The UbcH10 gene codes for a protein that belongs to the ubiquitin-conjugating enzyme family. Previous studies of our group suggest UbcH10 expression as a valid indicator of the proliferative and aggressive status of thyroid carcinomas. Therefore, to better understand the process of ovarian carcinogenesis, and to look for possible tools to be used as prognostic markers in these neoplasias, we decided to extend the analysis of the UbcH10 expression to the ovarian neoplastic disease. We found that the UbcH10 gene was upregulated in some ovarian carcinoma cell lines analysed. Then, immunohistochemical studies demonstrate that UbcH10 expression significantly correlates with the tumor grade and the undifferentiated histotype of the ovarian carcinomas. Furthermore, a significant relationship between UbcH10 expression and overall survival was observed. Finally, the block of UbcH10 protein synthesis by RNA interference inhibited the growth of ovarian carcinoma cell lines, suggesting a role of UbcH10 overexpression in ovarian carcinogenesis. Therefore, all these data taken together suggest the possibility to use UbcH10 detection as a marker for the diagnosis and prognosis of these neoplastic diseases and open the perspective of a therapy of some ovarian carcinomas based on the suppression of the UbcH10 synthesis and/or function.
Ovarian carcinomas have a great impact in human pathology, in fact ovarian cancer is the leading cause of death from gynecological neoplasias and the fifth most common cancer among women worldwide (Ozols et al., 2004; Jemal et al., 2005).
Research aimed to determine the specific genes involved in the development of ovarian cancers would help to understand how normal ovarian epithelial cells escape regulation of proliferation, apoptosis and senescence. It has been already (Welcsh and King, 2001) determined that approximately 10% of ovarian cancers arises in women who have inherited mutations in cancer-susceptibility genes such as BRCA1, BRCA2 and other DNA repair genes. Conversely, the vast majority of ovarian cancers are sporadic, presumably resulting from the accumulation of genetic damage over lifetime. Several genes involved in ovarian carcinogenesis have been identified, most notably the p53 tumor suppressor (Feki and Irminger-Finger, 2004).
Recently in our laboratory, by a microarray gene expression profiling, we found a gene, UbcH10, that was upregulated in thyroid anaplastic carcinoma samples vs the normal thyroid tissues, suggesting a correlation with the malignant progression (Pallante et al., 2005). A high expression of UbcH10 has also been found in carcinomas of different anatomic origin (Wagner et al., 2004).
The UbcH10 gene belongs to the E2 gene family and codes for a protein of 19.6 kDa that is involved in the ubiquitin-dependent proteolysis. In this pathway, ubiquitin-conjugating enzyme (E2), together with ubiquitin ligase (E3), transfers ubiquitin to specific substrate proteins (Hershko and Ciechanover, 1998; Joazeiro and Weissman, 2000).
The aim of our work was to investigate whether the UbcH10 expression might be a new useful indicator for the diagnosis and prognosis of ovarian cancer.
Therefore, we evaluated the expression of UbcH10 by reverse transcription–polymerase chain reaction (RT–PCR) and Western blot in ovarian carcinoma cell lines in comparison with the normal ovarian tissue. All of the carcinoma cell lines showed a high UbcH10 expression, which was barely detectable in the normal tissue (Figure 1a and b).
The same analyses, performed on ovarian carcinoma tissues, showed a different expression in the different histotypes analysed. In fact, as shown in Figure 1c and d, UbcH10 is expressed at low levels in clear cell carcinomas, endometrioid carcinomas and serous cystadenocarcinomas, whereas the undifferentiated carcinomas express high level of UbcH10.
Immunohistochemistry essentially confirms the data obtained by RT–PCR and Western blot analyses. Whereas benign ovary lacked UbcH10 expression (Figure 2f), the different histotypes of ovarian carcinomas showed certain variability in the range of neoplastic cells expressing UbcH10 (Figure 2a–e). However, there was a clear relationship between the loss of tumor differentiation and the gain of UbcH10 reactivity. This was evident both when examining tumor type and tumor grade: the average levels of UbcH10 expression were low in tumors belonging to the differentiated histotypes, as in serous (16.6% mean, range: 12.3–19.8%), clear cell (7.1% mean, range: 1.3–10.0%) and endometrioid (9.2% mean, range: 8.3–10.8%) carcinomas. Similarly the only case examined of the mucinous histotype showed a low level of expression (5%). On the contrary, higher levels (33.6% mean, range: 22.9–42.1%) of UbcH10 expression were observed in the undifferentiated histotype of ovarian cancer. All seven cases of undifferentiated ovarian carcinomas had levels of UbcH10 expression higher than the median value (P=0.015).
Similarly, UbcH10 expression was also related to the tumor grade, as significant differences were recorded among the different grades (P=0.002 Kruskall–Wallis test). In fact, as shown in Figure 2g, low levels of UbcH10 expression were recorded in the G1 well-differentiated tumors (0.8%), whereas there was a significant relationship between increasing cytological grading of malignancy and protein expression. In G2 and in G3 carcinomas, the cells expressing UbcH10 were, respectively, 12.3 and 22%, whereas G4 high-grade tumors showed staining in more than a third of neoplastic cells (34.7%). The correlation with tumor grade is further established if we compare G1 and G2 vs G3 and G4 grades and UbcH10 expression (P-value=0.023 Fisher's exact test) (data not shown).
As far as the overall survival is concerned, there is a significant relationship with UbcH10 expression (P-value=0.044 – log-rank test), as shown in Figure 3, only if we consider the mean as pooling method. Regarding the other pooling methods, as well as for relapse-free survival, no statistical significant correlation was found.
We asked whether UbcH10 overexpression had a role in the process of ovarian carcinogenesis by evaluating the growth rate of one ovarian carcinoma cell line, in which the synthesis of UbcH10 protein was suppressed by RNA interference. The SKOV-3 cell line was treated with siRNA duplexes targeting to the UbcH10 mRNA. After transfection, we observed an efficient knock down of the UbcH10 protein levels at 48 h after treatment (Figure 4a). The cell growth analysis, in the presence or absence of the UbcH10 siRNA duplexes, revealed that the block of the UbcH10 protein synthesis significantly inhibits ovarian carcinoma cell growth. In fact, as shown in Figure 4b, a significant reduction in cell growth rate was observed in SKOV-3 cell line treated with UbcH10 siRNA in comparison with the untreated cells or those treated with the control scrambled siRNA.
These results indicate a role of UbcH10 in neoplastic ovarian cell proliferation, for this reason we propose the UbcH10 expression as a possible tool to be used in the diagnosis and prognosis of ovarian carcinomas. Abundant expression of UbcH10 was detected in primary ovarian tumors compared with benign ovarian tissues and significantly correlated to tumor grade (P=0.0008) and undifferentiated histotype (P=0.015). These results are consistent with our previous published data (Pallante et al., 2005) and further support the involvement of UbcH10 in the differentiation process of several human epithelial tissues (Wagner et al., 2004).
UbcH10 is also a negative predictor of clinical outcome in our series, further suggesting its ability to confer a more aggressive phenotype to tumor cells. Studies are ongoing to assess this hypothesis on a larger series of patients with a longer follow-up.
Functional studies also demonstrate that the suppression of the UbcH10 expression by RNA interference reduced the growth of one ovarian carcinoma cell line indicating a role of UbcH10 overexpression in ovarian carcinogenesis, in particular, in influencing the hyperproliferative status of the most malignant cells.
In conclusion, all these data taken together suggest the possibility to use this gene as a marker for the diagnosis and prognosis of these neoplastic diseases.
Aldovini D, Demichelis F, Doglioni C, Di Vizio D, Galligioni E, Brugnara S et al. (2006). M-CAM expression as marker of poor prognosis in epithelial ovarian cancer. Int J Cancer Jun 27 [Epub ahead of print].
Benedet JL, Bender H, Jones III H, Ngan HY, Pecorelli S . (2000). FIGO staging classifications and clinical practice guidelines in the management of gynecologic cancers. FIGO Committee on Gynecologic Oncology. Int J Gynaecol Obstet 70: 209–262.
Feki A, Irminger-Finger I . (2004). Mutational spectrum of p53 mutations in primary breast and ovarian tumors. Crit Rev Oncol Hematol 52: 103–116.
Hershko A, Ciechanover A . (1998). The ubiquitin system. Annu Rev Biochem 67: 425–479.
Jemal A, Murray T, Ward E, Samuels A, Tiwari RC, Ghafoor A et al. (2005). Cancer statistics, 2005. CA Cancer J Clin 55: 10–30.
Joazeiro CA, Weissman AM . (2000). Ring finger proteins: mediators of ubiquitin ligase activity. Cell 1 102: 549–552.
Liu X, Minin V, Huang Y, Seligson DB, Horvath S . (2004). Statistical methods for analysing tissue microarray data. J Biopharm Stat 14: 671–685.
Masciullo V, Baldassarre G, Pentimalli F, Berlingieri MT, Boccia A, Chiappetta G et al. (2003). HMGA1 protein over-expression is a frequent feature of epithelial ovarian carcinomas. Carcinogenesis 24: 1191–1198.
Ozols RF, Bookman MA, Connolly DC, Daly MB, Godwin AK, Schilder RJ et al. (2004). Focus on epithelial ovarian cancer. Cancer Cell 5: 19–24.
Pallante P, Berlingieri MT, Troncone G, Kruhoffer M, Orntoft TF, Viglietto G et al. (2005). UbcH10 overexpression may represent a marker of anaplastic thyroid carcinomas. Br J Cancer 93: 464–471.
Pierantoni GM, Rinaldo C, Esposito F, Mottolese M, Soddu S, Fusco A . (2005). High Mobility Group A1 (HMGA1) proteins interact with p53 and inhibit its apoptotic activity. Cell Death Differ (9 December): 1–10 (Epub ahead of print).
R Development Core Team (2004). A Language and Environment for Statistical Computing. R Foundation for Statistical Computing: Vienna, Austria.
Tavassoli FA, Devilee P . (2003). World Health Oraganization Classification of Tumours. Tumours of the breast and female genital organs. IARC Press: Lyon.
Troncone G, Iaccarino A, Caleo A, Bifano D, Pettinato G, Palombini L . (2003). p27 Kip1 protein expression in Hashimoto's thyroiditis. J Clin Pathol 56: 587–591.
Vacher-Lavenu MC, Le Tourneau A, Duvillard P, Godefroy N, Pinel MC . (1993). Pathological classification and grading of primary ovarian carcinoma: experience of the ARTAC ovarian study group. Bull Cancer 80: 135–141.
Wagner KW, Sapinoso LM, El-Rifai W, Frierson HF, Butz N, Mestan J et al. (2004). Overexpression, genomic amplification and therapeutic potential of inhibiting the UbcH10 ubiquitin conjugase in human carcinomas of diverse anatomic origin. Oncogene 23: 6621–6629.
Welcsh PL, King MC . (2001). BRCA1 and BRCA2 and the genetics of breast and ovarian cancer. Hum Mol Genet 10: 705–713.
This work was supported by grants from the Associazione Italiana Ricerca sul Cancro (AIRC), Progetto Strategico Oncologia Consiglio Nazionale delle Ricerche, the Ministero dell'Università e della Ricerca Scientifica e Tecnologica (MIUR), and ‘Piani di Potenziamento della Rete Scientifica e Tecnologica’ CLUSTER C-04, the Programma Italia-USA sulla Terapia dei Tumori coordinated by Professor Cesare Peschle and ‘Ministero della Salute’. This work was supported from NOGEC-Naples Oncogenomic Center. We thank the Associazione Partenopea per le Ricerche Oncologiche (APRO) for its support.
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Berlingieri, M., Pallante, P., Guida, M. et al. UbcH10 expression may be a useful tool in the prognosis of ovarian carcinomas. Oncogene 26, 2136–2140 (2007). https://doi.org/10.1038/sj.onc.1210010
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