The numerical aberrations of chromosome 7 detected by fluorescence in situ hybridization in human breast cancers

PURPOSE: The objective of our study was to compare the methods used in the literature to analyze HER-2/neu status on archival breast cancer tissue. Therefore, a series of antibodies was evaluated to assess their immunohistochemical (IHC) sensitivity in correlation to gene amplification determined by fluorescence in situ hybridization (FISH). MATERIALS AND METHODS: HER-2/neu overexpression was studied on paraffin sections of 85 invasive breast cancers using a panel of five monoclonal (9G6, 3B5, CB11, TAB250, GSF-HER2) and two polyclonal antibodies (A8010, A0485) in addition to the HercepTest (DAKO, Glostrup, Denmark). HER-2/neu gene amplification was determined by FISH using a dual-color probe (PathVysion; Vysis, Stuttgart-Fasanenhof, Germany). RESULTS: HER-2/neu overexpression was demonstrated in 26% (9G6, TAB250, GSF-HER2), 27% (3B5, CB11), 33% (A8010) and 42% (A0485, HercepTest) of the tumors. FISH on paraffin sections identified gene amplification in 28% of the tumors. Strongly positive IHC results (3+) were always associated with gene amplification. Among the 16 tumors presented with weakly positive IHC results (2+) using the HercepTest, 12 (75%) lacked gene amplification. CONCLUSION: The comparison of IHC and FISH demonstrated an excellent correlation of high-level HER-2/neu overexpression (3+) with gene amplification; ie, FISH does not provide further information in these tumors. However, weakly positive IHC results (2+) obtained with the HercepTest share only a minor association with gene amplification.

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Assessment of Methods for Tissue-Based Detection of the HER-2/neu Alteration in Human Breast Cancer: A Direct Comparison of Fluorescence In Situ Hybridization and Immunohistochemistry

Journal of Clinical Oncology ◽

10.1200/jco.2000.18.21.3651 ◽

2000 ◽

Vol 18 (21) ◽

pp. 3651-3664 ◽

Cited By ~ 450

Author(s):

Giovanni Pauletti ◽

Suganda Dandekar ◽

HongMei Rong ◽

Lilllian Ramos ◽

HongJun Peng ◽

...

Keyword(s):

Breast Cancer ◽

In Situ Hybridization ◽

Fluorescence In Situ Hybridization ◽

Gene Amplification ◽

Survival Probability ◽

Human Breast Cancer ◽

Patient Survival ◽

Human Breast ◽

Her 2

PURPOSE: To compare the efficacy of fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC) in detecting the HER-2/neu alteration in human breast cancer. PATIENTS AND METHODS: Unselected stage I, II, and III breast cancer patients (N = 900) were tested for HER-2/neu gene amplification by FISH in paraffin-embedded, formalin-fixed archival material. Of these samples, 856 were tested for HER-2/neu overexpression by non–antigen-retrieval IHC with the polyclonal antibody R60, the sensitivity and specificity of which was preliminarily compared with the United States Food and Drug Administration–approved HercepTest (Dako Corp, Carpinteria, CA). Patient survival was analyzed in relation to the presence of the HER-2/neu alteration as determined by these two methodologies. RESULTS: A total of 189 (21%) of 900 patients were positive by FISH and 147 (17.2%) of 856 were positive by IHC. This discrepancy is consistent with expected loss of IHC sensitivity associated with tissue fixation/embedding. The HercepTest did not improve sensitivity and introduced false positives. Comparison of R60-based IHC with FISH demonstrates that patient survival is associated progressively to gene amplification level as determined by FISH, whereas for IHC an association is found only in the highest (3+) immunostaining group. Among FISH-negative tumors, 45 (6.6%) of 678 were IHC-positive, with a survival probability similar to that of FISH-negative/IHC-negative cases; FISH-positive/IHC-negative patients have a survival probability similar to that of FISH-positive/IHC-positive cases. CONCLUSION: IHC does not consistently discriminate patients likely to have a poor prognosis, whereas FISH provides superior prognostic information in segregating high-risk from lower-risk beast cancers. HER-2/neu protein overexpression in the absence of gene amplification occurs infrequently in breast cancer, in which case, patient outcome is similar to that of patients without the alteration.

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HER-2/neu gene amplification characterized by fluorescence in situ hybridization: poor prognosis in node-negative breast carcinomas.

Journal of Clinical Oncology ◽

10.1200/jco.1997.15.8.2894 ◽

1997 ◽

Vol 15 (8) ◽

pp. 2894-2904 ◽

Cited By ~ 539

Author(s):

M F Press ◽

L Bernstein ◽

P A Thomas ◽

L F Meisner ◽

J Y Zhou ◽

...

Keyword(s):

In Situ Hybridization ◽

Fluorescence In Situ Hybridization ◽

Gene Amplification ◽

Poor Prognosis ◽

Southern Hybridization ◽

Recurrent Disease ◽

Breast Cancers ◽

Node Negative ◽

Her 2

PURPOSE The HER-2/neu gene codes for a membrane receptor protein that is homologous, but distinct from the epidermal growth factor receptor. This investigation was performed to validate fluorescence in situ hybridization (FISH) as a sensitive and specific method for assessing HER-2/neu gene amplification in archival tissue and to test whether this alteration is associated with poor prognosis. MATERIALS AND METHODS HER-2/neu gene amplification was determined by FISH in 140 archival breast cancers, previously characterized for gene amplification by Southern hybridization or dot-blot hybridization, and for gene expression by Northern hybridization, Western immunoblot, or immunohistochemistry. A separate cohort of 324 node-negative breast cancers was assessed for amplification by FISH to determine the utility of HER-2/neu gene amplification. RESULTS Relative to solid-matrix blotting procedures, FISH analysis of HER-2/neu gene amplification showed a sensitivity of 98% and a specificity of 100% in 140 breast cancers. Among patients treated by surgery only, the relative risks (relative hazard) of early recurrence (recurrent disease within 24 months of diagnosis), recurrent disease (at any time), and disease-related death were statistically significantly associated with amplification. The prognostic information contributed by HER-2/neu amplification was independent of the other markers studied. CONCLUSION FISH was an alternative technique for determining gene amplification and had some distinct advantages over Southern hybridization. Our results demonstrate that HER-2/neu gene amplification in the absence of adjuvant therapy is an independent predictor of poor clinical outcome and is a stronger discriminant than tumor size. Women with small tumors that had gene amplification were at increased risk of recurrence and disease-related death.

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