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An Interleukin-6 Gene Promoter Polymorphism Influences the Biological Phenotype of Ovarian Cancer

Departments of Obstetrics and Gynecology, University of Vienna Medical School, A-1090 Vienna, Austria [L. A. H., C. G., A. R. R-R., S. L., R. Z., C. B. T.]; Friedrich-Alexander-University Erlangen, Nuernberg, Germany [L. A. H., S. A., S. M., M. W. B.]; and Martin-Luther-University Halle, Wittenberg, Germany [H. K.]

	ABSTRACT

Top ABSTRACT Introduction Materials and Methods Results and Discussion REFERENCES

 
Interleukin (IL)-6 is known to be involved in the pathogenesis of ovarian cancer. We investigated a common G/C polymorphism at position -174 of the IL-6 gene (IL6) promoter in 121 patients with ovarian cancer using pyrosequencing. Presence of at least one mutant allele was associated with early tumor stage as well as an expanded length of disease-free (DFS) and overall survival with a dose-dependent effect regarding the carriage of 0, 1, and 2 alleles. In a multivariate Cox regression model incorporating tumor stage and residual tumor mass, presence of the -174 C allele of IL6 was the best predictor of DFS. We conclude that the mutant -174 C allele of IL6 influences the biological phenotype of ovarian cancer because it is associated with early stage and improved DFS and overall survival.

	Introduction

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IL-6,² a phosphorylated glycoprotein containing 185 amino acids, is a pleiotropic cytokine involved in different physiological and pathophysiological processes, such as inflammation, bone metabolism, synthesis of CRP, and carcinogenesis (1 , 2) .

With respect to ovarian cancer, in vitro experiments have shown that various ovarian carcinoma cell lines produce IL-6, which is thought to be involved in ovarian carcinogenesis by mediating host immune responses to the disease (2, 3, 4) . In humans, IL-6 levels in ascitic fluid were found to be significantly elevated in patients with ovarian cancer compared with controls (5) . Ascitic IL-6 levels were also associated with peripheral CRP levels, which are thought to serve as additional prognostic parameters by themselves (6 , 7) . Furthermore, IL-6 serum levels have been associated with an increased tumor burden and impaired survival (8, 9, 10) . A possible use as additional tumor marker of serum IL-6 has been suggested.

The human IL-6 gene (IL-6) is mapped to chromosome 7p21–24 with an upstream promoter containing 303 bp (11) . A common G/C polymorphism of the IL-6 promoter on position -174 has been investigated in a wide variety of diseases, including multiple myeloma, coronary heart disease, and endometriosis (12 , 13) . Of note, the -174 C IL6 polymorphism has been shown to influence in vivo protein expression (6 , 14) .

Thus, based on the important role of IL-6 in ovarian carcinogenesis, IL-6 can be regarded as a candidate gene for ovarian cancer. In the present study, we hypothesized that the -174 C IL6 polymorphism is associated with the biological phenotype of ovarian cancer.

	Materials and Methods

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Patients.
The present study included examinations of the -174 C IL-6 polymorphism in 121 Caucasian patients with ovarian cancer Union Internationale Contra Cancrum stages I (n = 50), II (n = 16), and III (n = 55), who were treated between April 1986 and July 1999 at the University of Vienna Medical School, Department of Gynecology and Obstetrics. Lymph node status was available in 89 patients. Reasons for not performing lymphadenectomy were patients’ poor medical condition (n = 9), advanced disease (n = 17), and other not specified reasons (n = 6). Lymph node involvement was noted in 27 cases.

Median age at the time of diagnosis of ovarian cancer was 54 (range 26–86) years. Histologically, 67 tumors were graded as serous adenocarcinomas, 25 as mucinous adenocarcinomas, 25 as endometrioid adenocarcinomas, 9 as clear cell carcinomas, and 4 as undifferentiated carcinomas. Thirty-one tumors were graded as well differentiated, 34 as moderately differentiated, and 56 as poorly differentiated. The median (range) duration of follow-up was 38.1 (4–163) months. Forty-four patients developed recurrent disease; 23 patients died of cancer-related death during the observation period. CA 125 and CRP serum levels were evaluated before surgery. Patients were treated and followed up according to established protocols.

Genotyping.
Pyrosequencing was performed according to established protocols as described previously (15) . CA 125 and CRP serum levels were measured before surgery in all patients.

Statistics.
Where appropriate, ² analysis was performed. Odds ratios and 95% confidence intervals are given accordingly. Survival probabilities were calculated by the product limit method of Kaplan and Meier using the Log-rank test. The results were analyzed for the end point of DFS and OS. Survival times of patients still alive were censored with the last follow-up date. A multivariate Cox regression model was performed comprising tumor stage, lymph node involvement, residual tumor mass, and -174 C IL6 polymorphism. Ps < 0.05 were considered statistically significant. For statistical analysis, we used the SPSS statistical software system (SPSS 11.0; SPSS, Inc., Chicago, IL).

	Results and Discussion

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The distribution of -174 C IL-6 genotypes in patients with ovarian cancer was in Hardy-Weinberg equilibrium. Associations between the -174 C IL-6 polymorphism and several clinicopathological parameters are depicted in Table 1 . The presence of at least one mutant allele of IL6 was associated with early tumor stage [pT1 versus pT2 and pT3; odds ratio 0.33 (95% confidence interval 0.12–0.88); P = 0.036].

View larger version (19K): [in this window] [in a new window] [Download PPT slide]   Fig. 1. Kaplan-Meier analysis of disease-free survival of patients with ovarian cancer with 0, 1, and 2 mutant alleles of the common G/C polymorphism of the IL-6 gene (IL6).

View larger version (20K): [in this window] [in a new window] [Download PPT slide]   Fig. 2. Kaplan-Meier analysis of overall survival of patients with ovarian cancer with 0, 1, and 2 mutant alleles of the common G/C polymorphism of the IL-6 gene (IL6).

The number of genetic association studies in all fields of medicine has risen substantially in recent years. Testing of mutations and polymorphisms in candidate genes of ovarian cancer revealed a close association between the risk for and survival length of ovarian cancer and BRCA1, BRCA2, p53, and nm23-H1 (16, 17, 18) . As published previously for breast cancer, microarray techniques allowing for the simultaneous detection of a high number of polymorphism might provide prognostic information in addition to established clinical parameters (19) . In our present study, we ascertained a significant association between the presence of the mutant -174 C allele and early tumor stage. In addition, carriage of the mutant -174 C allele was associated with improved DFS and OS in a univariate and multivariate regression model. Of note, the -174C IL6 polymorphism was the best predictor of DFS in this model. Our data support the assumption that IL-6, most likely via host-immune response modification, is involved in local tumor growth and prognosis of ovarian cancer.

Overall, it can be stated that the -174 G/C promoter polymorphism of IL-6 influences the biological phenotype of ovarian cancer. These data confirm IL-6 as a candidate gene for ovarian cancer and suggest that the -174G/C polymorphism of IL6 should be included in future analyses of the genetic background of this disease.

	FOOTNOTES

 
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ To whom requests for reprints should be addressed, at Department of Obstetrics and Gynecology, University of Vienna Medical School, Waehringer Guertel 18-20, A-1090 Vienna, Austria. Phone: 43 1 40400 2962; Fax: 43 1 40400 2911; E-mail: lukas.hefler{at}akh-wien.ac.at

² The abbreviations used are: IL, interleukin; CRP, C-reactive protein; OS, overall survival; DFS, disease-free survival.

Received 2/ 8/03. Accepted 5/ 1/03.

	REFERENCES

Top ABSTRACT Introduction Materials and Methods Results and Discussion REFERENCES

 

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