Identifying Immune-Specific Subtypes of Adrenocortical Carcinoma Based on Immunogenomic Profiling

Figures

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  Figure 1. Hierarchical clustering of ACC tumors based on the enrichment of 23 immune signatures. (A) Clustering analyses uncovering two immune subtypes of ACC: Immunity-H and Immunity-L, which have high and low immune cell enrichment scores, respectively, consistently in five datasets. Comparisons of the enrichment scores of immunostimulatory signatures (T cell co-stimulation, activated B cell, cytolytic activity, and natural killer cell) and immunosuppressive signatures (T cell co-inhibition, immune checkpoint molecules, regulatory T cells, and myeloid-derived suppressor cells). (B) Ratios of immunostimulatory to immunosuppressive signatures (CD8+/PD-L1, CD8+/CD4+ regulatory T cells) between the two immune subtypes. (C) Immune scores evaluated by ESTIMATE [7]. (D) One-tailed Mann–Whitney U test (B,D) and Student’s t test (C) p-values are shown. ** p < 0.01, *** p < 0.001. It also applies to the following figures. (E) Prediction of the three immune subtypes of ACC by Random Forest based on the enrichment scores of 23 immune cell types. The 10-fold cross-validation results in the training set and prediction results in the other datasets are shown. (F) PCA confirms that ACCs can be clearly…

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  Figure 2. Comparisons of clinical outcomes between two immune subtypes of ACC. (A) Comparisons of overall survival (OS) and disease-free survival (DFS) rates between the immune subtypes by Kaplan–Meier curves. The log-rank test p-values are shown. (B) Comparisons of OS and DFS between high immune score (>median) and low immune score (<median). Comparisons of neoplasm status, the response rate to chemotherapy, Weiss score, and excess adrenal hormones, between the two immune subtypes in TCGA-ACC (C), and comparisons of grade between two immune subtypes in GSE10927 (D). The overlap between immune subtypes and CoC or C1A/C1B subtypes (E). The Fisher’s exact test p-values are shown (C–E).

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  Figure 3. Comparisons of phenotypic and molecular features between the immune subtypes of ACC. Comparisons of the tumor stemness and proliferation (A,B), intratumor heterogeneity (ITH) (C), comparisons of CNA scores and TMB between the two ACC subtypes (D), and logistic regression models predicting high immune signature scores (>median) versus low immune signature scores (<median) of ACCs using CNA scores and TMB. CNA: copy number alteration. TMB: tumor mutation burden. Immune signature: activated CD8 T cell (E), comparisons of DNA damage repair (DDR) pathways (mismatch repair (MMR), base excision repair (BER), nucleotide excision repair (NER), Fanconi anemia (FA), homologous recombination (HR), non-homologous end joining (NHEJ), translesion DNA synthesis (TLS), and damage sensor (DS), p-values are shown. ** p < 0.01, *** p < 0.001 (F). The one-tailed Mann–Whitney U test p-values are shown in (A–D,F).

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  Figure 4. Different mutation rates between the immune subtypes of ACC. Eleven genes more frequently mutated in Immunity-H than in Immunity-L ACC (Fisher’s exact test, p < 0.05).

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  Figure 5. Differently expressed proteins between the immune subtypes of ACC. Heatmap showing the proteins with significant expression differences between Immunity-H and Immunity-L in TCGA-ACC (two-tailed Student’s t test, FDR < 0.05).

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  Figure 6. Pathways and gene ontology (GO) enriched in the immune subtypes of ACC. The KEGG pathways highly enriched in Immunity-H and Immunity-L in TCGA-ACC (A,B). (C) Spearman correlations between the enrichment scores of pathways upregulated in Immunity-H, Immunity-L, and immune scores in ACC. The correlation coefficients (ρ) and p-values are shown. * p < 0.5, ** p < 0.01, *** p < 0.001. (D) Eight gene modules that significantly differentiated ACC by subtype, survival time, or survival status identified by WGCNA. (E) Comparisons of stromal scores evaluated by ESTIMATE [7] between the immune subtypes of ACC (one-tailed Mann–Whitney U test, p < 0.01).

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  Figure 7. A summary of molecular and clinical characteristics of the immune subtypes of ACC. The figure was created with BioRender.com.