Abstract LB123: Spatial immune architecture at single-cell resolution predicts response to atezolizumab plus bevacizumab in advanced HCC

Abstract Background: Atezolizumab+bevacizumab (atezo+bev) is a first-line therapy for advanced hepatocellular carcinoma (aHCC) with ∼30% objective responses. Single-cell studies identified responses linked to CD8⁺ T effector cells (CD8_Tex and Temra) and CXCL10⁺ macrophages (Macro_CXCL10), and resistance to TREM2⁺ macrophages (Macro_TREM2) and CD14⁺ monocytes (Mono_CD14). We hypothesized that, beyond cell composition, immune spatial organization within the tumor microenvironment (TME) determines response to atezo+bev. Methods: Single-cell spatial transcriptomics (ST) was performed on 27 aHCCs (9 atezo+bev responders and 18 resistant) using the Xenium Prime 5.000-gene panel (10x Genomics) customized with 100 additional genes. For local tissue organization, distances between cells were computed using the dbscan package [radius(r)=300µm]. Immune cell clusters were defined as groups of ≥3 immune cells within 100 µm, and were identified through a density-based algorithm. This spatial clustering was assessed using both ST and multiplexed immunohistochemistry (mIHC) data. Cell-to-cell interactions (neighborhoods) were analyzed (r=25µm). Ligand-receptor interactions were evaluated using a spatial neighborhood-based approach. Results: ST of 27 aHCCs yielded 4.6 million cells. In responders, CD8_Tex and Temra, and Macro_CXCL10 cells localized within the tumor and were closer to cancer cells (p=0.02, p=0.08, and p=0.1, respectively). Furthermore, in responders, tumor hepatocytes exhibited a significantly enriched lymphoid neighborhood composition and significantly higher rates of immune-recruting interactions, including PDL1-PD1, CXCL9-CXCR3 and CXCL10-CXCR3 (p<0.05). Non-responders were enriched in TREM2⁺ macrophage-containing clusters. Specifically, they showed Macro_TREM2-pure clusters (10% vs 0%, p=0.009), mixed Macro_TREM2-Mono_CD14 clusters (27% vs 5%, p=0.03), and mixed Macro_TREM2-CD8⁺ T-cell clusters (38% vs 22%, p=0.04). This was accompanied by enrichment of the CXCL12-CXCR4 chemokine axis, consistent with an immunosuppressive TME. Using mIHC, we confirmed enrichment of Macro_TREM2-pure (p=0.02) and Macro_TREM2-CD8⁺ mixed clusters (p=0.03). Conclusions: Atezo+bev response in HCC is marked by intratumoral effector CD8⁺ T cells and CXCL10_Macro, tumor hepatocytes enriched in lymphoid neighborhoods, and immune-recruiting ligand-receptor interactions, whereas resistance is driven by TREM2_Macro clusters and CXCL12-CXCR4 signaling. These findings identify spatial immune organization as a key determinant of response to atezo+bev. Citation Format: Anna Vila-Escoda, Marta Piqué-Gili, Marta Casado-Pelaez, Roser Pinyol, Ana Hernández de Sande, Verónica Davalos, Albert Gris-Oliver, Carla Montironi, Judit Peix, Daniela Grases, Ezequiel Mauro, Guillem Cano-Segarra, Sarah Cappuyns, Igor Figueiredo, Giorgio Ioannou, Edgar Gonzalez-Kozlova, Tim Meyer, Anja Lachenmayer, Jens U Marquardt, Helen L Reeves, Julien Edeline, Fabian Finkelmeier, Jörg Trojan, Sacha Gnjatic, Jean-Frederic Blanc, Richard A Hubner, Matthias Pinter, Tom Luedde, Arndt Vogel, Daniela Sia, Vincenzo Mazzaferro, Manel Esteller, Jeroen Dekervel, Eduard Porta-Pardo, Josep M Llovet. Spatial immune architecture at single-cell resolution predicts response to atezolizumab plus bevacizumab in advanced HCC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(8_Suppl):Abstract nr LB123.