Transcriptome-Wide Analysis of Bone Marrow CD19+ Cells As Well As Bone Marrow B Cell Clones of Waldenström’s Macroglobulinemia (WM) Vs. IgM Monoclonal Gammopathy of Undetermined Significance (IgM-MGUS)

• Published in: Blood 2014-12, Vol.124 (21), p.5176-5176
• Main Authors: Trojani, Alessandra, Greco, Antonino, Tedeschi, Alessandra, Lodola, Milena, Di Camillo, Barbara, Frustaci, Anna Maria, Morra, Enrica
• Format: Article
• Language: English
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood.V124.21.5176.5176

We performed a transcriptome-wide analysis of bone marrow (BM) CD19+ cells of WM vs. IgM-MGUS to find the genes and key pathways mostly involved in the comparison between WM and IgM-MGUS. We isolated BM CD19+ and CD138+ cells of 36 WM and 13 IgM-MGUS using Miltenyi Microbeads and we performed expression analysis with Affymetrix GeneChip HG U133 Plus 2.0 Array. Data was processed using RMA and analyzed using SAM and a false discovery rate threshold of 5% to select the differentially expressed genes. To further select a subset of robust biomarkers, SVMs were used in a Monte Carlo bootstrap resampling schema with B=100 external training/test splits to discriminate between WM and IgM-MGUS. 641 probes were selected by SAM on CD19+ cells. SVMs permitted the selection of 66 robust biomarker genes (Fig.1) with a MCC accuracy on external samples equal to 0.87. Functional enrichment analysis demonstrated the involvement of the following pathways: - Notch signaling pathway: ADAM17 (promoting cell growth) was upregulated while AGO1 and AGO4 (negative regulation of translation) were downregulated in WM - CAPRIN1 and SORT1 (pro-apoptosis) were underexpressed while CIAPIN1 (anti-apoptosis) was overexpressed in WM - Purine/pyrimidine metabolism (cell growth): ENTPD5 (cell proliferation) was overexpressed while NT5E (catabolic process) was underexpressed in WM - Sphingolipid pathway: ACER3, COL4A3BP, GBA3 were downregulated in WM - Rho-protein signal transduction: FARP2 (cell survival) was overexpressed in WM - Transcription: ITPRIPL2, L3MBTL4 were underexpressed while NFX1, LIMS1 (cell aging) USF1 were overexpressed in WM - Immune system: HLA-C was upregulated in WM - PRKCA was underexpressed in WM and involved in 85 pathways (MAPK-NGF-EGF-VEGF-ErbB-Ras-HIF-1-mTOR-PI3K-Akt) - MIR1204///PVT1 (oncogene), METTL3 (RNA-methylation), MINA (cell survival) were upregulated in WM As a second step of the study, we investigated 7 IgM-MGUS and 11 WM (2 symptomatic and 9 asymptomatic) newly diagnosed patients. We isolated the BM B cell clones (CD45+,CD38+,CD19+,LAIR-1-; CD27dim, IgM+, CD22dim, CD25+) using cell sorting with MoFlo XDP 3 laser cell sorter (Beckman Coulter). We identified clonally restricted B lymphocyte populations using BD LSRFortessa Flow Cytometer equipped with 5 lasers. We performed gene expression profiling (GEP) of the isolated BM cell clones of WM vs. IgM-MGUS using Affymetrix next-generation GeneChip HTA 2.0 to further investigate genomic complexity. The data wa