ERBB2 amplification status in 67 salivary duct carcinomas assessed by immunohistochemistry, fluorescence in situ hybridization, and targeted exome sequencing

Abstract

AbstractSalivary duct carcinoma (SDC) is an aggressive salivary gland malignancy with poor survival. Approximately 30% SDC harbor HER2 amplification and response to trastuzumab has been reported. However, a systematic approach for HER2 status assessment in this tumor type has not been established. A total of 67 tumor samples were evaluated for HER2 protein overexpression or ERBB2 gene amplification using at least 2 methods: immunohistochemistry (IHC), fluorescence in situ hybridization (FISH), and/or targeted exome next-generation sequencing (NGS). NGS assessed ERBB2 copy number fold change (FC) and total copy number (TCN). HER2 status was first determined by IHC/FISH according to the 2018 ASCO/CAP breast cancer guidelines. FISH results, the “gold standard”, were compared with the NGS results. All (15/15) IHC positive, 35% (6/17) equivocal, and no (0/19) IHC negative SDC were HER2 amplified by FISH. HER2 FISH signal/cell showed a good correlation with FC (Spearman correlation: 0.708, R2: 0.501, p < 0.0001) and TCN (Spearman correlation: 0.763, R2: 0.582, p < 0.0001). Receiver operating characteristics curve estimation showed an area under curve (AUC) of 0.975 for ERBB2 FC. FC cutoff of ≥1.8 corresponded to an accuracy of 95.2% for ERBB2 amplification (Youden's index: 0.84, sensitivity: 89.47%, specificity: 100%). FC < 1.3 could be reliably classified as ERBB2 not amplified and FC ≥ 1.3 and <1.8 as equivocal. TCN estimation showed AUC of 0.981. TCN cutoff of >6.0 corresponded to an accuracy of 92% for HER2 amplification (Youden's index: 0.81, sensitivity: 81.2%, specificity: 100%). TCN < 4 could be reliably classified as ERBB2 not amplified and TCN ≥ 4.0 and ≤6.0 as equivocal. FC and TCN were binarized with respective cutoffs of ≥1.8 and ≥6.0 and the proportion of agreement with FISH were 95% and 92%, respectively. The assessment of ERBB2 copy number by NGS is accurate and reliable with FC or TCN nearly equivalent to FISH in identifying HER2 amplified SDC.