Abstract 3297: IFNalpha by in vivo-engineered macrophages abates liver metastases and triggers counter regulatory responses limiting efficacy
Authors: Kerzel Thomas , Beretta Stefano , Scamardella Eloise , Balestrieri Chiara , Canu Tamara , Pedica Federica , Norata Rossana , Sergi Lucia Sergi , Genua Marco , Renato Ostuni , Kajaste-Rudnitski Anna , Esposito Antonio , Oshima Masanobu , Tonon Giovanni , Sanvito Francesca , Squadrito Mario Leonardo , Naldini Luigi 
Published: June 2022

Abstract
The liver hosts an immune suppressive environment favouring metastatic seeding and proliferation of cancer cells. Pharmacological treatments, including immunotherapies, fail in the presence of liver metastases (LMS). Therefore, identifying new interventional tools and key targetable players involved in the immunosuppressive environment is of pivotal importance. We developed a lentiviral vector (LV)-based platform for selective genetic engineering of resident and tumour-associated macrophages enabling locally-sourced delivery of therapeutic molecules to LMS. Selective transgene expression is driven by a macrophage specific promoter and fine-tuned by microRNA target sequences. Upon systemic delivery of the LV, we observed enhanced transgene expression in macrophages located in areas surrounding LMS. We then equipped the LV with an IFNα-coding sequence, a cytokine with pleiotropic immune effects. Long term analysis in mice showed LV dose-dependent, sustained and well-tolerated IFNα expression. To investigate the therapeutic efficacy, we employed a colorectal cancer (CRC) organoid-based syngeneic mouse model of LMS containing molecular and histopathological hallmarks of the human disease. IFNα LV treatment significantly delayed LMS growth reaching a complete response in up to 50%. Single cell omics of LMS from IFNα LV-treated mice showed upregulation of IFNα-responsive genes, macrophage skewing to an antigen presenting (M1-like) polarization state, and expansion as well as reduced exhaustion of LMS-associated antigen specific CD8 T cells. Employing spatial transcriptomics, we found that the interface between LMS and liver parenchymal tissue was the major site of IFNα action, which was associated with enhanced immune activation and antigen presentation. Furthermore, we observed decreased angiogenesis and hypoxia in IFNα LV-treated LMS. When comparing LMS of treatment responsive to resistant mice, we found accumulation of activated CD8 T-cells in responsive whereas a high number of immunosuppressive T regulatory type 1 (TR1)-like cells in resistant mice. Molecular analyses suggest that TR1-like cell infiltration was associated with increased IL10 signaling in resistant mice. Furthermore, we found a positive correlation between IFNα and TR1-like cell signatures in human LMS and primary CRC thus supporting the link between IFNα activation and expansion of TR1-like cells in cancer. In summary, we developed an innovative gene-based platform that upon a single well-tolerated intravenous LV infusion rapidly promotes a protective therapeutic response against LMS through enabling immune activation. However, we also found that TR1-like cells might promote tumor immune evasion in presence of IFNα signaling in this setting, suggesting targeting of TR1-like cells when facing resistance to cancer immunotherapies that trigger IFNα signaling.
Citation Format: Thomas Kerzel, Stefano Beretta, Eloise Scamardella, Chiara Balestrieri, Tamara Canu, Federica Pedica, Rossana Norata, Lucia Sergi Sergi, Marco Genua, Ostuni Renato, Anna Kajaste-Rudnitski, Antonio Esposito, Masanobu Oshima, Giovanni Tonon, Francesca Sanvito, Mario Leonardo Squadrito, Luigi Naldini. IFNalpha by in vivo-engineered macrophages abates liver metastases and triggers counter regulatory responses limiting efficacy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3297.