Classification of Brainstem Gliomas Based on Tumor Microenvironment Status

Abstract

The inter-tumor heterogeneity of the tumor microenvironment (TME) and how it correlates with clinical profiles and biological characteristics in brainstem gliomas (BSGs) remain unknown, dampening the development of novel therapeutics against BSGs. The TME status was determined with a list of pan-cancer conserved gene expression signatures using a single-sample gene set enrichment analysis (ssGSEA) and was subsequently clustered via consensus clustering. BSGs exhibited a high inter-tumor TME heterogeneity and were classified into four clusters: "immune-enriched, fibrotic", "immune-enriched, non-fibrotic", "fibrotic", and "depleted". The "fibrotic" cluster had a higher proportion of diffuse intrinsic pontine gliomas (p = 0.041), and "PA-like" tumors were more likely to be "immune-enriched, fibrotic" (p = 0.044). The four TME clusters exhibited distinct overall survival (p < 0.001) and independently impacted BSG outcomes. A four-gene panel as well as a radiomics approach were constructed to identify the TME clusters and achieved high accuracy for determining the classification. Together, BSGs exhibited high inter-tumor heterogeneity in the TME and were classified into four clusters with distinct clinical outcomes and tumor biological properties. The TME classification was accurately identified using a four-gene panel that can potentially be examined with the immunohistochemical method and a non-invasive radiomics method, facilitating its clinical application.

Keywords: brainstem glioma; classification; survival; tumor microenvironment.

Figure 1

BSGs are grouped into four clusters based on their TME status. (a) A heatmap for results of consensus clustering based on the TME-related GESs among 98 BSG cases; (b) comparison of activities between clusters in key components of the pan-cancer TME GESs; the activities were measured as scores of the according GESs. F: fibrotic; IEF: immune-enriched, fibrotic; D: depleted; IENF: immune-enriched, non-fibrotic.

Figure 1

BSGs are grouped into four clusters based on their TME status. (a) A heatmap for results of consensus clustering based on the TME-related GESs among 98 BSG cases; (b) comparison of activities between clusters in key components of the pan-cancer TME GESs; the activities were measured as scores of the according GESs. F: fibrotic; IEF: immune-enriched, fibrotic; D: depleted; IENF: immune-enriched, non-fibrotic.

Figure 2

The TME clusters are correlated with the clinical and molecular features of BSGs. Comparisons between clusters in patients’ ages (a); in proportions of DIPG (b), tumor locations (c), driver mutations (d), methylation clusters (e), and WHO grades (f). F: fibrotic; IEF: immune-enriched, fibrotic; D: depleted; IENF: immune-enriched, non-fibrotic; bold font: p < 0.05.

Figure 2

The TME clusters are correlated with the clinical and molecular features of BSGs. Comparisons between clusters in patients’ ages (a); in proportions of DIPG (b), tumor locations (c), driver mutations (d), methylation clusters (e), and WHO grades (f). F: fibrotic; IEF: immune-enriched, fibrotic; D: depleted; IENF: immune-enriched, non-fibrotic; bold font: p < 0.05.

Figure 3

The TME clusters predict outcomes in BSG patients. (a) The Kaplan–Meier analysis showing the survival differences among the TME clusters; (b) the multivariate Cox regression showing variables including DIPG diagnosis, the TME clusters, received radiotherapy, and WHO grades as independent risk factors for prognosis; (c) a nomogram based on WHO grades, DIPG diagnosis, received radiotherapy, and the TME clusters. F: fibrotic; IEF: immune-enriched, fibrotic; D: depleted; IENF: immune-enriched, non-fibrotic; * p < 0.05.

Figure 4

The TME clusters exhibit distinct molecular pathway activity and radiation sensitivity. (a) Comparisons of activities between clusters in the PROGENy pathways that are commonly implicated in tumors; (b) comparisons of radiosensitivity between clusters and the lower scores correlating with better outcomes following radiotherapy. F: fibrotic; IEF: immune-enriched, fibrotic; D: depleted; IENF: immune-enriched, non-fibrotic.

Figure 5

A four-gene panel determines the TME classification. (a) A heatmap showing distinct expression of the four genes among the TME clusters; (b) the immune-enriched or fibrotic scores based on the four-genes’ expressions showing good discrimination ability for the TME clusters; (c) ROC curves showing high accuracy of the immune-enriched or fibrotic scores for identifying the according clusters in six independent cohorts; (d) differences of four genes’ expressions between the TME clusters observed using multidimensional fluorescence staining; (e) representative multidimensional fluorescence staining images of four genes’ expressions: CD3E in red, Collagen III in green, MMP1 in tangerine, and CA9 in gold. * p < 0.05; ** p < 0.01; F: fibrotic; IEF: immune-enriched, fibrotic; D: depleted; IENF: immune-enriched, non-fibrotic.

Figure 6

Radiomics features related to the TME clusters. (a) The top 10 most important radiomics features selected with random forest algorithm for identifying the “fibrotic” clusters; (b) the top 10 most important radiomics features selected with random forest algorithm for identifying the “immune-enriched” clusters; (c) discrimination ability of generated radiomics models for identifying the “fibrotic” or “immune-enriched” clusters. * p < 0.05; ** p < 0.01; *** p < 0.001.