Journal: Cancer Research

Article Title: Integrating AI-Powered Digital Pathology and Imaging Mass Cytometry Identifies Key Classifiers of Tumor Cells, Stroma, and Immune Cells in Non–Small Cell Lung Cancer

doi: 10.1158/0008-5472.CAN-23-1698

Figure Lengend Snippet: Multidimensional analysis of the NSCLC tumor ecosystem by IMC. A, Schematic representation of the IMC workflow on a formalin-fixed, paraffin-embedded tissue microarray. Key steps include staining with metal-tagged antibodies, spot-by-spot laser ablation, and acquisition by a mass cytometer. High dimensional images are reconstructed, processed, and segmented at both cellular and tissue level, generating data for further analyses. B, Heat map showing the mean values of key lineage markers adopted for cell cluster annotation. Proteins and cell phenotypes are ordered by hierarchical clustering with the Pearson correlation distance. Protein expression is color-coded from blue (lower) to red (higher) and scaled by column. C, Representative matched pictures of a NSCLC specimen showing pan-cytokeratin–positive tumor cells (left) and the tissue segmentation resulting from the machine learning pixel classifier (right). D, Spatial distribution and quantification of immune cell populations as the absolute cell number per mm 2 (left) or as a percentage of total immune cells (right) in the tumor and the stroma. E, Heat map showing the normalized marker expression in each macrophage cluster. Markers and cell clusters are ordered by hierarchical clustering according to Pearson correlation distance. Mean values of marker expression are represented and color-coded from blue (lower) to red (higher) and scaled by column. Color code indicates cluster identity. F and G, UMAP projections of macrophage cells ( n = 46733) from NSCLC tumors showing 20 clusters ( F ) or the cell distribution according to tissue segmentation ( G ). Each dot represents an individual cell. H, S100A8 + Mϕ infiltrate both the stroma and the tumor nests of NSCLC tissues. Representative pictures of the distribution of Mϕ (defined as CD68 + cells) and the subpopulation of S100A8 + Mϕ within tumor nests of a NSCLC tissue.

Article Snippet: Briefly, the Ilastik v.1.4.0 software was used to interactively train a supervised random forest pixel classifier on multiple image crops, by manually drawing labels for nuclei, membrane or cytoplasm, and background, on the basis of the IMC Cell Segmentation Kit (Standard Biotools) staining.

Techniques: Formalin-fixed Paraffin-Embedded, Microarray, Staining, Cytometry, Expressing, Marker