarly, I developed a reliable and reproducible dPCR method to investigate the mESR1 status for four hotspot mutations in minute amounts of cfDNA. Finally, I also investigated the occurrence of ESR1-CCDC170 genomic rearrangements and showed that their presence are not predictive for outcome in breast cancer with advanced disease treated with endocrine treatment but rather related to the prognosis of breast cancer cases with primary disease. Overall, this thesis has delivered pre-analytical and analytical tools to improve the use of cfDNA for diagnostic purposes in metastatic cancer patients.