PB2098 IDENTIFICATION OF NOVEL REGULATORS OF PLASMA CELLS MIGRATION IN MULTIPLE MYELOMA PATIENTS
Authors: Vdovin A. , Jelinek T. , Hrdinka M. , Sevcikova T. , Hajek R. , Simicek M. 
Published: June 2019

Background:

High number of peripheral blood circulating aberrant plasma cells in the patients with multiple myeloma (MM) correlates with worse survival prognosis and likely contributes to the development of extramedullary lesions. Molecular mechanisms responsible for egress of tumor cells from the supportive bone marrow niche into the blood stream are largely unknown. In the presented study we focus on deubiquitinating enzymes (DUBs), a group of hydrolytic proteases removing ubiquitin modifications from proteins. DUBs are involved in numerous cellular processes and as such provide very promising drug targets.

Aims:

Through analysis of expression datasets and progression free survival (PFS) we seek to identify DUBs deregulated specifically in MM, highlight those with relevance to plasma cells migration and finally uncover the responsible molecular mechanisms.

Methods

The GSE2658 dataset was used to analyse PFS of MM patients and other hematological malignancies (GSE12417, GSE10846, GSE16131, GSE22762, GSE2350, GSE4475) in relation to expression levels of all and selected human DUBs, respectively. Pathological bone marrow residing plasma cells from 14 newly diagnosed MM patients identified by the expression pattern of following antigens (CD19, CD56, CD45, CD38) were sorted and total mRNA was isolated for gene‐specific qPCR analysis. In parallel, percentages of aberrant plasma cells circulating in the peripheral blood were determined by flow cytometry. Co‐immunoprecipitation of OTUD1 and Filamin A was performed using HEK293 T cells and specific antibodies. For the gene set enrichment analysis (GSEA) the GSE4581 dataset was used.

Results:

In the pilot analysis we compared PFS of a large cohort of MM patients (n = 559) in relation to expression levels of all currently annotated human DUBs (n = 100). We have indentified 10 DUBs whose expression significantly affected PFS of MM patients (p < 0,05). Low expression levels of 6 DUBs (USP22, USP48, USP49, YOD1, OTUD6B, FAM105A) correlated with good PFS, while low expression of another 4 DUBs (USP3, USP30, USP35, OTUD1) correlated with significantly worse prognosis. Only 4 of the indentified DUBs (USP22, USP30, USP35 and OTUD1) were affecting PFS exclusively in MM patients and not in other hematological malignancies (AML, CLL, DLBCL, FCL, BL). We focused specifically on OTUD1 as it is the only DUBs from our list that was previously implicated to suppress migration and metastasis (Zhang, 2017). Quantitative PCR revealed that low expression of OTUD1 in bone marrow plasma cells (in 14 newly diagnosed MM patients) correlates with significantly higher number of plasma cells in peripheral blood. We have also identified actin crosslinking protein Filamin A as a novel OTUD1 binding partner. Additionally, GSEA of the GSE4581 dataset reveled that low expression of OTUD1 is associated with induction of genes related to actin polymerization and ER stress response.

Summary/Conclusion:

Database analysis, patient and cell lines derived data indicate that downregulation of OTUD1 is associated with higher number of peripheral blood circulating plasma cells and significantly worse PFS in a large subset of MM patients. Mechanistical investigation unveils Filamin A as direct OTUD1 binding partner and highlights involvement OTUD1 in actin cytoskeleton remodeling and ER stress response.