Oncolytic bovine herpesvirus type 1 induces immune microenvironment remodeling and enhances treatment responses in multiple myeloma

Background: Despite recent therapeutic advances, multiple myeloma (MM) remains an incurable disease due to the high drug resistance and the severe immunosuppressive bone marrow (BM) microenvironment. Oncolytic virotherapy combines direct tumor cytolysis with immune activation, though human viruses’ efficacy is often limited by pre-existing antiviral immunity. Here, we evaluated the oncolytic and immunomodulatory effects of Bovine Herpesvirus type 1 (BoHV-1), a non-pathogenic virus to humans, as a possible therapeutic agent in MM. Methods: We assessed the expression of BoHV-1 attachment and entry receptors on MM cell lines and patient-derived BM mononuclear cells (BMMCs) by flow cytometry analysis. The effects of BoHV-1 infection on cell viability, apoptosis, transcriptional remodeling, and immune effector functions (activation, cytotoxicity, and cytokine release) were evaluated through cell-based assays, immunoblotting, RNA sequencing, and ELISA. Ex vivo infection assays were also conducted in combination with standard and immune-based anti-MM agents. Results: BoHV-1 efficiently infected MM cells, triggering mitochondrial apoptosis and extensive transcriptional rewiring. Key pro-survival programs, including MYC targets, oxidative phosphorylation, and the unfolded protein response, were suppressed. Infected tumor cells upregulated NK-activating ligands and downregulated MHC class I, enhancing susceptibility to NK-mediated cytotoxicity. In BMMCs, BoHV-1 selectively depleted malignant plasma cells and immunosuppressive myeloid subsets, while sparing lymphoid and hematopoietic progenitors. The virus induced activation of CD8⁺ T cells, NK cells, and monocytes, driving a shift toward a pro-inflammatory M1-like polarization. Monocyte depletion in BMMCs attenuated the BoHV-1 anti-MM effect, confirming their functional contribution. This pronounced immune remodeling was accompanied by an inflammatory cytokine storm, characterized by increased levels of IFN-α, IFN-γ, and TNF-α. Co-treatment of BoHV-1 with either bortezomib or lenalidomide increased anti-MM cytotoxicity compared to each agent alone. Similarly, sequential addition of elranatamab enhanced tumor cell clearance. Finally, BoHV-1 upregulated CD38 on both MM cells and immune effectors, thereby increasing sensitivity to the anti-CD38 daratumumab. Conclusions: BoHV-1 demonstrates a significant anti-MM oncolytic activity, remodels the BM immune landscape, and enhances the efficacy of both standard and emerging therapies. These findings establish BoHV-1 as an immunovirotherapy promising candidate, effective both as monotherapy and in combination strategies.