Epidermal Growth Factor Receptor Mutations in Resectable Non-Small Cell Lung Cancer Patients and their Potential Role in the Immune Landscape

Abstract

Background

The epidermal growth factor receptor (EGFR) is a therapeutic target for non-small cell lung cancer (NSCLC), but knowledge on gene mutations that contribute to NSCLC development and persistence is lacking. In this study, we investigated genetic variations in EGFR and their association with the clinical and pathological factors of NSCLC.

Material/Methods

Clinical cases (331 patients) and The Cancer Genome Atlas (TCGA) cases (1040 patients) were selected and analyzed using the refractory mutation systems cBioPortal and the Tumor Immune Estimation Resource (TIMER).

Results

EGFR mutation frequencies were 54.4% (180 of 331 patients) and 8.0% (83 of 1040 patients) in the clinical and TCGA cohorts, respectively. EGFR mutations were strongly associated with smoking and pathology (P≤0.05) in the clinical cohort, and with gender, smoking, and pathology (P=0.001, P<0.001, and P<0.001, respectively) in TCGA cohort. In cases of lung squamous carcinoma (LUSC), EGFR was overexpressed as a result of DNA amplification, but this amplified expression showed no association with the overall survival (OS) or progression-free survival of LUSC patients. EGFR gene alterations were, however, associated with worse OS in lung adenocarcinoma (LUAD) patients. Immune cell infiltrates from LUAD and LUSC tumors differed according to EGFR expression. EGFR mutations resulted in a decline of immune infiltration or a lack of infiltrating immune cells in the NSCLC microenvironment.

Conclusions

Mutational profiles of the EGFR in NSCLC patients provide useful information for the use of tyrosine kinase inhibitors for adjuvant or neoadjuvant therapy and immunotherapy.

Background

As one of the most lethal tumors globally, lung cancer ranks first and second in terms of mortality among males and females in China, respectively [1], with 5-year survival rates as low as 4% [2]. A range of driver gene mutations have been implicated in lung cancer development including epidermal growth factor receptor (EGFR) and ROS1 mutations [3–5]. Precision therapy using tyrosine kinase inhibitors (TKIs) can improve prognosis in those patients harboring specific genetic alterations [6], producing response rates of up to 80% in non-small cell lung cancer (NSCLC) patients with TKI-sensitive EGFR mutations [7,8].

The majority of studies have focused on TKI administration during early disease stages, and impressive responses and improved patient outcomes have been documented [9,10]. In addition, EGFR TKI neoadjuvant therapy in resectable NSCLC can diminish the surgical rate [9]. When used as adjuvant therapy for stage II–IIIA (N1–N2) NSCLC patients after complete resection (R0), EGFR TKI therapy can yield longer disease-free survival when compared with traditional platinum-based chemotherapy [10]. Thus, the clinical benefits of EGFR TKIs in resectable NSCLC are promising.

The mutation profile of the EGFR has been investigated widely in metastatic NSCLC. However, EGFR mutation profiles in resectable NSCLC are rare. Understanding the EGFR mutation profile and its correlation with clinicopathological factors will help guide EGFR TKI therapy in resectable NSCLC precisely in the future.

The recent discovery of immune checkpoints, including programmed cell death 1 ligand (PDL1) and its receptor (PD1), represents a breakthrough in lung cancer immunotherapy [11]. Inhibiting the PD1/PDL1 interaction is efficacious in NSCLC immunotherapy owing to immune cell effector reactivity on NSCLC [12]. Despite the promise of immunotherapy during cancer treatment [13,14], there is limited information on the immune signature of EGFR in NSCLC [15]. Furthermore, the relationship between EGFR-mediated signaling and the immune checkpoint molecules, PD1/PDL1, has not been studied in detail [16].

Herein, we investigated the altered EGFR profiles in resectable NSCLC patients and their potential role in shaping the tumor immune microenvironment to unveil the potential clinical importance of the EGFR immune signature and its association with PD1/PDL1.

Material and Methods

Clinical and The Cancer Genome Atlas (TCGA) cohort analyses of EGFR mutations

A single-center retrospective analysis was performed to assess the genetic spectrum of the EGFR in patients with resectable NSCLC from July 2016 to November 2018 in our hospital. Enrollment criteria in our clinical cohort were: 1) age over 18 years; 2) pathological NSCLC at stage 0–IIIA; and 3) EGFR genetic testing performed. The Wuxi People’s Hospital affiliated to Nanjing Medical University approved the study (no. HS2019013) and consent was obtained from all patients.

For The Cancer Genome Atlas (TCGA) cohort, the enrollment criteria were: 1) pathologic diagnosis confirmed as NSCLC at stage I–IIIA; and 2) exon 18–21 mutations in EGFR. A total of 21 hotspot mutations in EGFR in exons 18–21 in the clinical cohort were subject to mutation-based amplification (CFDA #. 3401228, AmoyDx, Xiamen, China). Based on the pre-design protocol, EGFR mutation information and corresponding patient demographic data (i.e., gender, age, stage, differentiation grade, pathological type, and smoking status) were recorded. Whole exome sequencing data (21 mutation sites in EGFR exons 18–21) and clinicopathological information were obtained from TCGA cohort (www.cbioportal.org) [17].

UCSC Xena functional genomics explorer analysis

In patients with lung squamous carcinoma (LUSC) or lung adenocarcinoma (LUAD), heat maps of EGFR copy number segments, mRNA expression, exon expression, and pan-cancer gene expression were examined by data mining in TCGA database using the UCSC Xena browser (http://xena.ucsc.edu/) [18]. Subsequently, Kaplan-Meier curves were plotted for overall survival (OS) and recurrence-free survival (RFS) in resectable NSCLC patients (stages I–IIIA). EGFR pan-cancer expression in each pathologic stage of the same patient cohort was verified.

cBioPortal for cancer genomics analyses

EGFR genetic and copy number alterations in TCGA-LUAD and TCGA-LUSC patients were examined using cBioPortal for Cancer Genomics [17]. Kaplan-Meier survival curves were used to assess the association between EGFR gene alterations and OS and progression-free survival (PFS) in LUAD and LUSC patients. The relationships between EGFR and PDCD1 (PD1) and CD274 (PDL1) in the same NSCLC cohort were also analyzed using cBioPortal.

Tumor Immune Estimation Resource (TIMER) analysis

Tumor Immune Estimation Resource (TIMER) quantifies the abundance of B cells, CD4 T cells, CD8 T cells, macrophages, neutrophils, and dendritic cells that infiltrate the tumor microenvironment (https://cistrome.shinyapps.io/timer/) [19]. EGFR expression in TCGA tumors versus healthy tissue was compared using the Exp module. The Gene module was used to correlate EGFR expression with immune cell infiltration. Changes in the rates of survival according to the levels of EGFR and immune cell infiltrates were obtained from Kaplan-Meier curves. TIMER was used to generate multivariate Cox proportional hazard models to identify the factors regulating survival. Mutation models were used to quantify the levels of immune cell infiltration according to somatic copy number alterations in EGFR according to the SCNA module.

Statistical analyses

All data were compared using SPSS 25.0 software. EGFR mutations and clinical outcomes were compared using Fisher’s or χ² tests. A multivariate logistic model was performed for specific factors. EGFR mRNA expression was compared between LUSC and LUAD patients using Welch’s t-test. Kaplan-Meier curves were used for survival analysis and compared between groups via the log-rank test. Spearman’s rank correlation, Pearson’s correlation, or a partial correlation were used for estimating correlations. Multivariate Cox proportional hazard models were used to assess and analyze patient prognosis. P-values <0.05 were considered statistically significant (confidence intervals [CI]: 95%).

Results

Clinical and TCGA cohort analyses

Within the NSCLC patient cohort, 331 resectable cases were deemed eligible. The presence of EGFR mutations was assessed by ARMS. Patient demographics are summarized in Table 1. Male patients accounted for 41.69% (138 of 331 cases), whereas 38.37% (127 of 331 cases) were 65 years of age or older, and 18.73% (62 of 331 cases) were smokers. LUAD and LUSC accounted for 95.77% (317 of 331 cases) and 3.63% (12 of 331 cases) of the lung cancers, respectively. The percentages of patients with low, medium, and high tumor differentiation were 22.05% (73 of 331 cases), 50.15% (166 of 331 cases), and 8.16% (27 of 331 cases), respectively. The percentages of patients in stages 0–I, II, and IIIA were 78.55% (260 of 331 cases), 9.06% (30 of 331 cases), and 12.39% (41 of 331 cases), respectively. Gender, smoking, pathology, and differentiation were significantly associated with EGFR mutation frequency in the univariate analysis (Table 1, P=0.013, P<0.001, P<0.002, P<0.001, respectively). The multivariate logistic regression showed that smoking (P=0.009) and pathology (P=0.017) were 2 independent factors associated with the frequency of EGFR mutations.

Table 1.

Subgroup analysis for EGFR mutations in the clinical cohort.

Subgroup		Positive	Negative	P-value*	P-value**	HR (95% CI)**
Chinese cohort
Total	331	180	151
Age				0.505
<64	204	108	96
≥65	127	72	55
Gender				0.013	0.717	0.882 (0.448–1.736)
Male	138	64	74
Female	193	116	77
Smoking				<0.001	0.009	0.346 (0.156–0.771)
No	259	152	107
Yes	62	18	44
Pathology				0.002	0.017	12.819 (1.567–104.892)
Adenocarcinoma	317	179	138
Squamous carcinoma	12	1	11
Differentiation				0.001
Low differentiation	73	32	41
Median differentiation	166	111	55		0.092	0.413 (0.148–1.154)
High differentiation	27	20	7		0.509	0.728 (0.284–1.866)
Stage				0.305
0–I	260	140	120
II	30	20	10
IIIA	41	20	21

^*Univariate analysis;

^**multivariate analysis.

EGFR – epidermal growth factor receptor; HR – hazard ratio; CI – confidence interval.

From TCGA, 1040 resectable NSCLC patients were deemed eligible. Baseline demographic data are summarized in Table 2. Female patients accounted for 41.73% (434 of 1040 cases) and 87.40% (909 of 1040 cases) were smokers. LUAD and LUSC accounted for 56.35% (586 of 1040 cases) and 43.65% (454 of 1040 cases) of the lung cancers, respectively. The percentages of patients in stages I, II, and IIIA were 55.29% (575 of 1040 cases), 29.52% (307 of 1040 cases), and 15.19% (158 of 1040 cases), respectively. The EGFR mutation frequency was 7.98% (83 of 1040 cases) in TCGA cohort. In the univariate analysis, gender, smoking, and pathology showed significant associations with the frequency of EGFR mutations (Table 2, P<0.001 for all). The multivariate logistic regression showed that gender, smoking, and pathology were 3 independent factors associated with EGFR mutation frequency (Table 2, P=0.001, P<0.001, and P<0.001, respectively).

Table 2.

Subgroup analysis for EGFR mutations in TCGA cohort.

Subgroup		Positive	Negative	P-value*	P-value**	HR (95% CI)**
TCGA cohort
Total		83	957
Gender				<0.001	0.001	0.386 (0.221–0.674)
Male		20	586
Female	434	63	371
Smoking				<0.001	<0.001	0.160 (0.094–0.272)
No		36	66
Yes	909	45	864
Pathology				<0.001	<0.001	8.153 (3.200–20.771)
Adenocarcinoma	586	78	508
Squamous carcinoma	454	5	449
Stage				0.587
I	575	47	528
II	307	21	286
IIIA	158	15	143

^*Univariate analysis;

^**multivariate analysis.

EGFR – epidermal growth factor receptor; TCGA – The Cancer Genome Atlas; HR – hazard ratio; CI – confidence interval.

EGFR amplification occurs in LUSC but not LUAD

EGFR expression was compared between healthy and tumor tissue in LUAD and LUSC using the DiffExp module of TIMER. In LUSC, the expression of EGFR was ~1.8-fold higher than healthy tissue and ~0.869-fold higher than LUAD (Figure 1). We extracted data for the EGFR copy number segments, gene expression, exon expression, and pan-cancer gene expression in TCGA-LUSC and TCGA-LUAD. Significantly higher EGFR expression was evident in LUSC versus LUAD (Figure 2A, 2B).

Figure 1.

Epidermal growth factor receptor expression in tumors versus adjacent normal tissue. * P<0.05, ** P<0.01, *** P<0.001.

Figure 2.

Epidermal growth factor receptor (EGFR) expression and copy number alterations in lung squamous carcinoma (LUSC) and lung adenocarcinoma (LUAD). (A) Heat maps of EGFR expression, pan-cancer expression, exon expression, and copy number segments in different tumor stages for both LUAD and LUSC. (B) Box plots of EGFR expression in LUAD and LUSC. (C) Genetic alterations of EGFR in 507 LUAD and 496 LUSC patients.

Figure 2C shows that EGFR mutations occurred in ~16% and ~9% of LUAD and LUSC cases, respectively. EGFR amplification was predominant in both cancer types as revealed from the EGFR copy numbers (Figure 3A, 3B).

Figure 3.

Box plots of epidermal growth factor receptor expression in (A) lung adenocarcinoma (LUAD) and (B) lung squamous cell carcinoma (LUSC) based on the genetic status for LUAD and LUSC according to The Cancer Genome Atlas.

Elevated EGFR levels do not correlate with poor OS or RFS in resectable LUAD or LUSC patients

Figure 4A shows that EGFR mRNA expression was unaffected by the pathological stage in resectable NSCLC (stages I–IIIA). Further analyses showed the same trends in both resectable LUAD and LUSC (Figure 4D, 4G). EGFR expression was not related to OS or RFS (Figure 4B, 4C). Subgroup analyses of LUAD (Figure 4E, 4F) and LUSC (Figure 4H, 4I) revealed no evidence that EGFR influenced OS or RFS.

Figure 4.

Epidermal growth factor receptor (EGFR) expression and survival in stage I–IIIA resectable non-small cell lung cancer (NSCLC). EGFR expression in the different pathological stages of (A) NSCLC, (D) lung adenocarcinoma (LUAD), and (G) lung squamous cell carcinoma (LUSC). EGFR expression and its association to overall survival (OS) and recurrence-free survival (RFS) in total NSCLC (B, C), LUAD (E, F), and LUSC (H, I) patients.

EGFR gene alterations are associated with worse OS in LUAD

LUAD patients with EGFR gene alterations had significantly worse OS (Figure 5A; P<0.01). However, there was no association of EGFR alterations with PFS in LUAD patients (Figure 5B). By contrast, EGFR gene alterations did not prominently affect OS or PFS in LUSC patients (Figure 5C, 5D).

Figure 5.

Association of epidermal growth factor receptor (EGFR) mutations and overall survival (OS) and progression-free survival (PFS) in lung adenocarcinoma (LUAD) (A, B) and lung squamous cell carcinoma (LUSC) (C, D) patients.

EGFR expression and its relationship to immune signatures

Kaplan-Meier assessments of EGFR expression and immune cell infiltrates were compared to determine their contribution to the survival outcomes. In LUAD patients, infiltrates of B and dendritic cells were significantly associated with prognosis. However, in LUSC patients, no survival difference related to the 6 immune cell infiltrates was observed (Figure 6). Pathologic stage II (hazard ratio [HR]: 2.309, 95% CI: 1.493–3.573, P<0.001) and stage III (HR: 2.562, 95% CI: 1.627–4.036, P<0.001) were independent prognostic factors for poor survival in LUAD as assessed through multivariate Cox proportional hazard models. However, LUAD patients who exhibited higher B cell infiltrates had better outcomes (HR: 0.040, 95% CI: 0.002–0.668, P=0.025). In LUSC patients, the multivariate analysis indicated that being male (HR: 1.725, 95% CI: 1.149–2.591, P=0.009) and pathologic stage III (HR: 1.884, 95% CI: 1.214–2.923, P=0.005) significantly affected survival.

Figure 6.

Kaplan-Meier curves of 6 subsets of immune cell infiltrates and epidermal growth factor receptor (EGFR) expression in lung adenocarcinoma (LUAD) and lung squamous cell carcinoma (LUSC) patients.

We confirmed the positive correlations by analyzing lung cancer patient data in the TIMER database. The levels of EGFR expression in LUAD patients correlated with the infiltration of all immune cells assessed (Figure 7). However, EGFR expression was negatively associated with the infiltration of B and CD8+ T cells in LUSC patients. These findings suggested that EGFR is closely related to immune cells in lung cancer.

Figure 7.

Relationship between epidermal growth factor receptor (EGFR) expression and the different subsets of immune cell infiltrates in lung squamous carcinoma (LUSC) and lung adenocarcinoma (LUAD) patients using the TIMER database.

In LUAD patients, those with mutated EGFR had significantly higher infiltration of B and dendritic cells (Figure 8). This implied that the immune response to LUAD in mutated and wild-type patients was disparate. Data were not available for LUSC patients in TIMER.

Figure 8.

Infiltration levels of 6 subsets of immune cells in mutant versus wild-type epidermal growth factor receptor (EGFR) patients. * P<0.05, ** P<0.01.

In different copy number subsets of EGFR mutations, the immune cell infiltration level decreased in deep deletion, arm-level deletion, arm-level gain, and high amplification mutations, as compared to diploid/normal, in both LUAD and LUSC patients (Figure 9). Further correlation analyses showed that EGFR expression was significantly positively correlated with CD274 (PDL1) in LUAD (Figure 10A; r=0.27, P<0.001), but was not related with PDCD1 (PD1) (Figure 10B). In LUSC patients, there was a significant negative correlation between EGFR and PDCD1 (PD1) (r=−0.19, P=0.002, Figure 10D). An inverse correlation of EGFR and CD274 (PDL1) was not observed (Figure 10C). These findings indicate that EGFR might be closely involved in the effect of immunotherapies targeting PD-1 and PD-L1 in lung cancer.

Figure 9.

Tumor immune cell infiltrates in those with disparate epidermal growth factor receptor (EGFR) copy numbers. LUAD, lung adenocarcinoma; LUSC, lung squamous cell carcinoma. * P<0.05, ** P<0.01, *** P<0.001, ● P<0.1.

Figure 10.

Correlation between epidermal growth factor receptor (EGFR) and programmed cell death 1 ligand (PDL1) or its receptor PD1. (A) EGFR with CD274 (PDL1) in lung adenocarcinoma (LUAD) patients. (B) EGFR with PDCD1 (PD1) in LUAD patients. (C) Correlation of EGFR and CD274 (PDL1) in lung squamous cell carcinoma (LUSC) patients. (D) Correlation of EGFR and PDCD1 (PD1) in LUSC patients. Analyses were performed using cBioPortal for Cancer Genomics using The Cancer Genome Atlas LUAD and LUSC data.

Discussion

Currently, the usage of EGFR TKIs in resectable NSCLC for neoadjuvant and adjuvant therapy is considered of great clinical value [20–22]. In this study, clinical and TCGA cohorts were used to assess EGFR mutation profiles and their clinicopathological significance. A larger number of EGFR mutations were observed in the clinical compared to TCGA cohort. Tumor immune cell infiltrates in LUAD and LUSC were different, and EGFR mutations may cause the decline or lack of immune infiltration in the NSCLC microenvironment.

Compared with TCGA cohort, a larger number of EGFR mutations occurred in clinical samples. Both cohorts highlighted the relationship between smoking and pathology for EGFR mutations. In a previous study, Liu et al. also found the occurrence rate was higher in the smoking and pathology subgroups, confirming our findings [23]. In our study, the EGFR mutation frequency in the clinical and TCGA cohorts had no relation with disease stage, indicating that EGFR mutations are likely to occur during the early stages of NSCLC development.

In the current study, we found that EGFR expression was remarkably increased in LUSC compared with adjacent normal tissues. However, EGFR expression had no prognostic ability on OS or PFS in resectable LUAD and LUSC patients based on a Kaplan-Meier survival analysis. Additionally, EGFR alterations in LUAD had an unfavorable influence on OS, but were not associated with changes in PFS. Meanwhile, no significant relationship in LUSC was found between EGFR alterations and OS or PFS. These results suggested that EGFR alterations might be a vital mechanism involved in long-term survival of LUAD patients.

Tumor-infiltrating immune cells are a key point of the anticancer effect in the immune system, and are involved in the response to therapy and cancer escape mechanisms [24,25]. Nevertheless, it remains challenging to confirm the association of immune cell infiltrates with tumor cells owing to the complexity of cancer genomics and the indeterminate immune system of patients. Cytolytic activity regulates the immune response to cancer and its treatment. This represents immunogenicity and acts as a predictor of the response to checkpoint blockade [26].

We speculated that immune evasion or dysfunction of tumor-infiltrating immune cells may be factors for inducing tumor progression and predicting outcomes. In untreated early-stage NSCLC, the immune microenvironment imposes a strong selection pressure on tumor evolution, producing complex mechanisms of immune evasion, namely neoantigen-directed immune escape, which might forecast decreased disease-free survival times [27]. Immune evasion before tumor invasion is also evident in early LUSC, and is associated with the dynamic evolution of pre-invasive bronchial cells and the relevant immune response [28]. A lack of infiltrating immune cells in premalignant lesions could produce tumor progression to LUSC owing to corresponding transcriptomic alterations [29]. Meanwhile, we identified different immune cell infiltrates in LUAD and LUSC and found that lower levels of infiltrating B and dendritic cells predicted poor prognosis in LUAD. Conversely, the level of immune cell infiltrates was not related to a significant survival disparity in LUSC.

The EGFR provides immuno-competence in lung cancer. In our study, positive associations in LUAD with EGFR expression were observed for the infiltration level of B cells, CD8+ T cells, CD8+ T cells, CD4+ T cells, neutrophils, and dendritic cells. However, we found a negative relationship between the infiltration of B and CD8+ T cells with EGFR expression in LUSC, in accordance with a previous study [30]. This trend was also demonstrated in B and dendritic cells of patients with mutated compared to wild-type EGFR (Figure 9), although the relationship did not reach statistical significance for CD4 T cells, CD8 T cells, macrophages, and neutrophils. In addition, to determine the role of copy number alterations of EGFR in the immune cell infiltration level in LUAD and LUSC patients, further analyses revealed that the levels in deep deletion, arm-level deletion, arm-level gain, and high amplification mutation subtypes were decreased compared to diploid/normal. Overall, these findings indicate that mutations of EGFR might induce a decline or lack of immune cell infiltration in the tumor microenvironment, reducing the anticancer effect of immune cells.

The PDL1/PD1 pathway plays essential roles in the immune evasion of tumor cells, and is a key immune co-inhibitory pathway for NSCLC immunotherapy [31]. Through correlation analyses, we observed that EGFR expression was positively correlated with PDL1 expression in LUAD, and negatively associated with PD1 in LUSC. However, the correlation between PDL1/PD1 expression and EGFR expression/mutation remains controversial. Azuma et al. highlighted the association of elevated PDL1 levels with EGFR mutations by immunohistochemistry in 164 surgically resected NSCLC specimens [32]. Similarly, Gatalica et al. showed that PDL1 positivity was related to the occurrence of EGFR mutations [33]. Ji et al. highlighted a negative correlation between PDL1 and EGFR mutational status in 100 resected patients with primary LUAD, with higher rates of mutations occurring in those with low levels of PDL1 expression [34]. This was confirmed by a meta-analysis showing that wild-type EGFR NSCLC was more likely to be PDL1-positive compared to mutant EGFR NSCLC [35].

TKIs inhibiting EGFR activity can reduce PDL1 expression by inhibiting NF-κB in EGFR mutant NSCLC [36]. Furthermore, NSCLC tumors can evade the immune response by upregulating PDL1 via EGFR activation [37]. Conversely, Mu et al. reported no significant correlation between PD-L1 and EGFR expression in stage I NSCLC patients [38]. Additionally, Zhang et al. found no significant relationship between PDL1 and EGFR expression in LUAD [39]. The expression of PDL1 was associated with the levels of wild-type EGFR, but no significant changes in PDL1 expression were observed according to EGFR mutational status [40].

First-line treatment with pembrolizumab (which blocks PD1) was ineffective in 10 patients with advanced NSCLC with mutated EGFR and positive PDL1 who did not receive TKI therapy in a phase II clinical trial [41]. Second-line therapy with immune checkpoint inhibitors did not improve survival over chemotherapy in EGFR-mutant advanced NSCLC [42]. High PDL1 expression correlated with poor responses to TKI therapy in those harboring EGFR mutations [43], whilst higher PDL1 expression was associated with TKI resistance in advanced LUAD patients harboring EGFR mutations [44]. Despite the poor efficacy of TKI therapy in those with high PDL1 expression upon first-line treatment, the efficacy improved in third generation treatment and appeared unaffected by PDL1 expression. Given these findings, EGFR mutant and PDL1-positive patients should receive TKIs targeting the EGFR as a first-line therapy.

There are several limitations in our study. Firstly, as a retrospective study, the interpretation of the data and the conclusions may be limited to a certain extent. PDL1 immunohistochemical analysis was not performed in the clinical cohort, and the correlation of PDL1 and EGFR mutations in resectable NSCLC must be performed. Secondly, this was a single center retrospective study. A multiple center study would provide more compelling clinical judgements. In addition, experiments were not conducted in our patients to detect the tumor immune cell infiltration level and immunity signature. Thus, the results of our study should be evaluated carefully, and additional research on the current topic is recommended.

Conclusions

This study demonstrated that EGFR mutations are frequent in resectable lung cancer and contribute to the long-term survival outcomes of LUAD patients. The levels of tumor immune cell infiltrates in LUAD and LUSC differed and, in part, were significantly related to EGFR expression. EGFR mutations decrease immune cell infiltration in the tumor microenvironment, decreasing the anticancer effect of immune cells. The correlation between PDL1/PD1 expression and EGFR expression/mutation must now be investigated to elucidate whether combination therapy with EGFR TKIs and immune checkpoint blockers is beneficial for subsets of NSCLC patients.