Studies on proteome profiling of pancreatic cancer cells and proteomic biomarker development for Intraductal Papillary Mucinous Neoplasm

Abstract

ABSTRACT Yikwon Kim Division of Cancer Biology Seoul National University Graduate School

Introduction: Pancreatic cancer (PC) is one of the most common causes of cancer mortality worldwide. The 5-year survival rate of PC is less than 3% because of the late diagnosis of this disease and poor responsiveness to chemotherapy. A better understanding of the cellular and molecular mechanisms involved in drug resistance is imperative for improving drug efficiency and patient outcomes. Further, much effort has been made in examining precursor lesions of PC to detect PC in the early stages.

Methods: (Chapter I) To investigate the underlying molecular mechanisms for drug resistance (gemcitabine resistance) in pancreatic cancer, we performed label-free quantification of protein expression in intrinsic gemcitabine-resistant and -sensitive human pancreatic adenocarcinoma cell lines using our improved proteomic strategy, combined with filter-aided sample preparation (FASP), single-shot liquid chromatography-mass spectrometry (single-shot LC-MS/MS), enhanced spectral counting, and a statistical method based on a power law global error model (PLGEM). (Chapter II) Further, we performed Multiple Reaction Monitoring (MRM) quantification to identify plasma-based biomarkers of precursor lesions of PC, especially Intraductal Papillary Mucinous Neoplasm (IPMN).

Results: In chapter I, we identified 1931 proteins and quantified 837 differentially expressed proteins in the BxPC3, PANC-1, and HPDE cell lines. Bioinformatics analysis identified 15 epithelial to mesenchymal transition (EMT) markers and 13 EMT-related proteins that were closely associated with drug resistance were differentially expressed. Interestingly, 8 of these proteins were involved in glutathione and cysteine/methionine metabolism. In chapter II, a robust MRM pipeline was applied to discovery and verify IPMN biomarker candidates in a large cohort of plasma samples (n = 184). Eleven proteins were selected as IPMN marker candidates with high confidence in 184 plasma samples, comprising a training (n = 84) and test set (n = 100). Further, to improve the discriminatory power, a 6-protein panel was constructed by combining marker candidates. The multimarker panel had high discriminatory power in distinguishing between IPMN and controls, including other benign diseases.

Conclusions: First, we identified 15 known EMT markers and 13 EMT-related proteins in the comparison between BxPC3 and PANC-1 cells, suggesting that the EMT-related proteins may confer gemcitabine chemoresistance. Second, 11 candidate markers and the multimarker panel of IPMN showed high abundance changes in IPMN from the heterogeneous control group, demonstrating its value as a diagnostic marker of IPMN.

* This work is published in Molecules and Cells/Journal of proteome research Journal. ------------------------------------------------------------------------------------------------------------- Keywords: Pancreatic cancer (PC), Drug resistance, Intraductal Papillary Mucinous Neoplasm (IPMN), Biomarker, Multiple Reaction Monitoring (MRM), LC-MS/MS, Proteomics, Student Number: 2008-21966