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Veracode Challenge Winner
Christopher Sears
AmberGen, Inc.
Watertown, MA
Gene-Assisted Serum Proteomics: Blood-based Colorectal Cancer Diagnostic and Prognostic Assay Development using Multiplexed VeraCode Beads.
Application Abstract
Panels of tumor associated autoantigens (TAAs) for blood-based clinical diagnosis and prognosis of colorectal cancer (CRC) will be developed using the unique multiplexing capability of the VeraCode Beads. The proposed six-month study will involve screening blood samples of CRC patients against targeted bead-protein libraries in order to detect autoantibody binding and thereby identify key TAA biomarkers that can be used in a multiplexed clinical assay. VeraCode protein-bead libraries can be produced rapidly and inexpensively using AmberGen's proprietary cell-free protein expression and protein-capture technology. The initial selection of proteins to be screened will be based on AmberGen's discovery of a panel of over 100 genes whose expression is highly correlated with the recurrence of CRC. Gene over-expression is linked to likelihood of autoantigenicity, as confirmed in our initial studies. The multiplexed VeraCode platform is ideal for this novel, targeted proteomic approach which we term Gene-Assisted Serum Proteomics. By leveraging the VeraCode technology, we expect to develop the first commercial TAA blood-based CRC cancer assays that can be used for early diagnosis, guiding therapy and developing anticancer vaccines.
Biological Question
The near-term objective of this proposal is to design a sensitive, low-cost blood test for colorectal cancer (CRC) diagnosis and prognosis, a disease which ranks as the second leading cause of cancer deaths in the Western world. A significant reduction in the high mortality of CRC could be achieved through effective screening for premalignant adenomas and early stage carcinomas. Furthermore, a test that predicts which patients will recur or not recur (e.g. develop metastases) would also be extremely useful to categorize patients which require aggressive treatment and those that do not. A promising area of cancer research is based on the discovery of cancer specific tumor-associated autoantigens (TAAs). A panel of such TAAs could form the basis for an effective non-invasive blood test for early CRC detection. Several reports have demonstrated the importance of TAA discovery as an immunological approach to cancer diagnostics. However, an important challenge is to find an effective method to select and screen the large number of potential TAAs which in principle includes the entire human proteome. Our approach to this problem is to utilize a highly effective method of genetic expression
profiling (GEP), developed at AmberGen, as a filter to produce a targeted protein sub-array that leverages the VeraCode Beads' unsurpassed sensitivity and multiplexing capability. The initial study is expected to validate this combined approach and lead to the identification of several TAA biomarkers which could be used as part of a multiplexed VeraCode Bead assay for early diagnosis, guiding therapy and developing anticancer vaccines.
Experimental Outline
The focus of the six-month study will be on the development of TAA signatures to predict CRC recurrence (with diagnostic assays developed beyond this study). VeraCode Beads will be used to screen blood serum/plasma samples from CRC patients against a panel of potential autoantigens. The panel will be selected on the basis of an earlier NIH-sponsored gene expression profiling (GEP) study which resulted in a highly correlated gene signature for CRC recurrence. The top 40 over-expressing genes from this study will be produced in a cell-free protein expression system using proprietary methods and reagents developed by AmberGen. This approach facilitates rapid expression and direct capture of the proteins in an ultra-pure form on VeraCode Beads and also facilitates normalization for variations in proteins bound from bead-to-bead using the two-laser detection of the BeadXpress system. The multiplex assay performance using the VeraCode system will first be evaluated using known CRC autoantigens and matching blood samples already validated by AmberGen. Blood samples (25 recurrent and 25 non-recurrent) from clinically annotated Stage I and II CRC patients which are matched to tumors from the GEP will then be screened on the VeraCode BeadXpress to identify putative autoantigens. Following screening of these first 40 candidate autoantigens, more will be tested as needed. Results will be analyzed using the PraxisTM software system from Ananomouse Inc. which provides a wide range of powerful statistical and graphical methods suitable for signature development. The resulting signature will be validated in a double blinded-study involving an additional cohort of CRC patients (50). Resources devoted to this project include a team of 12 scientists (6 Ph.D.) trained in protein expression, multiplex assay development, proteomics, biostatistics and biomarker discovery along with a fully equipped laboratory including a special focus on protein expression, gene expression and mutation profiling.
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