ABSTRACT
A challenge in the treatment of lung cancer is the lack of early diagnostics. Here, we describe the application of monoclonal antibody proteomics for discovery of a panel of biomarkers for early detection (stage I) of non-small cell lung cancer (NSCLC). We produced large monoclonal antibody libraries directed against the natural form of protein antigens present in the plasma of NSCLC patients. Plasma biomarkers associated with the presence of lung cancer were detected via high throughput ELISA. Differential profiling of plasma proteomes of four clinical cohorts, totaling 301 patients with lung cancer and 235 healthy controls, identified 13 lung cancer-associated (p < 0.05) monoclonal antibodies. The monoclonal antibodies recognize five different cognate proteins identified using immunoprecipitation followed by mass spectrometry. Four of the five antigens were present in non-small cell lung cancer cells in situ. The approach is capable of generating independent antibodies against different epitopes of the same proteins, allowing fast translation to multiplexed sandwich assays. Based on these results, we have verified in two independent clinical collections a panel of five biomarkers for classifying patient disease status with a diagnostics performance of 77% sensitivity and 87% specificity. Combining CYFRA, an established cancer marker, with the panel resulted in a performance of 83% sensitivity at 95% specificity for stage I NSCLC.
Subject(s)
Antibodies, Monoclonal/metabolism , Biomarkers, Tumor/blood , Carcinoma, Non-Small-Cell Lung/blood , Early Detection of Cancer/methods , Lung Neoplasms/blood , Proteome/metabolism , Adolescent , Adult , Aged , Aged, 80 and over , Animals , Antibodies, Monoclonal/chemistry , Antibody Specificity , Area Under Curve , Biomarkers, Tumor/immunology , Carcinoma, Non-Small-Cell Lung/diagnosis , Case-Control Studies , Complement Factor H/immunology , Complement Factor H/metabolism , Female , Glycoproteins/blood , Glycoproteins/immunology , Haptoglobins/immunology , Haptoglobins/metabolism , Humans , Immunoassay/methods , Lung Neoplasms/diagnosis , Male , Mice , Mice, Inbred BALB C , Middle Aged , Proteomics , ROC Curve , Young Adult , alpha 1-Antichymotrypsin/blood , alpha 1-Antichymotrypsin/immunologyABSTRACT
We define mAb proteomics as the global generation of disease specific antibodies that permit mass screening of biomarkers. An integrated, high-throughput, disease-specific mAb-based biomarker discovery platform has been developed. The approach readily provided new biomarker leads with the focus on large-scale discovery and production of mAb-based, disease-specific clinical assay candidates. The outcome of the biomarker discovery process was a highly specific and sensitive assay, applicable for testing of clinical validation paradigms, like response to treatment or correlation with other clinical parameters. In contrast to MS-based or systems biology-based strategies, our process produced prevalidated clinical assays as the outcome of the discovery process. By re-engineering the biomarker discovery paradigm, the encouraging results presented in this paper clearly demonstrate the efficiency of the mAb proteomics approach, and set the grounds for the next steps of studies, namely, the hunt for candidate biomarkers that respond to drug treatment.
