科研之友
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摘要
Objectives: Non-invasive tests for early detection of lung cancer are an important unmet clinical need. CIZ1b plasma biomarker can discriminate stage I lung cancer from within high-risk groups with clinically useful accuracy, with ROC AUCs in excess of 0.9 for two independent retrospective cohorts, and could therefore meet this need. Our aim was to characterise the native state of the biomarker and develop a quantitative immunoassay. Design and methods: Selective denaturation, preparative electrophoresis and mass spectrometry of human plasma were used to characterise the biomarker and interaction partners. A sandwich ELISA was generated, and specificity for CIZ1b biomarker tested on lung cancer patient plasma. Results: CIZ1b biomarker is a denaturation-resistant complex between a C-terminal fragment of CIZ1 bearing the CIZ1b epitope specified by alternative splicing of exonl4, and fibrinogen alpha chain. Reconstitution of the biomarker epitope with purified fibrinogen and CIZ1b, but not CIZla (non-alternatively spliced exon 14) confirmed the specificity of the results. The endogenous complex is highly stable in lung cancer plasma and can be quantified by pairing of a CIZ1b exon junction specific antibody with detection of fibrinogen. Application of this sandwich ELISA to a prospectively collected development set of plasmas reveals the same level of accuracy as the western blot used to validate the discriminatory capability of the biomarker. Conclusions: Unexpected and unusual molecular structure of CIZ1b in native plasma has complicated immunoassay design, and delayed translation of this promising biomarker. However, CIZ1b can now be measured using a high-throughput, hospital-friendly sandwich ELISA format, overcoming an important barrier to further clinical development and application of this blood test for early stage lung cancer.
- 出版日期2017-4
