Spatial clustering of tumor-infiltrating lymphocytes in symptomatic melanoma brain metastases

Half of patients with advanced cutaneous melanoma develop brain metastases (BrMs), resulting in a dismal prognosis. The brain’s neuroprotective environment, exacerbated by dexamethasone treatment in patients with symptomatic BrMs, may affect tumor-infiltrating lymphocytes. A clearer understanding of how tumor-infiltrating lymphocytes differ between BrMs and extracerebral metastases (ECMs) is essential for tailoring immunotherapeutic strategies in this population. From 55 patients with melanoma, BrMs (n=60) and paired ECMs (n=53) were analyzed with multiplex immunohistochemistry. Intratumoral lymphocyte densities were computed and two-dimensional coordinates of lymphocytes were used to analyze spatial lymphocyte clustering using the local correlation function. On a subset of BrMs (n=13) and paired ECMs (n=21), bulk transcriptomics was performed. BrMs exhibited a sixfold reduction in lymphocyte density compared with paired ECMs and showed markedly reduced expression of immune signatures. Despite lower abundance, lymphocytes in BrMs displayed a higher degree of spatial clustering. Low spatial clustering, particularly of CD8 T cells, correlated with favorable post-neurosurgery survival and response to immune checkpoint inhibitors. Long (> 30 days) dexamethasone exposure of BrMs correlated with reduced immune signatures and increased tumor-promoting signatures relative to shorter exposure. Melanoma BrMs harbor a distinct lymphocyte architecture from ECMs, characterized by sparse but spatially clustered lymphocytes. Spatial clustering, rather than density, was predictive for clinical outcomes and immunotherapy responsiveness. Hence, the spatial pattern of infiltration may be functionally more important than infiltration numbers. Additionally, the negative effects of prolonged dexamethasone treatment on BrMs should be taken into account and elucidated in follow-up studies.