Exploration of the Tumor-Suppressive Immune Microenvironment by Integrated Analysis in EGFR-Mutant Lung Adenocarcinoma

Abstract

Background: Clinical evidence has shown that few non-small cell lung cancer (NSCLC) patients with epidermal growth factor receptor (EGFR) mutations can benefit from immunotherapy. The tumor immune microenvironment (TIME) is a significant factor affecting the efficacy of immunotherapy. However, the TIME transformational process in EGFR-mutation patients is unknown.

Methods: The mRNA expression and mutation data and lung adenocarcinoma (LUAD) clinical data were obtained from The Cancer Genome Atlas (TCGA) database. Profiles describing the immune landscape of patients with EGFR mutations were characterized by differences in tumor mutation burden (TMB), ESTIMATE, CIBERSORT, and microenvironment cell populations-counter (MCP-counter).

Results: In total, the TCGA data for 585 patients were analyzed. Among these patients, 98 had EGFR mutations. The TMB was lower in the EGFR group (3.94 mut/Mb) than in the KRAS mutation group (6.09 mut/Mb, P < 0.001) and the entire LUAD (6.58 mut/Mb, P < 0.001). The EGFR group had a lower population of activated immune cells and an even higher score of immunosuppressive cells. A further inter-group comparison showed that differences in the TMB and tumor-infiltrating lymphocytes were only found between patients with oncogenic mutations and unknown mutation. Meanwhile, there were more myeloid dendritic cells (DCs) in EGFR 19del than in L858R-mutation patients and in common mutation patents than in uncommon mutation patients (P < 0.05). Additionally, we established a D score, where D = MCP-counter score for cytotoxic T lymphocytes (CTLs)/MCP-counter score for myeloid DCs. Further analysis revealed that lower D scores indicated immune suppression and were negatively related to several immunotherapy biomarkers.

Conclusions: The TIME of EGFR mutant NSCLC was immunosuppressive. Myeloid DCs gradually increased in EGFR 19del, L858R, and uncommon mutations. The potential role of CTLs and DCs in the TIME of patients requires further investigation.

Keywords: Bioinformatics & Computational Biology; cytotoxic T lymphocyte (CTL); epidermal growth factor receptor (EGFR) mutation; immune microenvironment; lung adenocarcinoma; myeloid dendritic cells (mDCs).

Figure 1

Flowchart of the study design.

Figure 2

Comparison of the immune landscape of EGFR-mutated group, KRAS-mutated group and the whole lung adenocarcinoma. The TIME of EGFR-mutated NSCLC show an immunosuppressive status with lower (A) TMB, a lower score of activated immune cell and an even higher score of immunosuppressive cell, estimated by (C) CIBERSORT and (D) MCP-counter. No difference was found in tumor purity among three group (B). *P < 0.05, **P < 0.01, and ***P < 0.0001, respectively. ns, not significant.

Figure 3

Comparison of the immune landscape among 19del, L858R, uncommon mutation and unknown mutation group. (A) TMB, (B) ESTIMATE, (C) CIBERSORT, (D) MCP-counter. Only significant difference in myeloid DC cells was presented among different groups estimated by (C) CIBERSORT and (D) MCP-counter. *P < 0.05, **P < 0.01, ***P < 0.001 and ****P < 0.0001, respectively. ns, not significant.

Figure 4

Analysis of differentially expressed genes (DEGs) from the D value of EGFR-mut group quartile 1 compared with quartile 3. (A) Volcano plot showing DEGs between higher D score group and lower D score group. (B) Top 10 enrichments of up-regulated DEGs by KEGG analyses. (C) Gene Ontology analyses of DEGs according to their biological process.

Figure 5

The D value of EGFR group was lower than rare mutation group except MET mutation and there was no significant difference among BRAF, ERBB2, KRAS and MET mutation. *P < 0.05 and ****P < 0.0001, respectively. ns, not significant.

Figure 6

Explore the role of D value in LUAD. Tumor tissue showed a lower D value than normal tissue (A). No difference in tumor purity was observed between high-D and low-D group (B). Less abundant of activated immune cell were observed in low-D group by CIBERSORT (C) as well as the result MCP-counter (D). *P < 0.05, **P < 0.01, ***P < 0.001 and ****P < 0.0001, respectively. ns, not significant.

Figure 7

Relationship between the D score and the expression of immune checkpoint. (A–F) the distribution of mRNA level of PDCD1, CD274, PDCD1LG2, LAG3, TIGIT and IDO1 in high- and low-D score groups.

Figure 8

A schematic summarizing the major findings of this study. The TIME of EGFR-mutated NSCLC show an immunosuppressive status with lower TMB, fewer cytotoxic T cells (CTLs) and more myeloid DC (M-DC) cells compared with KRAS-mutated NSCLC. The myeloid DCs increased in EGFR 19del compared with L858R mutation patients, and in common mutation patents compared with uncommon mutations.