High STK40 Expression as an Independent Prognostic Biomarker and Correlated with Immune Infiltrates in Low-Grade Gliomas

Figures & data

Table 1 Clinical Characteristics of the LGG Patients Based on TCGA

Characteristics	Levels	Overall
n		528
WHO grade, n (%)	G2	224 (48%)
	G3	243 (52%)
IDH status, n (%)	WT	97 (18.5%)
	Mut	428 (81.5%)
1p/19q codeletion, n (%)	Codel	171 (32.4%)
	Non-codel	357 (67.6%)
Primary therapy outcome, n (%)	PD	110 (24%)
	SD	146 (31.9%)
	PR	64 (14%)
	CR	138 (30.1%)
Gender, n (%)	Female	239 (45.3%)
	Male	289 (54.7%)
Race, n (%)	Asian	8 (1.5%)
	Black or African American	22 (4.3%)
	White	487 (94.2%)
Age, n (%)	≤40	264 (50%)
	>40	264 (50%)
Histological type, n (%)	Astrocytoma	195 (36.9%)
	Oligoastrocytoma	134 (25.4%)
	Oligodendroglioma	199 (37.7%)
Laterality, n (%)	Left	256 (48.9%)
	Midline	6 (1.1%)
	Right	261 (49.9%)
Age, median (IQR)		40.5 (32, 53)

Table 2 Correlation Between STK40 Expression and Clinicopathological Characteristics in LGG

Characteristic	Levels	Low Expression of STK40	High Expression of STK40	p	Method
n		264	264
WHO grade, n (%)	G2	147 (31.5%)	77 (16.5%)	< 0.001	Chisq.test
	G3	90 (19.3%)	153 (32.8%)
IDH status, n (%)	WT	13 (2.5%)	84 (16%)	< 0.001	Chisq.test
	Mut	249 (47.4%)	179 (34.1%)
1p/19q codeletion, n (%)	Codel	163 (30.9%)	8 (1.5%)	< 0.001	Chisq.test
	Non-codel	101 (19.1%)	256 (48.5%)
Primary therapy outcome, n (%)	PD	33 (7.2%)	77 (16.8%)	< 0.001	Chisq.test
	SD	80 (17.5%)	66 (14.4%)
	PR	34 (7.4%)	30 (6.6%)
	CR	79 (17.2%)	59 (12.9%)
Gender, n (%)	Female	125 (23.7%)	114 (21.6%)	0.382	Chisq.test
	Male	139 (26.3%)	150 (28.4%)
Race, n (%)	Asian	5 (1%)	3 (0.6%)	0.369	Fisher.test
	Black or African American	8 (1.5%)	14 (2.7%)
	White	244 (47.2%)	243 (47%)
Age, n (%)	≤40	124 (23.5%)	140 (26.5%)	0.192	Chisq.test
	>40	140 (26.5%)	124 (23.5%)
Histological type, n (%)	Astrocytoma	53 (10%)	142 (26.9%)	< 0.001	Chisq.test
	Oligoastrocytoma	59 (11.2%)	75 (14.2%)
	Oligodendroglioma	152 (28.8%)	47 (8.9%)
Laterality, n (%)	Left	125 (23.9%)	131 (25%)	0.602	Fisher.test
	Midline	2 (0.4%)	4 (0.8%)
	Right	134 (25.6%)	127 (24.3%)
Age, median (IQR)		41 (34, 53)	39 (31, 52.25)	0.151	Wilcoxon

Table 3 Logistic Analysis of the Association Between STK40 Expression and Clinical Characteristics

Characteristics	Total (N)	Odds Ratio (OR)	P value
WHO grade (G3 vs G2)	467	3.245 (2.228–4.760)	<0.001
1p/19q codeletion (non-codel vs codel)	528	51.644 (26.025–117.948)	<0.001
Primary therapy outcome (CR vs PD&SD&PR)	458	0.635 (0.423–0.948)	0.027
IDH status (Mut vs WT)	525	0.111 (0.058–0.199)	<0.001
Gender (Male vs Female)	528	1.183 (0.840–1.669)	0.336
Race (White vs Asian & Black or African American)	517	0.762 (0.355–1.596)	0.473
Age (>40 vs ≤40)	528	0.784 (0.557–1.104)	0.164
Laterality (Midline & Right vs Left)	523	0.919 (0.652–1.295)	0.630
Histological type (Oligodendroglioma vs Astrocytoma & Oligoastrocytoma)	528	0.160 (0.106–0.236)	<0.001

Figure 1 STK40 is significantly upregulated in LGG than normal tissues. (A) The difference expression of TLX1 in LUAD and unpaired normal lung tissues. (B) ROC curve. Significance markers: ***P<0.001.

Figure 2 Box plot evaluating STK40 expression of patients with LGG according to different clinical characteristics. (A) WHO grade. (B) 1p/19q codeletion. (C) IDH status. (D) primary therapy outcome. (E) histological type. Significance markers: **P<0.01; ***P<0.001.

Table 4 Univariate and Multivariate Cox Regression Analyses of Clinical Characteristics Associated with OS

Characteristics	Total (N)	Univariate Analysis		Multivariate Analysis
		Hazard Ratio (95% CI)	P value	Hazard Ratio (95% CI)	P value
WHO grade (G3 vs G2)	466	3.059 (2.046–4.573)	<0.001	1.735 (1.073–2.808)	0.025
1p/19q codeletion (non-codel vs codel)	527	2.493 (1.590–3.910)	<0.001	0.862 (0.377–1.975)	0.726
Primary therapy outcome (CR vs PD&SD&PR)	457	0.203 (0.094–0.437)	<0.001	0.276 (0.127–0.601)	0.001
IDH status (Mut vs WT)	524	0.186 (0.130–0.265)	<0.001	0.353 (0.215–0.580)	<0.001
Gender (Male vs Female)	527	1.124 (0.800–1.580)	0.499
Race (White vs Asian & Black or African American)	516	0.849 (0.395–1.823)	0.675
Age (>40 vs ≤40)	527	2.889 (2.009–4.155)	<0.001	3.315 (2.092–5.254)	<0.001
Histological type (Oligodendroglioma vs Astrocytoma & Oligoastrocytoma)	527	0.673 (0.471–0.961)	0.029	0.762 (0.457–1.272)	0.299
Laterality (Midline & Right vs Left)	522	0.776 (0.548–1.099)	0.154
STK40 (High vs Low)	527	3.071 (2.089–4.515)	<0.001	2.284 (1.125–4.637)	0.022

Figure 3 High expression of STK40 is associated with poor OS and DSS in patients with LGG. (A) The Kaplan–Meier curves and number at risk of OS in LGG. (B) The Kaplan–Meier curves and number at risk of PFS in LGG. (C) The Kaplan–Meier curves and number at risk of DSS in LGG.

Abbreviations: OS, over survival; PFS, progress free interval; DSS, disease specific survival.

Figure 4 Forest plot of the multivariate Cox regression analysis for LGG.

Figure 5 Nomogram for predicting the probability of patients with 1-, 3- and 5-year overall survival.

Table 5 Gene Sets Enriched in the High STK40 Expression Phenotype

Description	NES	P Adjust	FDR
REACTOME_CELL_CYCLE_MITOTIC	1.725	0.015	0.011
REACTOME_NEUTROPHIL_DEGRANULATION	1.949	0.015	0.011
REACTOME_SIGNALING_BY_RHO_GTPASES	1.548	0.015	0.011
REACTOME_SIGNALING_BY_INTERLEUKINS	2.249	0.015	0.011
REACTOME_M_PHASE	1.660	0.015	0.011
REACTOME_RHO_GTPASE_EFFECTORS	1.711	0.015	0.011
REACTOME_EXTRACELLULAR_MATRIX_ORGANIZATION	2.238	0.015	0.011
WP_PI3KAKT_SIGNALING_PATHWAY	1.637	0.015	0.011
NABA_SECRETED_FACTORS	1.748	0.015	0.011

Figure 6 Enrichment plots from gene set enrichment analysis (GSEA). (A) neutrophil degranulation, (B) signaling by interleukins, (C) GPCR-ligand binding, (D) G alpha (I) signaling events, (E) VEGFAVEGFR-2 signaling pathway, (F) Class A 1 Rhodopsin-Like Receptors, (G) naba secreted factors, (H) PI3K-Akt signaling pathway, and (I) Focal Adhesion-PI3K-Akt-mTOR-signaling pathway, were differentially enriched in STK40-related OC.

Abbreviations: NES, normalized ES; FDR, false discovery rate.

Table 6 The Correlation Between STK40 Expression and Immune Cells Detected by Spearman Correlation Method

Gene Name	Cell Type	Correlation Coefficient (Spearman)	P value (Spearman)
STK40	aDC	0.357	<0.001
STK40	B cells	0.136	0.002
STK40	CD8 T cells	−0.047	0.284
STK40	Cytotoxic cells	0.301	<0.001
STK40	DC	−0.174	<0.001
STK40	Eosinophils	0.517	<0.001
STK40	iDC	0.345	<0.0018
STK40	Macrophages	0.57	<0.001
STK40	Mast cells	−0.044	0.316
STK40	Neutrophils	0.475	<0.001
STK40	NK CD56bright cells	−0.305	<0.001
STK40	NK CD56dim cells	0.179	<0.001
STK40	NK cells	0.346	<0.001
STK40	pDC	−0.357	<0.001
STK40	T cells	0.311	<0.001
STK40	T helper cells	0.315	<0.001
STK40	Tcm	−0.001	0.981
STK40	Tem	0.088	0.044
STK40	TFH	−0.033	0.455
STK40	Tgd	0.228	<0.001
STK40	Th1 cells	0.094	0.031
STK40	Th17 cells	0.282	<0.001
STK40	Th2 cells	0.313	<0.001
STK40	Treg	−0.298	<0.001

Figure 7 Correlation of STK40 expression levels with immune infiltration (Spearman r).

Abbreviations: DC, dendritic cells; aDC, activated DC; iDC, immature DC; pDC, plasmacytoid DC; Tem, T effector memory; Tgd, T gamma delta.