Genomic molecular markers to monitor minimal residual disease with a non invasive liquid biopsy in breast cancer patients

Background Circulating cell free DNA (cfDNA) is one of the most intriguing and developing topic in the field of precision and personalized medicine. From 1977, when Leon and colleagues reported an increase in cfDNA in cancer patients, several studies have tried to explain and understand deeper how cfDNA is produced, lasts into the bloodstream, and what kind of information it contains and can be useful to detect and/or manage the disease. Liquid biopsy has already been approved to determine those patients that will benefit from an epidermal growth factor receptor (EGFR)-targeted therapy in non-small cell lung cancer (NSCLC). Nowadays, several studies on clinical applicability and clinical trials are on-going, in order to determine if information obtained from cfDNA can drive clinical decisions. Homologous recombination (HR) is a high-fidelity DNA repair mechanism involved in double-strand DNA (dsDNA) break repair. Recent studies have highlighted a broader involvement of HR status in BRCA wild type breast cancers (BC), in particular in triple negative BC (TNBC). Identification of mutations in the HR pathway can suggest the administration of a specific therapy, such as platinum agents or PARP inhibitors (PARPi). Material & Methods In this work we have collected fresh tissue and blood from 6 BC patients. Plasma was separated with two consecutive centrifugations to completely remove cell and cellular debris. We have extracted genomic DNA (gDNA) from fresh tissue and cfDNA from plasma using commercial kits. We have analysed in parallel both DNAs with the Homologous Recombination Solution™ kit (Sophia™ Genetics) that allows the identification of mutations in exonic regions of 16 genes involved in HR. Results All samples passed target enrichment and sequencing quality controls (QC). In 3 out of 6 patients no mutations were detected in both gDNA and cfDNA. Patients 1 and 6 had mutations in gDNA not detected in cfDNA. Patient 2 had 2 mutations in gDNA and only one of the two was detected in cfDNA. Conclusions This kit can be used to analyse cfDNA, as confirmed by QC reports. Interestingly, the identification of the mutation in TP53 in patient 2’s cfDNA supports the possibility to detect mutations with this kit. The higher variant fraction (VF) in cfDNA compared to gDNA can sustain the possibility to have a broader detection of heterogeneity. The non-recognition of mutations in cfDNA in patients carrying gDNA mutations can be ascribed to the reduced amount of cfDNA analysed and to the fact that tumoral DNA is only a fraction of the total cfDNA. Thus, the sample analysed can be unrepresentative of the circulating DNA pool. The analysis of a higher number of samples will give us a clearer idea of the applicability of this kit to monitor HR mutational status in cfDNA of BC patients, giving us statistical significance of detection.