A spatial profiling approach to evaluating the prognostic impact of heterogeneity in the triple-negative breast cancer immune microenvironment

Triple-negative breast cancer (TNBC), the breast cancer subtype defined by the absence of hormone receptors and no amplified HER2 receptors, is the most aggressive breast cancer subtype. Targeted treatment remains a challenge with TNBC. Non-targeted chemotherapy regimens have been the longstanding treatment modality, recently supplemented by immune checkpoint inhibitors (ICIs) targeting the PD-1/PDL1 pathway. TNBC's high tumor mutational burden, PD-1 expression, and immunogenicity make it a good candidate for immunotherapy. However, further studies are needed to determine the individuals that benefit most from this treatment. Factors like the spatial patterns of immune cell infiltration, immune-tumour cell and immune-immune cell relationships impact the immune microenvironment's anti-tumour response and ICI response. The high-plex spatial analysis of archival patient tissue samples-an abundant but underutilised resource for tumor characterization-has remained a challenge. However, using the GeoMx Digital Spatial Profiler (DSP), the spatial immune heterogeneity of 40 immune markers were quantified in a cohort of archival treatment-naïve TNBC tumours (n=140). The heterogeneity of 40 immune markers identifying immune cell lineages and sub lineages, activation and exhaustion states was quantified across separate stromal spatial regions of immune infiltration (n=486 from 140 patients). Tumours with overall low immune cell infiltration were significantly associated with higher spatial immune heterogeneity. Notably, low immune infiltration was also associated with a significantly higher spatial immune heterogeneity and total expression of multiple immunosuppressive markers, some of which are ICI targets This suggests that immunosuppression is spatially localized in stromal niches, which would impact immunotherapy response. High spatial immune heterogeneity was also associated with a trend of poorer patient survival.