Immunohistochemical Antibodies in Breast Cancer HER2 Diagnostics

Abstract

Overexpression and/or gene amplification of the HER2 oncogene predicts worse prognosis and altered sensitivity to chemotherapy. Trastuzumab is capable of improving prognosis of HER2-positive breast cancer, but for the success of treatment appropriate HER2 testing is essential. Our aim was to determine the value of immunohistochemical (IHC) screening prior to fluorescence in situ hybridization (FISH). We assessed five conventional IHC assays (NCL-CB11, Pathway CB11, CBE356, CBE1, HercepTest) and the novel rabbit monoclonal antibody, RM-4B5, combined with FISH on 199 invasive breast cancer cases. Taking FISH as the endpoint, we calculated sensitivity, specificity, positive and negative predictive values (PPV, NPV) and accuracy for all IHC assays with either taking both 2+/3+ cases or only 3+ cases as IHC positives. With 2+/3+ cases HercepTest showed 100% sensitivity and NPV, while the highest specificity, PPV and accuracy was associated with RM-4B5 (97.36, 80 and 95.34%, respectively). The second highest values belonged to either NCL-CB11 or Pathway CB11. When calculating only with 3+ cases, the results were reversed with increased specificity, PPV and accuracy. Our findings suggest that improving sensitivity by using two parallel IHC reactions might be beneficial; we recommend primarily HercepTest and Pathway CB11. Nevertheless, we may consider performing FISH analysis without prior IHC screening.

Immunohistochemical analysis of the monoclonal antibody 4B5 in breast tissue expressing human epidermal growth factor receptor 4 (HER4)

“…These suboptimal conditions probably resulted in a decreased sensitivity of 4B5, and may have contributed to the lower number of level 2+ cases. However, 4B5, under the conditions tested, showed substantial overall accuracy, with a higher specificity and positive predictive value than FISH . Staining results with 4B5 have been highly comparable with those of P‐CB11: the difference between P‐CB11 and 4B5 involves 4B5 scoring fewer level 2+ cases, thus yielding a higher concordance with FISH .…”

“…These results further support 4B5 as a clinically relevant assay for HER2 expression in breast carcinoma. Studies comparing 4B5 and FISH results in breast cohorts have demonstrated that 4B5 has a higher concordance with FISH than other pharmacodiagnostic HER2 antibodies . In one comparison of 4B5 with HercepTest, the results for 4B5, used according to the manufacturer's protocol, differed little from those reported for HercepTest (correlation P < 0.005) .…”

“…Numerous studies have compared the concordance between FISH and these HER2 IHC assays. Variation occurs in the distribution of breast cancer cases in the non‐amplified, equivocal and amplified categories among the various HER2 IHC assays compared, but, in all studies to date, 4B5 has demonstrated the highest concordance with FISH . The higher proportion of positive cases with 4B5, particularly in one study where 4B5 identified more level 2+ cases than HercepTest, has been hypothesized by Schrohl et al .…”

“…To date, three tests have been approved by the US Food and Drug Administration for IHC analysis of HER2: HercepTest (Dako, Denmark A/S, Glostrup, Denmark), the mouse monoclonal PATHWAY HER2 CB11 (P‐CB11) (Ventana Medical Systems, Tucson, AZ, USA), and the rabbit monoclonal replacement for P‐CB11, PATHWAY HER2 4B5 (4B5). Additional anti‐HER2 antibodies are also used, including other commercially available forms of the CB11 clone (Table ) . 4B5 and all commercialized types of the CB11 clone are monoclonal antibodies of rabbit or mouse origin, respectively, that bind to the intracellular domain of HER2.…”

“…The suggestion that affinity for HER4 impacts on the detection of HER2 by 4B5 in IHC evaluation of breast cancer is not supported, given the numerous studies demonstrating excellent concordance wth FISH . However, in order to assess whether 4B5 can detect membrane‐bound HER4 in IHC of FFPE human breast carcinoma, and any impact that this staining might have on the 4B5 score for HER2 expression, an HER4 IHC assay for membrane staining was developed.…”