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MicroRNA-21 is a candidate driver gene for 17q23-25 amplification in ovarian clear cell carcinoma.

Hirata Y, Murai N, Yanaihara N, Saito M, Saito M, Urashima M, Murakami Y, Matsufuji S, Okamoto A - BMC Cancer (2014)

Bottom Line: The patients with 17q23-25 amplification had significantly shorter progression-free and overall survival than those without 17q23-25 amplification (log-rank test: p = 0.0496; p = 0.0469, respectively).A significant correlation was observed between miR-21 overexpression and endometriosis.Aberrant expression of miR-21 by chromosomal amplification might play an important role in CCC carcinogenesis through the regulation of the PTEN tumor suppressor gene.

View Article: PubMed Central - PubMed

Affiliation: Department of Obstetrics and Gynecology, The Jikei University School of Medicine, 3-25-8, Nishi-Shinbashi, Minato-ku, Tokyo 105-8461, Japan. yanazou@jikei.ac.jp.

ABSTRACT

Background: Epithelial ovarian cancer (EOC) is the most common cause of gynecological malignancy-related mortality. Ovarian clear cell carcinoma (CCC) has unique clinical characteristics and behaviors that differ from other histological types of EOC, including a frequent association with endometriosis and a highly chemoresistant nature, resulting in poor prognosis. However, factors underlying its malignant behavior are still poorly understood. Aberrant expression of microRNAs has been shown to be involved in oncogenesis, and microRNA-21 (miR-21) is frequently overexpressed in many types of cancers. The aim of this study was to investigate the role of miR-21 in 17q23-25 amplification associated with CCC oncogenesis.

Methods: We identified 17q23-25 copy number aberrations among 28 primary CCC tumors by using a comparative genomic hybridization method. Next, we measured expression levels of the candidate target genes, miR-21 and PPM1D, for 17q23-25 amplification by real-time RT-PCR analysis and compared those data with copy number status and clinicopathological features. In addition, immunohistochemical analysis of PTEN (a potential target of miR-21) was performed using the same primary CCC cases. We investigated the biological significance of miR-21 overexpression in CCC using a loss-of-function antisense approach.

Results: 17q23-25 amplification with both miR-21 overexpression and PTEN protein loss was detected in 4/28 CCC cases (14.2%). The patients with 17q23-25 amplification had significantly shorter progression-free and overall survival than those without 17q23-25 amplification (log-rank test: p = 0.0496; p = 0.0469, respectively). A significant correlation was observed between miR-21 overexpression and endometriosis. Both PTEN mRNA and PTEN protein expression were increased by miR-21 knockdown in CCC cells. We also confirmed that miR-21 directly bound to the 3'-untranslated region of PTEN mRNA using a dual-luciferase reporter assay.

Conclusions: MiR-21 is a possible driver gene other than PPM1D for 17q23-25 amplification in CCC. Aberrant expression of miR-21 by chromosomal amplification might play an important role in CCC carcinogenesis through the regulation of the PTEN tumor suppressor gene.

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Analysis of clinical CCC specimens. Of the 9 tumors with 17q23-25 amplification, 7 (77.7%) showed miR-21 overexpression. Of the 19 tumors (58.8%) without 17q23-25 amplification, 10 showed miR-21 overexpression. Of all the 28 17q23-25 amplification cases, both miR-21 overexpression and PTEN protein loss were detected in 4 (14.2%).
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Fig2: Analysis of clinical CCC specimens. Of the 9 tumors with 17q23-25 amplification, 7 (77.7%) showed miR-21 overexpression. Of the 19 tumors (58.8%) without 17q23-25 amplification, 10 showed miR-21 overexpression. Of all the 28 17q23-25 amplification cases, both miR-21 overexpression and PTEN protein loss were detected in 4 (14.2%).

Mentions: CGH array profiles of chromosome 17 in 28 primary CCCs revealed that 9 out of 28 patients (32%) showed 17q23-25 amplification that included miR-21 (Figure 1). MiR-21 and PPM1D mRNA expression were then measured by real-time RT-PCR analysis (Additional file 1: Figure S1). We defined standardized value as each median value of miR-21 and PPM1D expression without 17q23-25 amplification. Overexpression of miR-21 and PPM1D were found in 60% and 57% of these tumors, respectively. Seven of 9 tumors (77.7%) with 17q23-25 amplification showed miR-21 overexpression, and 10 of 19 tumors (52.6%) without 17q23-25 amplification also showed miR-21 overexpression. In addition, 6 of 9 tumors (66.6%) with 17q23-25 amplification showed PPM1D overexpression, and 10 of 19 tumors (52%) without 17q23-25 amplification showed PPM1D overexpression (Additional file 1: Figure S1). We next evaluated the relationship between 17q23-25 amplification and either miR-21 or PPM1D overexpression. No significant correlation between the amplification and overexpression was observed for either gene. Next, immunohistochemical analysis of PTEN (a potential target of miR-21) was performed on samples from the same primary CCC patients. Loss of PTEN protein was observed in 13 of 28 patients (46.4%) (Additional file 2: Figure S2) and in 6 of 17 tumors (35.3%) with miR-21 overexpression. No significant correlation was observed between miR-21 overexpression and loss of PTEN expression. To further confirm these results, we added 15 CCC samples from an additional cohort, performing real-time RT-PCR of miR21 and IHC of PTEN. Again, no significant correlation was observed between miR-21 overexpression and loss of PTEN expression (date not shown). In total, as shown in Figure 2, the occurrence of 17q23-25 amplification with both miR-21 overexpression and PTEN protein loss was detected in 4 out of 28 CCC patients (14.2%) (Figure 2).Figure 1


MicroRNA-21 is a candidate driver gene for 17q23-25 amplification in ovarian clear cell carcinoma.

Hirata Y, Murai N, Yanaihara N, Saito M, Saito M, Urashima M, Murakami Y, Matsufuji S, Okamoto A - BMC Cancer (2014)

Analysis of clinical CCC specimens. Of the 9 tumors with 17q23-25 amplification, 7 (77.7%) showed miR-21 overexpression. Of the 19 tumors (58.8%) without 17q23-25 amplification, 10 showed miR-21 overexpression. Of all the 28 17q23-25 amplification cases, both miR-21 overexpression and PTEN protein loss were detected in 4 (14.2%).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4289307&req=5

Fig2: Analysis of clinical CCC specimens. Of the 9 tumors with 17q23-25 amplification, 7 (77.7%) showed miR-21 overexpression. Of the 19 tumors (58.8%) without 17q23-25 amplification, 10 showed miR-21 overexpression. Of all the 28 17q23-25 amplification cases, both miR-21 overexpression and PTEN protein loss were detected in 4 (14.2%).
Mentions: CGH array profiles of chromosome 17 in 28 primary CCCs revealed that 9 out of 28 patients (32%) showed 17q23-25 amplification that included miR-21 (Figure 1). MiR-21 and PPM1D mRNA expression were then measured by real-time RT-PCR analysis (Additional file 1: Figure S1). We defined standardized value as each median value of miR-21 and PPM1D expression without 17q23-25 amplification. Overexpression of miR-21 and PPM1D were found in 60% and 57% of these tumors, respectively. Seven of 9 tumors (77.7%) with 17q23-25 amplification showed miR-21 overexpression, and 10 of 19 tumors (52.6%) without 17q23-25 amplification also showed miR-21 overexpression. In addition, 6 of 9 tumors (66.6%) with 17q23-25 amplification showed PPM1D overexpression, and 10 of 19 tumors (52%) without 17q23-25 amplification showed PPM1D overexpression (Additional file 1: Figure S1). We next evaluated the relationship between 17q23-25 amplification and either miR-21 or PPM1D overexpression. No significant correlation between the amplification and overexpression was observed for either gene. Next, immunohistochemical analysis of PTEN (a potential target of miR-21) was performed on samples from the same primary CCC patients. Loss of PTEN protein was observed in 13 of 28 patients (46.4%) (Additional file 2: Figure S2) and in 6 of 17 tumors (35.3%) with miR-21 overexpression. No significant correlation was observed between miR-21 overexpression and loss of PTEN expression. To further confirm these results, we added 15 CCC samples from an additional cohort, performing real-time RT-PCR of miR21 and IHC of PTEN. Again, no significant correlation was observed between miR-21 overexpression and loss of PTEN expression (date not shown). In total, as shown in Figure 2, the occurrence of 17q23-25 amplification with both miR-21 overexpression and PTEN protein loss was detected in 4 out of 28 CCC patients (14.2%) (Figure 2).Figure 1

Bottom Line: The patients with 17q23-25 amplification had significantly shorter progression-free and overall survival than those without 17q23-25 amplification (log-rank test: p = 0.0496; p = 0.0469, respectively).A significant correlation was observed between miR-21 overexpression and endometriosis.Aberrant expression of miR-21 by chromosomal amplification might play an important role in CCC carcinogenesis through the regulation of the PTEN tumor suppressor gene.

View Article: PubMed Central - PubMed

Affiliation: Department of Obstetrics and Gynecology, The Jikei University School of Medicine, 3-25-8, Nishi-Shinbashi, Minato-ku, Tokyo 105-8461, Japan. yanazou@jikei.ac.jp.

ABSTRACT

Background: Epithelial ovarian cancer (EOC) is the most common cause of gynecological malignancy-related mortality. Ovarian clear cell carcinoma (CCC) has unique clinical characteristics and behaviors that differ from other histological types of EOC, including a frequent association with endometriosis and a highly chemoresistant nature, resulting in poor prognosis. However, factors underlying its malignant behavior are still poorly understood. Aberrant expression of microRNAs has been shown to be involved in oncogenesis, and microRNA-21 (miR-21) is frequently overexpressed in many types of cancers. The aim of this study was to investigate the role of miR-21 in 17q23-25 amplification associated with CCC oncogenesis.

Methods: We identified 17q23-25 copy number aberrations among 28 primary CCC tumors by using a comparative genomic hybridization method. Next, we measured expression levels of the candidate target genes, miR-21 and PPM1D, for 17q23-25 amplification by real-time RT-PCR analysis and compared those data with copy number status and clinicopathological features. In addition, immunohistochemical analysis of PTEN (a potential target of miR-21) was performed using the same primary CCC cases. We investigated the biological significance of miR-21 overexpression in CCC using a loss-of-function antisense approach.

Results: 17q23-25 amplification with both miR-21 overexpression and PTEN protein loss was detected in 4/28 CCC cases (14.2%). The patients with 17q23-25 amplification had significantly shorter progression-free and overall survival than those without 17q23-25 amplification (log-rank test: p = 0.0496; p = 0.0469, respectively). A significant correlation was observed between miR-21 overexpression and endometriosis. Both PTEN mRNA and PTEN protein expression were increased by miR-21 knockdown in CCC cells. We also confirmed that miR-21 directly bound to the 3'-untranslated region of PTEN mRNA using a dual-luciferase reporter assay.

Conclusions: MiR-21 is a possible driver gene other than PPM1D for 17q23-25 amplification in CCC. Aberrant expression of miR-21 by chromosomal amplification might play an important role in CCC carcinogenesis through the regulation of the PTEN tumor suppressor gene.

Show MeSH
Related in: MedlinePlus