The importance of biological factors (bcl-2, bax, p53, PCNA, MI, HPV and angiogenesis) in invasive cervical cancer

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Abstract

Objective: The present study was designed to analyse the relationship between apoptosis related proteins (bcl-2 and bax), tumour suppressor protein p53, proliferation markers (PCNA and mitotic index), human papillomavirus (HPV) and angiogenesis in cervical cancer and their impact on clinical outcome. Study Design: Tumours from 111 patients were assessed by immunohistochemistry for the expression of bcl-2, bax, p53 and PCNA, by PCR for the presence of HPV-DNA, for the quantification of the mitotic index and the microvessel density (CD 31). The results were correlated with various histopathologic characteristics and survival. Results: The multiple Cox’s regression analysis for overall survival of all prognostic variables gave as best model: bcl-2 (P<0.001), lymphovascular permeation (P=0.004), mitotic index (P=0.019), tumour grade (P=0.048) and FIGO stage (P=0.070). Subanalysis was performed for the patients where the lymph node status was known (n=79). Adding the lymph node status gave as best model for overall survival bcl-2 (P=0.001), lymphovascular permeation (P=0.003) and mitotic index (P=0.044). However, they hardly influenced the association. Conclusion: In the apoptotic pathway of cervical cancer, bcl-2 is one of most important proteins. It can probably not only mediate cell death but also regulate cell growth. A better understanding of their relations will probably provide the basis for more rational cancer therapies in the future.

Introduction

The clinical behaviour of invasive uterine cervix carcinoma is diverse and includes tumours of both relatively slowly and rapidly progressing diseases. The development and progression of cervical tumours is likely to be associated with alterations in apoptosis (programmed cell death), disturbance in immune surveillance, increased cell growth and/or loss of growth suppression (uncontrolled proliferation) [1], [2].

The bcl-2 gene family seems to act as a regulator of the apoptotic pathway [3], [4], [5]. The two most important apoptosis regulating proteins of this family are most likely bcl-2 and bax. The former is a member of the anti-apoptotic family and latter of the pro-apoptotic family [6]. Together they probably act as a rheostat for the cell death program [4], [6].

The normal or wild type p53 protein plays a role in the regulation of the cell cycle. Mutation of this gene is associated with carcinogenesis and tumour progression. In eliminating cells with damaged DNA, p53 plays an important role not only by inducing apoptosis but probably also by increasing the sensitivity of the tumour cell to apoptosis [7].

One of the oldest ways to assess tumour proliferation is by counting the number of mitotic figures [8], [9], [10]. Another way to study proliferation is by immunohistochemical assessing of the proliferating cell nuclear antigen (PCNA) [10]. Wild-type p53 can inhibit cell proliferation by blocking entry into the S-phase of the cell cycle [11]. There is a good correlation between tumour-cell kinetics measured by the PCNA index and the p53 gene in several malignancies [12].

Human papillomaviruses infections, especially types 16 and 18, are strongly associated with the development of cervical cancer. The E6 protein produced by HPV 16 and 18 forms a complex with wild-type p53 protein and rapidly degrades its functions [13]. The inactivation of the wild-type p53 protein and/or mutations of the p53 gene could play a leading role in the malignant transformation and tumour progression of cervical cancer.

Neovascularisation is essential for tumour growth beyond 1–2 mm3 [14], [15], [16]. A correlation between apoptosis and tumour angiogenesis might be expected in human solid tumours. Because angiogenesis inhibitors can induce and sustain the dormancy of experimental primary tumours and micrometastases by elevating the incidence of apoptosis in tumour cells, while the proliferation rate remains unchanged [17], [18], [19]. Also the incidence of spontaneous apoptosis in tumour cells is inversely related to the extent of neovascularisation [20].

The current study was performed to evaluate the correlation between bcl-2, bax, p53, PCNA, mitotic index (MI), HPV type and microvessel density with clinicopathological findings in cervical cancer. Furthermore, the relationship between these biological variables and their impact on clinical outcome c.q. survival is investigated.

Section snippets

Patients and follow-up

One hundred and eleven patients with primary cervical cancer were included in the study. All patients were evaluated at the University Hospital of Antwerp or at the General Hospital Saint Camillus, Saint Augustinus, between 1979 and 1995. The mean age was 52 years (range 24–82). Representative tumour specimens were used for immunohistochemistry and PCR analysis. The histopathologic features of the surgical specimens were classified according to the World Health Organisation criteria and the

Results

The characteristics of this patient cohort, including age, stage, histology, lymph node status, grade and vascular-lymphatic space involvement are listed in Table 1.

Discussion

The significance of the apoptotic pathway in the development and progression of human malignant tumours has become a major topic of discussion during the last years [2]. One of the proteins that have a regulating role in the programmed cell death is encoded by the bcl-2 gene. Previous studies regarding bcl-2 and invasive cervical cancer have underlined the cell survival-promoting activity of this protein and showed in accordance with the current study that bcl-2 is a strong independent

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    Presented at the Wertheim Operation Centenary Scientific Meeting 1898–1998, University of Vienna, Austria, 19 September 1998 and awarded with the Wertheim Award.

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