PURPOSE

While HER2 testing is well established in directing appropriate treatment for breast cancer, a small percentage of cases show equivocal results by IHC and FISH. Alternative probes may be used in equivocal cases. We present a single community-based institution's experience in further evaluating these cases.

PATIENTS AND METHODS

Between 2014-2016, 4255 samples were submitted for HER2 amplification testing by alternative probes, TP53, RAI1, and RARA. Of patients tested by FISH, 505/3908 (12.9%) also had IHC data.

RESULTS

Most (73.9%) FISH equivocal cases remained equivocal after IHC testing. Duplicate testing by IHC using a different paraffin block resulted in poor reproducibility as 44% of IHC negative became equivocal and 40% of IHC equivocal became negative. Of the equivocal cases by IHC, 52.3% became positive by alternative FISH. However, 50.5% of cases equivocal by conventional FISH were classified as HER2 amplified by alternative probes. Most cases were positive by more than one probe; 78% of positive cases were positive by RAI1 and 73.9% by TP53. There was a significant difference between IHC and FISH alternative testing (P<0.0001) among the equivocal cases by conventional FISH testing, 44% of IHC negative cases became positive while 36% of the positive IHC cases became negative by alternative FISH testing. Available data showed that 41% of patients were treated with palbociclib and were positive by alternative FISH.

CONCLUSION

The prevalence of double HER2 equivocal cases and the discrepancy between IHC and alternative FISH testing suggest that FISH alternative testing using both RAI1 and TP53 probes is necessary for conclusive classification. Due to the significant difference between IHC repeat testing, reflexing on IHC alone in this group of cases may not be reproducible. Because almost half of FISH equivocal cases converted to HER2 amplified upon alternative testing, clinical studies to determine benefit of anti-HER2 therapy in these patients is urgently needed.

Alternative FISH IHC Score 0 Negative IHC Score 1 Negative IHC Score 2 Equivocal IHC Score 3 Positive 
 No No No No 
Negative 14 63.6 54 54.5 178 47.7 36.4 
Positive 36.4 45 45.5 195 52.3 63.6 
Total 22 100.0 99 100.0 373 100.0 11 100.0 
Alternative FISH IHC Score 0 Negative IHC Score 1 Negative IHC Score 2 Equivocal IHC Score 3 Positive 
 No No No No 
Negative 14 63.6 54 54.5 178 47.7 36.4 
Positive 36.4 45 45.5 195 52.3 63.6 
Total 22 100.0 99 100.0 373 100.0 11 100.0 

Citation Format: Sally Agersborg, Christopher Mixon, Thanh Nguyen, Sramila Aithal, Forrest Blocker, Lawrence Weiss, Robert Gasparini, Shiping Jiang, Wayne Chen, Gregory Hess, Maher Albitar. Immunohistochemistry and alternative FISH testing in breast cancer with HER2 equivocal amplification [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4524.