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Prostate Cancer: Protocol Optimization study for biomarker identification in clinical tissues


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TITLE: Prostate Cancer: Protocol Optimization study for biomarker identification in clinical tissues


Formalin Fixed Paraffin Embedded (FFPE) tumor tissue

ØPreferred method of tissue preservation in clinical practice
ØValuable source of research material since large cohorts are archived in hospitals’ biobanks
ØNotorious for suboptimal Protein quantity and quality
ØSample quality evaluation is key for optimizing downstream processing for proteomics analysis
ØProtocol optimization for the proteomics analysis of FFPE tissues to obtain proteins of good quality and quantity to reliably identify biomarkers able to predict the clinical outcome of Prostate Cancer
ØSeven protein extraction protocols including combinations of homogenization means (beads, sonication, boiling) and buffers (Tris-HCl and Urea-Thiourea) were applied
ØThe quantity and quality of the extracted proteins were evaluated by SDS-PAGE and GELC-MS/MS
ØA combination of cell lysis by FASP buffer with bead homogenization, sonication and boiling, provided efficient protein extraction from FFPE tissues based on gel image analysis
ØLC-MS/MS analysis of FFPE and FF samples prepped with the selected protocol showed a relatively high number of total proteins identifications in both cases (FF IDs: 2019 , FFPE IDs:1463)
ØProtein identification and quantification by LC-MS/MS showed a high overlap between the FF and FFPE samples proving the efficacy of the optimized protocol
In Conclusion, our study presents a focused effort to optimize and establish high resolution protocols for analysis of clinical FFPE tissues. This study is expected to open up multiple research avenues considering the availability of FFPE tissue and enhance the ongoing efforts of Prostate Cancer prognostic biomarker identification.