High-Plex and High-Throughput Digital Spatial Profiling of Non-Small-Cell Lung Cancer (NSCLC)

Simple Summary Characterizing the tumour microenvironment (TME) has become increasingly important to understand the cellular interactions that may be at play for effective therapies. In this study, we used a novel spatial profiling tool, the Nanostring GeoMX Digital Spatial Profiler (DSP) technology, to profile non-small-cell lung cancer (NSCLC) for protein markers across immune cell typing, immune activation, drug targets, and tumour modules. Comparative analysis was performed between the tumour, adjacent tissue, and microenvironment to identify markers enriched in these areas with spatial resolution. Our study reveals that this methodology can be a powerful tool for determining the expression of a large number of protein markers from a single tissue slide. Profiling the tumour microenvironment (TME) has been informative in understanding the underlying tumour-immune interactions. Multiplex immunohistochemistry (mIHC) coupled with molecular barcoding technologies have revealed greater insights into the TME. In this study, we utilised the Nanostring GeoMX Digital Spatial Profiler (DSP) platform to profile a non-small-cell lung cancer (NSCLC) tissue microarray for protein markers across immune cell profiling, immuno-oncology (IO) drug targets, immune activation status, immune cell typing, and pan-tumour protein modules. Regions of interest (ROIs) were selected that described tumour, TME, and normal adjacent tissue (NAT) compartments. Our data revealed that paired analysis (n = 18) of matched patient compartments indicate that the TME was significantly enriched in CD27, CD3, CD4, CD44, CD45, CD45RO, CD68, CD163, and VISTA relative to the tumour. Unmatched analysis indicated that the NAT (n = 19) was significantly enriched in CD34, fibronectin, IDO1, LAG3, ARG1, and PTEN when compared to the TME (n = 32). Univariate Cox proportional hazards indicated that the presence of cells expressing CD3 (hazard ratio (HR): 0.5, p = 0.018), CD34 (HR: 0.53, p = 0.004), and ICOS (HR: 0.6, p = 0.047) in tumour compartments were significantly associated with improved overall survival (OS). We implemented both high-plex and high-throughput methodologies to the discovery of protein biomarkers and molecular phenotypes within biopsy samples, and demonstrate the power of such tools for a new generation of pathology research.