Abstract 5253: Immune landscape analysis of solid tumors <i>via</i> spatially guided cell isolation for enhanced immunotherapeutic development

Abstract The tumor-immune microenvironment (TIME) in solid tumors is a complex ecosystem consisting of diverse interactions between cellular populations. Deeper investigation into the TIME reveals a prominent role of immune cells in regulating tumor progression- a novel phenomenon that has encouraged the evaluation of the crosstalk between cell types. The development of spatial omic technologies has thus enabled the exploration of these nuances at previously inaccessible resolutions, but the currently available platforms come at the cost of introducing bias, excluding molecular information, and incompatibility with single cell technologies. Here, we leverage a photoactivatable (PA) dye Janelia-Fluor-646 in combination with a digital light processing (DLP) microscope to address these issues by developing a platform that enables the spatially guided isolation of cell populations from a region of interest (ROI) for downstream multiomic and single cell analyses. As proof of concept, we show that ROIs identified via immunofluorescence could successfully be barcoded in mouse spleen and tumor sections using the dye and distinguished from the background by fluorescence activated cell sorting (FACS). We also demonstrate the ability to extract high quality DNA, RNA, and protein- as well as maintain high cell viability post-sorting. We have also developed a pipeline for the analysis of the sequenced RNA to identify differential patterns of gene expression present in immune cells from spatially distinct regions. Moving forward, we hope to establish this as a platform for immune landscape analysis within the TIME to uncover unique targets that could be explored within the context of tumor-immune interactions as well as immunotherapeutic development. Our findings will also help us generate the first single-cell atlas of niche-specific immune cells. Citation Format: Elie Abi Khalil, Yi-Chien Wu, Jingtian Zheng, Joshua Plank, Alexander Lippert, Steve Lee. Immune landscape analysis of solid tumors via spatially guided cell isolation for enhanced immunotherapeutic development [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 5253.