Automated ERCC1 immunochemistry on hybrid cytology/tissue microarray of malignant effusions: evaluation of antibodies 8F1 and D-10

Our plain-English summary of Automated ERCC1 immunochemistry on hybrid cytology/tissue microarray of malignant effusions: evaluation of antibodies 8F1 and D-10, by A. Soltermann, S. Kilgus-Hawelski, S. Behnke, et al (Journal of Clinical Bioinformatics, 2011).

The study

• Authors: A. Soltermann, S. Kilgus-Hawelski, S. Behnke, M. Storz, H. Moch, B. Bode
• Journal: Journal of Clinical Bioinformatics
• Published: 2011;1:25
• Cited by: 6 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Background: The excision repair cross-complementation group 1 (ERCC1) protein is the key enzyme of the nucleotide excision repair (NER) pathway. Loss of protein expression on immunohistochemistry is predictive for platinum-based chemotherapy response. Frequently, the diagnosis of malignancy is made on cytologic effusion samples. Therefore, we evaluated the staining quality of monoclonal anti-ERCC1 antibodies 8F1 and D-10 on microarrays of malignant pleural and peritoneal effusions by automated immunochemistry.

Methods: Cores from effusion cell blocks of 117 patients with > 40 malignant cell clusters per whole section (pleural n = 75, peritoneal n = 42) were assembled together with 30 histologic control cores from large tissue blocks (lung, breast and ovarian carcinoma, each n = 10) on hybrid cytology-tissue microarrays (C/TMA). Four immunochemistry protocols (Mab 8F1 and D-10, CC1-mono Ventana and H2-60 Bond automat) were performed. Immunoreactivity was semi-quantitatively scored for intensity and intensity multiplied by percentage staining (H-score).

Results: Tumors were classified into female genital tract carcinoma (n = 39), lung adenocarcinoma (n = 23), mesothelioma (n = 15), unknown primary (n = 14), breast carcinoma (n = 10), gastro-intestinal carcinoma (n = 12) and other (n = 4). On both platforms, reproducible nuclear ERCC1 immunoreactivity was achieved with both antibodies, although D-10 was slightly weaker and presented more background staining as well as more variation in the low expression range. No significant differences were found between cytologic and histologic cores. Using the 8F1 CC1-mono protocol, lung and breast carcinomas had lower ERCC1 expression in comparison to the other entities (p-value < 0.05).

Conclusions: Cytology microarrays (CMA) are suitable for investigation of clinical biomarkers and can be combined with conventional TMA's. Dichotomization of ERCC1 immunoreactivity scores is most suitable for patient stratification since definition of negativity is antibody-dependent.

In plain English

This laboratory study, from the Institute of Surgical Pathology at University Hospital Zurich, did not study asbestos exposure. It tested how well two antibodies, 8F1 and D-10, stain the DNA-repair protein ERCC1, each run on two automated staining machines. The samples were cell blocks made from fluid that had collected around the lungs (pleural, 75 patients) or in the abdomen (peritoneal, 42 patients), taken between 2005 and 2010. The 117 tumors included 15 mesotheliomas. They were compared with 30 tissue samples of lung, breast and ovarian carcinoma. Both antibodies stained cell nuclei. D-10 was generally weaker and produced unwanted background staining. Fluid and tissue samples did not differ significantly. Among the 117 fluid-sample tumors, counting scores of 0 to 1 as low and 2 to 3 as high, 43.6% to 82.9% would be classed as ERCC1 high, depending on antibody and method. Inflammatory cells mixed in with the tumor cells also stained, which complicated measurement. The authors concluded that these cell-block arrays are suitable for studying clinical biomarkers and that 8F1 performed better than D-10.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory comparison of antibody staining methods on archived cytology cell blocks and tissue samples
• Who was studied: Cell blocks from malignant pleural (n = 75) or peritoneal (n = 42) effusions of 117 patients (77 female, 40 male; mean age 66). The samples were collected 2005-2010 at University Hospital Zurich, out of 125 enrolled (8 lost in processing). Tumor types included 15 mesotheliomas. There were also 30 histologic control cores (lung, breast and ovarian carcinoma, 10 each).
• Exposure: No asbestos or other environmental exposure was studied. The study tested laboratory methods: two anti-ERCC1 antibodies (8F1 and D-10), each run with two automated staining protocols (CC1-mono on the Ventana Benchmark platform and H2-60 on the Leica Bond platform).
• Main results:
  • Both antibodies produced distinct staining of cell nuclei. D-10 also showed focal but strong background staining of the cell membrane and of mucin vacuoles.
  • D-10 was generally weaker than 8F1 under the same protocols.
  • The 8F1 antibody showed much less variation in the low expression range than D-10.
  • Comparing cytologic (fluid) cores with histologic (tissue) control cores, the average of all mean scores was slightly lower in the controls. The difference was not statistically significant.
  • Among the 117 effusion cell-block tumors, scores of 0 to 1 were counted as low and 2 to 3 as high. Under that split, the share that would be classed as ERCC1 high ranged from 43.6% (D-10, CC1-mono, H-score) to 82.9% (8F1, H2-60, intensity score).
  • With 8F1 and the CC1-mono protocol, lung adenocarcinoma and breast carcinoma had significantly lower ERCC1 scores. This relation was not found with any of the other scoring combinations.
  • Female genital tract carcinoma and mesothelioma had average score means above 2.
  • Inflammatory and reactive cells mixed with the tumor cells also expressed ERCC1. The authors concluded that this makes quantitative measurement difficult.
• The authors' conclusion: The authors concluded that cell-block cytology microarrays are suitable for investigating clinical biomarkers and can be combined with tissue microarrays. They also concluded that on both automated platforms 8F1 performed better than D-10 in staining quality and in staying confined to the nucleus.
• Limitations the authors state:
  • The authors noted that formalin fixation time can differ significantly between cytologic and histologic samples. They judged its influence to be of minor importance.
  • The authors stated that automated quantitative intensity measurement would be difficult for ERCC1, because background inflammatory cells also stain.

Topics

Medical terms in this study, explained in our Lexicon: Cytology, Immunohistochemistry, Chemotherapy, Monoclonal Antibody, Cell Block, Paraffin Block, Carcinoma, Lung Adenocarcinoma.

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