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Synaptonemal complex protein 3 is a prognostic marker in cervical cancer.

Cho H, Noh KH, Chung JY, Takikita M, Chung EJ, Kim BW, Hewitt SM, Kim TW, Kim JH - PLoS ONE (2014)

Bottom Line: Functional studies using NIH3T3 cells demonstrated that the C-terminal region of human SCP3 is important for AKT activation and its oncogenic potential.High expression of SCP3 was significantly associated with tumor stage (P = 0.002) and tumor grade (P<0.001), while SCP3 expression was positively associated with pAKT protein level in cervical neoplasias.Survival times for patients with cervical cancer overexpressing both SCP3 and pAKT (median, 134.0 months, n = 68) were significantly shorter than for patients with low expression of either SCP3 or pAKT (161.5 months, n = 108) as determined by multivariate analysis (P = 0.020).

View Article: PubMed Central - PubMed

Affiliation: Department of Obstetrics and Gynecology, Gangnam Severance Hospital, Yonsei University College of Medicine, Seoul, Republic of Korea; Institute of Women's Life Medical Science, Yonsei University College of Medicine, Seoul, Republic of Korea.

ABSTRACT
Synaptonemal complex protein 3 (SCP3), a member of Cor1 family, is up-regulated in various cancer cells; however, its oncogenic potential and clinical significance has not yet been characterized. In the present study, we investigated the oncogenic role of SCP3 and its relationship with phosphorylated AKT (pAKT) in cervical neoplasias. The functional role of SCP3 expression was investigated by overexpression or knockdown of SCP3 in murine cell line NIH3T3 and human cervical cancer cell lines CUMC6, SiHa, CaSki, and HeLa both in vitro and in vivo. Furthermore, we examined SCP3 expression in tumor specimens from 181 cervical cancer and 400 cervical intraepithelial neoplasia (CIN) patients by immunohistochemistry and analyzed the correlation between SCP3 expression and clinicopathologic factors or survival. Overexpression of SCP3 promoted AKT-mediated tumorigenesis both in vitro and in vivo. Functional studies using NIH3T3 cells demonstrated that the C-terminal region of human SCP3 is important for AKT activation and its oncogenic potential. High expression of SCP3 was significantly associated with tumor stage (P = 0.002) and tumor grade (P<0.001), while SCP3 expression was positively associated with pAKT protein level in cervical neoplasias. Survival times for patients with cervical cancer overexpressing both SCP3 and pAKT (median, 134.0 months, n = 68) were significantly shorter than for patients with low expression of either SCP3 or pAKT (161.5 months, n = 108) as determined by multivariate analysis (P = 0.020). Our findings suggest that SCP3 plays an important role in the progression of cervical cancer through the AKT signaling pathway, supporting the possibility that SCP3 may be a promising novel cancer target for cervical cancer therapy.

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hSCP3 increases oncogenic potentials of human cervical cancer cells in vitro and in vivo.(A) Western blot analysis of levels of SCP3 in human cervical cancer cell lines CUMC6, SiHa, CaSki, and HeLa cells. Numbers below western blots refer to the relative values of the intensity normalized to CUMC6 control. (B) Western blot analysis of expression of SCP3, pAKT, and AKT in CaSki cells retrovirally transduced with a pMSCV vector encoding hSCP3 (CaSki/hSCP3). CaSki/no insert cells were used as a control. Numbers below western blots refer to the relative values of the intensity normalized to no insert control. (C) In vitro growth curves of CaSki/hSCP3 cells. Cells were counted after trypan blue staining to exclude dead cells. (D) Colony-forming capacity of CaSki/hSCP3 cells in soft agar; (Top) Representative colony images of each group; (Bottom) Bar graph representing the number of colonies with diameters greater than 2 mm in soft agar (scale bar: 1 mm). (E) Tumorigenicity of CaSki/hSCP3 cells. Balb/c Nude mice (n = 5) were inoculated with CaSki/no insert or CaSki/hSCP3 cells (1×106 cells/mouse) subcutaneously. (F) Western blot analysis of expression of SCP3, pAKT, AKT in HeLa/shSCP3 cells. HeLa/shGFP were used as a control. Numbers below western blots refer to the relative values of the intensity normalized to shGFP control. (G) In vitro growth curves of HeLa/shSCP3 cells. (H) Colony-forming capacity of HeLa/hSCP3 cells in soft agar (scale bar: 1 mm). (I) Tumorigenicity of HeLa/shSCP3 cells. Balb/c Nude mice (n = 5) were inoculated subcutaneously with 1×106 cells/mouse of HeLa/shGFP or HeLa/shSCP3 cells. Error bars represent the mean ± SD.
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pone-0098712-g004: hSCP3 increases oncogenic potentials of human cervical cancer cells in vitro and in vivo.(A) Western blot analysis of levels of SCP3 in human cervical cancer cell lines CUMC6, SiHa, CaSki, and HeLa cells. Numbers below western blots refer to the relative values of the intensity normalized to CUMC6 control. (B) Western blot analysis of expression of SCP3, pAKT, and AKT in CaSki cells retrovirally transduced with a pMSCV vector encoding hSCP3 (CaSki/hSCP3). CaSki/no insert cells were used as a control. Numbers below western blots refer to the relative values of the intensity normalized to no insert control. (C) In vitro growth curves of CaSki/hSCP3 cells. Cells were counted after trypan blue staining to exclude dead cells. (D) Colony-forming capacity of CaSki/hSCP3 cells in soft agar; (Top) Representative colony images of each group; (Bottom) Bar graph representing the number of colonies with diameters greater than 2 mm in soft agar (scale bar: 1 mm). (E) Tumorigenicity of CaSki/hSCP3 cells. Balb/c Nude mice (n = 5) were inoculated with CaSki/no insert or CaSki/hSCP3 cells (1×106 cells/mouse) subcutaneously. (F) Western blot analysis of expression of SCP3, pAKT, AKT in HeLa/shSCP3 cells. HeLa/shGFP were used as a control. Numbers below western blots refer to the relative values of the intensity normalized to shGFP control. (G) In vitro growth curves of HeLa/shSCP3 cells. (H) Colony-forming capacity of HeLa/hSCP3 cells in soft agar (scale bar: 1 mm). (I) Tumorigenicity of HeLa/shSCP3 cells. Balb/c Nude mice (n = 5) were inoculated subcutaneously with 1×106 cells/mouse of HeLa/shGFP or HeLa/shSCP3 cells. Error bars represent the mean ± SD.

Mentions: We further investigated the oncogenic role of SCP3 and its relationship with phosphorylated AKT (pAKT) in human cervical cancer cell lines. For this, the expression of SCP3 was measured in human cervical cancer cell lines CUMC6, SiHa, CaSki, and HeLa using western blot analysis. As shown in Figure 4A, expression of SCP3 was observed in all of cervical cancer cells although that of SCP3 was most profound in HeLa cells. In contrast, CaSki cells exhibited lowest expression of SCP3 protein. Next, to evaluate the effects of SCP3 on cell proliferation and tumorigenicity in cervical cancer cells, we established the CaSki cell lines expressing hSCP3 or no insert (empty vector) using a retroviral transduction system CaSki/hSCP3 or CaSki/no insert, respectively. Conversely, we established the HeLa cell lines expressing shRNA targeting SCP3 (shSCP3) or GFP (irrelevant negative control, shGFP) using a shRNA expressing vector (pSilencer 3.1-H1 puro), HeLa/shSCP3 or HeLa/shGFP, respectively, after puromycin selection. First of all, SCP3, pAKT, AKT protein levels in the CaSki cells were measured by western blotting. As shown in Figure 4B, the expression of pAKT was significantly increased in CaSki/SCP3 cells compared with control CaSki/no insert cells. The cell growth assay revealed that cell growth rate in the SCP3-transduced cells was significantly higher than control cells (Figure 4C). Similar increase was also observed in colony formation assay (Figure 4D). Consistent with in vitro data, the tumor-forming ability of CaSki cells was significantly increased after introducing SCP3 (Figure 4E). In contrast, knock-down of SCP3 in HeLa cells significantly decreased pAKT level, in vitro cell growth rate and colony formation efficacies and in vivo tumor growth rate compared with shGFP control group (Figure 4F–4I). These data demonstrate that SCP3 has a key role in cell proliferation and tumorigenicity of cervical cancer cells.


Synaptonemal complex protein 3 is a prognostic marker in cervical cancer.

Cho H, Noh KH, Chung JY, Takikita M, Chung EJ, Kim BW, Hewitt SM, Kim TW, Kim JH - PLoS ONE (2014)

hSCP3 increases oncogenic potentials of human cervical cancer cells in vitro and in vivo.(A) Western blot analysis of levels of SCP3 in human cervical cancer cell lines CUMC6, SiHa, CaSki, and HeLa cells. Numbers below western blots refer to the relative values of the intensity normalized to CUMC6 control. (B) Western blot analysis of expression of SCP3, pAKT, and AKT in CaSki cells retrovirally transduced with a pMSCV vector encoding hSCP3 (CaSki/hSCP3). CaSki/no insert cells were used as a control. Numbers below western blots refer to the relative values of the intensity normalized to no insert control. (C) In vitro growth curves of CaSki/hSCP3 cells. Cells were counted after trypan blue staining to exclude dead cells. (D) Colony-forming capacity of CaSki/hSCP3 cells in soft agar; (Top) Representative colony images of each group; (Bottom) Bar graph representing the number of colonies with diameters greater than 2 mm in soft agar (scale bar: 1 mm). (E) Tumorigenicity of CaSki/hSCP3 cells. Balb/c Nude mice (n = 5) were inoculated with CaSki/no insert or CaSki/hSCP3 cells (1×106 cells/mouse) subcutaneously. (F) Western blot analysis of expression of SCP3, pAKT, AKT in HeLa/shSCP3 cells. HeLa/shGFP were used as a control. Numbers below western blots refer to the relative values of the intensity normalized to shGFP control. (G) In vitro growth curves of HeLa/shSCP3 cells. (H) Colony-forming capacity of HeLa/hSCP3 cells in soft agar (scale bar: 1 mm). (I) Tumorigenicity of HeLa/shSCP3 cells. Balb/c Nude mice (n = 5) were inoculated subcutaneously with 1×106 cells/mouse of HeLa/shGFP or HeLa/shSCP3 cells. Error bars represent the mean ± SD.
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pone-0098712-g004: hSCP3 increases oncogenic potentials of human cervical cancer cells in vitro and in vivo.(A) Western blot analysis of levels of SCP3 in human cervical cancer cell lines CUMC6, SiHa, CaSki, and HeLa cells. Numbers below western blots refer to the relative values of the intensity normalized to CUMC6 control. (B) Western blot analysis of expression of SCP3, pAKT, and AKT in CaSki cells retrovirally transduced with a pMSCV vector encoding hSCP3 (CaSki/hSCP3). CaSki/no insert cells were used as a control. Numbers below western blots refer to the relative values of the intensity normalized to no insert control. (C) In vitro growth curves of CaSki/hSCP3 cells. Cells were counted after trypan blue staining to exclude dead cells. (D) Colony-forming capacity of CaSki/hSCP3 cells in soft agar; (Top) Representative colony images of each group; (Bottom) Bar graph representing the number of colonies with diameters greater than 2 mm in soft agar (scale bar: 1 mm). (E) Tumorigenicity of CaSki/hSCP3 cells. Balb/c Nude mice (n = 5) were inoculated with CaSki/no insert or CaSki/hSCP3 cells (1×106 cells/mouse) subcutaneously. (F) Western blot analysis of expression of SCP3, pAKT, AKT in HeLa/shSCP3 cells. HeLa/shGFP were used as a control. Numbers below western blots refer to the relative values of the intensity normalized to shGFP control. (G) In vitro growth curves of HeLa/shSCP3 cells. (H) Colony-forming capacity of HeLa/hSCP3 cells in soft agar (scale bar: 1 mm). (I) Tumorigenicity of HeLa/shSCP3 cells. Balb/c Nude mice (n = 5) were inoculated subcutaneously with 1×106 cells/mouse of HeLa/shGFP or HeLa/shSCP3 cells. Error bars represent the mean ± SD.
Mentions: We further investigated the oncogenic role of SCP3 and its relationship with phosphorylated AKT (pAKT) in human cervical cancer cell lines. For this, the expression of SCP3 was measured in human cervical cancer cell lines CUMC6, SiHa, CaSki, and HeLa using western blot analysis. As shown in Figure 4A, expression of SCP3 was observed in all of cervical cancer cells although that of SCP3 was most profound in HeLa cells. In contrast, CaSki cells exhibited lowest expression of SCP3 protein. Next, to evaluate the effects of SCP3 on cell proliferation and tumorigenicity in cervical cancer cells, we established the CaSki cell lines expressing hSCP3 or no insert (empty vector) using a retroviral transduction system CaSki/hSCP3 or CaSki/no insert, respectively. Conversely, we established the HeLa cell lines expressing shRNA targeting SCP3 (shSCP3) or GFP (irrelevant negative control, shGFP) using a shRNA expressing vector (pSilencer 3.1-H1 puro), HeLa/shSCP3 or HeLa/shGFP, respectively, after puromycin selection. First of all, SCP3, pAKT, AKT protein levels in the CaSki cells were measured by western blotting. As shown in Figure 4B, the expression of pAKT was significantly increased in CaSki/SCP3 cells compared with control CaSki/no insert cells. The cell growth assay revealed that cell growth rate in the SCP3-transduced cells was significantly higher than control cells (Figure 4C). Similar increase was also observed in colony formation assay (Figure 4D). Consistent with in vitro data, the tumor-forming ability of CaSki cells was significantly increased after introducing SCP3 (Figure 4E). In contrast, knock-down of SCP3 in HeLa cells significantly decreased pAKT level, in vitro cell growth rate and colony formation efficacies and in vivo tumor growth rate compared with shGFP control group (Figure 4F–4I). These data demonstrate that SCP3 has a key role in cell proliferation and tumorigenicity of cervical cancer cells.

Bottom Line: Functional studies using NIH3T3 cells demonstrated that the C-terminal region of human SCP3 is important for AKT activation and its oncogenic potential.High expression of SCP3 was significantly associated with tumor stage (P = 0.002) and tumor grade (P<0.001), while SCP3 expression was positively associated with pAKT protein level in cervical neoplasias.Survival times for patients with cervical cancer overexpressing both SCP3 and pAKT (median, 134.0 months, n = 68) were significantly shorter than for patients with low expression of either SCP3 or pAKT (161.5 months, n = 108) as determined by multivariate analysis (P = 0.020).

View Article: PubMed Central - PubMed

Affiliation: Department of Obstetrics and Gynecology, Gangnam Severance Hospital, Yonsei University College of Medicine, Seoul, Republic of Korea; Institute of Women's Life Medical Science, Yonsei University College of Medicine, Seoul, Republic of Korea.

ABSTRACT
Synaptonemal complex protein 3 (SCP3), a member of Cor1 family, is up-regulated in various cancer cells; however, its oncogenic potential and clinical significance has not yet been characterized. In the present study, we investigated the oncogenic role of SCP3 and its relationship with phosphorylated AKT (pAKT) in cervical neoplasias. The functional role of SCP3 expression was investigated by overexpression or knockdown of SCP3 in murine cell line NIH3T3 and human cervical cancer cell lines CUMC6, SiHa, CaSki, and HeLa both in vitro and in vivo. Furthermore, we examined SCP3 expression in tumor specimens from 181 cervical cancer and 400 cervical intraepithelial neoplasia (CIN) patients by immunohistochemistry and analyzed the correlation between SCP3 expression and clinicopathologic factors or survival. Overexpression of SCP3 promoted AKT-mediated tumorigenesis both in vitro and in vivo. Functional studies using NIH3T3 cells demonstrated that the C-terminal region of human SCP3 is important for AKT activation and its oncogenic potential. High expression of SCP3 was significantly associated with tumor stage (P = 0.002) and tumor grade (P<0.001), while SCP3 expression was positively associated with pAKT protein level in cervical neoplasias. Survival times for patients with cervical cancer overexpressing both SCP3 and pAKT (median, 134.0 months, n = 68) were significantly shorter than for patients with low expression of either SCP3 or pAKT (161.5 months, n = 108) as determined by multivariate analysis (P = 0.020). Our findings suggest that SCP3 plays an important role in the progression of cervical cancer through the AKT signaling pathway, supporting the possibility that SCP3 may be a promising novel cancer target for cervical cancer therapy.

Show MeSH
Related in: MedlinePlus