Abstract 2075: Combining Vizgen’s MERSCOPE spatial transcriptomics profiling and InSituPlex (ISP) multiplex immunofluorescence (mIF) to unravel the complexity of the tumor-immune microenvironment (TiME)

Abstract Despite remarkable results for many patients, cancer therapies have still a poor response rate. Biomarkers used to select patients for treatment are few and only single-biomarker assays are clinically approved. Such single-biomarker assays are unable to capture the complexity and dynamic nature of the TiME and fail to improve patients’ clinical response rate. Here we aim to enable a comprehensive understanding of the molecular, morphological and functional landscape of tumors using a multi-omics approach combining spatial transcriptomics and proteomics assays. Vizgen’s MERSCOPE® platform allows for high-resolution, spatially resolved analysis of up to 1000 genes, enabling the study of molecular and cellular signatures associated with the TiME while Vizgen’s spatial proteomics ISP assays allow for ultrasensitive and high-throughput detection of tumor and immune cell interactions, functions and spatial distribution. We used these approaches to generate datasets from multiple tumors and evaluated the complementary nature of analytical spatial detection methods to elucidate how local microenvironment influence gene expression and protein activity. Formalin-fixed paraffin-embedded (FFPE) specimens from multiple tumors were cut into 5µm sections for both technologies. For mIF experiments, slides were stained with a cocktail of primary antibodies using Leica Biosystems BOND RX autostainer, amplified simultaneously, and imaged on the Zeiss AxioScan.Z1 using multiple rounds of detection. Images were co-registered and exported for analysis using Ultivue’s STARVUETM image data science platform. Serial sections were profiled with Vizgen’s MERSCOPE Platform and the 500 Gene Immune-Oncology (IO) Panel with MERFISH 2.0 chemistry. The resulting spatial transcriptomic data was segmented using Cellpose2 and analyzed using both UMAP and spatial projections for visualization. Concordance of RNA and protein results were established by evaluating the expression of markers present in both the 500-gene IO panel and the ISP probes. We then evaluated immune responses measured by gene signatures for multiple functions and the corresponding protein markers. We similarly examined changes between the tumor boundary and within the tumor mass. We demonstrate the ability to orthogonally validate findings and expand on the biomarker-driven protein data with pathway-level data from the transcriptomic data. The integration of gene expression profiling enabling screening of a broad range of targets, and mIF assays for target validation, allows for a holistic understanding of spatial and molecular insights of the TiME. These complementary technologies can be used in parallel for biomarker discovery and IO pathways interrogation providing synergistic insights into the complex and dynamic TiME. Citation Format: Angela Vasaturo, Cassandra Kysilovsky, Lauren Duro, Sudhir Tattikota, Renchao Chen, Justin He, Manisha Ray, Jiang He He, George Emmanuel. Combining Vizgen’s MERSCOPE spatial transcriptomics profiling and InSituPlex (ISP) multiplex immunofluorescence (mIF) to unravel the complexity of the tumor-immune microenvironment (TiME) [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 2075.