Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2023 Mar 16;15(6):1801.
doi: 10.3390/cancers15061801.

PRAME Promotes Cervical Cancer Proliferation and Migration via Wnt/β-Catenin Pathway Regulation

Xin Chen  1 , Mengying Jiang  1 , Shengjie Zhou  2 , Hong Chen  1 , Gendi Song  1 , Yichen Wu  1 , Xueqiong Zhu  1   2
Affiliations

PRAME Promotes Cervical Cancer Proliferation and Migration via Wnt/β-Catenin Pathway Regulation

Xin Chen et al. Cancers (Basel). .

Abstract

A significant burden is placed on the lives of females due to cervical cancer, which is currently the leading cause of cancer death among women. Preferentially expressed antigen in melanoma (PRAME) belongs to the CTA gene family and was found to be abnormally expressed among different types of cancers. Our previous research also indicated that PRAME was highly expressed in cervical cancer compared with normal tissues. However, the roles and detailed mechanisms of PRAME have not been explored in cervical cancer. In the present study, the expression of PRAME in cervical tissues and cells was detected by immunohistochemistry (IHC), qRT-PCR, and Western blotting. Additionally, CCK-8, BrdU, scratch, transwell, and flow cytometry assays were conducted to explore the function of PRAME in regulating the malignant biological behaviors of cervical cancer cells. Nude mice were used to confirm the role of PRAME in tumor growth in vivo. Furthermore, the Wnt inhibitor MSAB was used to verify the role of PRAME in regulating the Wnt/β-catenin pathway both in vitro and in vivo. The results of IHC, qRT-PCR, and Western blotting showed that PRAME was highly expressed in cervical cancer tissues and cells. PRAME knockdown attenuated cell growth, migration, and invasion; induced G0/G1 arrest; and increased cell apoptosis in C33A and SiHa cells through Wnt/β-catenin signaling regulation. However, the upregulation of PRAME exhibited the opposite effects accordingly, which could be partly reversed via MSAB treatment. The growth rate of xenograft tumors was enhanced when PRAME was overexpressed via Wnt/β-catenin signaling activation. Taken together, PRAME is associated with cervical cancer occurrence and progression mediated by Wnt/β-catenin signaling, suggesting that PRAME might be a factor in manipulating cervical carcinogenesis and a potential therapeutic target.

Keywords: PRAME; Wnt/β-catenin signaling; cervical cancer; proliferation; tumorigenesis.

PubMed Disclaimer

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Differential expression of PRAME in cervical cancer and normal cervix, and the role of PRAME expression in cell viability and proliferation in cervical cancer cells. (A) The expression of PRAME protein in adjacent normal tissues and cervical cancer tissues as shown by IHC staining. (B) The IHC score of PRAME protein. (Adjacent normal tissues: 22 cases, Cervical cancer tissues: 22 cases). (C) The expression of PRAME mRNA in normal cervical epithelium cells and cervical cancer cells as shown by qRT-PCR. (D) The expression of PRAME protein in normal cervical epithelium cells and cervical cancer cells as shown by Western blotting. (E) The quantitative analysis of (D). (F) PRAME-silenced C33A and SiHa cells were constructed via the two shPRAME sequences and confirmed by Western blotting. (G) The effect of PRAME knockdown on cell viability in C33A and SiHa cells was detected via CCK-8 assay. (H) The effect of PRAME knockdown on cell proliferation in C33A and SiHa cells was measured via BrdU assay. (I) The quantitative results of (H). (J) PRAME-overexpressed C33A and SiHa cells were constructed and confirmed via Western blotting. (K) The role of PRAME overexpression in cell viability in C33A and SiHa cells was detected via CCK-8 assay. (L) The role of PRAME overexpression on cell proliferation in C33A and SiHa cells was measured via BrdU assay. (M) The quantitative results for (L). * p < 0.05, ** p < 0.01, *** p < 0.001.
Figure 2
Figure 2
The effect of PRAME expression on cell cycle distribution of C33A and SiHa cells. The cell cycle profiles were analyzed and exported by FlowJo. The bimodal images represented the proportion of cells at different phases of the cell cycle distribution, with purple peaks representing the cells in G0/G1 phase, green peaks representing the cells in G2/M phase, and the cells in S phase labeled yellow. (A) The cell cycle profiles in PRAME-silenced C33A and SiHa cells. (B) The quantitative results for (A). (C) The cell cycle profiles in PRAME-overexpressed C33A and SiHa cells. (D) The quantitative results for (C). * p < 0.05, ** p < 0.01, *** p < 0.001.
Figure 3
Figure 3
The effect of PRAME expression on cell apoptosis. The sum of cells in the upper right quadrant (UR, late apoptotic cells) and the lower right quadrant (LR, early apoptotic cells) was considered to be the apoptotic population. (A) The percentage of apoptotic cells in PRAME-silenced C33A and SiHa cells. (B) The quantitative results for (A). (C) The percentage of apoptotic cells in PRAME-overexpressed C33A and SiHa cells. (D) The quantitative results for (C). ** p < 0.01, *** p < 0.001.
Figure 4
Figure 4
The role of PRAME expression on cell migration and invasion (A,B). The wound-healing rate in PRAME-silenced C33A and SiHa cells. (C,D) The quantitative results for (A,B). (E,F) The wound-healing rate in PRAME-overexpressed C33A and SiHa cells. (G,H) The quantitative results for (E,F). (I) The number of migratory and invasive cells in PRAME-knockdown SiHa cells. (J) The quantitative results for (I). (K) The number of migratory and invasive cells in PRAME-overexpressed SiHa cells. (L) The quantitative results for (K). * p < 0.05, *** p < 0.001.
Figure 5
Figure 5
Regulation of EMT-related proteins and Wnt/β-catenin pathway by PRAME expression in cervical cancer cells. (A) The change in E-cadherin and N-cadherin expression in PRAME-knockdown SiHa and PRAME-overexpressed C33A cells. (B) Western blotting analysis of Wnt/β-catenin signaling pathway-related protein expression after PRAME knockdown in SiHa cells and PRAME overexpression in C33A cells. (C) Western blotting analysis of Wnt3a and β-catenin expression in PRAME-overexpressed C33A and SiHa cells after MSAB treatment. The results of gray value quantification are displayed below the corresponding bands. (D) The cell viability in PRAME-overexpressed and control C33A and SiHa cells with MSAB treatment. (E) Left panel: The cell apoptosis in PRAME-overexpressed and control C33A and SiHa cells with MSAB treatment. Right panel: The quantitative results. *** p < 0.001.
Figure 6
Figure 6
The change in migratory and invasive abilities of PRAME-overexpressed C33A and SiHa cells after MSAB treatment. (A,B) The wound-healing rate of control C33A and SiHa cells as well as PRAME-overexpressed C33A and SiHa cells after treatment with or without MSAB. (C) The quantitative results for (A). (D) The quantitative results for (B). (E) The number of migratory and invasive cells in control SiHa groups, PRAME-overexpressed SiHa groups, control SiHa groups with MSAB treatment, and PRAME-overexpressed SiHa groups with MSAB treatment. (F) The quantitative results for (E). * p < 0.05, ** p < 0.01, *** p < 0.001.
Figure 7
Figure 7
The effect of PRAME on tumor growth via Wnt/β-catenin pathway in vivo. (A) The image of xenografts injected with PRAME-knockdown SiHa cells and control SiHa cells. (B) The weight of the xenografts in PRAME-knockdown group and control group. (C) The growth curves of xenograft tumors of PRAME-knockdown group and control group. (D) The immunofluorescence staining of Ki67 in tumor slices. (E) The immunofluorescence staining of TUNEL assay in tumor slices. (F) The image of xenografts injected with PRAME-overexpressed C33A cells and control C33A cells after the MSAB therapy. (Tumor groups as G1: Control cDNA+DMSO, G2: PRAME cDNA+DMSO, G3: Control cDNA+MSAB, and G4: PRAME cDNA+MSAB). (G) The weight of the xenografts in PRAME-overexpressed group and control group with or without MSAB therapy. (H) The growth curves of the xenografts in PRAME-overexpressed group and control group with or without MSAB therapy. (I) The TUNEL and IHC staining of tumor tissues. * p < 0.05, ** p < 0.01, *** p < 0.001.
See this image and copyright information in PMC

References

    1. Sung H., Ferlay J., Siegel R.L., Laversanne M., Soerjomataram I., Jemal A., Bray F. Global Cancer Statistics 2020: GLOBOCAN Estimates of Incidence and Mortality Worldwide for 36 Cancers in 185 Countries. CA Cancer J. Clin. 2021;71:209–249. doi: 10.3322/caac.21660. - DOI - PubMed
    1. Nicolas I., Marimon L., Barnadas E., Saco A., Rodriguez-Carunchio L., Fuste P., Marti C., Rodriguez-Trujillo A., Torne A., Del Pino M., et al. HPV-negative tumors of the uterine cervix. Mod. Pathol. 2019;32:1189–1196. doi: 10.1038/s41379-019-0249-1. - DOI - PubMed
    1. Ikeda H., Lethe B., Lehmann F., van Baren N., Baurain J.F., de Smet C., Chambost H., Vitale M., Moretta A., Boon T., et al. Characterization of an antigen that is recognized on a melanoma showing partial HLA loss by CTL expressing an NK inhibitory receptor. Immunity. 1997;6:199–208. doi: 10.1016/S1074-7613(00)80426-4. - DOI - PubMed
    1. Al-Khadairi G., Decock J. Cancer Testis Antigens and.d Immunotherapy: Where Do We Stand in the Targeting of PRAME? Cancers. 2019;11:984. doi: 10.3390/cancers11070984. - DOI - PMC - PubMed
    1. Stanojevic M., Hont A.B., Geiger A., O’Brien S., Ulrey R., Grant M., Datar A., Lee P.H., Lang H., Cruz C.R.Y., et al. Identification of novel HLA-restricted preferentially expressed antigen in melanoma peptides to facilitate off-the-shelf tumor-associated antigen-specific T-cell therapies. Cytotherapy. 2021;23:694–703. doi: 10.1016/j.jcyt.2021.03.001. - DOI - PMC - PubMed