Mass Spectrometry-Based Glycoproteomic Profiling Identifies SIRP alpa as a Potential Protein Biomarker in Primary Mediastinal Large B-Cell Lymphoma
Noah A Brown, Delphine Rolland, Damian Fermin, Venkatesha Basrur, Dafydd Thomas, Farah Keyoumarsi, Kevin Conlon, Kojo SJ Elenitoba-Johnson, Megan S Lim. University of Michigan, Ann Arbor, MI
Background: Primary mediastinal large B-cell lymphoma (PMBCL) is currently recognized as a non-Hodgkin B-cell lymphoma with pathologic, clinical and molecular characteristics which are distinct from other diffuse large B-cell lymphomas (DLBCL) and from classical Hodgkin lymphoma (cHL). However, overlapping histologic and immunophenotypic features of these lymphomas make this diagnostic distinction challenging in some cases. Since a majority of N-linked glycoproteins are either localized on the membrane or secreted, these proteins are attractive targets for the identification of biomarkers that could aid in the distinction of these three lymphomas.
Design: Using a proteomic strategy targeting N-linked glycosylated proteins, we characterized the glycoproteomic profiles of PMBCL, DLBCL, and cHL. Cell lines derived from PMLBL (K1106 and MEDB1), DLBCL (OCILY2), and cHL (L428, L1236 and KMH2) were used for solid-phase extraction of N-linked glycopeptides which were identified and quantified using liquid chromatography and tandem mass spectrometry. A protein selectively expressed in PMBCL was selected for validation using immunohistochemistry of tissue microarrays constructed from 27 cases of PMBCL, 70 DLBCL and 120 cHL.
Results: Glycopeptides from over 250 proteins were identified from each cell line using false discovery rate of < 4.5 %. Over 90% of the peptides contained the NXST motif which identifies distinct sites of asparagines glycosylation. Importantly, mass spectrometry data correlated strongly with known proteins expressed by cHL (CD30) and PMBCL/DLBCL (CD18, CD19, CD22, CD45, CD79A, and CD79B). In addition, a novel glycoprotein signal regulatory protein alpha (SIRP alpha) was found to be selectively expressed by PMBCL. SIRP alpha is a transmembrane signaling protein expressed in monocytes, dendritic cells and granulocytes. Binding to its ligand (CD47) results in negative regulation of leukocyte adhesion and transmigration, macrophage fusion, phagocytosis. Immunohistochemical analysis of tissue microarrays using a monoclonal antibody for SIRP alpha demonstrated expression in 55.6% of PMBCL compared to 22.9% of DLBCL (p<0.01) and 6.7% of cHL (p<0.0001).
Conclusions: We characterized the glycoproteomics signature of PMBCL, DLBCL or cHL using mass spectrometry. In doing so, we identified a potential biomarker more commonly expressed in PMBCL than in either DLBCL or cHL. This study demonstrates that mass spectrometry-based glycoproteomic profiling can be used for discovery of novel biomarkers and phenotypic profiling.
Category: Special Category - Pan-genomic/Pan-proteomic approaches to Cancer
Tuesday, March 20, 2012 9:30 AM
Poster Session III # 291, Tuesday Morning