USP13 dictates Ran turnover and vulnerability to ferroptosis in diffuse large B cell lymphoma (DLBCL)

Abstract Diffuse large B-cell lymphoma (DLBCL) is one of the most common and lethal B-cell malignancies worldwide, with high relapse rate after standard treatment, and the relapsed cases are often difficult to treat. Ubiquitination has a pivotal role in cellular protein homeostasis and tumorigenic deubiquitinases (DUBs) stabilize oncoproteins during carcinogenesis. In this study, we have identified USP13 as one of the abnormally-overexpressed and stage-related DUBs in DLBCL critical to disease pathogenesis. Employing LC-MS/MS, Ran GTPase was characterized as potential substrate of USP13, with the interaction between Ran and USP13 modeled by Alphafold3. USP13 co-overexpressed, co-localized and correlated with Ran at protein level in DLBCL patient samples and cell lines. Pharmacological inhibition and genetic manipulation of USP13 modulated Ran protein stability and K11-linked ubiquitination level of Ran. By RNA-seq, pharmacological inhibition of USP13 via Spautin-1 treatment in DLBCL revealed downstream targets in NF-κB and Notch pathways. Suggested by the DRESIS Database, NF-κB and Notch signaling cascades are key to the resistance of Doxorubicin (Dox) and cyclophosphamide (CTX) within the standard R-CHOP regimen of DLBCL hinting benefits of combining Spautin-1 with Dox or CTX. Further in-vitro and in-vivo experiments confirmed Spautin-1 synergized with Dox or CTX to initiate ferroptosis in DLBCL with few toxicities to main organs by mediating ROS, GSH, MDA and ultimately LPO. Altogether, these results demonstrate USP13 deubiquitinates Ran in DLBCL, and Spautin-1 well synergizes with Dox or CTX to initiate ferroptosis. Combinational treatment with Spautin-1 and Dox or CTX may represent an effective approach and therapeutic hope to combat with DLBCL.