Detection of TP53 and PIK3CA Mutations in Circulating Tumor DNA Using Next-Generation Sequencing in the Screening Process for Early Breast Cancer Diagnosis

Abstract

Circulating tumor DNA (ctDNA) has emerged as a non-invasive “liquid biopsy” for early breast cancer diagnosis. We evaluated the suitability of ctDNA analysis in the diagnosis of early breast cancer after mammography findings, comparing PIK3CA and TP53 mutations between tumor biopsies and pre-biopsy circulating DNA. Matched plasma and frozen fresh tissue biopsies from patients with Breast Imaging-Reporting and Data System (BIRADS) 4c/5 mammography findings and subsequent diagnosis of primary breast cancer were analyzed using NGS TruSeq Custom Amplicon Low Input Panel (Illumina) and plasma SafeSEQ (Sysmex Inostics). The same plasma and tumor mutations were observed in eight of 29 patients (27.6%) with four in TP53 and five in PIK3CA mutations. Sequencing analysis also revealed four additional ctDNA mutations (three in TP53 and one in PIK3CA) previously not identified in three patients tissue biopsy. One of these patients had mutations in both genes. Age, tumor grade and size, immunohistochemical (IHC) subtype, BIRADS category, and lymph node positivity were significantly associated with the detectability of these blood tumor-derived mutations. In conclusion, ctDNA analysis could be used in early breast cancer diagnosis, providing critical clinical information to improve patient diagnosis.

The Polemic Diagnostic Role of TP53 Mutations in Liquid Biopsies from Breast, Colon and Lung Cancers

“…Later, another meta-analysis was able to demonstrate the diagnostic utility of TP53 mutations in cfDNA for advanced breast cancer with a diagnostic performance of 0.94 [ 43 ]. Frequencies of TP53 mutation reported in these studies agree with some recent ones reporting frequencies of 34% in tissue [ 44 , 45 ] and of 31% [ 46 ] and 32.5% [ 47 ] in plasma of primary breast cancer patients. Nevertheless, mutation frequency is higher in some other studies, showing a TP53 mutation prevalence of 41% [ 45 ], 50% [ 7 ] and 51.7% [ 24 ].…”

“…Of note, the region of study for a particular gene is also of great importance. Most early studies only analyzed TP53 mutations in exons 4 and 8, comprising the majority of the known mutations [ 92 ] and later sequencing studies of the whole gene revealed lower concordances between liquid and solid biopsies, possibly due to the fact that rare mutations might be present at extremely low VAFs to be detected in both specimens [ 44 ]. In addition, genetic alterations such as methylation may also impact on TP53 gene expression without the need for identifying point mutations [ 93 ], thus inclusion of further biomarkers in both solid and liquid biopsies might improve their diagnostic and prognostic utility.…”

“…Madic et al, 2015 also demonstrated 81% concordant results between the two types of biopsies [ 48 ]. Later, 50% discordances for TP53 mutations were found by different authors [ 40 ], [ 50 ], and lower concordant results (36.4% and 22%) were reported by Chae et al, 2017 [ 51 ] and Rodriguez et al, 2019 [ 44 ], respectively. Finally, Delmonico et al, 2019 found no concordance for TP53 mutations in paired cfDNA/tissue samples [ 46 ].…”

Quantitative analysis of the BRAF V595E mutation in plasma cell-free DNA from dogs with urothelial carcinoma

“…We found that the ctDNA concentration in patients with BRAF-mutated urothelial carcinoma is increased compared to that in wild-type patients, demonstrating that the BRAF mutation in urothelial carcinoma may be detectable in the plasma of these canines. CtDNAs with mutations in several genes have been detected in human patients with cancer, and ctDNA analysis provides critical clinical information to improve diagnosis in these patients [6,7,17].…”

Challenges and achievements of liquid biopsy technologies employed in early breast cancer

“…In a recent study, Jimenez-Rodriguez et al. [30] compared the mutational profiles of solid tumours and plasma samples from early breast cancer patients using the amplicon-based SafeSEQ (Sysmex Inostics) technology and a fixed-gene panel for plasma sequencing. Interestingly, the authors observed that plasma DNA sequencing permitted the identification of clonal mutations not detected in tumour biopsy sequencing.…”