GNPNAT1 is a Biomarker That Predicts a Poor Prognosis of Breast Cancer

Figures & data

Table 1 Baseline Characteristics of the GNPNAT1 High Expression Set and the Low Expression Set in the TCGA Cohort

Clinicopathologic Factors	High Expression	Low Expression	P value
Race			< 0.001***
Asian	23	37
White	373	380
Others	119	62
Age (years)			0.411
<=60	293	308
>60	248	234
Histological type			< 0.001***
Infiltrating Ductal Carcinoma	356	416
Infiltrating Lobular Carcinoma	128	77
PR status			< 0.001***
Negative	208	134
Indeterminate	1	3
Positive	311	377
ER status			< 0.001***
Negative	162	78
Indeterminate	0	2
Positive	359	434
HER2 status			0.005**
Negative	294	264
Indeterminate	5	7
Positive	60	97
Menopause status			0.074
Pre	99	130
Peri	20	20
Post	365	338
T stage (AJCC7)			0.640
T1	137	140
T2	318	311
T3	72	67
T4	14	21
N stage (AJCC7)			0.013*
N0	283	231
N1	168	190
N2	47	69
N3	37	39
M stage (AJCC7)			1.000
M0	438	464
M1	10	10
Pathologic stage (AJCC7)			0.103
Stage I	98	83
Stage II	323	296
Stage III	106	136
Stage IV	10	8
Total	541	542

Notes: *p-value< 0.05, **p-value<0.01, ***p-value<0.001.

Abbreviations: GNPNAT1, glucosamine-phosphate N-acetyltransferase 1; AJCC7, American Joint Committee on Cancer 7th edition.

Table 2 Baseline Characteristics of the GNPNAT1 High Expression Set and the Low Expression Set in the Validated Cohort

Clinicopathologic Factors	High Expression	Low Expression
Age (years)
<=60	11	15
>60	7	7
PR status
Negative	18	18
Positive	0	4
ER status
Negative	17	16
Positive	1	6
HER2 status
Negative	16	11
Positive	2	11
Menopause status
Pre	6	13
Post	12	9
T stage (AJCC7)
T1	7	13
T2	10	7
T3	1	2
N stage (AJCC7)
N0	13	13
N1	5	9
M stage (AJCC7)
M0	18	22
Pathologic stage (AJCC7)
Stage I	4	5
Stage II	11	15
Stage III	3	2
Total	18	22

Abbreviations: GNPNAT1, glucosamine-phosphate N-acetyltransferase 1; AJCC7, American Joint Committee on Cancer 7th edition.

Figure 1 The expression level of GNPNAT1 in breast cancer. (A) The GNPNAT1 expression level of different cancers in the TCGA dataset. (B) GNPNAT1 expression of different breast cancer subtypes in TCGA+GTEx dataset. (C) GNPNAT1 expression in breast cancer invalidated cohort. *P<0.05, **p<0.01, ***p<0.001.

Table 3 The Correlation Between GNPNAT1 Expression and Clinicopathologic Factors in the TCGA Cohort

Characteristics	Total (N)	OR (95% CI)	P value
Race (Others vs Asian)	994	0.558 (0.323–0.948)	0.033*
Age (>60 vs <=60)	1083	0.898 (0.706–1.141)	0.377
Menopause status (Post vs Pre & Peri)	972	0.735 (0.553–0.974)	0.032*
Histological type (Lobular vs Ductal)	977	0.515 (0.374–0.705)	<0.001***
PR status (Positive vs Negative)	1030	1.882 (1.447–2.454)	<0.001***
ER status (Positive vs Negative)	1033	2.511 (1.858–3.416)	<0.001***
HER2 status (Positive vs Negative)	715	1.800 (1.257–2.596)	0.001**
T stage (T3&4 vs T1&2)	1080	1.032 (0.746–1.429)	0.848
N stage (N1&2and3 vs N0)	1064	1.449 (1.138–1.846)	0.003**
M stage (M1 vs M0)	922	0.944 (0.384–2.322)	0.899
Pathologic stage (III&IV vs I&II)	1060	1.379 (1.042–1.829)	0.025*

Notes: *p-value< 0.05, **p-value<0.01, ***p-value<0.001. The stages were graded according to AJCC7.

Abbreviations: GNPNAT1, glucosamine-phosphate N-acetyltransferase 1; AJCC7, American Joint Committee on Cancer 7th edition.

Figure 2 (A–H) GNPNAT1 expression level in Race, Menopause status, Pathological stage, N stage, Histological type, ER status, PR status, and HER2 status based on different clinical features. *P<0.05, **p<0.01, ***p<0.001.

Figure 3 Diagnostic value of the expression of GNPNAT1 and the correlation between SND1 gene expression and the survival prognosis of cancers in TCGA. (A) The expression level of GNPNAT1 in different breast cell lines. (B) ROC curve for differentiating normal people and breast cancer patients. (C and D) The survival map (C) and survival curves (D) of GNPNAT1 in breast cancer from the GEPIA2 dataset. *P<0.05, **p<0.01.

Table 4 Univariate COX Regression Analysis for Overall Survival in the TCGA Cohort

Characteristics	Total (N)	HR (95% CI)	P value
T stage (T3&4 vs T1&2)	1079	1.608 (1.110–2.329)	0.012*
N stage (N1&2and3 vs N0)	1063	2.239 (1.567–3.199)	<0.001***
M stage (M1 vs M0)	922	4.254 (2.468–7.334)	<0.001***
Pathologic stage (III&IV vs I&II)	1059	2.391 (1.703–3.355)	<0.001***
Race (Others vs Asian)	993	1.362 (0.432–4.289)	0.598
Age (>60 vs <=60)	1082	2.020 (1.465–2.784)	<0.001***
Histological type (Lobular vs Ductal)	977	0.827 (0.526–1.299)	0.410
PR status (Positive vs Negative)	1029	0.732 (0.523–1.024)	0.068
ER status (Positive vs Negative)	1032	0.712 (0.495–1.023)	0.066
HER2 status (Positive vs Negative)	715	1.593 (0.973–2.609)	0.064
Menopause status (Post vs Pre & Peri)	971	2.348 (1.428–3.860)	<0.001***
GNPNAT1 (High vs Low)	1082	1.570 (1.137–2.168)	0.006**

Notes: *p-value< 0.05, **p-value<0.01, ***p-value<0.001. The stages were graded according to AJCC7.

Abbreviations: GNPNAT1, glucosamine-phosphate N-acetyltransferase 1; AJCC7, American Joint Committee on Cancer 7th edition.

Table 5 Univariate COX Regression Analysis for Disease Free Survival in the TCGA Cohort

Characteristics	Total (N)	HR (95% CI)	P value
T stage (T3&4 vs T1&2)	1059	1.902 (1.174–3.081)	0.009**
N stage (N1&2and3 vs N0)	1044	3.797 (2.222–6.489)	<0.001***
M stage (M1 vs M0)	903	7.454 (3.988–13.931)	<0.001***
Pathologic stage (III&IV vs I&II)	1041	3.519 (2.274–5.444)	<0.001***
Race (Others vs Asian)	974	0.804 (0.252–2.560)	0.712
Age (>60 vs <=60)	1062	1.445 (0.941–2.219)	0.093
Histological type (Lobular vs Ductal)	958	0.471 (0.226–0.982)	0.045*
PR status (Positive vs Negative)	1010	0.519 (0.334–0.807)	0.004**
ER status (Positive vs Negative)	1013	0.559 (0.351–0.891)	0.015*
HER2 status (Positive vs Negative)	704	1.477 (0.740–2.948)	0.269
Menopause status (Post vs Pre & Peri)	961	1.560 (0.871–2.794)	0.135
GNPNAT1 (High vs Low)	1062	1.564 (1.016–2.408)	0.042*

Notes: *p-value< 0.05, **p-value<0.01, ***p-value<0.001. The stages were graded according to AJCC7.

Abbreviations: GNPNAT1, glucosamine-phosphate N-acetyltransferase 1; AJCC7, American Joint Committee on Cancer 7th edition.

Table 6 Multivariate COX Regression Analysis for Overall Survival in the TCGA Cohort

Characteristics	Total (N)	HR (95% CI)	P value
T stage (T3&4 vs T1&2)	1079	1.000 (0.609–1.643)	0.999
N stage (N1&2and3 vs N0)	1063	1.562 (1.015–2.403)	0.043*
Pathologic stage (III&IV vs I&II)	1059	1.760 (1.057–2.930)	0.030*
GNPNAT1 (High vs Low)	1082	1.422 (1.006–2.010)	0.046*

Notes: *p-value< 0.05. The stages were graded according to AJCC7.

Abbreviations: GNPNAT1, glucosamine-phosphate N-acetyltransferase 1; AJCC7, American Joint Committee on Cancer 7th edition.

Figure 4 A nomogram for predicting the 1-year, 3-year, and five-year survival probability of breast cancer patients.

Table 7 Connection Between the Expression Level of GNPNAT1 and the Immune Infiltration in the Tumor Microenvironment

Immune Cell	Spearman Correlation	P value
aDC	−0.091	0.003**
B cells	−0.147	<0.001***
CD8 T cells	−0.243	<0.001***
Cytotoxic cells	−0.241	<0.001***
DC	−0.185	<0.001***
Eosinophils	0.149	<0.001***
iDC	−0.128	<0.001***
Macrophages	−0.038	0.208
Mast cells	0.022	0.471
Neutrophils	−0.124	<0.001***
NK CD56bright cells	−0.109	<0.001***
NK CD56dim cells	−0.143	<0.001***
NK cells	−0.187	<0.001***
pDC	−0.417	<0.001***
T cells	−0.122	<0.001***
T helper cells	0.305	<0.001***
TCM	0.270	<0.001***
Tem	−0.090	0.003**
TFH	−0.064	0.034*
Tgd	0.014	0.644
Th1 cells	−0.127	<0.001***
Th17 cells	−0.002	0.939
Th2 cells	0.353	<0.001***
TReg	−0.119	<0.001***

Notes: *p-value< 0.05, **p-value<0.01, ***p-value<0.001.

Abbreviations: aDC, activated dendritic cell; DC, dendritic cell; iDC, immature dendritic cell; Macrophages; Mast cells; pDC, Plasmacytoid dendritic cell; TCM, T central memory; Tem, T effector memory; Tfh, T follicular helper; Tgd, T gamma delta; Treg, T regulatory cell.

Figure 5 Correlations between the expression of GNPNAT1 expression and immune infiltration. (A) The bar graph is about the immune infiltration level of different immune infiltration cells. (B) The lollipop figure is about the immune infiltration level of different immune infiltration cells. (C) The scatter diagrams about the immune infiltration level of different immune infiltration cells. (D) The heatmap about the correlation between different immune infiltration cells and breast cancer. *P<0.05, **p<0.01, ***p<0.001.

Figure 6 Mutation feature of GNPNAT1 in TCGA. (A) The alteration frequency GNPNAT1 with mutation type in different cancers. (B) The alteration frequency with mutation site in breast cancer. (C) The 3D structure of GNPNAT1 in breast cancers. (D) The alteration frequency GNPNAT1 with mutation type in whole cancers.

Table 8 The Basic Information of IHC Slice

Tissue Type	ID	Age	Gender
Normal	2773	23	Female
Tumor	2565	51	Female

Figure 7 GNPNAT1-related gene enrichment analysis. (A) (B) Heat map about the top 5 GNPNAT1-related gene enrichment. (C) The scatter diagrams about the top 5 GNPNAT1-related gene enrichment. ***p<0.001.

Figure 8 GNPNAT1-binding proteins enrichment analysis. (A) The difference in GNPNAT1 total protein between breast tumor and normal breast tissues. (B) The connection net of the top 50 GNPNAT1-binding proteins. (C) A Venn diagram about intersection analysis of the top 50 proteins and the top 100 genes.

Figure 9 GO and KEGG enrichment analyses of GNPNAT1. (A) GO enrichment analyses of GNPNAT1. (B) GO enrichment interactive network colored by cluster-ID (C) and P-value. (D) KEGG enrichment analyses of GNPNAT1.

Figure 10 Protein–protein interaction network and IHC slices of GNPNAT1. (A) Protein-protein interaction network of GNPNAT1. (B) IHC slices of GNPNAT1.

Figure 11 (A) Reactome mitotic telophase cytokinesis, (B) WP OMEGA9 FA synthesis, (C) Reactome cohesion loading chromatin, and (D) Reactome establishment of sister chromatid cohesion were the top four signaling pathways of GNPNAT1-related.