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CHD1L Protein is overexpressed in human ovarian carcinomas and is a novel predictive biomarker for patients survival
© He et al.; licensee BioMed Central Ltd. 2012
Received: 16 January 2012
Accepted: 26 September 2012
Published: 29 September 2012
Our recent studies suggested that the chromodomain helicase DNA binding protein 1-like (CHD1L) gene plays an oncogenic role in human hepatocellular carcinoma. However, the status of CHD1L protein expression in ovarian cancer and its clinical/prognostic significance are obscure.
In this study, immunohistochemistry (IHC) for CHD1L was performed on a tissue microarray (TMA) containing 102 primary ovarian carcinomas and 44 metastatic lesions (omental metastasis). Receiver-operator curve (ROC) analysis was used to evaluate patients’ survival status.
There is an augmented tendency of CHD1L expression in ovarian carcinoma metastasis than in primary lesions (P<0.05). A significant association was found between positive expression of CHD1L and tumors histological type (P <0.05). By univariate survival analysis of the ovarian carcinoma cohorts, positive expression of CHD1L was significantly correlated with shortened patient survival (mean 66.7 months versus 97.4 months, P<0.05). Moreover, CHD1L expression was evaluated to be a significant and independent prognostic factor in multivariate analysis (P<0.05).
These findings provide evidence that positive expression of CHD1L protein is significantly correlated with the metastasis proceeding of ovarian carcinoma, and CHD1L protein expression, as examined by IHC, may act as a novel prognostic biomarker for patients with ovarian carcinoma.
Ovarian cancer, the leading cause of death from a gynecological malignancy, has increasing incidence recently in China and Singapore . The onset age for ovarian cancer is about 45 years old . On account of the absence of early diagnosis methods and the absence of characteristic symptoms, the majority of ovarian cancer patients (70%) were diagnosed at FIGO III/IV, and had a unfavorable prognosis with a 5-year survival rate of <20% . Recently, ovarian cancer was considered as a more complicated genetic change and multiple-step patho-physiological change . Clinically, most ovarian cancers are ovarian carcinoma. It is prevalent to discovery a tumor marker for the diagnosis and prognosis of ovarian carcinoma, as well as developing targeted therapy, predicting the survival, and predicting relapse.
Recently, we reported that the chromodomain helicase DNA binding protein 1-like (CHD1L) gene is a putative oncogene mapped to chromosome 1q21 and it is associated with hepatocellular carcinoma (HCC) tumorigenesis . CHD1L belongs to SNF-2family, which contains a domain (approximately 400 amino acids) with highly conserved helicase motifs . The full-length mRNA (2.98kb) of CHD1L encodes an 897aa protein. Most SNF2-like proteins participate in various nuclear activities, such as transcriptional activation or repression, DNA repair and recombination [6, 7]. In HCC patients, over 50% cases have overexpression of CHD1L, which is associated with the carcinoma progression and metastasis and predict the poor prognosis of the survival status . To date, despite these facts, the status of CHD1L protein in ovarian carcinoma tissue and its prognostic significance has not been elucidated.
In this study, therefore, we examined the expression status of CHD1L protein by IHC in a cohort of ovarian carcinoma tissues (including 102 primary lesions and 44 corresponding metastatic lesions). The clinical/prognostic significance of CHD1L expression in our ovarian carcinoma cohort was assessed.
Patients and tissue specimens
Association of CHD1L protein expression with clinic-pathological parameter
CHD1L protein expression
Age at surgery (years)
≤ 51 b
Construction of tissue microarrays (TMA)
The tissue microarray was constructed as previously described [9–11]. The formalin-fixed, paraffin-embedded tissue blocks and the corresponding histological HE-stained slides were overlaid for tissue TMA sampling. Our senior pathologist reviewed the slides and marked the representative areas of tumor tissues. We used a tissue arraying instrument (Beecher Instruments, Silver Spring, MD, USA) to punch triplicate 0.6-mm-diameter cylinders of tissue from selected cancer areas of individual donor tissue block and re-embed into a recipient paraffin block at a predefined position. The TMA blocks included 102 ovarian carcinomas and 44 corresponding metastasis lesions. Subsequently, multiple sections (5-um thick) were cut from the TMA tissue array.
The immunohistochemical study of CHD1L was performed using a standard streptavidin-peroxidase method demonstrated previously . The endogenous peroxidase activity was blocked with 3% H2O2 for 10 minutes. For the antigen retrieval, slides were immersed in 10 mM citrate buffer (pH 6.0) and boiled for 15 minutes in a microwave oven. Non-specific binding was blocked by 5% normal goat serum for 10 minutes. The slides were incubated with a 1:100 dilution of monoclonal antibody against CHD1L (Abcam) at 4°C overnight in a moist chamber. The slides were sequentially incubated with biotinylated goat anti-mouse IgG (1:100 dilution; Santa Cruz Biotechnology) and then streptavidin-peroxidase conjugate, each for 30 minutes at room temperature. Isotope-matched human IgG was used in each case as a negative control. Finally, the 3, 5-diaminobenzidine (DAB) Substrate Kit (Dako) was used for color development followed by Mayer hematoxylin counterstaining.
For the evaluation of CHD1L staining, a semi-quantitative scoring criterion was used, in which both staining intensity and positive cells percentage were contained [12, 13]. A staining index (with values from 0 to 12) was obtained as the intensity of CHDIL staining (0=negative, 1=weakly positive, 2=positive, 3=strongly positive) times the proportion of immunopositive tumor cells (0%=0;<10%=1;10%≤ to<50%=2;50%≤to <75%=3;≥75%=4 ). All histological evaluations were carried out in a double-blind manner by two expert pathologists (M-YC and DX).
Statistical analysis was executed with the SPSS statistical software package (SPSS Standard version 13.0, SPSS Inc.). We assess the association of CHD1L protein expression with clinico-pathological features by the Chi-square test or Fishers’ exact test. Wilcoxon Signed Ranks Test is used to compare the CHD1L protein expression between the primary ovarian carcinomas and metastasis. Multivariate survival analysis was performed on all factors which have been proved to be significant on univariate analysis by the Cox regression model. For univariate survival analysis, Kaplan–Meier analysis is used. We utilized Log rank test to compare different survival curves. ROC analysis is used to compare the clinico-pathological features for estimation of the survival prediction. P value of <0.05 was considered statistically significant.
CHD1L protein expression in primary ovarian carcinomas and corresponding metastatic lesions
The expression of CHD1L protein in ovarian primary lesions and corresponding metastatic lesions
CHD1L protein expression score
ovarian primary lesions
corresponding metastatic lesions
Association of CHD1L protein expression with clinic-pathological parameters
In this study, according to the staining index above, protein expression with a scoring index of ≥ 4 (median score of CHD1L protein expression in the primary ovarian lesions) is defined as positive expression. The associations between CHD1L expression in primary ovarian carcinomas and several clinico-pathological variables are assessed and displayed in Table 1. The positive expression of CHD1L protein expression increasingly presented from mucinous/serous ovarian carcinoma to others types of tumor, including undifferentiated ovarian carcinoma. There was no significant difference between CHD1L protein expression and other clinicopathological features, such as patients’ age, histological grade, FIGO stage and residual tumor (P>0.05, Table 1).
Relationship between clinicpathologic variables, CHD1L protein expression, and ovarian carcinoma patient survival: univariate survival analysis
Clinical pathogical parameters and expression of CHD1L for prognosis of 102 patients with ovarian carcinoma by univariate survival analysis (log-rank test)
Mean survival (months)
Median survival (months)
Age at surgery
Satisfactory cytoreductive operation
Unsatisfactory cytoreductive operation
Cyclophosphamide + Adriamycin + Carbo
CHD1L protein expression
Independent prognostic factors of epithelial ovarian carcinoma: multivariate cox regression analysis
Multivariate analysis on overall survival (Cox regression model)
95% confidence interval
1.395 - 6.456
1.253 - 5.089
Relationship between clinicpathologic variables, CHD1L protein expression, and ovarian carcinoma patient survival: receiver operating characteristic curve (ROC) analysis
CHD1L, the candidate oncogene, has been isolated from 1q21 amplicon and found frequently amplified in hepatocellular carcinoma (HCC). Amplification of 1q21 is the most frequent genetic alteration in human HCC . It plays an important role in the development and metastasis of HCC. Gains of 1q21-q25 have been linked to a more aggressive phenotype with metastatic potential in renal clear cell carcinoma and squamous cell carcinomas of lung [15, 16]. In ovarian carcinoma and neuroblastoma, 1q21-q22 amplification indicated a drug-resistant phenotype [17, 18]. CHD1L is activated by poly (ADP-ribose) (PAR) polymerase-1 (PARP-1), a DNA nick sensor enzyme that is activated by DNA breaks, during the posttranslational modification process termed “PARylation” . CHD1L normally loosens tightly packaged chromatin to allow for repairing of DNA damage. In cancer cells, however, too much CHD1L can overly relax the chromatin, making the DNA vulnerable to mistake and thus raising the risk of cancer. However, to date, the expression dynamics of CHD1L in ovarian carcinomas has not been investigated, and its clinico-pathological and/or prognostic value in ovarian carcinomas remains unknown.
With regards to the gene function and oncogenic role of CHD1L, it is still not thoroughly characterized and clarified so far. Our recent in vitro and in vivo functional studies in mice showed that CHD1L contributed to HCC cell aggressiveness by increasing cell motility and inducing filopodia formation and epithelial-mesenchymal transition (EMT) via ARHGEF9-mediated Cdc42 activation. Silencing ARHGEF9 expression by RNAi lagely abolished the invasive and metastatic abilities of CHD1L in mice . Furthermore, our investigation of clinical HCC specimens showed that CHD1L and ARHGEF9 were markedly overexpressed in metastatic HCC tissue compared with healthy tissue . These findings suggested that CHD1LARHGEF9-Cdc42-EMT might be a novel pathway involved in HCC progression and/or metastasis. However, the potential oncogenic role and underling mechanisms of CHD1L in ovarian carcinoma have not been investigated. Clearly, further studies are needed to elucidate the mechanisms by which CHD1L promoting the development and metastasis of ovarian carcinomas in detail.
In summary, we have shown here that CHD1L protein expression was significantly higher in in metastatic lesions than in the primary lesions and that CHD1L expression was significantly correlated with survival. Therefore, CHD1L is not only a biomarker associated with ovarian cancer, but it also appears to be a prognostic factor associated with adverse outcome.
This study was supported by the Project of Guangdong Science and Technology Agency (No. 2010B060900098 ), the 973 Project of China (2010CB912802 and 2010CB529400).
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