Abstract 5317: Multimodal spatial analysis of colon cancer tissue reveals emergence of an immunosuppressive tumor maintenance mechanism

Abstract Background: Colorectal cancer is the second most fatal cancer worldwide, accounting for 9% of all reported cancer deaths, and is the number one cause of death among people under the age of 50. Rapid technical advances in the field of spatial biology are allowing us to gain new insight into colorectal cancer biology and patient heterogeneity, which can help to guide the development of immunotherapies. Using a combination of modalities such as sequential multiplex immunofluorescence (mIF) and spatial transcriptomics (STx) on a colon cancer sample we determined the spatial distribution, phenotype, function, and a cytokine and chemokine profile within the colon cancer tissue. Experimental Procedure: 5 um FFPE sections were prepared from a colon cancer sample exhibiting moderate to poorly differentiated adenocarcinoma of T3, N0 staging. mIF was performed using Lunaphore COMET with a 20-antibody panel, followed by H&E staining and imaging. An adjacent section was used for STx with the 10x Genomics Visium CytAssist platform. Data was analyzed with Lunaphore Horizon®, 10x Genomics Loupe Browser software and PredxBio SpaceIQ™ unbiased cell typing, microdomain discovery and network biology platform that co-analyzes mIF and STx datasets. Results: The tumor showed an aberrant growth pattern and a highly proliferated crypt region. Three distinct microdomains in the tumor tissue were discovered: one with high levels of PD-L1+ cells, a second with high levels of NK cells and a third with low levels of infiltrating cells in the lumen and crypt. There was significant presence of VISTA+ and IDO1+ cells throughout the lumen. These cells interacted with CD4+ and CD8+ cells which were found to be present in both lumen and crypt regions. PD-L1 was prominent in the cells found in both lumen and crypt. Finally, NK cells were abundant throughout the lumen and in regions adjacent to the crypt. Conclusions: Using multimodal spatial imaging and spatial co-analysis of mIF and STx data, we were able to elucidate relationships between different cell types in a complex tumor tissue and assign context to their activities. Despite high levels of NK cells, indicative of a possible active anti-tumor immune response, the tumor has developed an immune evasion mechanism due to the presence of cells expressing high levels of the anti-tumor immune response markers such as PD-L1, IDO1, and VISTA. Based on these results, therapies targeting more than one immune checkpoint protein may yield improved response due to an abundance of the existing NK cells. The transcriptomic profiling reveals a complex relationship between various cytokines secreted by different cell populations which alter the landscape of the tumor microenvironment, facilitating the emergence of an immunosuppressive phenotype. Citation Format: Gaurav Joshi, Nicholas Sciascia, Michael Yang, Brian Falkenstein, Raymond Yan, Shannon Quinn, Brenna Fearey, Junya Yoshioka, Arif Burak Tosun, S. Chakra Chennubhotla, Filippo Pullara, Fumiki Yanagawa. Multimodal spatial analysis of colon cancer tissue reveals emergence of an immunosuppressive tumor maintenance mechanism [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5317.