Characterization of Aberrations in DNA Damage Repair Pathways in Gastrointestinal Stromal Tumors: The Clinicopathologic Relevance of gamma H2AX and 53BP1 in Correlation with Heterozygous Deletions of CHEK2, BRCA2, and RB1

Genetic aberrations involving DNA damage repair (DDR) remain underexplored in gastrointestinal stromal tumors (GISTs). We characterized DDR abnormalities using targeted next-generation sequencing and multiplex ligation-dependent probe amplification, and performed immunofluorescence (IF) and immunohistochemistry (IHC) analyses of gamma H2AX and 53BP1. Consistent with IF-validated nuclear co-localization, gamma H2AX and 53BP1 showed robust correlations in expression levels, as did both biomarkers between IF and IHC. Without recurrent pathogenic single-nucleotide variants, heterozygous deletions (HetDels) frequently targeted DNA damage-sensing genes, with CHEK2-HetDel being the most prevalent. Despite their chromosomal proximity, BRCA2 and RB1 were occasionally hit by HetDels and were seldom co-deleted. HetDels of CHEK2 and BRCA2 showed a preference for older age groups, while RB1-HetDel predominated in the non-gastric, high-risk, and 53BP1-overexpressing GISTs. Higher risk levels were consistently related to gamma-H2AX or 53BP1 overexpression (all p < 0.01) in two validation cohorts, while only 53BP1 overexpression was associated with the deletion of KIT exon 11 (KITex11-del) among genotyped GISTs. Low expressers of dual biomarkers were shown by univariate analysis to have longer disease-free survival (p = 0.031). However, higher risk levels, epithelioid histology, and KITex11-del retained prognostic independence. Conclusively, IHC is a useful surrogate of laborious IF in the combined assessment of 53BP1 and gamma-H2AX to identify potential DDR-defective GISTs, which were frequently aberrated by HetDels and a harbinger of progression.