Because of the long history of the use of formalin as the standard fixative for tissue processing in histopathology, there are a large number of archival formalin-fixed and paraffin-embedded (FFPE) tissue banks worldwide. These FFPE tissue collections, with attached clinical and outcome information, present invaluable resources for conducting retrospective protein biomarker investigations. In addition to sample amount constraints imposed by current proteome techniques including two-dimensional polyacrylamide gel electrophoresis and multidimensional liquid chromatography system, the lack of optimized methodologies for retrieving proteins from FFPE tissues further restricts the ability to perform the molecular analysis of archival tissues. By collaborating with Drs. Shan-Rong Shi and Clive R. Taylor from the University of Southern California (USC) Keck School of Medicine during the R41 Phase I studies, the combination of antigen retrieval (AR) with Gemini proteomic technologies not only accomplished the rigorous evaluation of the quality and the reproducibility of proteins retrieved from FFPE tissues for the optimization of AR methodology, but also demonstrated significant opportunities in the pursuit of biomarker discovery using archived FFPE tissue collections. The proposed synergistic efforts between Calibrant and the USC team during the R42 Phase II project aim to generate proteotypic peptide libraries among model and tumor FFPE tissues. These proteotypic peptide libraries represent the first step toward globally cataloging antigens retrievable from FFPE tissues and presenting the available epitope database for subsequent immunohistochemistry (IHC) antibody development. Besides providing guidelines for practitioners of IHC to select the optimized AR condition/antibody combination, the comparative proteomic and validation studies involving the use of proteotypic peptide libraries and associated antibodies will provide further enhancements in the reproducibility and the sensitivity of quantitative IHC measurements. The demand for quantitative IHC continues to escalate due to the widespread utilization of IHC in clinical diagnosis/prognosis and translational cancer research. Furthermore, the greatest expectations for targeted proteomics research using enriched and selected cells from high quality specimens reside in the identification of diagnostic, prognostic, and predictive biological markers in the clinical setting and during preclinical testing and clinical trials, as well as the discovery and validation of new protein targets in the biopharmaceutical industry. The Critical Path Opportunity Report released by FDA in March 2006 not only serves as the first specific blueprint for the Critical Path Initiative, an effort to streamline the drug-approval process by applying new strategies and technologies, but also highlights biomarker development as one of the """"""""most important areas for improving medical product development."""""""" Public Health Relevance Statement: By joining Calibrant's unique tissue proteome capabilities with the expertise of Dr. Clive R. Taylor at the University of Southern California in antigen retrieval-immunohistochemistry, the proposed research not only aims to evaluate and optimize quantitative immunohistochemistry measurements through the creation of proteotypic peptide libraries, but also focuses on further development and demonstration of a novel biomarker discovery paradigm for enabling comprehensive and comparative proteomic analysis of archived formalin-fixed and paraffin-embedded tissue collections in support of cancer research, diagnosis, and treatment.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Small Business Technology Transfer (STTR) Grants - Phase II (R42)
Project #
5R42CA122715-05
Application #
7915328
Study Section
Special Emphasis Panel (ZCA1-SRRB-C (M1))
Program Officer
Rahbar, Amir M
Project Start
2006-09-01
Project End
2012-08-31
Budget Start
2010-09-01
Budget End
2012-08-31
Support Year
5
Fiscal Year
2010
Total Cost
$750,000
Indirect Cost
Name
Calibrant Biosystems, Inc.
Department
Type
DUNS #
616684036
City
Gaithersburg
State
MD
Country
United States
Zip Code
20878
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Fang, Xueping; Wang, Chenchen; Lee, Cheng S (2013) Capillary electrophoresis-based proteomic techniques for biomarker discovery. Methods Mol Biol 919:181-7
Chien, Yuchen; Scuoppo, Claudio; Wang, Xiaowo et al. (2011) Control of the senescence-associated secretory phenotype by NF-?B promotes senescence and enhances chemosensitivity. Genes Dev 25:2125-36
Shi, Shan-Rong; Shi, Yan; Taylor, Clive R (2011) Antigen retrieval immunohistochemistry: review and future prospects in research and diagnosis over two decades. J Histochem Cytochem 59:13-32
Becker, Karl-Friedrich; Taylor, Clive R (2011) ""Liquid morphology"": Immunochemical analysis of proteins extracted from formalin-fixed paraffin-embedded tissues: Combining proteomics with immunohistochemistry. Appl Immunohistochem Mol Morphol 19:1-9
Jinawath, Natini; Vasoontara, Chanont; Jinawath, Artit et al. (2010) Oncoproteomic analysis reveals co-upregulation of RELA and STAT5 in carboplatin resistant ovarian carcinoma. PLoS One 5:e11198
Balgley, Brian M; Guo, Tong; Zhao, Kejia et al. (2009) Evaluation of archival time on shotgun proteomics of formalin-fixed and paraffin-embedded tissues. J Proteome Res 8:917-25