- Proteomic analysis of primary and metastatic breast cancers reveals a wide range of expression of the folate receptor, a potential drug target In collaboration with: Joyce O’Shaughnessy, Texas Oncology – Baylor Charles A. Sammons Cancer Center.
- Quantitative measurement of HER2 levels by multiplexed mass spectrometry to predict survival in gastric cancer patients treated with trastuzumab In collaboration with: Chan-Young Ock, Do-Youn Oh, Yung-Jue Bang, Seoul National University Hospital.
- HER2 quantification benefit from anti-HER2 therapy in HER2 positive breast cancer In collaboration with: Paolo Nuciforo and Javier Cortes, Vall D’Hebron Institute of Oncology, and Maurizio Scaltriti, Memorial Sloan Kettering Cancer Center.
San Antonio Breast Cancer Symposium
- Quantitative measurement of HER2 levels by multiplexed mass spectrometry from FFPE tissue predicts survival in patients treated with anti-HER2 based therapy. Paolo Nuciforo et al.
The 26th EORTC-NCI-AACR Symposium
- A novel clinical tool that provides quantitative and accurate measurement of Met protein. (Presented by Fabiola Cecchi, Ph.D. at the 26th EORTC-NCI-AACR Symposium on Molecular Targets and Cancer Therapeutics).
- Development and Clinical Validation of a Quantitative Mass Spectrometric Assay for PD-L1 Protein in FFPE NSCLC Samples. (Presented by Sheeno Thyparambil, Ph.D. at the 26th EORTC-NCI-AACR Symposium on Molecular Targets and Cancer Therapeutics).
The 2014 Chicago Multidisciplinary Symposium in Thoracic Oncology
- Quantification of ALK from Non-Small Cell Lung Cancer (NSCLC) FFPE Tissue by Targeted Mass Spectrometry.(Presented by Wei-Li Liao, Ph.D. at the 2014 Chicago Multidisciplinary Symposium in Thoracic Oncology). PLoS ONE 9 (7): e100586.
- Absolute Quantitation of Met Using Mass Spectrometry for Clinical Application: Assay Precision, Stability, and Correlation with MET Gene Amplification in FFPE Tumor Tissue.
The 2014 ASCO Annual Meeting
- Nuciforo, Paolo, et al. Correlation of high levels of HER2 measured by multiplex mass spectrometry with increased overall survival in patients treated with anti-HER2-based therapy. ASCO 2014. Chicago, IL
Literature Supporting the OncoPlexDx Technology and Platform
- Hembrough T, Thyparambil S, Liao WL, Darfler M, Abdo J, Bengali K, Taylor P, Tong J, Lara-Guerra H, Waddell TK, Moran MF, Tsao MS, Krizman DB, Burrows J. SRM analysis of EGFR in formalin fixed tumor tissue: Application to personalized medicine.Clinical Proteomics, 9:5, 2012
- Bateman NW, Sun M, Bhargava R, Hood BL, Darfler MM, Kovatich AJ, Hooke JA, Krizman DB, Conrads TP. Differential proteomic analysis of late-stage and recurrent breast cancer from formalin-fixed paraffin-embedded tissues. J. Proteome Res. 10:1323–1332, 2011
- Desouza LV, Krakovska O, Darfler MM, Krizman DB, Romaschin AD, Colgan TJ, Siu KW. mTRAQ-based quantification of potential endometrial carcinoma biomarkers from archived formalin-fixed paraffin-embedded tissues. Proteomics10(17):3108-16, 2010
- Huang SK, Darfler MM, Nicholl MB, You J, Bemis KG, Tegeler TJ, Wang M, Wery JP, Chong KK, Nguyen L, Scolyer RA, Hoon DS. LC/MS-based quantitative proteomic analysis of paraffin-embedded archival melanomas reveals potential proteomic biomarkers associated with metastasis. PLoS One. 2009;4(2):e4430
- Jain MR, Liu T, Hu J, Darfler M, Fitzhugh V, Rinaggio J, Li H. Quantitative Proteomic Analysis of Formalin Fixed Paraffin Embedded Oral HPV Lesions from HIV Patients. Open Proteomics J. 2008;1:40-45.
- Cheung W, Darfler MM, Alvarez H, Hood BL, Conrads TP, Habbe N, Krizman DB, Mollenhauer J, Feldman G, Maitra A. Application of a Global Proteomic Approach to Archival Precursor Lesions: Deleted in Malignant Brain Tumors 1 (DMBT1) and Tissue Transglutaminase-2 (TG2) are upregulated in pancreatic cancer precursors. Pancreatology 8(6):608-616, 2008
- Patel V, Hood BL, Molinoli A, Lee NH, Conrads TP, Braisted JC, Krizman DB Veenstra TD, Gutkind JS. Proteomic Analysis of Laser-Captured Paraffin-Embedded Tissues: A Molecular Portrait of Head and Neck Cancer Progression. Clinical Cancer Research 14(4):1002-1014, 2008
- Barron JA, Young HD, Ringeisen BR, Dlott DD, Darfler MM, Krizman DB. Printing of Protein Microarrays Via a Capillary-Free Fluid Jetting Mechanism. Proteomics 4:4136-4144, 2005
- Hood BL, Darfler MM, Guiel TG, Furusato B, Lucas DA, Ringeisen BR, Sesterhenn IA, Conrads TP, Veenstra TD, Krizman DB. Proteomic analysis of formalin fixed prostate cancer tissue. Mol Cell Proteomics 4(11):1741-1753, 2005.
- Prieto DA, Hood BL, Darfler MM, Guiel TG, Lucas DA, Conrads TP, Veenstra TD, Krizman DB. Liquid Tissue™: Proteomic profiling of formalin-fixed tissues. Biotechniques June, 2005
Quoted Statements from EPI Peer-Reviewed Manuscripts
Hembrough T, Thyparambil S, Liao WL, Darfler M, Abdo J, Bengali K, Taylor P, Tong J, Lara-Guerra H, Waddell TK, Moran MF, Tsao MS, Krizman DB, Jon Burrows J. SRM analysis of EGFR in formalin fixed tumor tissue: Application to personalized medicine. Clinical Proteomics, 9:5, 2012.
- SRM methodology provides a quantitative, sensitive, and reproducible approach for analysis of proteins in any biological sample and this report demonstrates the application of SRM methodology to measuring EGFR protein levels in FFPE biological samples, including patient tumor tissue.
- The application of this assay to a cohort of FFPE tissues surgically resected from NSCLC patients that participated in an EGFR-targeted therapeutic trial indicates its ability to monitor EGFR levels directly in patient-derived tumor tissue.
- Clinical application of this assay could provide for patient selection to optimize EGFR-targeted therapy strategies, predict patient outcome, and determine the potential for therapeutic resistance to other receptor tyrosine kinase inhibitor molecules.
- Combining the current EGFR assay in a multiplex fashion with similar assays for IGF-1R and cMet could provide for optimal therapeutic choice and a drug resistance biomarker panel to drive targeted therapeutic efforts in NSCLC, as well as other cancers.
Cheung W, Darfler MM, Alvarez H, Hood BL, Conrads TP, Habbe N, Krizman DB, Mollenhauer J, Feldman G, Maitra A. Application of a Global Proteomic Approach to Archival Precursor Lesions: Deleted in Malignant Brain Tumors 1 (DMBT1) and Tissue Transglutaminase-2 (TG2) are upregulated in pancreatic cancer precursors. Pancreatology8(6):608-616, 2008.
- Global proteomics analysis using the Liquid Tissue® workflow is a feasible approach for unbiased biomarker discovery in limited archival material, particularly applicable to precursor lesions of cancer.
- Two significant candidate marker proteins for IPMN were identified by MS, Deleted in Malignant Brain Tumors 1 (DMBT1) and Tissue Transglutaminase 2 (TG2).
- We have confirmed the upregulation of two of the expressed proteins (DMBT1 and TG2) in IPMNs versus normal pancreatic tissues by immunohistochemistry, underscoring the validity of our approach for tumor marker discovery in precursor lesions.
Hood BL, Darfler MM, Guiel TG, Furusato B, Lucas DA, Ringeisen BR, Sesterhenn IA, Conrads TP, Veenstra TD, Krizman DB. Proteomic analysis of formalin fixed prostate cancer tissue. Mol Cell Proteomics 4(11):1741-1753, 2005.
- This study represents the first differential large scale proteome analysis from FFPE archived tissue.
- Furthermore, this study provides a technological framework by which the vast archive of pathologically characterized FFPE clinical samples may be analyzed utilizing state of the art MS-based proteomic technologies.
- In contrast to existing tissue microdissection approaches, the novel microdissection technology described here has been developed and optimized for procurement of cells directly from standard formalin fixed tissue sections for downstream protein analysis.
- In addition, this tissue microdissection technology is a noncontact method that explodes cells directly downward into a collection tube providing the added benefit of CAD/CAM automation for high throughput tissue microdissection capabilities.
- Data presented demonstrate the potential for identification of proteins and their relative expression levels and relate this information to a particular disease state from FFPE tissue.
- Development of the capability to conduct large scale analyses of tissues using proteomics approaches analogous to the scale and throughput of high throughput gene expression analysis could have far reaching implications for protein biomarker investigations of disease through interrogation of the vast archived FFPE tissue collections.
Prieto DA, Hood BL, Darfler MM, Guiel TG, Lucas DA, Conrads TP, Veenstra TD, Krizman DB. Liquid Tissue™: Proteomic profiling of formalin-fixed tissues. Biotechniques June, 2005.
- This report demonstrates the analysis of peptides obtained from formalin-fixed tissue utilizing a variety of MS platforms, including microcapillary reversed-phase liquid chromatography (µRPLC) tandem mass spectrometry (MS/MS), matrix assisted laser desorption ionization (MALDI) tandem time-of-flight (TOF-TOF), and surface-enhanced laser desorption ionization time-of-flight (SELDI-TOF).
- Development of methods to use state-of-the-art proteomic discovery tools to analyze FFPE tissue provides an exciting new opportunity to identify disease-specific biomarkers in pathologically defined samples.
- Liquid Tissue® MS protein preparation allows researchers to utilize the vast, already existing, collections of formalin-fixed paraffin-embedded (FFPE) tissues for the procurement of peptides and the analysis across a variety of MS platforms.
- Results demonstrate that the Liquid Tissue® methodology provides the ability to unlock the proteome of the world’s vast reservoir of archived tissue for large-scale discovery and validation of biomarkers to improve disease diagnosis and therapy.
Quoted Statements from Manuscripts Using Liquid Tissue Technology (no EPI authorship)
Güzel C, Ursem NT, Dekker LJ, Derkx P, Joore J, van Dijk E, Ligtvoet G, Steegers EA, Luider TM. Multiple reaction monitoring assay for pre-eclampsia related calcyclin peptides in formalin fixed paraffin embedded placenta. J Proteome Res. 2011 Jul 1;10(7):3274-82.
- Data presented show that SRM on laser microdissected material from FFPE tissue material is possible.
- The developed MRM assay to study quantitative levels of proteins in FFPE laser microdissected cells using nonisotopic-labeled chemical analogs of mass tagged internal references showed that in pre-eclamptic patients elevated levels of calcyclin is observed in placental trophoblast cells compared to normal trophoblast cells.
- By immunohistochemistry, we were able to confirm this observation in a qualitative manner.
Rodriguez FJ, Gamez JD, Vrana JA, Theis JD, Giannini C, Scheithauer BW, Parisi JE, Lucchinetti CF, Pendlebury WW, Bergen HR 3rd, Dogan A. Immunoglobulin derived depositions in the nervous system: novel mass spectrometry application for protein characterization in formalin-fixed tissues. Lab Invest. 2008 Oct;88(10):1024-37.
- Proteinaceous deposits encountered in biopsies of the nervous system often represent diagnostic challenges even after a thorough clinicopathologic work-up.
- Our present study extends the use of MS to the characterization of these deposits in FFPE. There was an excellent concordance with immunohistochemical results, although MS confirmed light-chain restriction and the nature of intracellular deposits in at least one case with noncontributory immunostaining.
- Proteomics is beginning to have an important role, not only in research into human disease, but also in molecular diagnostics as well.
- The use of novel exquisitely sensitive and specific practical techniques, including MS, in the analysis of protein accumulations enhances the diagnostic precision afforded by routine histology and immunohistochemistry.
- We employed this technique (Liquid Tissue) in a focused study of immunoglobulin-derived deposits, but we believe in its potential to find widespread applications in diagnostic surgical pathology.
Confirmatory Studies/Reviews of FFPE Tissue by Key Opinion Leaders
Gallien S, Duriez E, Domon B. Selected reaction monitoring applied to proteomics. J Mass Spectrom. 2011 Mar;46(3):298-312.
- The SRM technique is ideally suited for precise quantification, and under specific conditions for absolute quantification.
- Over the past few years, the SRM technology has rapidly developed to broaden its field of application. While initially limited to the quantification of a relative small number of peptides in biological samples, the new scheduling capabilities have enabled large screens allowing the detection and quantification of large sets of analytes (several hundreds).
Ostasiewicz P, Zielinska DF, Mann M, Wiśniewski JR. Proteome, phosphoproteome, and N-glycoproteome are quantitatively preserved in formalin-fixed paraffin-embedded tissue and analyzable by high-resolution mass spectrometry. J Proteome Res. 2010 Jul 2;9(7):3688-700.
- In many experiments, we employed high mass accuracy measurements not only in the MS but also in the MS/MS dimension. This demonstrates that very high data quality can be obtained from proteomes extracted from FFPE tissue material.
- Both qualitatively and quantitatively we were not able to detect differences in protein levels between fresh or fixed and paraffin embedded proteomes. This indicates that the protein modifications induced by FFPE are not permanent.
- The FFPE method described here has the potential for opening new perspectives in the proteomic exploration of archival clinical samples at the proteome level.