Dataset Information

Kinome profiling of diffuse large B-cell lymphoma cells in response to dasatinib

ABSTRACT: Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous disease, with at least one-third of its patients not responding to the current chemotherapy regimen, R-CHOP. By gene expression profiling, patients with DLBCL can be categorized into two clinically relevant subtypes: activated B-cell (ABC) DLBCL and germinal center B-cell (GCB). Patients with ABC DLBCL have a worse prognosis, and are defined by chronic, overactive signaling through the B-cell receptor and NF-κB pathways. We examined the effects of the Src family kinase (SFK) inhibitor dasatinib in a panel of ABC and GCB DLBCL cell lines, and found that the ABC DLBCL cell lines are much more sensitive to dasatinib than the GCB DLBCL cell lines. However, using multiplexed inhibitor bead coupled to mass spectrometry (MIB/MS) kinome profiling competition and western blot analysis, both subtypes display inhibition of the SFKs in response to dasatinib after both short- and long-term treatment. MIB/MS analyses revealed several cell cycle kinases, including CDK4, CDK6, and the Aurora kinases, are inhibited by dasatinib treatment in the ABC DLBCL subtype, but not in the GCB DLBCL subtype. The present findings have important implications for the clinical use of dasatinib for the treatment of ABC DLBCL, either alone or in combination with other agents.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): B Cell, Cell Culture

SUBMITTER: Laura Herring

LAB HEAD: Laura E. Herring

PROVIDER: PXD009380 | Pride | 2018-12-14

REPOSITORIES: Pride

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			Action	DRS
	20160809_PC159-Lawrence-Cann-MIBS-HT-C1.raw	Raw
	20160809_PC159-Lawrence-Cann-MIBS-HT-C2.raw	Raw
	20160809_PC159-Lawrence-Cann-MIBS-HT-C3.raw	Raw
	20160809_PC159-Lawrence-Cann-MIBS-HT-D1.raw	Raw
	20160809_PC159-Lawrence-Cann-MIBS-HT-D2.raw	Raw

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Publications

Dasatinib Is Preferentially Active in the Activated B-Cell Subtype of Diffuse Large B-Cell Lymphoma.

Cann Marissa L ML Herring Laura E LE Haar Lauren L LL Gilbert Thomas S K TSK Goldfarb Dennis D Richards Kristy L KL Graves Lee M LM Lawrence David S DS

Journal of proteome research 20181221 1

Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous disease, and at least one-third of its patients relapse after treatment with the current chemotherapy regimen, R-CHOP. By gene-expression profiling, patients with DLBCL can be categorized into two clinically relevant subtypes: activated B-cell (ABC) and germinal center B-cell (GCB) DLBCL. Patients with the ABC subtype have a worse prognosis than those with GCB, and the subtype is defined by chronic, over-active signaling through the B-cell ...[more]

PMID: 30540191

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Project description:The ABC subtype of diffuse large B cell lymphoma (DLBCL) remains the least curable form of this lymphoma despite recent advances in therapy. We have combined structural and functional genomics to triangulate on new oncogenic mechanisms and devise new therapeutic strategies. RNA interference screen revealed a dependence of ABC DLBCL cell lines on MYD88 and IRAK1. High throughput resequencing of RNA (RNA-Seq) revealed frequent somatic mutations in MYD88 that preferentially occurred in the ABC DLBCL subtype. Remarkably, one third of ABC DLBCL tumor samples harbored the same amino acid substitution, L265P, in the MYD88 TIR domain at an evolutionarily invariant residue in its hydrophobic core. This mutation was rare or absent in two other DLBCL subtypes, but was observed in 9% of MALT lymphomas. At a lower frequency, multiple other mutations were observed in the MYD88 TIR domain, occurring in both the ABC and GCB subtypes of DLBCL. Survival of ABC DLBCL lines bearing the L265P mutation was sustained by the mutant but not wild type MYD88 isoform, demonstrating that this MYD88 mutant is oncogenic and gain-of-function. The MYD88 L265P mutant assembled a protein complex that spontaneous triggers the phosphorylation of IRAK1, leading to NF-kB signaling, secretion of the cytokines IL-6, IL-10 and interferon-b, and JAK kinase signaling. These findings demonstrate that the MYD88 signaling pathway is integral to the pathogenesis of ABC DLBCL, providing a genetic rationale for therapeutic targeting of the MYD88 signaling pathway in this lymphoma subtype. To generate a gene expression signature of MYD88 signaling in ABC DLBCL, the HBL-1 cell line was transduced with retroviral vectors expressing either shMYD88-4 or shMYD88-7. Following puromycin selection, shRNA expression was induced for 24 or 48 hours and gene expression was measured, comparing uninduced (Cy3) to induced (Cy5) cells, using genome-wide Agilent 4x44K oligonucleotide microarrays. A signature of NF-kB signaling in ABC DLBCL was generated by treating HBL-1 cells with the IkB kinase beta inhibitor MLN120B for 2h, 3h, 4h, 6h, 8h, 12h, 16h, and 24h (Cy5), and comparing their gene expression to untreated cells (Cy3). A signature of JAK signaling in ABC DLBCL was generated by treating HBL-1 cells with JAK inhibitor I (5 micromolar; Calbiochem) for 2h, 4h, 6h, and 8h (Cy5) and comparing their gene expression to vehicle-treated cells (DMSO, Cy3). RNA-Seq data not provided.