Development and Validation of an ERCC1 Immunohistochemistry Assay for Solid Tumors.
BACKGROUND
- Excision repair cross-complementation 1 (ERCC1) is a key enzyme in nuclear excision repair pathway and has a critical role in helping remove DNA adducts caused by cross-linking agents, such as platinum-containing cancer chemotherapies and other DNA-damaging therapeutic modalities. ERCC1 expression, evaluated by techniques such as immunohistochemistry, has been associated with clinical response; ERCC1+tumors are more resistant to cisplatin treatment than are ERCC1-tumors. Although several immunohistochemistry, anti-ERCC1 antibodies are available, the 8F1 clone, in particular, has been used in many studies. Recent evidence has suggested that the 8F1 antibody cross-reacts with at least one other protein, raising concerns about the specificity of this clone.


OBJECTIVE
- To design an immunohistochemistry assay to detect ERCC1 levels that show dynamic range and consistent analytic performance.


METHODS
- Two different primary antibodies to ERCC1, clones 4F9 and D6G6, were evaluated on formalin-fixed, paraffin-embedded tissue. We then performed a fit-for-purpose assay validation with the 4F9 clone, which included sensitivity assessment across several solid tumor types and evaluation of analytic parameters, such as precision and reproducibility.


RESULTS
- The 4F9 clone was consistently superior to the D6G6 clone in the optimization phase. A range of expression was seen in ovarian, head and neck, non-small cell lung, and esophageal cancer samples when tested with the 4F9 clone. The antibody showed acceptable reproducibility (31.02%) and precision (16.06%).


CONCLUSIONS
- This assay can be used to assess ERCC1 levels during clinical studies of patient tumors from a variety of tumor types.
From the Departments of Translational Medicine (Ms Bahamon and Dr Danaee) and Biostatistics (Dr Gao), Takeda Pharmaceuticals Inc, Cambridge, Massachusetts. Dr Gao is now with the Department of Biostatistics, Biogen, Cambridge.
Brittany N
BN
Bahamon
Feng
F
Gao
Hadi
H
Danaee
D000230:Adenocarcinoma / Q000175:diagnosis / Q000378:metabolism / Q000473:pathology; D000911:Antibodies, Monoclonal / Q000378:metabolism*; D000918:Antibody Specificity; D014408:Biomarkers, Tumor / Q000235:genetics / Q000378:metabolism; D002277:Carcinoma / Q000175:diagnosis / Q000378:metabolism / Q000473:pathology; D002289:Carcinoma, Non-Small-Cell Lung / Q000175:diagnosis / Q000378:metabolism / Q000473:pathology; D045744:Cell Line, Tumor; D002999:Clone Cells; D015331:Cohort Studies; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D004720:Endonucleases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006258:Head and Neck Neoplasms / Q000175:diagnosis / Q000378:metabolism / Q000473:pathology; D006801:Humans; D007150:Immunohistochemistry; D008175:Lung Neoplasms / Q000175:diagnosis / Q000378:metabolism / Q000473:pathology; D008297:Male; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D009369:Neoplasms / Q000175:diagnosis / Q000378:metabolism* / Q000473:pathology; D010051:Ovarian Neoplasms / Q000175:diagnosis / Q000378:metabolism / Q000473:pathology; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D000911:Antibodies, Monoclonal; D014408:Biomarkers, Tumor; D004268:DNA-Binding Proteins; D009363:Neoplasm Proteins; C071447:ERCC1 protein, human; D004720:Endonucleases