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

Soltermann, Alex; Kilgus-Hawelski, Sandra; Behnke, Silvia; Storz, Martina; Moch, Holger; Bode, Beata

doi:10.1186/2043-9113-1-25

Cited by 8 publications

(5 citation statements)

References 35 publications

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“…Currently, there is no clinically approved ERCC1 IHC diagnostic assay available, likely due to the lack of a high quality anti-ERCC1 monoclonal antibody. Studies from a number of labs have shown that the 8F1 clone gives the best performance on NSCLC tissue in IHC assays [18,34]. However, Bhagwat, et al claimed that 8F1 is not mono-specific [19].…”

Section: Discussionmentioning

confidence: 99%

Using protein microarray technology to screen anti-ERCC1 monoclonal antibodies for specificity and applications in pathology

Baruch

Shu

et al. 2012

BMC Biotechnol

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BackgroundAn antibody with cross-reactivity can create unexpected side effects or false diagnostic reports if used for clinical purposes. ERCC1 is being explored as a predictive diagnostic biomarker for cisplatin-based chemotherapy. High ERCC1 expression is linked to drug resistance on cisplatin-based chemotherapy. 8F1 is one of the most commonly used monoclonal antibodies for evaluating ERCC1 expression levels in lung cancer patient tissues, but it has been noted that this antibody cross-reacts with an unknown protein.ResultsBy using a high density protein microarray chip technology, we discovered that 8F1 not only reacts with its authentic target, ERCC1, but also cross-reacts with an unrelated nuclear membrane protein, PCYT1A. The cross-reactivity is due to a common epitope presented on these two unrelated proteins. Similar to the subcellular localization of ERCC1, IHC tests demonstrated that PCYT1A is localized mainly on nuclear membrane. In this study, we also discovered that the PCYT1A gene expression level is significantly higher than the ERCC1 gene expression level in a certain population of lung cancer patient tissue samples. To develop the best monoclonal antibody for ERCC1 IHC analysis, 18 monoclonal antibodies were generated and 6 of them were screened against our protein microarray chip. Two clones showed high mono-specificity on the protein microarray chip test and both worked for the IHC application.ConclusionIn summary, the 8F1 clone is not suitable for ERCC1 IHC assay due to its cross-reactivity with PCYT1A protein. Two newly generated monoclonal antibodies, 4F9 and 2E12, demonstrated ultra-specificity against ERCC1 protein and superior performance for IHC analyses.

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Section: Discussionmentioning

confidence: 99%

Using protein microarray technology to screen anti-ERCC1 monoclonal antibodies for specificity and applications in pathology

Baruch

Shu

et al. 2012

BMC Biotechnol

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“…The effusion liquids were prepared and processed as previously described [32]. Briefly, after low-speed centrifugation, the upper white phase of the sediment was aspirated and transferred into a micro tube.…”

Section: Preparation Of Cellblocks and Cytologic Microarray (C-tma)mentioning

confidence: 99%

Prognostic Immune Cell Profiling of Malignant Pleural Effusion Patients by Computerized Immunohistochemical and Transcriptional Analysis

Mairinger

Casanova

et al. 2019

Cancers

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Malignant pleural effusion (MPE) is a severe condition of advanced tumors without effective therapy. We used digitalized immunohistochemical and transcriptional approaches to investigate the prognostic influence of immune cells and expression variance of associated immunomodulatory molecules in MPE. Cytology tissue microarrays were constructed from MPE cell blocks of 155 patients with five tumor entities. Immune cells lineage markers were quantified by computational cytopathology on immunohistochemistry. mRNA expression analysis of nine lineage markers and 17 immunomodulators was performed by NanoString. Immunohistochemically quantified high B cells to leukocytes ratio (hazard ratio (HR) = 0.70, p-value = 0.043) and low neutrophils to leukocytes ratio (HR = 1.78, p-value = 0.003) were favorable prognosticators for overall survival independent of tumor entity. Correspondingly, patients with high B cells but low neutrophils gene expression signature showed longer median overall survival of 500 days (HR = 2.29, p-value = 0.009). Regarding targetable molecule expressions, lung adenocarcinomas were characterized by high PD-L1, but mesothelioma by high LAG-3. Ovarian carcinoma was least immunogenic. Independent of tumor entity, the condition of the immune system in MPE liquids is able to provide additional prognostic cytologic information. Combined analysis of lineage specific markers and related immunomodulators may direct immune-based therapeutic decisions.

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“…As described previously ( 29 ), the effusion liquids were centrifuged and the sediments were transferred into a microtube. Subsequently, thrombin and plasma were added for clot formation.…”

Section: Methodsmentioning

confidence: 99%

Lung adenocarcinoma patients with malignant pleural effusions in hot adaptive immunity status have a longer overall survival

Casanova

Mairinger

et al. 2022

Front. Oncol.

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Malignant pleural effusion (MPE) is a common complication of lung adenocarcinoma (LADC) which is associated with a dismal prognosis. We investigated the prognostic role of PD-L1 and other immunomodulators expression in the immune compartment of MPE immune composition. MPE cytologic cell blocks of 83 LADC patients were analysed for the mRNA expression of 770 cancer-immune genes by the NanoString nCounter platform. The expression of relevant immune cell lineage markers was validated by immunohistochemistry (IHC) using quantitative pathology. The mRNA immune profiling identified four MPE patient clusters (C). C1/2 (adaptive+, hot) showed better overall survival (OS) than C3/4 (adaptive-, cold). Additionally, cold immunity profiles (adaptive-), C4 (innate+) were associated with worse OS than C3 (innate-). High PD-L1 expression was linked to the regulation of T cell activation and interferon signalling pathways. Genes of pattern recognition receptor and type I interferon signalling pathways were specifically upregulated in the long-survival (≥90 days) patient group. Moreover, immunomodulators were co-activated and highly expressed in hot adaptive immunity patient clusters, whereas CD274 (PD-L1), TNFRSF9 (4-1BB), VEGFA (VEGF-A) and CD276 (B7-H3) were upregulated in the groups referred as cold. The patient cluster, age and PD-L1 expression were independent prognosticators for LADC MPE patients (p-value < 0.05). Our study sheds light on the variances of immune contexture regarding different PD-L1 expression and survival conditions. It revealed four distinct prognostic patient clusters with specific immune cell components and immunomodulator expression profiles, which, collectively, is supportive for future therapeutic and prognosis for cancer management.

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

Cited by 8 publications

References 35 publications

Using protein microarray technology to screen anti-ERCC1 monoclonal antibodies for specificity and applications in pathology

Using protein microarray technology to screen anti-ERCC1 monoclonal antibodies for specificity and applications in pathology

Prognostic Immune Cell Profiling of Malignant Pleural Effusion Patients by Computerized Immunohistochemical and Transcriptional Analysis

Lung adenocarcinoma patients with malignant pleural effusions in hot adaptive immunity status have a longer overall survival

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