P53 AND Ki67 IN ADRENOCORTICAL TUMORS
J. Arola, K. Salmenkivi, J.Liu, A.I.Kahri and P.Heikkilä Department.of Pathology Helsinki University Central Hospital, P.O.Box 21, FIN-00014 University of Helsinki, Finland
ABSTRACT
The p53 tumor-supressor gene has been reported as the most frequent genetic abnormality seen in human malignancies. Here we studied immunohistochemically the expression of p53 in a large series of adrenocortical tumors. The proliferative activity was assessed by the expression of Ki67. Tumor material consisted of 60 adrenocortical adenomas and 27 adrenocortical carcinomas. A tumor was scored as positive for p53 if more than 10% of the cells showed nuclear staining. All adrenocortical adenomas were negative for p53 and the percentage of Ki67 positive cells was mostly 1-2% but never exceeded 5%. Hormonal activity did not reflect the proliferation index. Adrenocortical carcinomas, however, behaved differently depending on hormonal activity. 10/13 of non-functional , 0/3 Conn’s, 3/7 Cushing’s and 3/4 virilizing carcinomas were positive for p53. The proliferative activity was also higher in non-fuctional carcinomas compared with hormonally active tumors. Our data show that majority of adrenocortical carcinomas are positive for p53, whereas all adenomas are negative. Hormonal activity of carcinomas reflects both p53 status and proliferation index. Thus, immunohistochemical levels of p53 and Ki67 are higher in hormonally inactive adrenocortical carcinomas.
INTRODUCTION
The p53 tumor-supressor gene has been reported as the most frequent genetic abnormality seen in human malignancies. The role of p53 in adrenocortical tumorigenesis has recently been a subject of great interest. Ten years ago, germline mutations in p53 were identified in families with the Li- Fraumeni syndrome 1. This tumor syndrome has a high incidence of adrenocortical carcinomas. Mutations of p53 gene and overexpression of protein have been reported in adrenocortical tumors 2,3,4. P53 positivity has mainly been found in adrenocortical carcinomas 5, however, high levels of expression in aldosterone producing adenomas has also been suggested 6.
Here we studied immunohistochemically the expression of p53 in adrenocortical tumors. The role of malignancy and steroidogenesis was estimated since our tumor material consisted of large number of hormonally active and inactive, as well as benign and malignant tumors. The proliferative activity was assessed by the expression of Ki67.
MATERIALS
Tissue materials were obtained during the operations performed at the Department of Surgery, Helsinki University Central Hospital. Adrenocortical tumours included 15 nonfunctional adrenocortical adenomas, 20 Conn’s adenomas, 20 Cushing’s adenomas, 6 virilizing adenomas, 13 nonfunctional adrenocortical carcinomas, 3 Conn’s carcinomas, 7 Cushing’s carcinomas and 4 virilizing carcinomas. Malignancy of the adrenocortical tumors was diagnosed according to the criteria of Weiss. 7
METHODS
Sections were cut from paraffin blocks and deparaffinized in xylene
and rehydrated in a series of graded alcohols. Sections were pretreated in a microwave oven in 10 mmol/l citrate buffer, pH 6.0, at 600 W for 20 minutes. Endogenous peroxidase activity was blocked in 0,5% H2O2 for 30 minutes. The primary monoclonal antibodies for p53 (Santa Cruz, Santa Cruz, CA, U.S.A.) and Ki67 (Immunotech, Marseille, France) at 1:50 dilutions were incubated over night. The detections were performed using Vecastain ABC kit (Vector Laboratories, Burlingame, CA) according to the manufacturer’s instructions. Sections were counterstained with haematoxyline. Immunoreactivity was assessed by two trained pathologists (J.A. and K.S.) A tumor was scored as positive for p53 if more than 10% of the cells showed nuclear staining, where as proliferation index was reported as percetage of positive staining nuclei.
RESULTS
All adrenocortical adenomas, regadless of functional status were negative for p53. Only occasional positive nuclei were detected, but the precentage of positive staining nuclei never exceeded 1%. The percentage of Ki67 positive cells was mostly 1-2% but never exceeded 5%. Hormonal activity did not reflect the proliferation index either (Table).
Adrenocortical carcinomas, however, behaved differently depending on the hormonal activity. Highest p53 expressions were found in non-functional and virilizing carcinomas. 10/13 of non-functional and 3/4 virilizing carcinomas were positive for p53. None of the Conn’s carcinomas (n=3) were p53 positive and only 3/7 Cushing’s carcinomas were positive.
The proliferative activity was higher in non-fuctional carcinomas compared with hormonally active tumors. Proliferation index was from 10- 50%, median being 30% in non-functional carcinomas, whereas, all the hormonally active carcinomas, except one Cushings carcinoma, expressed only 10-20% Ki67 positive tumor cells.
| n | P53+ | Ki67+ | |
|---|---|---|---|
| Adrenocortical adenomas | |||
| Conn's | 20 | 0 | 1-2% |
| Cushing's | 20 | 0 | 1-5% |
| Virilizing | 6 | 0 | 1-3% |
| Non-functional | 15 | 0 | 1-2% |
| Adrenocortical carcinomas | |||
| Conn's | 3 | 0 | 10-20% |
| Cushing's | 7 | 3 | 10-40% |
| Virilizing | 4 | 3 | 10-20% |
| Non-functional | 13 | 10 | 10-50% |
DISCUSSION
Our data show that majority of adrenocortical carcinomas are positive for p53, whereas all adenomas regardless of functional status are negative. Hormonal activity of carcinomas reflects p53 status of the tumor. Thus, immunohistochemical positivity of p53 is high in non-functional carcinomas as well as in androgen-producing adrenocortical carcinomas. This suggests that p53 may have a major role in the neoplastic process of these tumors. It also serves as an additional tool for histopathologist to diagnose between benign versus malignant adrenocortical tumor.
We also report that nonfunctional adrenocortical carcinomas have higher proliferative activity compared with hormonally active carcinomas.
This may mean that hormonally inactive tumors grow more rapidly and thus are more aggressive.
ACKNOWLEDGEMENTS
Ms Eija Heilio is thanked for her technical assistance. This work was supported by the Helsinki University Central Hospital Research Contract No TYH 9107.
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