Karlijn Hummelink

Chapter 2 - Commentary by Anagnostou V and Luke JJ 283 A Figure 1. Impact and roles for spatially resolved high-plex assays in immunotherapy treatment and biomarker discovery. Spatially resolved high-plex methods, including quantitative immunofluorescence and digital spatial profiling, are high-throughput approaches that allow for simultaneous identification of multiple biomarkers in their spatial context. These methods have the unique potential to provide insights in the phenotype and spatial localization of immune cell subsets and thus serve as biology-informed biomarkers reflecting the quality and architecture of antitumor immune responses. As such, they can be incorporated in patient selection strategies for cancer immunotherapy and be used as a platform for novel biomarker discovery. (Adapted from an image created with BioRender. com.) Similarly, spatial phenotyping by reconstruction of cellular neighborhoods has pointed towards local enrichment in immune cell subpopulations and differential organization of the TME that is reflective of distinct antitumor immunity states19. Implementation of photo-cleavable oligonucleotide tags attached to antibodies or RNA probes has further increased the multiplexing capacity, dynamic range and level of detection of digital spatial profiling approaches. Spatial transcriptomics represent another avenue of interrogation of immune cell spatial heterogeneity, with neoantigen-reactive T-cell clones shown to harbor unique transcriptomic profiles that are further differentiated in the TME of ICI responsive tumors20. While spatially resolved and high-plex assays may uniquely assess the immune contexture of tumors at a single-cell resolution, further standardization is required to generate analytical platforms that allow for measurement of complex spatial associations. Notably, these approaches are more likely to succeed when representative of spatial and functional interactions, following the paradigm of the study by Hummelink and colleagues that relied on interrogation of a TIL subset previously functionally characterized and found to be tumor-reactive2.

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