Qing Kay Li1,2, Yingwei Hu3, Lijun Chen3, Michael Schnaubelt3, Daniel Cui Zhou4, Yize Li4, Rita Jui-Hsien Lu4, Mathangi Thiagarajan5, Galen Hostetter6, Chelsea J Newton6, Scott D Jewell6, Gil Omenn7, Ana I Robles8, Mehdi Mesri8, Oliver F Bathe9, Bing Zhang10, Li Ding4, Ralph H Hruban3,11, Daniel W Chan3,11, Hui Zhang12,13. 1. Department of Pathology, the Johns Hopkins University, 400 N Broadway, Smith Bldg Rm 4011, Baltimore, MD, 21231, USA. qli23@jhmi.edu. 2. Department of Oncology, Sidney Kimmel Cancer Center, Johns Hopkins Medical Institutions, 600 N. Wolfe Street, Baltimore, MD, USA. qli23@jhmi.edu. 3. Department of Pathology, the Johns Hopkins University, 400 N Broadway, Smith Bldg Rm 4011, Baltimore, MD, 21231, USA. 4. Department of Oncology, Washington University at Saint Louis, St Louis, MO, USA. 5. Leidos Biomedical Research Inc, Frederick National Laboratory for Cancer Research, Frederick, MD, USA. 6. Van Andel Research Institute, Grand Rapids, MI, USA. 7. Department of Computational Medicine & Bioinformatics, University of Michigan, Ann Arbor, MI, USA. 8. Office of Cancer Clinical Proteomics Research, National Cancer Institute, Rockville, MD, USA. 9. Department of Surgery and Oncology, Cumming School of Medicine, University of Calgary, Calgary, AB, Canada. 10. Lester and Sue Smith Breast Center, Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX, USA. 11. Department of Oncology, Sidney Kimmel Cancer Center, Johns Hopkins Medical Institutions, 600 N. Wolfe Street, Baltimore, MD, USA. 12. Department of Pathology, the Johns Hopkins University, 400 N Broadway, Smith Bldg Rm 4011, Baltimore, MD, 21231, USA. huizhang@jhu.edu. 13. Department of Oncology, Sidney Kimmel Cancer Center, Johns Hopkins Medical Institutions, 600 N. Wolfe Street, Baltimore, MD, USA. huizhang@jhu.edu.
Abstract
BACKGROUND: The identification of differentially expressed tumor-associated proteins and genomic alterations driving neoplasia is critical in the development of clinical assays to detect cancers and forms the foundation for understanding cancer biology. One of the challenges in the analysis of pancreatic ductal adenocarcinoma (PDAC) is the low neoplastic cellularity and heterogeneous composition of bulk tumors. To enrich neoplastic cells from bulk tumor tissue, coring, and laser microdissection (LMD) sampling techniques have been employed. In this study, we assessed the protein and KRAS mutation changes associated with samples obtained by these enrichment techniques and evaluated the fraction of neoplastic cells in PDAC for proteomic and genomic analyses. METHODS: Three fresh frozen PDAC tumors and their tumor-matched normal adjacent tissues (NATs) were obtained from three sampling techniques using bulk, coring, and LMD; and analyzed by TMT-based quantitative proteomics. The protein profiles and characterizations of differentially expressed proteins in three sampling groups were determined. These three PDACs and samples of five additional PDACs obtained by the same three sampling techniques were also subjected to genomic analysis to characterize KRAS mutations. RESULTS: The neoplastic cellularity of eight PDACs ranged from less than 10% to over 80% based on morphological review. Distinctive proteomic patterns and abundances of certain tumor-associated proteins were revealed when comparing the tumors and NATs by different sampling techniques. Coring and bulk tissues had comparable proteome profiles, while LMD samples had the most distinct proteome composition compared to bulk tissues. Further genomic analysis of bulk, cored, or LMD samples demonstrated that KRAS mutations were significantly enriched in LMD samples while coring was less effective in enriching for KRAS mutations when bulk tissues contained a relatively low neoplastic cellularity. CONCLUSIONS: In addition to bulk tissues, samples from LMD and coring techniques can be used for proteogenomic studies. The greatest enrichment of neoplastic cellularity is obtained with the LMD technique.
BACKGROUND: The identification of differentially expressed tumor-associated proteins and genomic alterations driving neoplasia is critical in the development of clinical assays to detect cancers and forms the foundation for understanding cancer biology. One of the challenges in the analysis of pancreatic ductal adenocarcinoma (PDAC) is the low neoplastic cellularity and heterogeneous composition of bulk tumors. To enrich neoplastic cells from bulk tumor tissue, coring, and laser microdissection (LMD) sampling techniques have been employed. In this study, we assessed the protein and KRAS mutation changes associated with samples obtained by these enrichment techniques and evaluated the fraction of neoplastic cells in PDAC for proteomic and genomic analyses. METHODS: Three fresh frozen PDAC tumors and their tumor-matched normal adjacent tissues (NATs) were obtained from three sampling techniques using bulk, coring, and LMD; and analyzed by TMT-based quantitative proteomics. The protein profiles and characterizations of differentially expressed proteins in three sampling groups were determined. These three PDACs and samples of five additional PDACs obtained by the same three sampling techniques were also subjected to genomic analysis to characterize KRAS mutations. RESULTS: The neoplastic cellularity of eight PDACs ranged from less than 10% to over 80% based on morphological review. Distinctive proteomic patterns and abundances of certain tumor-associated proteins were revealed when comparing the tumors and NATs by different sampling techniques. Coring and bulk tissues had comparable proteome profiles, while LMD samples had the most distinct proteome composition compared to bulk tissues. Further genomic analysis of bulk, cored, or LMD samples demonstrated that KRAS mutations were significantly enriched in LMD samples while coring was less effective in enriching for KRAS mutations when bulk tissues contained a relatively low neoplastic cellularity. CONCLUSIONS: In addition to bulk tissues, samples from LMD and coring techniques can be used for proteogenomic studies. The greatest enrichment of neoplastic cellularity is obtained with the LMD technique.
Authors: Ralph H Hruban; Matthias M Gaida; Elizabeth Thompson; Seung-Mo Hong; Michaël Noë; Lodewijk Aa Brosens; Martine Jongepier; G Johan A Offerhaus; Laura D Wood Journal: J Pathol Date: 2019-04-15 Impact factor: 7.996
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Authors: Daniel Öhlund; Abram Handly-Santana; Giulia Biffi; Ela Elyada; Ana S Almeida; Mariano Ponz-Sarvise; Vincenzo Corbo; Tobiloba E Oni; Stephen A Hearn; Eun Jung Lee; Iok In Christine Chio; Chang-Il Hwang; Hervé Tiriac; Lindsey A Baker; Dannielle D Engle; Christine Feig; Anne Kultti; Mikala Egeblad; Douglas T Fearon; James M Crawford; Hans Clevers; Youngkyu Park; David A Tuveson Journal: J Exp Med Date: 2017-02-23 Impact factor: 14.307