Pan-cancer multi-omic ERBB2-HER2 characterization using next-generation sequencing and quantitative proteomics

The recent successes of HER2-targeting agents, even in tumors characterized by FDA-approved molecular testing as HER2-negative or non-amplified, have underscored the limitations of current diagnostic approaches for accurately identifying patients with actionable HER2/EGFR activation/phosphorylation. We therefore performed a multi-omic investigation integrating clinical next-generation sequencing with a CLIA-certified reverse-phase protein array (RPPA) assay and laser microdissection-enriched tumor samples to characterize ERBB2/HER2 at the DNA, RNA, protein, and phosphoprotein level in patients with advanced pan-cancer solid tumor malignancies. Functional pathway activation mapping by RPPA revealed several patients with ERBB2 genomic or transcriptomic alterations and/or HER2^Total-positivity by immunohistochemistry who exhibited no significant HER2^Y1248 activation/phosphorylation. In contrast, other patients lacking ERBB2 genomic/transcriptomic alterations demonstrated significant HER2^Y1248 activation/phosphorylation with co-activation of EGFR^Y1173, a marker associated with prognostic significance. Our results highlight the weak concordance between ERBB2 genomic/transcriptomic alterations and downstream activation of HER family signaling and support the inclusion of functional proteomic/phosphoproteomic analysis as an essential component of precision oncology pipelines to more accurately guide selection of HER2- and EGFR-targeted therapies.