High expression of CDKN3 promotes migration and invasion of gastric cancer cells by regulating the p53/NF-κB signaling pathway and inhibiting cell apoptosis

doi:10.12122/j.issn.1673-4254.2025.04.21

Abstract

Abstract:

Objective To investigate the expression of CDKN3 in gastric cancer and its impact on prognosis of gastric cancer patients. Methods We analyzed CDKN3 expression in clinical specimens from 114 gastric cancer patients and assessed its association with 5-year postoperative survival of the patients. GO and KEGG enrichment analyses were used to predict the biological function and possible mechanism of CDKN3. The effects of lentivirus-mediated CDKN3 knockdown on biological behaviors of gastric cancer cells were evaluated using Transwell assay, CCK-8 assay, TUNEL staining, flow cytometry, and Western blotting. Results CDKN3 expression was significantly higher in gastric cancer tissues than in the adjacent tissues with significant correlations with CEA level, CA19-9 level, and T and N staging (P<0.05). High CDKN3 expression was an independent risk factor affecting 5-year postoperative survival of the patients and predictive for long-term prognosis (P<0.01). Enrichment analyses suggested a probable association of CDKN3 with apoptosis. In MGC-803 cells, CDKN3 knockdown significantly lowered migration and invasion capacities of the cells, while CDKN3 overexpression produced the opposite effects. TUNEL staining revealed a significantly lower level of cell apoptosis in gastric cancer tissues than in adjacent tissues (P<0.01). CDKN3 knockdown obviously inhibited proliferation and increased apoptosis of MGC-803 cells. CDKN3 overexpression down-regulated the expressions of p53, p21 and Bax and up-regulated the expressions of p-p65 and Bcl-2. Conclusion CDKN3 is highly expressed in gastric cancer tissues and affects patient prognosis. CDKN3 overexpression promotes proliferation, invasion and migration and suppressed apoptosis of gastric cancer cells possibly through the p53/NF-κB signaling pathway.

Key words: gastric cancer, CDKN3, apoptosis, proliferation, prognosis, p53/NF-κB

Yi ZHANG, Yu SHEN, Zhiqiang WAN, Song TAO, Yakui LIU, Shuanhu WANG. High expression of CDKN3 promotes migration and invasion of gastric cancer cells by regulating the p53/NF-κB signaling pathway and inhibiting cell apoptosis[J]. Journal of Southern Medical University, 2025, 45(4): 853-861.

Figures/Tables 11

Fig.1 Expression of CDKN3 in gastric cancer (GC) and adjacent tissues. A: Immunohistochemistry of CDKN3 in gastric cancer (GC) and adjacent tissues. B: Relative IOD values of CDKN3 expression. **P<0.01.

Tab.1 Relationship between the expression level of CDKN3 in GC tissues and clinicopathological parameters of the patients

Clinicopatholological parameters	n	CDKN3		χ²	P
Clinicopatholological parameters	n	Low expression (n=57)	High expression (n=57)	χ²	P
Gender
Male	81	43 (53.09%)	38 (46.91%)	1.066	0.302
Female	33	14 (42.42%)	19 (57.58%)	1.066	0.302
Age (year)
<60	46	22 (47.83%)	24 (52.17%)	0.146	0.703
≥60	68	35 (51.47%)	33 (48.53%)	0.146	0.703
CEA (μg/L)
<5	69	46 (66.67%)	23 (33.33%)	19.422	<0.001
≥5	45	11 (24.44%)	34 (75.56%)	19.422	<0.001
CA19-9 (kU/L)
<37	86	53 (61.63%)	33 (38.37%)	18.937	<0.001
≥37	28	4 (14.29%)	24 (85.71%)	18.937	<0.001
Tumor size (cm)
<5	67	38 (56.72%)	29 (43.28%)	2.932	0.087
≥5	47	19 (40.43%)	28 (59.57%)	2.932	0.087
Histological type
Adenocarcinoma	104	53 (50.96%)	51 (49.04%)	0.438	0.508
Other	10	4 (40.00%)	6 (60.00%)	0.438	0.508
Grade of differentiation
Well	10	4 (40.00%)	6 (60.00%)	4.001	0.135
Moderate	38	24 (63.16%)	14 (36.84%)
Poor	66	29 (43.94%)	37 (56.06%)
T Stage
T₁-T₂	37	24 (64.86%)	13 (35.14%)	4.842	0.028
T₃-T₄	77	33 (42.86%)	44 (57.14%)	4.842	0.028
N Stage
N₀-N₁	71	41 (57.75%)	30 (42.25%)	4.518	0.034
N₂-N₃	43	16 (37.21%)	27 (62.79%)	4.518	0.034

Fig.2 Correlation analysis between the expression level of CDKN3 in GC and the levels of CA19-9 and CEA in peripheral blood. A: Correlation analysis between CDKN3 and CA19-9. B: Correlation analysis between CDKN3 and CEA.

Fig.3 Correlation of the expression level of CDKN3 in gastric cancer tissues with long-term prognosis of the patients.

Tab.2 Univariate and multivariate analyses of the factors affecting the 5-year survival rate after radical gastrectomy for gastric cancer patients

Factors	Univariate analysis		Multivariate analysis
Factors	Log-rank χ²	P	HR	95% CI	P
Gender (female vs male)	0.108	0.742	-	-	-
Age (≥60 years vs <60 years)	0.162	0.687	-	-	-
CDKN3 expression (high vs low)	34.036	<0.001	2.819	1.396-5.692	0.004
CEA(≥5 μg/L vs <5 μg/L)	20.961	<0.001	1.954	1.084-3.522	0.026
CA19-9 (≥37 kU/L vs <37 kU/L)	22.695	<0.001	1.847	1.048-3.257	0.034
Tumor size (≥5 cm vs <5 cm)	6.139	0.013	0.978	0.552-1.730	0.938
Histological type (other vs adenocarcinoma)	0.309	0.578	-	-	-
Grade of differentiation (well vs moderate vs poor)	5.623	0.060	-	-	-
T Stage (T₃-T₄vs T₁-T₂)	13.586	<0.001	2.438	1.160-5.120	0.019
N Stage (N₂-N₃vs N₀-N₁)	20.178	<0.001	2.099	1.177-3.744	0.012
HR: Hazard ratio; CI: Confidence interval.

Fig.4 Assessment of the prognostic value of CDKN3 for gastric cancer patients.

Fig.5 CDKN3 overexpression promotes gastric cancer cell migration and invasion. A,B: The expression of CDKN3 protein after lentivirus transfection was detected by Western blotting. C:Transwell experiment for analyzing the impact of CDKN3 on the migration and invasion abilities of MGC803 cells. D: Quantitative analysis of the number of migrated cells. E: Quantitative analysis of the number of invaded cells. *P<0.05, **P<0.01 vs control.

Fig.6 KEGG and GO enrichment analysis of CDKN3. A: KEGG analysis of CDKN3 in gastric cancer. B: GO analysis of CDKN3 in gastric cancer.

Fig.7 TUNEL staining was used to detect cell apoptosis in gastric cancer and adjacent tissues. A,B: TUNEL staining and scoring of apoptosis in gastric cancer tissues sections and adjacent tissues sections. **P<0.01vs gastric cancer. GC: Gastric cancer.

Fig.8 Effect of CDKN3 on apoptosis and proliferation of MGC803 cells. A,B: Cell apoptosis rate detected by flow cytometry after lentivirus transfection. C: Expression of Bcl-2 and Bax in MGC803 cells. D: Quantitative analysis of Bcl-2/Bax ratio. E: Effect of CDKN3 on proliferation of MGC-803 cells. *P<0.05, **P<0.01 vs control.

Fig.9 Effect of CDKN3 on p53/NF-κB signaling in MGC803 cells. A: Expression of p53, p21, p65 and p-p65 in MGC803 cells. B, C: Quantitative analysis of p53, p21, p65 and p-p65 ratio. *P<0.05, **P<0.01 vs control.

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