OPTIMIZING CHEMO-IMMUNOTHERAPY AND MODULATING RNA SPLICING: EXPERIMENTAL STRATEGIES AGAINST HEPATOCELLULAR CARCINOMA

Hepatocellular carcinoma (HCC) is a malignant cancer and, despite recent advances in systemic therapies, the prognosis for patients with advanced stage HCC remains poor. Its multifaceted pathogenesis has prompted to investigate innovative therapeutic targets and to explore new molecular pathways involved in HCC onset and progression. Given that immune system dysfunction plays a pivotal role in tumor progression, immunotherapy has emerged as a promising and innovative strategy for its treatment1,2. Among the immune checkpoints, Programmed Death-Ligand 1 (PD-L1) is frequently overexpressed in various cancers, where its interaction with the Programmed cell Death Protein-1 (PD-1) receptor suppresses T-cell–mediated antitumor responses3. Immune checkpoint inhibitors have taken a central role in the management of advanced HCC, with the combination of atezolizumab (a monoclonal antibody binding PD-L1) and bevacizumab (anti-vascular endothelial growth factor) representing the first-line treatment in patients with unresectable HCC4,5. An important aspect in many tumors is the activation of epithelial-mesenchymal transition (EMT), a dynamic process that contributes to higher migration and invasion of cancer cells. Several studies underline the strong correlation between EMT and immune evasion due to the bidirectional regulation between EMT and PD-L16,7. Studies have shown that, despite the therapeutic improvement achieved with the combination of atezolizumab and bevacizumab compared to previous therapeutic approaches, immunotherapy alone is not effective in many HCC patients, often resulting in poor outcomes. This highlights the need to develop novel strategies that integrate immunotherapy with chemotherapeutic agents in order to increase the population of patients who can benefit and enhance overall response rates8,9. With this project, we investigated the anticancer effect of an immunoliposomal formulation (called SIL) obtained by conjugating liposomal doxorubicin with the Fab fragment of atezolizumab, to target PD-L1 expressed not only in cancer cells but also in other components of the tumor microenvironment (TME), like exhausted immune cells and M2 macrophages. We investigated the immunomodulatory effects of the SIL liposomal formulation and its impact on the TME using both 2D in vitro models with human HepG2 and Huh-7 cell lines, and more advanced 3D spheroid models obtained with HepG2 cells.In conclusion, these results indicate that the therapeutic efficacy of SIL may rely not only on its cytotoxic activity, due to the PD-L1 targeting, but also on its capacity to modulate the TME at different levels. Moreover, these findings highlight the dual chemo-immunotherapeutic activity of SIL, supporting its potential as a promising strategy to enhance immune responses and counteract tumor progression in HCC.