IDDF2023-ABS-0251 Epigenetic remodeling of endothelial cells underlying immune checkpoint resistance in hepatocellular carcinoma

BackgroundAlthough immune checkpoint blockade (ICB) has shown promise in cancers, only a modest proportion of hepatocellular carcinoma (HCC), patients showed desirable and durable responses. The success of combining ICB and anti-vascular endothelial growth factor as the first-line treatment for advanced-stage HCC highlights the great potential of endothelial cell (EC) normalization in enhancing immunotherapy response. Epigenetic regulation by bromodomain-containing protein 4 (BRD4) plays critical roles in cell differentiation implicated in cancer progression. Therefore, we aim to investigate the epigenetic remodeling of EC underlying immune checkpoint resistance in HCC and explore the therapeutic approach for enhancing HCC immunotherapy.MethodsSingle-cell RNA sequencing (scRNA-seq) was performed in Phase II clinical trial of pembrolizumab (anti-Programmed cell death 1(PD-1)) in hepatitis B virus-related HCC patients (NCT03419481). An ICB-resistant HCC mouse model generated via repeated selection was used. Immunofluorescence staining was conducted for EC assessment and BRD4 expression. Immune profiles were determined by high-dimensional flow cytometry.ResultsscRNA-seq analysis in anti-PD-1-treated patients revealed that the proportion of macrovascular-like endothelial cells (MaVEC) highly expressing disorganization signatures was significantly higher in non-responders when compared to responders. While the MaVEC proportion was increased in non-responders upon anti-PD-1 treatment, the proportion of liver-sinusoidal endothelial cells (LEC) with normal EC signatures was reduced. Moreover, RNA velocity analysis showed a strong directional flow from LEC to MaVEC in non-responders. BRD4 was significantly highly expressed in MaVEC compared to LEC in both patients and the ICB-resistant mouse model. Notably, a clinically-trialed BRD4 specific inhibitor AZD5153 synergized with PD-L1 antibody to suppress tumor growth and ameliorated immunosuppressive microenvironment, which was accompanied by significant reduction and increase in MaVEC and LEC densities, respectively.ConclusionsOur data demonstrated that epigenetic reversion of LEC-to-MaVEC trans-differentiation by BRD4 inhibition may be a potential strategy for improving HCC immunotherapy. Funded by RGC GRF (14120621), CUHK Strategic Seed Funding for Collaborative Research Scheme, AstraZeneca Preclinical Programme and Li Ka Shing Foundation.