Evidence for Unified Assessment Criteria of HER2 IHC in Colorectal Carcinoma.

HER2 expression is an important biomarker for the management of RAS wild-type metastatic colorectal carcinoma (CRC). Immunohistochemistry (IHC) with reflex in situ hybridization (ISH) is accepted as a standard method of assessment, yet there are currently two sets of criteria used to interpret results: the HERACLES criteria, and the My Pathway criteria. The HERACLES criteria require ISH confirmation when IHC staining is 3+ in 10-49% of cells, while the My Pathway criteria mirror those for gastric HER2 assessment and do not recommend ISH confirmation in the previously referenced scenario. We aimed to assess the prevalence of HER2 3+ heterogeneity and its association with ERBB2 copy number amplification (CNA) to evaluate the necessity of ISH testing when IHC staining is 3+ in <50% of cells. Next-generation sequencing (NGS) of DNA (592-gene panel or whole exome sequencing; WES) was performed for 13 208 colorectal carcinoma (CRC) tumors submitted to Caris Life Sciences (Phoenix, AZ). HER2 (4B5) expression was tested by IHC. A subset of tumors was tested for ERBB2 amplification via chromogenic ISH and/or via NGS (copy number amplification [CNA]). X²/Fisher-Exact tests were applied where appropriate, with p-values adjusted for multiple comparisons (p< 0.05). Of 13 208 CRCs with HER2 IHC, 87.4% (11 541/13 208) were negative for HER2 expression (intensity ≤3+ and < 10% tumor-cell staining) and 11.2% (1 473/13 208) demonstrated at least low HER2 expression (1-2+ and ≥10%). Only 1.5% (194/13 208) of all tested tumors were either positive or heterogeneously positive for HER2 overexpression (3+, ≥10%). Of these, 14% (28/194) had heterogenous HER2 overexpression (3+ staining of 10-49% of cells). Among 22 HER2-positive/heterogenous cases with successful ISH testing, 100% (22/22) demonstrated amplification via ISH. Because the classification of tumors as HER2-positive/heterogenous by IHC correlated very closely with ISH positivity, our results suggest that ISH is likely unnecessary for CRCs with 3+ HER2 overexpression in 10-49% of neoplastic cells.