A new rabbit monoclonal antibody (4B5) for the immunohistochemical (IHC) determination of the HER2 status in breast cancer:: Comparison with CB11, fluorescence in situ hybridization (FISH), and interlaboratory reproducibility

The 2 methodologies in current clinical use to assess HER2 status in breast cancer are: fluorescence in situ hybridization (FISH) (gene amplification) and immunohistochemistry (protein over-expression). A consistent finding has been that 3% to 15% of breast cancers over-express HER2 protein without evidence for gene amplification. Accurate determination of the HER2 status has implications for selecting patients most likely to respond to trastuzumab. We report here our preliminary experience with a new anti-HER2 rabbit monoclonal antibody, 4135. The evaluation of HER2 status in 2 different cohorts of breast cancer cases (Single Institution (SI) and Multinational (MN)) with a total of 322 breast cancer cases was performed on an automated staining system (Ventana Medical Systems, Inc, Tucson, AZ) and scored by 3 pathologists (0-3+), for comparison with CBII staining results (PATHWAY) and FISH (PathVysion). Interlaboratory reproducibility of automated staining results and interpretation was determined on a subset of the SI cohort at 3 separate laboratories. Rabbit monoclonal 4135 demonstrated sharper membrane staining with less cytoplasmic and stromal background staining than CB11. In the SI cohort, the staining results for 4135 were highly comparable with those obtained for CBII with an overall concordance of 93.3%. In the multinational cohort, the overall concordance with CBI I was 84.7%. This lower level of concordance was associated with a much higher overall agreement of 4135 with FISH (89.5%), compared with agreement of CBII with FISH (81.2%). The difference in the performance of CBII in the MN cohort versus the SI cohort may be due to differences in tissue fixation and processing in a centralized, high volume laboratory in an academic medical center versus multiple sites in the international community with potentially nonstandardized techniques. The staining results with 4B5 indicate that it has a more robust performance than CB11 because the correlation of 4135 with FISH was nearly equivalent (88.2% MN; 89.3% SI) in both cohorts. Interlaboratory reproducibility was also excellent (K 1.0). RMoAb 4135 provides excellent sensitivity, specificity, and interlaboratory reproducibility for the detection of HER2 status in breast cancer.