Abstract
BackgroundThe 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.MethodsCores 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).ResultsTumors 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).ConclusionsCytology 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.
Highlights
The excision repair cross-complementation group 1 (ERCC1) protein is the key enzyme of the nucleotide excision repair (NER) pathway
We have previously investigated the 3 anti-ERCC1 antibodies Mab 8F1, Mab D-10 and Rab FL-297 on a retrospective non-small cell lung cancer (NSCLC) patient cohort assembled on a tissue microarray (TMA) [18]
ERCC1 protein expression on whole sections In order to check for surface homogeneity of immunoreactivity, we first stained 4 μm thick whole sections of squamous cell lung carcinoma (Figure 1)
Summary
The excision repair cross-complementation group 1 (ERCC1) protein is the key enzyme of the nucleotide excision repair (NER) pathway. Cisplatin forms DNA adducts, thereby causing interand intra-strand cross links, comparable to alkylating agents. If not repaired, this DNA damage will lead to apoptotic cell death or mutation. In the NER system, the heterodimer ERCC1-XPF functions as a structure-specific endonuclease to make the 5’-incision on the damaged strand. This step is claimed to be the key factor [1,2,3]. It was deduced that tumors with low nuclear ERCC1 expression better respond to platinum-containing CT because of reduced repair capability for DNA adducts [4,5]. A valid predictor of this widely used regimen is of high clinical importance, because response rates in e.g. unselected non-small cell lung cancer (NSCLC) patients range from only 16 to 30% [6,7]
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