Landscape of mutation profiles in lung squamous cell carcinoma
We downloaded the somatic mutation profiling, RNA-seq transcriptome profiling and corresponding clinicopathological information of 491 patients with lung squamous cell carcinoma from TCGA. Missense mutation accounts for the most fraction in all types of mutation (Fig. 1), and single nucleotide polymorphism (SNP) is more common than insertion or deletion in lung squamous cell carcinoma (Fig. 1). What’s more, C > T and C > A were most common in single nucleotide variants (SNV) (Fig. 1). Top 10 most common mutated genes were TTN(68%), TP53(77%), MUC16(36%༉, CSMD3(40%), RYR2(35%), LRP1B(30%), USH2A(30%), SYNE1(29%), ZFHX4(26%), KMT2D(22%) (Fig. 1). Waterfall plot showed the mutation information and the different mutation types of each gene in each sample (Fig. 2). The association between different mutated genes were shown in Fig. 3, we can intuitively see TTN is co-occurrence with PAPPA2, ZFHX4, MUC16, and CSMD3 is co-occurrence with RYR3, NAV3, USH2A. The mutually exclusive relationships can be saw between PAPPA2 and TP53.
The correlation between TMB and overall survival, clinical characteristics
We calculated TMB score and classified samples into the low-TMB group and the high-TMB group based on median value. Kaplan-Meier method showed that compared with low-TMB group, patients in high-TMB group revealed better one-year survival outcomes (Fig. 4A). However, high-TMB correlated with advanced pathological stages(Figure 4B༉.
Differentially expressed genes between high- and low- TMB groups and enrichment analysis
Differentially expressed genes between high- and low- TMB groups can be seen in Table1, such as CYSLTR2, MS4A1, FAM107A, IGLL1.
Table 1
Differential expressed genes between low TMB and high TMB groups
gene
|
logFC
|
pValue
|
CYSLTR2
|
-1.6898991
|
0.00010269
|
MS4A1
|
-1.1441338
|
0.00058734
|
FAM107A
|
-1.0200621
|
7.73E-05
|
IGLL1
|
-1.7638948
|
0.00533523
|
LRRC55
|
-1.785876
|
0.00055621
|
MS4A8
|
-1.8295919
|
0.00021727
|
C20orf85
|
-1.1821732
|
0.00062144
|
SELE
|
-1.2084087
|
0.00472061
|
TNFSF8
|
-1.1427382
|
0.0032476
|
NR5A1
|
1.9226092
|
7.87E-06
|
CADM3
|
-1.2047907
|
5.53E-05
|
FCRL2
|
-1.159438
|
0.0008174
|
BPIFB1
|
-1.4510685
|
0.00011556
|
ADH1B
|
-1.1416372
|
9.09E-06
|
INHA
|
1.85340517
|
0.00362178
|
SCGB1A1
|
-1.1115467
|
1.63E-05
|
PIGR
|
-1.0412167
|
0.00022015
|
C1orf189
|
-1.1341047
|
0.00026482
|
WFDC12
|
-1.2491682
|
0.00164705
|
FAM216B
|
-1.3900118
|
2.55E-06
|
HS3ST4
|
-1.504201
|
0.00026443
|
PIP
|
-1.0171485
|
0.00026387
|
CD22
|
-1.0772369
|
0.00013723
|
FCER2
|
-1.3194606
|
0.00204346
|
C2orf40
|
-1.2412141
|
3.72E-05
|
CCL19
|
-1.073972
|
0.00058595
|
TLR10
|
-1.0166526
|
0.00302647
|
C1orf194
|
-1.0624677
|
0.00026991
|
APOA1
|
6.52638412
|
0.00622402
|
SMIM24
|
1.10145976
|
0.00793992
|
Then We conducted the KEGG enrichment analysis and the results showed that these DEGs were mainly involved in lymphocyte activation, lymphocyte proliferation, leukocyte proliferation and mononuclear cell proliferation(Figure 5).
GSEA revealed prominent enrichment of signatures related in the regulation of transcription involved in G1/S transition of mitotic cell cycle, DNA replication, cell meiosis cell cycle process, telomere maintenance via semi-conservative replication, and meiotic cell cycle process in the high TMB group (Fig. 6;Table 2). And in the low TMB group, regulation of microglial cell activation, regulation of B cell activation, humoral immune response and leukocyte migration were enriched group(Figure 7; Table 3).
Table 2
Gene Set Enrichment Analysis (GSEA)in high- TMB groups
NAME
|
ES
|
NES
|
NOM p-value
|
FDR q-value
|
GO_REGULATION_OF_TRANSCRIPTION_INVOLVED_
IN_G1_S_TRANSITION_OF_MITOTIC_CELL_CYCLE
|
0.784
|
2.134
|
0.000
|
0.491
|
GO_DNA_REPLICATION_INITIATION
|
0.784
|
2.119
|
0.004
|
0.304
|
GO_MEIOSIS_I_CELL_CYCLE_PROCESS
|
0.579
|
2.103
|
0.006
|
0.257
|
GO_DNA_POLYMERASE_COMPLEX
|
0.766
|
2.082
|
0.000
|
0.251
|
GO_TELOMERE_MAINTENANCE_VIA_SEMI_
CONSERVATIVE_REPLICATION
|
0.796
|
2.078
|
0.000
|
0.211
|
GO_MEIOTIC_CELL_CYCLE_PROCESS
|
0.538
|
2.076
|
0.002
|
0.181
|
GO_CATALYTIC_ACTIVITY_ACTING_ON_A_TRNA
|
0.637
|
2.052
|
0.006
|
0.207
|
GO_CHAPERONE_COMPLEX
|
0.692
|
2.045
|
0.004
|
0.195
|
GO_EUCHROMATIN
|
0.657
|
2.039
|
0.002
|
0.187
|
GO_REGULATION_OF_CHROMOSOME_SEPARATION
|
0.640
|
2.034
|
0.006
|
0.179
|
GO_ENDONUCLEASE_COMPLEX
|
0.683
|
2.034
|
0.000
|
0.163
|
GO_MEIOTIC_CHROMOSOME_SEGREGATION
|
0.579
|
2.030
|
0.004
|
0.155
|
GO_NCRNA_3_END_PROCESSING
|
0.653
|
2.028
|
0.000
|
0.146
|
GO_RNA_3_END_PROCESSING
|
0.606
|
2.019
|
0.006
|
0.151
|
GO_FEMALE_MEIOTIC_NUCLEAR_DIVISION
|
0.660
|
2.017
|
0.004
|
0.145
|
GO_HISTONE_EXCHANGE
|
0.686
|
2.015
|
0.002
|
0.138
|
GO_CHROMATIN_REMODELING_AT_CENTROMERE
|
0.780
|
2.007
|
0.002
|
0.142
|
GO_MITOTIC_SPINDLE_ASSEMBLY
|
0.638
|
2.006
|
0.008
|
0.136
|
GO_MITOTIC_SPINDLE_ORGANIZATION
|
0.598
|
2.006
|
0.008
|
0.130
|
GO_CHROMOSOME_SEGREGATION
|
0.557
|
2.004
|
0.014
|
0.125
|
Table 3
Gene Set Enrichment Analysis (GSEA)in low- TMB groups
NAME
|
ES
|
NES
|
NOM p-value
|
FDR q-value
|
GO_TRANSFORMING_GROWTH_FACTOR_BETA_BINDING
|
-0.842
|
-2.267
|
0.000
|
0.126
|
GO_EXTERNAL_SIDE_OF_PLASMA_MEMBRANE
|
-0.677
|
-2.265
|
0.002
|
0.065
|
GO_REGULATION_OF_MICROGLIAL_
CELL_ACTIVATION
|
-0.783
|
-2.256
|
0.000
|
0.052
|
GO_POSITIVE_REGULATION_OF_
B_CELL_ACTIVATION
|
-0.751
|
-2.245
|
0.000
|
0.046
|
GO_PEPTIDE_CROSS_LINKING
|
-0.741
|
-2.230
|
0.000
|
0.045
|
GO_POSITIVE_REGULATION_OF_
VASCULATURE_DEVELOPMENT
|
-0.574
|
-2.220
|
0.000
|
0.046
|
GO_ANTIGEN_BINDING
|
-0.829
|
-2.217
|
0.002
|
0.042
|
GO_FC_RECEPTOR_MEDIATED_STIMULATORY_
SIGNALING_PATHWAY
|
-0.724
|
-2.214
|
0.002
|
0.039
|
GO_INTEGRIN_BINDING
|
-0.633
|
-2.210
|
0.002
|
0.037
|
GO_HUMORAL_IMMUNE_RESPONSE
|
-0.631
|
-2.206
|
0.000
|
0.034
|
GO_LEUKOCYTE_MIGRATION
|
-0.608
|
-2.202
|
0.002
|
0.034
|
GO_B_CELL_RECEPTOR_SIGNALING_PATHWAY
|
-0.842
|
-2.202
|
0.000
|
0.031
|
GO_NEGATIVE_REGULATION_OF_SMOOTH_
MUSCLE_CELL_PROLIFERATION
|
-0.593
|
-2.195
|
0.000
|
0.033
|
GO_PHOSPHORUS_OXYGEN_LYASE_ACTIVITY
|
-0.746
|
-2.194
|
0.000
|
0.032
|
GO_HUMORAL_IMMUNE_RESPONSE_MEDIATED_BY_
CIRCULATING_IMMUNOGLOBULIN
|
-0.820
|
-2.192
|
0.002
|
0.030
|
GO_CELLULAR_EXTRAVASATION
|
-0.705
|
-2.188
|
0.002
|
0.031
|
GO_MEMBRANE_INVAGINATION
|
-0.741
|
-2.185
|
0.002
|
0.031
|
GO_REGULATION_OF_B_CELL_ACTIVATION
|
-0.715
|
-2.185
|
0.002
|
0.029
|
GO_PHAGOCYTOSIS
|
-0.658
|
-2.181
|
0.004
|
0.029
|
GO_CELL_RECOGNITION
|
-0.600
|
-2.181
|
0.002
|
0.028
|
Immune cells infiltration between high- and low- TMB groups
Violin plot showed different immune cells infiltration between high-TMB and low- TMB groups in in lung squamous cell carcinoma. Infiltration levels of CD8 + T cell, M1 macrophages, follicular helper T cells and activated NK cells in high-TMB group were higher than that in low-TMB group. And infiltration levels of plasma cells, activated CD4 + T memory cells, activated NK cells and M0 macrophages were lower in high-TMB group, compared with low-TMB group(Figure 8).