p53/ Status Assessment in Gastroesophageal Adenocarcinoma.

Chromosomal instability (CIN) is very frequent in gastroesophageal adenocarcinoma (GEA) and it is characterized by deletions/mutations resulting in p53 nuclear accumulation, as revealed by immunohistochemistry (IHC), which considers the cases with "high" staining levels to be positive. Aiming to improve aberrant detection, droplet digital PCR (ddPCR) was used to evaluate deletion in formalin-fixed, paraffin-embedded DNA (FFPE-DNA) and cell-free DNA (cfDNA). To further investigate the mutational profile, next-generation sequencing (NGS) was performed in a subset of FFPE samples. After combining "low" and "high" IHC staining level groups, the proportion of deletion events was significantly higher compared to the "intermediate" group (72.9% vs. 47.5%, -value = 0.002). The ddPCR deletion assay was feasible for cfDNA but only had good agreement (72.7%, Cohen's kappa = 0.48) with the assay performed with FFPE-DNA of the "low-level" group. NGS analysis confirmed that, in the "low-level" group, a high percentage (66.7%) of cases were aberrant, with disruptive mutations that probably led to p53 loss. Data suggested that p53 IHC alone underestimates the CIN phenotype in GEA and that molecular analysis in both solid and liquid biopsies could be integrated with it; in particular, in cases of completely negative staining.