Abstract B062: The role of oncogenic MYC signaling in shaping the canine metastatic osteosarcoma tumor immune microenvironment

Abstract Osteosarcoma (OS) is the most common primary malignant bone tumor in both humans and dogs. Canine and human OS share significant similarities in clinical presentation, histopathologic features, and molecular aberrations which have pioneered the dog as a translationally relevant model to accelerate treatment discovery in human OS. Despite aggressive treatment, up to 40% of children and >90% of dogs with OS die due to multi-drug-resistant metastatic disease, underscoring an urgent need to identify biomarkers which predict metastasis and/or response to therapy. Copy number gains in the cMYC oncogene have recently emerged as a potential biomarker of aggressive disease in human OS, and its genomic amplification has been recently described in primary canine OS tumors and cell lines. MYC is a nuclear transcription factor that drives a variety of tumor intrinsic hallmarks, but more recently has also been linked to regulation of the tumor immune microenvironment (TIME) in rodent models. We therefore hypothesize MYC overexpression in metastatic OS contributes to an immunosuppressive environment by driving expression of chemokines that promote tumor associated macrophage (TAM) enrichment and inhibit T cell recruitment. To characterize the role of oncogenic MYC signaling in the metastatic OS TIME, 42 archived formalin-fixed paraffin-embedded lung metastatic canine OS samples were evaluated for MYC copy number variation (CNV), mRNA, and protein expression via ddPCR, Nanostring analysis, and immunohistochemistry (IHC). 7 samples also underwent GeoMX spatial profiling to more specifically evaluate T cell and macrophage transcriptional profiles based on MYC status. To determine the role of MYC target modulation as a potential therapeutic option, canine and human OS cell lines were treated with a novel MYC inhibitor (MYCi975) and assessed for effects on survival, proliferation, and cytokine profiles. We demonstrate that CNV gains are not a key driver of MYC hyperactivity in canine metastatic OS. However, stratification based on MYC protein expression demonstrates that ‘MYC-high’ tumors are associated with downregulation of cytotoxic effector T-cell associated transcripts and upregulation of TAM and extracellular matrix remodeling transcripts. We also report that MYCi975 treatment of canine and human OS cell lines results in significant inhibition of OS cell survival and proliferation at concentrations which are pharmacologically achievable in mice. Furthermore, we demonstrate MYC inhibition by MYCi975 is associated with reduced pro-metastatic cytokine secretion in OS cell culture models. While MYC overactivity in metastatic canine OS may not be genomically driven, other mechanisms that lead to increased MYC protein expression are associated with transcriptomic profiles supportive of local immunosuppression. Furthermore, pharmacologic targeting of MYC may serve as a strategy to bolster immunotherapeutic options in canine and human metastatic OS. Citation Format: Rebecca Makii, Téa Ned, Eric Palmer, Kathryn Cronise, Mary Jane Underdown, Breelyn Wilky, Daniel Regan. The role of oncogenic MYC signaling in shaping the canine metastatic osteosarcoma tumor immune microenvironment [abstract]. In: Proceedings of the AACR IO Conference: Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2025 Feb 23-26; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2025;13(2 Suppl):Abstract nr B062.