Single-Cell Characterization of the Immune Microenvironment of Melanoma Brain and Leptomeningeal Metastases

Abstract

Purpose: Melanoma brain metastases (MBM) and leptomeningeal melanoma metastases (LMM) are two different manifestations of melanoma CNS metastasis. Here, we used single-cell RNA sequencing (scRNA-seq) to define the immune landscape of MBM, LMM, and melanoma skin metastases. Experimental Design: scRNA-seq was undertaken on 43 patient specimens, including 8 skin metastases, 14 MBM, and 19 serial LMM specimens. Detailed cell type curation was performed, the immune landscapes were mapped, and key results were validated by IHC and flow cytometry. Association analyses were undertaken to identify immune cell subsets correlated with overall survival. Results: The LMM microenvironment was characterized by an immune-suppressed T-cell landscape distinct from that of brain and skin metastases. An LMM patient with long-term survival demonstrated an immune repertoire distinct from that of poor survivors and more similar to normal cerebrospinal fluid (CSF). Upon response to PD-1 therapy, this extreme responder showed increased levels of T cells and dendritic cells in their CSF, whereas poor survivors showed little improvement in their T-cell responses. In MBM patients, therapy led to increased immune infiltrate, with similar T-cell transcriptional diversity noted between skin metastases and MBM. A correlation analysis across the entire immune landscape identified the presence of a rare population of dendritic cells (DC3) that was associated with increased overall survival and positively regulated the immune environment through modulation of activated T cells and MHC expression. Conclusions: Our study provides the first atlas of two distinct sites of melanoma CNS metastases and defines the immune cell landscape that underlies the biology of this devastating disease.

Spatially resolved transcriptomics reveals the architecture of the tumor-microenvironment interface

“…Finally, we investigated whether “interface”-like cells are also found in human melanoma. We chose to take advantage of a recently published scRNA-seq dataset of 29,247 cells isolated from 43 human patients with metastatic melanoma 39 , as this dataset contains significantly more cells than other commonly used human metastatic melanoma scRNA-seq datasets 40 . This new dataset contains mostly tumor, myeloid, and immune cells 39 (Supplementary Fig.…”

“…The counts matrix was obtained from GEO (GSE174401). All analysis was done using R version 3.6.3 and Seurat version 3.1.4 21 , based on the analyses done in the original publication 39 . A Seurat object was created with default parameters, keeping all genes expressed in three or more cells and all cells expressing 200 or more genes, and filtering out any cells with more than 20% expression of mitochondrial genes, resulting in a final object containing 29,247 cells.…”

High-Dimensional Single-Cell Transcriptomics in Melanoma and Cancer Immunotherapy

“…In recent years, a wide array of single-cell transcriptomic technologies has emerged with a range of biological and clinical applications in melanoma and cancer immunotherapy, demonstrating the ability to characterise rare cellular phenotypes and specific cellular responses in an unbiased manner with precision [46,[48][49][50]. Several recent studies have highlighted the impact of single-cell transcriptomics for identifying new potential molecular targets and the effect of checkpoint inhibitors on tumour cells and the TME [3,[51][52][53]. These studies have fully characterised the cellular composition and function of the TME, T cell and immunosuppressive cell states, transcriptional programs and checkpoints associated with disease progression and response to treatment [3,17,[51][52][53][54].…”

“…Several recent studies have highlighted the impact of single-cell transcriptomics for identifying new potential molecular targets and the effect of checkpoint inhibitors on tumour cells and the TME [3,[51][52][53]. These studies have fully characterised the cellular composition and function of the TME, T cell and immunosuppressive cell states, transcriptional programs and checkpoints associated with disease progression and response to treatment [3,17,[51][52][53][54]. The single-cell transcriptomic sequencing technologies are broadly classified by their respective cell isolation methods (Table 1); (i) droplet encapsulation, (ii) microwell encapsulation, and (iii) fluorescenceactivated cell sorting (FACS).…”

Brain Microenvironment Heterogeneity: Potential Value for Brain Tumors

“…Notably, tumor location influences microenvironmental landscapes (63,64). Meningiomas have higher TAM infiltration and less presence of T regs than gliomas (64), while in secondary brain tumors, scRNAseq revealed a distinct immune-suppressed T-cell microenvironment in leptomeningeal metastasis compared with brain metastasis derived from melanoma (63).…”