Analysis of HER-2/neu amplification in endometrial carcinoma by chromogenic in situ hybridization. Correlation with fluorescence in situ hybridization, HER-2/neu, p53 and Ki-67 protein expression, and outcome

Abstract

Fluorescence in situ hybridization (FISH) is the most widely used technique to detect HER-2/neu gene amplification; however, it is only available in some institutions. In contrast, chromogenic in situ hybridization (CISH) can be evaluated by routine light microscopy. In endometrial carcinoma there are few data concerning HER-2/neu status and prognosis. Therefore, we determined HER-2/neu gene status by CISH using a digoxigenin-labelled probe on 60 formalin-fixed paraffin-embedded endometrial carcinomas. The data were compared with the immunohistochemistry of HER-2/neu (A0485, TAB250), p53, Ki-67, clinicopathological factors, and survival. By conventional light microscopy, HER-2/neu amplification (X6 copies 450% cancer cells) was detected in 14% (8/59) tumours, HER-2/neu overexpression (410% cells moderate/strong complete membrane staining) in 22% (13/60) for A0485, and 18% (11/60) for TAB250, p53 (410% þ cells) in 61% (36/59), and Ki-67 (450% þ cells) in 50% (30/60). Discordant cases for CISH and immunohistochemistry, as well as all (2 þ ) were further analysed by FISH (Vysis). Among 10 cases (2 þ ) and not amplified by CISH, two showed lowlevel amplification by FISH. Significant correlation was found between amplification and protein overexpression (Pp0.001), and a trend with nonendometrioid type, higher grade, and older age. A better outcome (KaplanMeier) was observed for patients with nonamplified (1-5 copies per nucleus) or low-level (6-10 copies) amplification tumours, low Ki-67 expression, age o50 years, endometrioid type, low FIGO (International Federation of Obstetrics and Gynaecology) grade and stage, superficial myometrial infiltration, and no lymphvascular invasion (Pp0.036), but only as a trend for HER-2/neu protein negative (P ¼ 0.13). Cox analysis revealed age, FIGO grade and stage, myometrial infiltration, and lymph-vascular invasion to be independent prognostic factors (Pp0.05), and a trend for HER-2/neu gene copy number (0.18). In endometrial carcinoma, HER-2/neu gene status can be readily assessed by CISH in routine clinical practice, and it gives more prognostic information than HER-2/neu by immunohistochemistry. FISH analysis in (2 þ ) cases but negative by CISH may detect additional tumours with low-level amplification.

Chromogenic in-situ hybridization: a viable alternative to fluorescence in-situ hybridization in the HER2 testing algorithm

“…These findings are consistent with the results of previous studies, which have shown concordance rates between CISH and FISH and/or immunohistochemistry to be in the range of 90-100%. [15][16][17][18][19][20][21] A recent study by Gong et al 21 evaluated the HER2 status of 80 archival invasive breast carcinoma specimens by CISH and FISH, and also assessed the interobserver reproducibility of CISH results between three pathologists. A 91% agreement in CISH results between pathologists was reported and all these cases had HER2 status matching the corresponding FISH results.…”

Toward standard HER2 testing of endometrial serous carcinoma: 4-year experience at a large academic center and recommendations for clinical practice

“…30,31 Many other studies also included various histological subtypes and grades of endometrial carcinoma, and even malignant mixed mullerian tumors (carcinosarcomas) in the HER2 testing of endometrial serous carcinoma same analysis, making the interpretation of data difficult. [14][15][16]34,35 Several previous studies used non-FDA-approved HER2 antibodies, ie, c-erbB-2 clone A0485 (DAKO), 24,35,36 c-erbB-2/ Her-2/neu Ab17 (monoclonal antibodies e2-4001 and 3B5; Neomarkers) 8 and clone TAB250 (Zymed). 16,35 Although most studies used the FDA scoring criteria for HER2 immunohistochemistry, 8,9,[13][14][15]18,22,24,30,35,36 many of them considered both the 2 þ and 3 þ immunohistochemical scores positive for HER2 overexpression.…”

“…[14][15][16]34,35 Several previous studies used non-FDA-approved HER2 antibodies, ie, c-erbB-2 clone A0485 (DAKO), 24,35,36 c-erbB-2/ Her-2/neu Ab17 (monoclonal antibodies e2-4001 and 3B5; Neomarkers) 8 and clone TAB250 (Zymed). 16,35 Although most studies used the FDA scoring criteria for HER2 immunohistochemistry, 8,9,[13][14][15]18,22,24,30,35,36 many of them considered both the 2 þ and 3 þ immunohistochemical scores positive for HER2 overexpression. 8,13,15,30,35,36 Others used selfdeveloped semiquantitative immunohistochemical scoring systems, not currently approved for any other tumor types.…”

Amplifications of EGFR gene and protein expression of EGFR, Her-2/neu, c-kit, and androgen receptor in phyllodes tumor of the prostate

“…25 Overexpression of Her-2/neu is present in a variety of carcinomas, including those of the breast, bladder, ovary, endometrium, and lung, and is generally associated with a poor prognosis. [26][27][28][29] Recently, Montironi et al 30 found that Her-2/neu is overexpressed in androgen independent prostate cancer and is not associated with gene amplification. In our study, Her-2/neu staining was not present in any of the stromal components, and this protein was only weakly expressed in the epithelial components in some cases.…”