Validation of Dako DuoCISH HER2 Gene Amplification Assay
NA Granger, CM Haynes, A Quinn, X Wang, P Tang, L McMahon, H Yaziji, D Hicks, LM Schiffhauer. University of Rochester, Rochester; Vitro Molecular Laboratories, Miami; SUNY Syracuse, Syracuse; Stanford University, Stanford
Background: Over-expression and amplification of HER2 have been identified as poor prognostic factors in breast cancer and as predictive markers of response to targeted therapy. Standard laboratory methods used to assess HER2 status include immunohistochemistry (IHC), fluorescent in situ hybridization (FISH) and chromogenic in situ hybridization (CISH). Until now, the only commercially available kit for CISH used single color visualization of a labeled HER2 probe and lacked a probe for chromosome 17. Inclusion of a chromosome 17 probe is important to distinguish pseudo-amplification of the HER2 gene (polysomy of chromosome 17) from true gene amplification. The aim of this study was to validate a new commercially available CISH HER2 gene amplification assay (DuoCISHTM, Dako) that allows for dual color chromogenic visualization of signals from both HER2 and chromosome 17 probes.
Design: Breast cancer and normal tissue microarrays (TMAs) containing cores from 131 patients at one University Hospital were constructed. HER2 status was determined by validated IHC (HerceptestTM, Dako) and FISH (PathVysion Vysis), and compared with CISH (DuoCISHTM, Dako). All assays were performed and interpreted (blinded to other assays) according to the manufacturers guidelines. The results of CISH were compared to both IHC and FISH.
Results: A comparison of CISH and IHC (71) showed 100% agreement between CISH amplified and IHC 2+-3+, n=7; and 100% agreement between CISH not amplified and IHC 0-1+ (negative), n=59. Some cores read as CISH equivocal were IHC 0-1+, n=7. Comparing CISH and FISH (63) demonstrated 100% agreement between CISH not amplified and FISH ratio <1.8 (not amplified), n=51; and 71% agreement between CISH amplified and FISH ratio >2.2 (amplified), n=5/7. 5 cores read as CISH equivocal were FISH ratio <1.8 (not amplified). Comparing IHC and FISH (83) there was 100% agreement between IHC 0-1+ (negative) and FISH ratio < 1.8 (negative), n=76; and 100% agreement between IHC 3+ (positive) and FISH ratio >2.2 (amplified), n=4. Of the 3 cores read as IHC 2+, 2 were FISH ratio <1.8 (negative) and 1 was FISH ratio >2.2 (amplified).
Conclusions: The DuoCISH HER2 gene amplification assay (Dako) showed high concordance with IHC and FISH and should be considered for use in the assessment of HER2 status in breast cancer. CISH equivocal cases may need further evaluation.
Monday, March 9, 2009 9:30 AM
Poster Session I Stowell-Orbison/Autopsy Award # 31, Monday Morning