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The 2013 update of the American Society of Clinical Oncology-College of American Pathologists (ASCO-CAP) human epidermal growth factor receptor 2 (HER2) testing guidelines recommend using an alternative chromosome 17 probe assay to resolve HER2 results determined to be equivocal by immunohistochemistry (IHC) or fluorescence in-situ hybridization (FISH). However, it is unclear if cases considered HER2-positive (HER2 + ) by the alternative probe method are similar to those classified as HER2 +  by traditional IHC and FISH criteria and benefit the same from HER2-targeted therapies. We studied the clinical and pathologic features of all 31 breast cancers classified as HER2 +  by the alternative probe method at our institution since 2013 and determined their PAM50 intrinsic molecular subtypes. For comparison, we analyzed 19 consecutive cases that were classified as HER2 +  by traditional FISH criteria during the same time period. Thirty (97%) cancers in the alternative probe cohort were estrogen receptor (ER)-positive (ER + ), while only 9/19 (47%) of traditional HER2 controls were ER +  ( p  = 0.0002). Sufficient tissue for intrinsic subtype analysis was available for 20/31 cancers in the alternative probe cohort and 9/19 in the traditional HER2 +  group. None (0%) of the 20 alternative probe-positive cases were of the HER2-enriched intrinsic subtype, while 8/9 (89%) of those HER2 +  by traditional FISH criteria were HER2-enriched ( p  = 0.0001). These findings suggest that breast cancers classified as HER2 +  only by the alternative probe method are biologically distinct from those classified as HER2 +  by traditional criteria, and raises questions as to whether or not they derive the same benefit from HER2-targeted therapies.

Intrinsic molecular subtypes of breast cancers categorized as HER2-positive using an alternative chromosome 17 probe assay