Abstract A039: Oncogenic Ras Dosage Alters Senescent Immune Response and Influences Tumour Initiation

Abstract In classical Oncogene Induced Senescence (OIS) model, cells secrete a milieu of signaling molecules referred to as the senescence-associated secretory phenotype (SASP). This SASP attracts immune effector cells to the proximity of senescent cells to undertake the elimination, thus impeding their potential progression into fully fledged malignancy1. Achieving such senescent immune clearance typically requires a high level of oncogenic stress, yet the phenotype provoked by lower oncogenic dosage remains unclear. Recently, we developed oncogenic RAS-dose escalation models in vitro and in vivo, revealing a RAS-dose-driven non-linear continuum of downstream phenotypes2. In a hepatocyte OIS model in vivo, ectopic expression of NRASG12V fails to induce tumours in part due to OIS-driven immune clearance. scRNA-seq analyses reveal distinct hepatocyte clusters with typical OIS or progenitor-like features, corresponding to high- and intermediate-NRASG12V levels, respectively. Remarkably, when titered down, NRASG12V-expressing hepatocytes become immune-resistant, and develop tumours. Time-series monitoring at single-cell resolution identifies two distinct tumour types: early-onset aggressive undifferentiated and late-onset differentiated hepatocellular carcinoma (HCC). The molecular signature of each murine tumour type is associated with different progenitor features and enriched in distinct human HCC subclasses. Our results demonstrate that the nature of oncogene dose heterogeneity has implications on the efficiency of senescent immune surveillance and Hepatocellular Carcinoma (HCC) initiation and progression. Particularly, Tumor-Initiating Cells (TICs) expressing low levels of Ras escape senescent immune surveillance, prompting our focus on the TICs microenvironment. Using spatial multiplex staining in conjunction with scRNA-seq, we found 1) Overall impaired effector T-cell response; 2) Expansion of Tregs and presence of PD-L1-expressing Myeloid Derived Suppressor Cells (MDSCs) were observed in close spatial proximity to Low-Ras TICs. Remarkably, treatment with an anti-PD-L1 immune checkpoint inhibitor significantly enhances immune clearance of Low-Ras TICs. Understand the intricate dynamics of immune response is crucial for devising therapeutic strategies to re-engage the immune system in eliminating TICs.1. Zhu H† & Blake S†, Kusuma F, Pearson RB, Kang J, Chan KT (2020) Oncogene-induced Senescence: From Biology to Therapy. Mechanisms of Ageing and Development. 2. Chan A† & Zhu H † , Narita M, Cassidy L, Bermejo-Rodriguez C, Janowska A, Oshimori N, Zender L, Aitken S, Hoare M, Narita M (2024). Titration of RAS alters senescent state and influences tumour initiation. Nature Citation Format: Haoran Zhu, Adelyne S.L. Chan, Masako Narita, Andrew R.J. Young, Aleksandra T. Janowska, Lars Zender, Sarah J. Aitken, Matthew Hoare, Masashi Narita. Oncogenic Ras Dosage Alters Senescent Immune Response and Influences Tumour Initiation [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2024 Oct 18-21; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2024;12(10 Suppl):Abstract nr A039.