A gene-protein assay for human epidermal growth factor receptor 2 (HER2): brightfield tricolor visualization of HER2 protein, the HER2 gene, and chromosome 17 centromere (CEN17) in formalin-fixed, paraffin-embedded breast cancer tissue sections

被引:45
|
作者
Nitta, Hiroaki [1 ]
Kelly, Brian D. [2 ]
Padilla, Mary [3 ]
Wick, Nikolaus [4 ]
Brunhoeber, Patrick [5 ]
Bai, Isaac [6 ]
Singh, Shalini [5 ]
Ranger-Moore, Jim [6 ]
Bieniarz, Chris [2 ]
Tsuda, Hitoshi [7 ]
Grogan, Thomas M. [1 ]
机构
[1] Ventana Med Syst Inc, Med Innovat, Tucson, AZ USA
[2] Ventana Med Syst Inc, Technol & Appl Res, Tucson, AZ USA
[3] Roche Diagnost SL, Roche Tissue Diagnost, Barcelona, Spain
[4] Ventana Med Syst Inc, Sci Affairs, Tucson, AZ USA
[5] Ventana Med Syst Inc, Med Affairs, Tucson, AZ USA
[6] Ventana Med Syst Inc, Biostat & Data Management, Tucson, AZ USA
[7] Natl Canc Ctr, Dept Pathol, Tokyo, Japan
关键词
Gene-protein assay; Dual color in situ hybridization; Immunohistochemistry; HER2; Breast cancer; IN-SITU HYBRIDIZATION; ADJUVANT TRASTUZUMAB; IMMUNOHISTOCHEMISTRY; EXPRESSION; HER-2/NEU; BENZIDINE; SINGLE;
D O I
10.1186/1746-1596-7-60
中图分类号
R36 [病理学];
学科分类号
100104 ;
摘要
Background: The eligibility of breast cancer patients for human epidermal growth factor receptor 2 (HER2)-directed therapies is determined by the HER2 gene amplification and/or HER2 protein overexpression status of the breast tumor as determined by in situ hybridization (ISH) or immunohistochemistry (IHC), respectively. Our objective was to combine the US Food and Drug Administration (FDA)-approved HER2 & chromosome 17 centromere (CEN17) brightfield ISH (BISH) and HER2 IHC assays into a single automated HER2 gene-protein assay allowing simultaneous detection of all three targets in a single tissue section. Methods: The HER2 gene-protein assay was optimized using formalin-fixed, paraffin-embedded (FFPE) samples of the xenograft tumors MCF7 [HER2 negative (non-amplified gene, protein negative)] and Calu-3 [HER2 positive (amplified gene, protein positive)]. HER2 IHC was performed using a rabbit monoclonal anti-HER2 antibody (clone 4B5) and a conventional 3,3'-diaminobenzidine IHC detection. The HER2 & CEN17 BISH signals were visualized using horseradish peroxidase-based silver and alkaline phosphatase-based red detection systems, respectively with a cocktail of 2,4-dinitrophenyl-labeled HER2 and digoxigenin-labeled CEN17 probes. The performance of the gene-protein assay on tissue microarray slides containing 189 randomly selected FFPE clinical breast cancer tissue cores was compared to that of the separate HER2 IHC and HER2 & CEN17 BISH assays. Results: HER2 protein detection was optimal when the HER2 IHC protocol was used before (rather than after) the BISH protocol. The sequential use of HER2 IHC and HER2 & CEN17 BISH detection steps on FFPE xenograft tumor sections appropriately co-localized the HER2 protein, HER2 gene, and CEN17 signals after mitigating the silver background staining by using a naphthol phosphate-containing hybridization buffer for the hybridization step. The HER2 protein and HER2 gene status obtained using the multiplex HER2 gene-protein assay demonstrated high concordance with those obtained using the separate HER2 IHC and HER2 & CEN17 BISH assays, respectively. Conclusions: We have developed a protocol that allows simultaneous visualization of the HER2 IHC and HER2 & CEN17 BISH targets. This automated protocol facilitated the determination of HER2 protein and HER2 gene status in randomly selected breast cancer samples, particularly in cases that were equivocal or exhibited tumor heterogeneity. The HER2 gene-protein assay produced results virtually equivalent to those of the single FDA-approved HER2 IHC and HER2 & CEN17 BISH assays. Virtual slides: The virtual slides for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/2041964038705297
引用
收藏
页数:14
相关论文
共 50 条
  • [31] Quantitative RT-PCR assay of HER2 mRNA expression in formalin-fixed and paraffin-embedded breast cancer tissues
    Park, Sangjung
    Wang, Hye-Young
    Kim, Sunghyun
    Ahn, Sungwoo
    Lee, Dongsup
    Cho, Yoonjung
    Park, Kwang Hwa
    Jung, Dongju
    Kim, Seung Il
    Lee, Hyeyoung
    INTERNATIONAL JOURNAL OF CLINICAL AND EXPERIMENTAL PATHOLOGY, 2014, 7 (10): : 6752 - 6759
  • [32] Draft recommendations for human epidermal growth factor receptor 2 (HER2) testing in breast cancer will decrease HER2 positivity rates
    Johnson, Eric
    Gulbahce, Evin
    CANCER RESEARCH, 2018, 78 (04)
  • [33] Utility of simultaneous HER2 protein and gene assessment for the evaluation of discrepancy and intratumoral heterogeneity of HER2 status and the prediction of prognosis in invasive breast cancer using the gene-protein assay (GPA)
    Kurozumi, Sasagu
    Padilla, Mary
    Kurosumi, Masafumi
    Matsumoto, Hiroshi
    Hayashi, Yuji
    Tozuka, Katsunori
    Nagai, Shigenori E.
    Inoue, Kenichi
    Oba, Hanako
    Horiguchi, Jun
    Takeyoshi, Izumi
    Ranger-Moore, Jim
    Dennis, Eslie
    Nitta, Hiroaki
    CANCER RESEARCH, 2015, 75
  • [34] Simultaneous detection of HER2/neu gene amplification and protein overexpression in paraffin-embedded breast cancer.
    Szollosi, Z
    Egervari, K
    Nemes, Z
    Kaczur, V
    JOURNAL OF PATHOLOGY, 2005, 207 (01): : 119 - 120
  • [35] A Novel Proximity Assay for the Detection of Proteins and Protein Complexes: Quantitation of HER1 and HER2 Total Protein Expression and Homodimerization in Formalin-fixed, Paraffin-Embedded Cell Lines and Breast Cancer Tissue
    Shi, Yining
    Huang, Weidong
    Tan, Yuping
    Jin, Xueguang
    Dua, Rajiv
    Penuel, Elicia
    Mukherjee, Ali
    Sperinde, Jeff
    Pannu, Herjit
    Chenna, Ahmed
    DeFazio-Eli, Lisa
    Pidaparthi, Sailaja
    Badal, Youssouf
    Wallweber, Gerald
    Chen, Lili
    Williams, Steve
    Tahir, Hasan
    Larson, Jeff
    Goodman, Laurie
    Whitcomb, Jeannette
    Petropoulos, Christos
    Winslow, John
    DIAGNOSTIC MOLECULAR PATHOLOGY, 2009, 18 (01) : 11 - 21
  • [36] Comprehensive assessment of truncated HER2 in formalin-fixed paraffin embedded breast cancer tissue by chromogenic duplex immunohistochemistry
    Pickl, M.
    Rutz, C.
    Nicolai, H.
    Hasmann, M.
    Brockhoff, G.
    Feuerhake, F.
    JOURNAL OF CLINICAL ONCOLOGY, 2011, 29 (15)
  • [37] Monosomy of chromosome 17 in breast cancer during interpretation of HER2 gene amplification
    Brunelli, Matteo
    Nottegar, Alessia
    Bogina, Giuseppe
    Calio, Anna
    Cima, Luca
    Eccher, Albino
    Vicentini, Caterina
    Marcolini, Lisa
    Scarpa, Aldo
    Pedron, Serena
    Brunello, Eleonora
    Knuutila, Sakari
    Sapino, Anna
    Marchio, Caterina
    Bria, Emilio
    Molino, Annamaria
    Carbognin, Luisa
    Tortora, Giampaolo
    Jasani, Bharat
    Miller, Keith
    Merdol, Ibrahim
    Zanatta, Lucia
    Laurino, Licia
    Wirtanen, Tiina
    Zamboni, Giuseppe
    Marconi, Marcella
    Chilosi, Marco
    Manfrin, Erminia
    Martignoni, Guido
    Bonetti, Franco
    AMERICAN JOURNAL OF CANCER RESEARCH, 2015, 5 (07): : 2212 - 2221
  • [38] Genetic alterations and protein expression of HER2 and chromosome 17 polysomy in breast cancer
    Zhu, Xiaoli
    Lu, Yongming
    Lu, Hongfen
    Yang, Wentao
    Tu, Xiaoyu
    Cai, Xu
    Zhou, Xiaoyan
    HUMAN PATHOLOGY, 2011, 42 (10) : 1499 - 1504
  • [39] HER2 Expression and Gene copy analysis by Immunofluorescence and Fluorescence in situ Hybridization, on a Single formalin-fixed paraffin-embedded tissue section
    Ha, T.
    Seppo, A.
    Ginty, F.
    Kenny, K.
    Henderson, D.
    Kyshtoobayeva, A.
    Gerdes, M.
    Larriera, A.
    Liu, X.
    Corwin, A.
    Zingelewicz, S.
    Lazare, M.
    Jun, N.
    Kyshtoobayeva, A.
    Chow, C.
    Al-Kofahi, Y.
    Hollman, D.
    Bloom, K.
    CANCER RESEARCH, 2012, 72
  • [40] Assessment of HER2 Gene Status in Breast Carcinomas With Polysomy of Chromosome 17
    Vranic, Semir
    Teruya, Bryan
    Repertinger, Susan
    Ulmer, Pamela
    Hagenkord, Jill
    Gatalica, Zoran
    CANCER, 2011, 117 (01) : 48 - 53