ZWINT promotes the proliferation, migration, and invasion of cervical cancer cells by regulating the p53/p21 signaling pathway

Zhe Ma; Yufei Cai; Chenchen Tian

doi:10.4103/cjop.CJOP-D-23-00001

ORIGINAL ARTICLE

Year : 2023 | Volume : 66 | Issue : 5 | Page : 372-378

ZWINT promotes the proliferation, migration, and invasion of cervical cancer cells by regulating the p53/p21 signaling pathway

Zhe Ma, Yufei Cai, Chenchen Tian
Department of Gynaecology and Obstetrics, Affiliated Hospital of Beihua University, Jilin, Jilin Province, China

Date of Submission	18-Jan-2023
Date of Decision	28-Apr-2023
Date of Acceptance	04-May-2023
Date of Web Publication	31-Aug-2023

Correspondence Address:
Dr. Yufei Cai
Department of Gynaecology and Obstetrics, Affiliated Hospital of Beihua University, No. 12, Jiefang Middle Road, Jilin, Jilin Province
China

Source of Support: None, Conflict of Interest: None

DOI: 10.4103/cjop.CJOP-D-23-00001

Abstract

Cervical cancer leads to 300,000 deaths annually and the mechanism of cervical carcinogenesis remains unclear. Zeste White 10-interacting kinetochore protein (ZWINT) is uniquely elevated in malignancies, promoting proliferation, migration, and colony formation of cancer cells. To investigate the role of ZWINT in proliferation, migration, invasion of cervical cancer, and evaluate the potential ability of ZWINT as a therapeutic target. First, ZWINT expression in cervical cancer was analyzed using the bioinformatic methods and assessed in several cervical cancer cell lines. The cell viability and colony formation assays were used to evaluate cell proliferation. Then, transwell assay was performed to investigate cell migration and invasion. Moreover, western blot was used to measure the expression level of ZWINT, matrix metalloproteinase 9 (MMP-9), N-cadherin, E-cadherin, p53 and p21 in CaSki and HeLa cells with ZWINT overexpression or knockdown. The bioinformatic analysis and western blot assay revealed the expression of ZWINT was significantly increased in cervical cancer. The cell viability and colony formation analysis illustrated that cell proliferation could be promoted by ZWINT overexpression and suppressed by ZWINT knockdown. Moreover, ZWINT promoted migration and invasion of CaSki and HeLa cells, through regulating the expression of MMP-9, N-cadherin, and E-cadherin. Furthermore, ZWINT attenuated the expression of p53 and p21 in cervical cancer cells. In summary, ZWINT functions in promoting cell proliferation, migration, and invasion of cervical cancer cells by suppressing p53/p21 signaling pathway, which indicated ZWINT is a potential therapeutic target for cervical cancer treatment.

Keywords: Cervical cancer, invasion, migration, p21, p53, proliferation, Zeste White 10-interacting kinetochore protein

How to cite this article:
Ma Z, Cai Y, Tian C. ZWINT promotes the proliferation, migration, and invasion of cervical cancer cells by regulating the p53/p21 signaling pathway. Chin J Physiol 2023;66:372-8

How to cite this URL:
Ma Z, Cai Y, Tian C. ZWINT promotes the proliferation, migration, and invasion of cervical cancer cells by regulating the p53/p21 signaling pathway. Chin J Physiol [serial online] 2023 [cited 2023 Dec 4];66:372-8. Available from: https://www.cjphysiology.org/text.asp?2023/66/5/372/384767

Introduction

Cervical cancer is one of the most common malignancies which is seriously threatening the health for women.^[1] So far, the standard strategies for cervical cancer patients treatment contain surgical resection, chemotherapy, and radiotherapy.^[2] Intensified histopathological examination and improvements in early diagnosis have strongly reduced the mortality of cervical cancer over the past decades.^[3] However, there are still approximately 300,000 deaths each year.^[4] Although some molecular mechanisms of cervical carcinogenesis have been studied, the specific process of cervical carcinogenesis remains mostly unclear.^[5]

Zeste White 10-interacting kinetochore protein (ZWINT) is a component of centromeric complex, which is involved in centromeric function and cell growth.^[6] ZWINT interacts with another centromeric protein, Zeste White 10 (ZW10), which makes ZWINT to regulate the interaction of ZW10 with centromeres.^[7] It has been reported that ZWINT serves as a biomarker for cancers because its expression is uniquely elevated in human malignancies including glioblastoma, breast, ovarian, and lung cancers.^{[8],[9],[10],[11]} Furthermore, knockdown of ZWINT weakened the proliferation, migration, invasion, and colony-forming abilities of pancreatic cancer cells.^[12] Moreover, ZWINT promoted p53 ubiquitination to regulate pancreatic cancer cell proliferation.^[13] Therefore, ZWINT plays a key role in cancers.

However, the effect of ZWINT on cervical cancer is still unclear. In this work, the expression of ZWINT in cervical cancer was analyzed using online database, and its expression was measured in several cervical cancer cell lines. Further studies were performed to evaluate the role of ZWINT in proliferation, migration, and invasion in cervical cancer cells. Moreover, the regulation of p53/p21 signaling pathway by ZWINT was investigated as well.

Materials and Methods

Cell culture

HcerEpic is a normal human cervical epithelial cell and CaSki is a cervical cancer cell line, which were cultivated in RPMI-1640 medium plus 10% fetal bovine serum (FBS) (Gibco, Grand Island, NY, USA). Another three cervical cancer cell lines including HeLa, SiHa, and C-33A were cultivated in DMEM containing 10% FBS, respectively. All these cells were housed in a 37°C incubator supplied with 5% CO₂.

Overexpression or knockdown of ZWINT

ZWINT coding sequence was amplified using forward primer: 5'-CGCGGATCCACCATGGAGGCAGCGGAGACA-3' and reverse primer: 5'-CGCGTCGACTCATGGCAAATT TACATC-3', and the target fragment was inserted into pcDNA3.1 vector. The ZWINT-specific siRNA (5'-CCA GAGGAAACGGACACAA-3' and 5'-UUGUGU CCGUUUCCUCUGG-3') was used for ZWINT knockdown. The overexpression plasmid of ZWINT or siRNA were transfected into CaSki or HeLa cells with Lipofectamine 3000 kit (Invitrogen, CA, USA). The expression level of ZWINT was measured 48 h later to assess the efficiency of overexpression or knockdown.

Bioinformatic analysis

The analysis of ZWINT expression was performed in cervical cancer tissues via Gene Expression Profiling Interactive Analysis (GEPIA) database (http://gepia.cancer-pku.cn/) and The Cancer Genome Atlas (TCGA) on the University of Alabama at Birmingham CANcer data analysis portal (UALCAN) which is available at http://ualcan.path.uab.edu.

Western blot

The cellular protein was extracted by RIPA lysis buffer (89901, Thermo Scientific, Carlsbad, CA, USA). The lysates were processed for immunoblot with the primary antibodies listed in [Table 1], followed by incubation with HRP-conjugated goat anti-rabbit IgG (B900210, ProteinTech Group; 1:5000). Finally, the target bands were visualized with ECL reagents (Solarbio Life Sciences, Beijing, China). For the quantification of western blot signal, the relative intensity of each band was measured by ImageJ software (USA), and the relative expression of target proteins was normalized to GAPDH, respectively.

Table 1: Antibodies information

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Cell proliferation assay

CaSki or HeLa cells were seeded into 96-well plates and transfected with ZWINT overexpression plasmid or siRNA of ZWINT. The supernatant was replaced with new medium supplemented with 10 μl CCK-8 solution (Dojindo Molecular Technologies, Japan) in each well and the cells were cultured in incubator with 5% CO₂ at 37°C for 0, 24, 48, 72 and 96 h. Absorbance was detected at 450 nm wavelength, and the cell viability ratio was calculated.

Colony formation assay

Single cell of CaSki or HeLa was suspended and seeded in separated 6-well plates. Each well was transfected with ZWINT overexpression plasmid or siRNA of ZWINT. When the ocular cell clusters were formed, they were fixed with 4% paraformaldehyde followed by 0.1% violet crystal staining (548-62-9, Sigma-Aldrich, Shanghai, China). The numbers of visible colonies were counted.

Analysis of cell cycle by flow cytometry

HeLa cells treated with ZWINT overexpression or knockdown were fixed with 70% ethanol and incubated with 100 μL propidium iodide solution (500 μg/mL) for 15 min in the dark. The cell cycle was then analyzed by FACS LSR II (BD Biosciences, San Jose, CA, USA). The data were analyzed further by FlowJo software.

Transwell assay

CaSki or HeLa cells were transfected with ZWINT overexpression plasmid or siRNA of ZWINT and seeded into an invasion chamber, in which the lower chamber was supplemented with media containing 10% FBS but the upper one was filled with serum-free medium. Then, the cells which could passed through polycarbonate membrane were stained with 0.1% crystal violet. For each sample, more than 10 fields were selected randomly and the mean number of stained cells was counted.

Matrix metalloproteinase-9 activity assays

Matrix metalloproteinase 9 (MMP-9) activity was analyzed using QuickZyme MMP-9 Activity Assay Kit (QZBMMP9H, BioVendor, Czech Republic) following the manual instruction. Briefly, the supernatant of CaSki or HeLa cells was collected. 100 μl supernatant was added to 96-well plate and mixed with 50 μl APMA solution (1 mM) provided in the kit. Then, 50 μl asssy buffer was added into each well, incubated at 37°C for 1.5 h, and followed by 50 μl of detection reagent. The OD value at 405 nm was measured at 0, 1, and 4 h, respectively.

Quantification and statistical analysis

Data were presented as mean ± standard deviation from three biological replicates, and the differences between any two groups were calculated by unpaired t-tests. Multiple group comparisons were analyzed with ANOVA. *, ^#P < 0.05, **, ^##P < 0.01, ***, ^###P < 0.001 indicate the differences were statistically significant.

Results

ZWINT is upregulated in cervical cancer cells

To evaluate the ZWINT expression level in cervical cancer, an analysis from GEPIA databank was performed which showed that ZWINT expression in cervical cancer tissues was elevated [Figure 1]a. Another analysis of RNA-seq data derived from TCGA on UALCAN web portal was also completed, which revealed that the expression of ZWINT was upregulated in 305 samples of primary tumors [Figure 1]b. To verify that ZWINT expression was upregulated in cervical cancer, western blot was performed to test ZWINT expression in four cell lines of cervical cancer including CaSki, HeLa, SiHa, C-33A, and one normal cell line HcerEpic. The data revealed that the expression of ZWINT in CaSki, HeLa, SiHa, and C-33A was much higher than that in HcerEpic [Figure 1]c. To sum up, ZWINT was highly expressed in cervical cancer.

Figure 1: The expression of ZWINT is upregulated in cervical cancer cells. (a) Analysis from GEPIA databank showed ZWINT expression in cervical cancer (CESC) tissues was upregulated than in normal tissues. Totally, 306 tumor samples were collected for this analysis. (b) Analysis of RNA-seq data derived from TCGA on UALCAN web portal indicated the expression of ZWINT was increased in 305 samples of primary tumors compared to normal tissues. (c) Western blot was performed to measure the ZWINT protein level in the four cell lines of cervical cancer including CaSki, HeLa, SiHa, C-33A, and the normal cell line HcerEpic. At least three repeats were used for statistical analysis. *P < 0.05, ***P < 0.001. ZWINT: Zeste White 10-interacting kinetochore protein, GEPIA: Gene Expression Profiling Interactive Analysis, TCGA: The Cancer Genome Atlas.

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ZWINT promotes cervical cancer cell proliferation

Given that increased expression of ZWINT in cervical cancer, the cell proliferation in cervical cancer cells with ZWINT overexpression or knockdown was studied. Specifically, CaSki and HeLa cells were selected to be transfected with ZWINT overexpression (marked as ZWINT) or siRNA of ZWINT (marked as siZWINT). Firstly, western blot was performed to evaluate ZWINT expression, which showed that ZWINT expression was elevated in ZWINT group compared with vector or control group, while reduced in siZWINT group compared to siNC [Figure 2]a. Furthermore, the data of cell viability illustrated that ZWINT overexpression in ZWINT group efficiently promoted cell proliferation, but the cell proliferation was reduced significantly in siZWINT group compared to siNC [Figure 2]b. Moreover, colony formation assay was performed to test cell proliferation. As shown in CaSki and Hela cells, ZWINT overexpression increased the count of colony formation, while siZWINT potently reduced colony formation in contrast to siNC [Figure 2]c. Furthermore, the effect of ZWINT on cell cycle was studied using flow cytometry. The result revealed that ZWINT knockdown increased proportion at G1 phase and reduced S phase proportion, indicating that ZWINT knockdown led to cell cycle arrest at G1 phase [Supplementary Figure 1]b. Therefore, ZWINT played a positive role in proliferation of cervical cancer cells.

Figure 2: ZWINT promotes cervical cancer cell proliferation. (a) Western blot was performed to test the ZWINT expression in CaSki and HeLa cells. (b) The cell viability of CaSki and HeLa was studied to assess the effect of ZWINT overexpression or knockdown on cell proliferation. (c) Colony formation assay was performed to evaluate cell proliferation. Control, no transfection; Vector, transfected with empty plasmid; ZWINT, transfected with ZWINT overexpression plasmid; siZWINT, ZWINT knockdown; siNC, negative control to siZWINT. Three replicate experiments were conducted for statistical analysis. *ZWINT versus Vector, ^#siZWINT versus siNC, *, ^#P < 0.05, **, ^##P < 0.01, ***, ^###P < 0.001. ZWINT: Zeste White 10-interacting kinetochore protein.

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ZWINT promotes migration and invasion of cervical cancer cells

The effect of ZWINT on migration and invasion was investigated here. The data from the transwell assay revealed that ZWINT overexpression facilitated the migration and invasion than the control; however, ZWINT knockdown by siZWINT significantly weakened the migration and invasion of CaSki or HeLa cells compared with the shNC group [Figure 3]a.

Figure 3: ZWINT promotes migration and invasion of cervical cancer cells. (a) Transwell assay was performed to investigate the effect of ZWINT expression on migration and invasion of CaSki or HeLa cells. (b) Western blot was performed to evaluate the expression of MMP-9, N-cadherin and E-cadherin. (c) MMP-9 Activity Assay was performed to analyze MMP-9 activity of CaSki and HeLa cells. Vector, negative control for ZWINT overexpression; ZWINT, ZWINT overexpression; siZWINT, ZWINT knockdown; siNC, negative control to siZWINT. The statistical analysis was based on at least three replicate data. *ZWINT versus Vector, ^#siZWINT versus siNC, *, ^#P < 0.05, **, ^##P < 0.01, ***, ^###P < 0.001. ZWINT: Zeste White 10-interacting kinetochore protein, MMP-9: Matrix metalloproteinase 9.

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Further, western blot was performed to evaluate the expression of migration-related regulatory proteins including MMP-9, N-cadherin and E-cadherin. The results demonstrated that these proteins were regulated by ZWINT expression in CaSki and HeLa cells. For MMP-9 and N-cadherin, their expressions were elevated in ZWINT overexpression group, but decreased in siZWINT-transfected cells. For E-cadherin, its expression was reduced in ZWINT overexpression group, but increased in siZWINT group. High expression of MMP-9 and N-cadherin, or low level of E-cadherin induced by ZWINT overexpression facilitated the cell migration and invasion [Figure 3]b. Furthermore, the MMP-9 activity of cell supernatant was analyzed, and the results revealed that MMP-9 activity was elevated by ZWINT overexpression, while was reduced by ZWINT knockdown in CaSki and HeLa cells. Thus, ZWINT played a positive role in migration and invasion of cervical cancer cells.

ZWINT regulates the p53/p21 signaling pathway

It has been reported that ZWINT regulates cell proliferation through targeting p53 in pancreatic cancer.^[13] So, it is necessary to clarify whether ZWINT also regulates p53 in cervical cancer cells. The protein level of p53 and p21 was measured by western blot. Consistent with what was reported in pancreatic cancer cells, overexpression of ZWINT efficiently decreased the protein level of p53 and p21, while knockdown of ZWINT in siZWINT significantly elevated p53 and p21 expression in CaSki or Hela cells compared to siNC [Figure 4]. In addition, the protein level of murine double minute 2 (MDM2) that is another key molecule in p53/p21 pathway was evaluated using western blot, which showed knockdown of ZWINT significantly decreased MDM2 expression in HeLa cells [Supplementary Figure 1]a. In summary, ZWINT regulated p53/p21 signaling pathway in cervical cancer cells.

Figure 4: ZWINT regulates the p53/p21 signaling pathway. Western blot was used to measure the effect of ZWINT on the expression of p53 and p21 in CaSki or HeLa cells. Control, no transfection; Vector, transfected with empty plasmid; ZWINT, transfected with ZWINT overexpression plasmid; siZWINT, ZWINT knockdown; siNC, negative control to siZWINT. The statistical analysis contains three replicate data. *ZWINT versus Vector, ^#siZWINT versus siNC, *P < 0.05, ***, ^###P < 0.001. ZWINT: Zeste White 10-interacting kinetochore protein.

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ZWINT knockdown inhibits proliferation, migration and invasion of cervical cancer cell by upregulating p53/p21 pathway

Given that ZWINT is negatively associated with p53 expression, it is necessary to investigate the cell growth, migration and invasion in cells with double knockdown of ZWINT and p53. Western blots were performed to measure the expression level of p53 and p21 in HeLa cells, and the data showed that p53 and p21 expression was reduced in cells with double knockdown of ZWINT and p53 (siZWINT + sip53) compared to siZWINT [Figure 5]a. Then, the cell viability curves demonstrated that cell proliferation in cells transfected with siZWINT + sip53 was enhanced compared to cells transfected with siZWINT, which was recovered to the level similar with siNC [Figure 5]b. Meanwhile, the migration and invasion of cells in siZWINT + sip53 group was significantly increased compared to that in siZWINT group [Figure 5]c. All the data above illustrated that ZWINT did lose the function in HeLa cells once p53 was knockdown, meaning that knockdown of ZWINT inhibited proliferation, migration and invasion of cervical cancer cells through regulating p53/p21 pathway.

Figure 5: ZWINT knockdown inhibits proliferation, migration, and invasion of cervical cancer cell by regulating p53/p21 pathway. (a) Western blots were performed to measure the expression level of p53 and p21 in HeLa cells. (b) The cell viability of HeLa cells with knockdown of ZWINT and p53. (c) Transwell assays were conducted to evaluate the migration and invasion of HeLa cells with ZWINT and p53 knockdown. *siZWINT versus siNC, ^#siZWINT + sip53 versus siZWINT, *, ^#P < 0.05, **, ^##P < 0.01, ***, ^###P < 0.001. ZWINT: Zeste White 10-interacting kinetochore protein.

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Discussion

Cervical cancer is one the most common malignancies which is seriously threatening the health for women.^[1] It is necessary to clarify the cervical carcinogenesis or discover tumor suppressor genes for clinical treatment.^[4] ZWINT expression was elevated in some human malignancies including glioblastoma, breast, ovarian, and lung cancers.^{[8],[9],[10],[11]} Predictably, the bioinformatics analysis showed the expression of ZWINT was upregulated in cervical cancer. And western blot on several cervical cancer cell lines confirmed the expression level of ZWINT was significantly elevated in cervical cancer cells [Figure 3]c. Thus, it is reasonable to conclude that ZWINT is highly expressed in cervical cancer.

Previous studies reported that overexpression of ZWINT promoted the proliferation and colony-forming abilities of pancreatic cancer.^[12] Similar study was performed in this work, the cell viability and colony formation analysis illustrated that cell proliferation could be promoted by ZWINT overexpression and suppressed by ZWINT knockdown. As is known that ZWINT promoted p53 ubiquitination to mediate pancreatic cancer cell proliferation.^[13] So, it makes sense that ZWINT promotes proliferation of cervical cancer cells.

The migration and invasion were also investigated in this work. Transwell assay revealed that ZWINT played a positive role in migration and invasion of CaSki or HeLa cells. And the expression of MMP-9, N-cadherin and E-cadherin was also closely associated with ZWINT. It has been reported that ZWINT overexpression promoted pancreatic cancer cell migration, invasion and tumor development.^[12] MMP-9, N-cadherin and E-cadherin are the markers of epithelial-mesenchymal transition which is important for cancer cells migration and invasion.^[14] As is reported that ZWINT regulated the downstream molecules associated with cell adhesion and extracellular matrix including MMP-9, E-cadherin and N-cadherin, the expression levels of which were increased in cancer cells with ZWINT knockdown.^[15] ZWINT might regulate the expression of these cellular proteins via affecting cell cycles closely related with spindle assembly checkpoint.^[16] It is easily to conclude that ZWINT promotes migration and invasion of cervical cells via regulating expression of MMP-9, N-cadherin and E-cadherin.

Previous report showed that p53 signal pathway is the key regulator in ZWINT function, and ZWINT interacts with p53 and thereby regulating pancreatic cancer cell proliferation via promoting p53 ubiquitination, and they presumed ZWINT weakened p53 protein stability to accelerate its degradation via restraining MDM2.^[13] p53 overexpression or aberrant activity has been considered as a molecular hallmark of cancer. p53 could directly regulated p21 activity which arrested cell cycle at G1/S phase via suppressing cyclin D1 and cyclin/cyclin-dependent kinases (CDK).^[17] In melanoma cells, centrosomal protein 55 reduced p21 expression and so promoted Cyclin D1 and CDK1 expression, antagonizing cell cycle arrest.^[18] In pancreatic cancer cells, ZWINT reduced p21 expression and transcriptional activity, which contributed to pancreatic cancer cell progression by repressing p53/p21 signaling pathway.^[13] Consistent with previous reports, western blot in this study certified that ZWINT attenuated the expression of p53 and p21 in cervical cancer cells, indicating that ZWINT functions in promoting cell proliferation, migration and invasion in cervical cancer by inhibiting p53/p21 signaling pathway.

Post-translation modifications including acetylation, ubiquitylation and phosphorylation might affect transcriptionally activity and expression of p53. For example, sevoflurane ameliorated lipopolysaccharide (LPS) induced inflammatory injury of HK-2 cells through Sirtuin1 (SIRT1)/NF-κB pathway, as LPS inhibited SIRT1 but increased expression of acetylated p53.^[19] This work clarified that p53 protein level was attenuated by ZWINT in cervical cells. However, the specific mechanism remains unclear. More efforts should be concentrated on elucidating these details such as which regulators catalyzing p53 modifications or pathways transporting cellular degraded p53.

Conclusion

In summary, this study demonstrated ZWINT plays a vital role in promoting cell proliferation, migration and invasion in cervical cancer cells by inhibiting p53/p21 signaling pathway. Therefore, ZWINT is expected to be considered as a novel therapeutic target for cervical cancer treatment.

Acknowledgements

Thanks to the editor of Englishgo for editing the grammar and language of the manuscript.

Data availability statement

The authors declare that all data supporting the findings of this study are available within the paper and any raw data can be obtained from the corresponding author upon request.

Author contributions

Zhe Ma, Yufei Cai and Chenchen Tian designed the experiments, carried them out, analyzed and interpreted the data, and prepared the manuscript. All authors have read and approved the manuscript.

Financial support and sponsorship

This work was supported by Health and Hygiene Technology Innovation Project of Jilin Province (Grant No. 2020J008).

Conflicts of interest

There are no conflicts of interest.

References

1.	Jemal A, Bray F, Center MM, Ferlay J, Ward E, Forman D. Global cancer statistics. CA Cancer J Clin 2011;61:69-90.
2.	Brucker SY, Ulrich UA. Surgical treatment of early-stage cervical cancer. Oncol Res Treat 2016;39:508-14.
3.	Bansal N, Herzog TJ, Shaw RE, Burke WM, Deutsch I, Wright JD. Primary therapy for early-stage cervical cancer: Radical hysterectomy versus radiation. Am J Obstet Gynecol 2009;201:485.e1-9.
4.	Ferlay J, Soerjomataram I, Dikshit R, Eser S, Mathers C, Rebelo M, et al. Cancer incidence and mortality worldwide: Sources, methods and major patterns in GLOBOCAN 2012. Int J Cancer 2015;136:E359-86.
5.	Sun Q, Yang Z, Li P, Wang X, Sun L, Wang S, et al. A novel miRNA identified in GRSF1 complex drives the metastasis via the PIK3R3/AKT/NF-κB and TIMP3/MMP9 pathways in cervical cancer cells. Cell Death Dis 2019;10:636.
6.	Peng F, Li Q, Niu SQ, Shen GP, Luo Y, Chen M, et al. ZWINT is the next potential target for lung cancer therapy. J Cancer Res Clin Oncol 2019;145:661-73.
7.	Woo Seo D, Yeop You S, Chung WJ, Cho DH, Kim JS, Su Oh J. Zwint-1 is required for spindle assembly checkpoint function and kinetochore-microtubule attachment during oocyte meiosis. Sci Rep 2015;5:15431.
8.	Yang L, Han N, Zhang X, Zhou Y, Chen R, Zhang M. ZWINT: A potential therapeutic biomarker in patients with glioblastoma correlates with cell proliferation and invasion. Oncol Rep 2020;43:1831-44.
9.	Zhou G, Shen M, Zhang Z. ZW10 binding factor (ZWINT), a direct target of mir-204, predicts poor survival and promotes proliferation in breast cancer. Med Sci Monit 2020;26:e921659.
10.	Zhao S, Yu M. Identification of MMP1 as a potential prognostic biomarker and correlating with immune infiltrates in cervical squamous cell carcinoma. DNA Cell Biol 2020;39:255-72.
11.	Yi M, Li T, Qin S, Yu S, Chu Q, Li A, et al. Identifying tumorigenesis and prognosis-related genes of lung adenocarcinoma: Based on weighted gene coexpression network analysis. Biomed Res Int 2020;2020:4169691.
12.	Kim JH, Youn Y, Lee JC, Kim J, Hwang JH. Involvement of the NF-κB signaling pathway in proliferation and invasion inhibited by Zwint-1 deficiency in pancreatic cancer cells. J Cancer 2020;11:5601-11.
13.	Chen P, He Z, Wang J, Xu J, Jiang X, Chen Y, et al. Hypoxia-induced ZWINT mediates pancreatic cancer proliferation by interacting with p53/p21. Front Cell Dev Biol 2021;9:682131.
14.	Yun JA, Kim SH, Hong HK, Yun SH, Kim HC, Chun HK, et al. Loss of E-cadherin expression is associated with a poor prognosis in stage III colorectal cancer. Oncology 2014;86:318-28.
15.	Mou K, Zhang J, Mu X, Wang L, Liu W, Ge R. Zwint facilitates melanoma progression by promoting c-Myc expression. Exp Ther Med 2021;22:818.
16.	Ying H, Xu Z, Chen M, Zhou S, Liang X, Cai X. Overexpression of Zwint predicts poor prognosis and promotes the proliferation of hepatocellular carcinoma by regulating cell-cycle-related proteins. Onco Targets Ther 2018;11:689-702.
17.	Wagner KD, Wagner N. The senescence markers p16INK4A, p14ARF/p19ARF, and p21 in organ development and homeostasis. Cells 2022;11:1966.
18.	Hua H, Yang H. Centrosomal protein 55 enhances melanoma cell proliferation, invasion and migration through PI3K/AKT signaling. Trop J Pharm Res 2022;21:2065-70.
19.	Peipei W, Ping W, Miaomiao Y, Shuo W. Sevoflurane ameliorates LPS-induced inflammatory injury of HK-2 cells through Sirtuin1/NF-κB pathway. Allergol Immunopathol (Madr) 2022;50:115-23.