The Use of Itraq and Mass Spectrometry To Study Multiple Myeloma Associated with Bortezomib Resistance
JH Zhao, J Chen, K Evans, H Chang. University Health Network and University of Toronto, Toronto, Canada
Background: Bortezomib, a novel proteasome inhibitor, represents a promising new clinical strategy presently deployed in clinical trials of relapsed and refractory multiple myeloma (MM). However, only 30-40% of MM patients respond to this treatment. The mechanism of resistance is not understood. Our previous study on the molecular cytogenetic profiles of MM showed that the response to bortezomib in MM patients is independent of high-risk genomic aberrations. In order to gain insights into the mechanisms of bortezomib resistance, we examined the proteome of bortezomib sensitive and resistant MM cells by comparative global proteomic analysis.
Design: Two myeloma cell lines, 8226/S (bortezomib senbsitive) and 8226/R5 (bortezomib resistance) were used in this study to determine differentially expressed proteins possibly directly involved in the acquisition of bortezomib resistance. The proteins of bortezomib sensitive and resistant cells were simultaneously identified and quantified by using isobaric tag peptide labeling (iTRAQ) technology, followed by hybrid quadruple time-of-flight mass spectrometry (MS). The MS based multiple-reaction-monitoring (MRM) technique was used to independently verify the quantitative differences of the protein expression levels between the two cell lines.
Results: The iTRAQ labeling combined with MS approach identified a large population of 665 proteins from MM cells. Among those proteins, 176 proteins (27%) were found differentially expressed between 8226/S and 8226/R5. Of the 176 proteins, 115 (65%) were up-regulated whereas 61 (35%) were down-regulated. These proteins can be classified into several major cellular pathways involving cell proliferation, differentiation, nuclear acid metabolism, molecular transport and protein synthesis. Several selected highly expressed proteins such as DEN1C, Ubiqulin-4, SCFD1 may be considered as potential protein biomarkers for bortezomib resistance.
Conclusions: iTRAQ in concert with MS is a powerful approach to identify differentially expressed proteins related to drug resistance in MM. Further verification and functional analysis of the proteins of interest in MM bortezomib resistance are ongoing.
Category: Special Category for 2009 - Pan-genomic/Pan-proteomic approaches to Cancer
Tuesday, March 10, 2009 9:30 AM
Poster Session III # 226, Tuesday Morning