Expression signature, prognosis value, and immune characteristics of Siglec-15 identified by pan-cancer analysis

Figures & data

Table 1. Patient clinical parameters and their association with Siglec-15 expression.

Clinical parameters	n (%)	Siglec-15		p-Value
		Positive	Negative
T classification
T1	55(53.4)	6	49	0.149
T2	39(37.9)	10	29
T3+ T4	9(8.8)	1	8
N classification
N0	62(60.2)	9	53	0.303
N1	6(5.8)	0	6
N2	35(34.0)	8	27
Clinical stage
I	48(46.6)	6	42	0.275
II	18(17.5)	2	16
III	37(35.9)	9	28
Smoking index
<400	70(68.0)	13	57	0.572
≥400	33(32.0)	4	29
Age (years)
≤60	58(56.3)	10	48	0.819
>60	45(43.7)	7	38
Gender
Males	42(40.8)	5	37	0.419
Females	61(59.2)	12	49

Table 3. Univariate survival analysis of Siglec-15 expression in subgroups with different clinical parameters.

Clinical parameters	n	PFS		OS
		Hazard ratio (95%CI)	p-Value	Hazard ratio (95%CI)	p-Value
T classification
T1	56	1.761(0.471–6.590)	0.290	0.538(0.112–2.591)	0.542
T2+ T3+ T4	47	2.962(1.120–7.833)	0.001	2.060(0.753–5.634)	0.080
N classification
N0	62	3.156(0.895–11.120)	0.005	2.026(0.497–8.260)	0.207
N1+ N2	41	2.125(0.773–5.843)	0.053	1.431(0.474–4.318)	0.472
Clinical stage
I+ II	66	2.788(0.758–10.25)	0.018	1.432(0.350–5.858)	0.566
III	37	1.980(0.778–5.042)	0.073	1.534(0.538–4.374)	0.361
Smoking index
<400	70	2.488(1.036–5.973)	0.005	1.738(0.635–4.754)	0.196
≥400	33	3.221(0.453–22.90)	0.052	1.625(0.266–9.941)	0.527
Age (years)
≤60	58	2.941(1.001–8.639)	0.003	2.167(0.666–7.046)	0.100
>60	45	2.552(0.728–8.943)	0.034	1.140(0.315–4.127)	0.834
Gender
males	42	5.040(0.803–31.63)	<0.001	1.723(0.379–7.845)	0.386
females	61	2.065(0.842–5.063)	0.043	1.742(0.586–5.180)	0.231

Note. Bold font indicates p < 0.05.

Figure 1. The transcription levels of Siglec-15 in human cancers.

(a) The mRNA expression of Siglec-15 between tumor and normal tissues was assessed using tissues from TCGA and GTEx. (b) Association of Siglec-15 mRNA expression and different pathological stages in patients with different cancers from TCGA. (c) The expression of Siglec-15 in different molecular subtypes of cancers via TISIDB. (. p-value ≤ 0.1; *p-value ≤ 0.05; **p-value ≤ 0.01; *** p-value ≤ 0.001; Siglec-15 gene symbol is SIGLEC15).

Figure 2. CNA and DNA methylation of Siglec-15 in human cancers.

(a) CNA and mutation frequency data of Siglec-15 in different cancer studies were accessed from cBioPortal. (b) The relationship between Siglec-15 expression and relative liner copy-number values from TCGA. (c) Negative correlations between DNA methylation and mRNA expression of Siglec-15 were shown from TCGA. (d) DNA-methylation beta values ranging from 0 (unmethylated) to 1 (fully methylated) were determined by UALCAN. (hypermethylation [beta-value: 0.7–0.5] or hypo-methylation [beta-value: 0.3–0.25]).

Figure 3. The prognostic significance of Siglec-15 assessed by Kaplan-Meier analysis.

(a) High Siglec-15 mRNA expression correlated with good prognosis in BLCA, BRCA, HNSC, LIHC, OV, THCA, and UCEC from TCGA (HR < 1, p-value <0.05). (b) High Siglec-15 mRNA expression correlated with bad prognosis in KIRC, PAAD, and SARC from TCGA (HR > 1, p-value <0.05). (c) High Siglec-15 mRNA expression correlated with good prognosis in BRCA-luminal A, BRCA-luminal B, and OV-serous, and STAD-diffuse from GEO and other projects (HR < 1, p-value <0.05). (d) High Siglec-15 mRNA expression correlated with bad prognosis in LUAD, BRCA-Basal, BRCA-Her2, STAD-intestinal, and STAD-mixed from GEO and other projects (HR < 1, p-value <0.05).

Figure 4. Correlation analysis between Siglec-15 expression and tumor-infiltrating immune cell.

(a) Correlation analysis of Siglec-15 mRNA expression with 22 types of immune cells were investigated across cancers from TCGA by CIBERSORT (*p-value ≤ 0.05, |Spearman r| > 0.2). (b) Correlation analysis of Siglec-15 mRNA expression with 22 types of immune cells were investigated across cancers from GEO by CIBERSORT. (c) Siglec-15 mRNA expression in different immune subtypes in LUAD, SARC, BRCA, UCEC, BLCA, MESO, KICH, PCPG, and TGCT via TISIDB.

Figure 5. Significant pathways influenced by Siglec-15.

(a) Relationships between Siglec-15 and KEGG pathways in cancers from TCGA analyzed by GSEA. (b) Relationships between Siglec-15 and GO BP terms in cancers from TCGA analyzed by GSEA (c) Relationships between Siglec-15 and GO MF terms in cancers from TCGA analyzed by GSEA. (d) Relationships between Siglec-15 and GO CC terms in cancers from TCGA analyzed by GSEA. (NES ≥ 1.0, FDR <0.25)

Figure 6. Preliminary experimental verification of Siglec-15 signature in LUAD.

(a) Siglec-15 expression on 20 paired LUAD tissues and adjacent normal tissues by RT-qPCR. (b) “Chemokine signaling pathway” associated with Siglec-15 in LUAD. “Cytokine production” and “regulation of cytokine production” included in the TOP20 GO terms in LUAD (NES ≥ 1.5, FDR <0.05). (c) The association between the key genes in chemokine signaling pathway and Siglec-15 was detected by RT-qPCR in 20 LUAD tissue samples (r: Pearson’s correlation coefficient). (d) Representative negative and positive samples were taken at 200x and 400x magnification for IHC staining of Siglec-15. (e) Survival analysis of patients stratified into positive group (red) and negative group (blue) according to the membrane staining of Siglec-15 by IHC in a cohort of 103 LUAD specimens. (f) The relative levels of immune cells infiltration investigated for Siglec-15 negative and positive samples. (g) Immunohistochemical staining of DCs (CD11 c) and B cells (CD20) in Siglec-15 positive and negative group (400x).

Table 2. Univariate survival analysis of clinical parameters and Siglec-15 expression with PFS and OS in patients with LUAD.

Clinical parameters	n	PFS		OS
		Hazard ratio (95%CI)	p-Value	Hazard ratio (95%CI)	p-Value
T classification
T1	56	2.346	0.001	2.566	0.003
T2+ T3+ T4	47	(1.393–3.952)		(1.375–4.789)
N classification
N0	62	2.450	<0.001	2.889	<0.001
N1+ N2	41	(1.434–4.187)		(1.505–5.544)
Clinical stage
I+II	66	3.380	<0.001	3.050	<0.001
III	37	(2.031–5.625)		(1.581–5.887)
Smoking index
<400	70	1.427	0.218	1.120	0.742
≥400	33	(0.839–2.427)		(0.580–2.162)
Age (years)
≤60	58	1.112	0.679	0.8703	0.656
>60	45	(0.670–1.845)		(0.471–1.610)
Gender
Males	42	0.838	0.501	1.074	0.821
Females	61	(0.504–1.393)		(0.575–2.008)
Siglec-15
Positive	17	2.755	<0.001	1.734	0.138
Negative	86	(1.219 − 6.229)		(0.717–4.195)

Note. Bold font indicates p < 0.05.

Figure 7. Analysis explanation with a detailed flow chart of this study. The study comprises 4 parts: Ⅰ Siglec-15 mRNA expression characteristics investigated by TCGA, GTEx, and Ocomine; Ⅱ Siglec-15 prognostic value landscape in TCGA and GEO; Щ Tumor infiltrating pattern and associated pathways about Siglec-15 in TCGA and GEO; Ⅳ Experiment verification in LUAD patients.