Eep profiling of CD45-enriched regions from the invasive margin and tumor center of MSS and MSI tumors have different immunosuppressive and activated immune phenotypes. Comparing colorectal tumors characterized as MSS, DSP was able to differentiate immune hot and cold tumors regardless of MSS status. Additional evaluation applying segment profiling of tumors versus stroma also identified certain immune proteins and RNA pathways that have been distinctly associated with every single compartment and were different involving MSI and MSS tumors. Conclusions Our outcomes suggests DSP has the prospective to become utilized to predict patients’ response to PD-1 immune checkpoint blockade with higher sensitivity than typical MSS/MSI profiling, and moreover DSP may perhaps allow identification of distinctive localized immune characteristics that would guide mixture therapeutic approaches. P429 Integrative spatially-resolved, high-plex digital profiling enables characterization of complex immune biology within the tumor microenvironment of mesothelioma Carmen Ballesteros Merino, PhD1, Moritz Widmaier, PhD2, Sarah Church3, Thomas Herz, PhD2, Alexei Budco, MSC2, Dasa Medrikova, PhD2, Ivan Kanchev, PhD2, Andrew White, BSc3, Douglas Hinerfeld, PhD3, Shawn Jensen, PhD1, John Handy, MD1, Rachel Sanborn, MD1, Carlo Bifulco, MD1, Sarah Warren, PhD3, Joseph Beechem, PhD3, Bernard A. Fox, PhD1 1 Providence Portland Cancer Center, Portland, OR, USA; 2Definiens, Munich, Germany; 3NanoString Technologies, Seattle, WA, USA Correspondence: Bernard A. Fox ([email protected]) CLEC-2 Proteins Biological Activity Journal for ImmunoTherapy of Cancer 2018, six(Suppl 1):P429 Background Malignant mesothelioma is an aggressive cancer with poor prognosis and few powerful therapies. Since mesothelioma is derived from the mesothelium of the lung, we hypothesize that immune cells within the tumor microenvironment (TME) may perhaps behave differently than other solid tumors. In our preceding studies, using multi- plexed immunofluorescence, we did not find immune phenotypes related with improved patient survival. Here we describe a novel combination of two technologies to spatially characterize the Complement Component 4 Binding Protein Beta Proteins Synonyms interface in between mesothelioma cells, stroma and immune cells within the TME within a highplex capacity. Techniques Ten FFPE mesothelioma tumors had been characterized by Definiens’ Immune-Oncology Profiling (IOP) and NanoString Digital Spatial Profiling (DSP). Three alternating sequential sections had been stained with Definiens’ IOP (CD8/PD-1/FOXP3, CD68/PD-L1/CD3, Granzyme B). Definiens analysis was combined to recognize localization of every marker within the tumor center, invasive margin or stroma. Twelve regions-ofinterest (ROIs) were then selected depending on the Definiens evaluation for high-plex evaluation on DSP on the interleaving slide: 4 CD68-enriched, 6 CD8- enriched and 2 CD3-low. For DSP analysis, every single slide was stained with a combination of fluorescent-labeled antibodies (pancytokeratin, CD3, CD68) as well as a panel of 38-antibodies each conjugated to a one of a kind UV- photocleavable DNA barcode. ROIs from Definiens’ defined analysis were overlayed on DSP fluorescent scans,followed by UV excitation of the defined ROIs, which releases the DNA barcodes for downstream quantitation on the NanoString nCounterplatform. Outcomes We found robust correlation among Definiens and NanoString analysis of T cell and macrophage markers in selected regions. Typically, patients with longer survival (6 months) had elevated density of immune infiltrates which includes higher density of T cells, T-cell activation markers (.