Proteome alterations in peripheral immune cells of DLBCL patients and evidence of cancer extracellular vesicles involvement.

Diffuse large B-cell lymphoma (DLBCL) is an aggressive disease and a frequent form of non-Hodgkin lymphoma. Given the primary localization of DLBCL and the effect of tumors on the systemic immune response, we investigated the proteome of DLBCL patients' and healthy donors (HDs') peripheral immune cells (PICs). Since the ubiquitin-proteasome system has a vital role in proteome regulation and immune cells' functions, this study also explores the potential impact of DLBCL secretome on the polyubiquitination level in PICs. PICs from DLBCL patients and HDs were isolated and analyzed by mass spectrometry-based proteomics. The analysis resulted in 135 down and 51 upregulated proteins (adjusted p-value <0.05). Unsupervised principal component analysis revealed distinct proteomic profiles between DLBCL and HDs. Functional enrichment analysis for comparison between DLBCL and HDs-PICs proteome identified immune-related pathways such as innate immune system, specifically neutrophil degranulation, Fcγ receptor-dependent phagocytosis, and JAK-STAT signaling after IL-12 stimulation as downregulated. Proteomics analysis of DLBCL-PICs also showed dysregulation of proteostasis factors. This prompted the investigation of the effect of tumor secretome on viability and polyubiquitination level in mononuclear immune cells. Therefore, human HD peripheral blood mononuclear cells (PBMCs) were cultured in the presence of DLBCL cell line-derived soluble factors, small-EVs, and large-EVs in vitro. Our results revealed that exposure of mainly small-EVs, and large-EVs to HD PBMCs increased the polyubiquitination in PBMCs and decreased PIC viability. These findings suggest impaired immune responses in DLBCL-PICs, with tumor secretome-inducing polyubiquitination and reduced PIC viability.