Comparison of HER2 Testing by IHC/FISH and RT-PCR in Estrogen Receptor (ER) Negative or Borderline Patients (pts) with Early Stage Breast Cancer (ESBC)
Baljit Singh, Nelli Ziguridis, Steve Butler, Farid Jamshidian, Diana Cherbavaz, Amy Sing. New York University School of Medicine, New York, NY; Genomic Health, Inc., Redwood City, CA
Background: Accurate determination of HER2 status is critical for making HER2 targeted therapy treatment (Tx) decisions. HER2 concordance between analytical methods (IHC, FISH & RT-PCR) is ≥95% (Baehner JCO 2012, Baehner ASCO Breast 2008). Lack of agreement between IHC/FISH and RT-PCR has been recently reported (Dabbs JCO 2011). A study to evaluate new CAP-mandated IHC positive cutoffs for ER protein expression using ER mRNA expression assessed by RT-PCR was undertaken in consecutive ESBC pts from a single institution. Here we report the agreement of HER2 assessments by standardized IHC, FISH and RT-PCR.
Design: Consecutive IHC-defined ER-negative (<1% positive cells) or ER-borderline (1-10% positive cells) ESBC pts were identified at NYU School of Medicine. HER2 protein assessment used standard IHC (4B5, Ventana) with reflex testing of HER2 2+ cases by FISH (defined as ≥2.0) using the FDA-approved HER2 DNA Probe Kit (Vysis). Hybridization results were recorded and analyzed by BioView Duet system (Allegro Plus Automated Scanner) with HER2 application software (BV-HER2-AF). Quantitative HER2 mRNA assessment used the Oncotype DX® assay, reference-normalized expression measurements ranged from 0 to 15, where each 1-unit increase reflects about a two-fold increase in RNA and HER2 categories used pre-specified reference-normalized values (positive≥11.5, equivocal ≥10.7 to <11.5, negative <10.7).
Results: Of 140 evaluable samples, 106 (76%) were ER(-) and 34 (24%) were ER-borderline by IHC. Pts characteristics included: 69.3% were >50 yrs; 83.6% had poor differentiation; 31.2% had >1 positive nodes; 34.3% were > 2.0 cm; 26.4% were HER2(+) by IHC/FISH. There was 83.8% positive agreement (PPA) between HER2 (+) pts by IHC/FISH and RT-PCR (88.6% PPA by ASCO/CAP guidelines). Percent negative agreement (PNA) was 99.0% between HER2 (-) cases by IHC/FISH and RT-PCR. Of the 32 HER2-IHC 3+ pts, 93.8% were HER2 (+) and 6.2% were equivocal by RT-PCR. Table shows cross classification of HER2 status by RT-PCR versus IHC/FISH.
Conclusions: Accurate and precise measurement of HER2 is important for Tx decision making. In this subset analysis of ESBC pts with borderline or negative ER by IHC there is a high degree of PPA and PNA between HER2 assessment by standardized methods by IHC/FISH and quantitative RT-PCR.