RASSF1A methylation and K-ras and B-raf mutations and recurrent endometrial cancer.

BACKGROUND: Aberrations in mediators of Ras signaling may increase the risk of developing recurrent endometrial carcinoma. PATIENTS AND METHODS: Primary tumors of patients with (n = 44) and without (n = 44) recurrent stage I endometrioid endometrial carcinoma were compared regarding the presence of K-ras mutations (codons 12 and 13), B-raf mutations (V599), and RASSF1A gene promoter methylation. RESULTS: K-ras mutations were present in 18% of the patients independent of recurrent disease. No B-raf mutations were found. RASSF1A methylation was demonstrated in 85% of endometrial carcinomas, independent of recurrence. The presence of K-ras mutations and RASSF1A promoter methylation were not related, either directly or inversely. Analysis in premenopausal endometrial carcinomas demonstrated K-ras mutations in 40%, no B-raf mutations, and RASSF1A promoter methylation in 70% of the cases. RASSF1A methylation was also observed in samples of cyclic (n = 14), hyperplastic (n = 8), and atrophic (n = 13) endometrial tissues in 21%, 50% and 38%, respectively. CONCLUSIONS: RASSF1A methylation was observed in a high frequency in endometrioid endometrial carcinoma whereas K-ras and B-raf mutations were observed in a low frequency. No association was observed with the development of recurrent disease. High-frequency RASSF1A methylation in premenopausal carcinomas and an increased frequency in endometrial hyperplasia indicate that this may be an early event in endometrial carcinogenesis.

Aberrations in the progesterone receptor gene and the risk of recurrent endometrial carcinoma.

A case-control study was performed in order to determine whether expression of the progesterone receptor (PR) and/or aberrations of the PR gene contribute to the development of recurrent endometrial carcinoma. Primary tumours from 44 patients with recurrence of stage I endometrial carcinoma (patients) within 3 years after initial treatment were compared with tumours from 44 matched patients who were free of recurrence for a minimum of 3 years (controls). Paraffin wax-embedded primary tumours (n = 88) and recurrent tumours (n = 32) were analysed immunohistochemically for PR expression. A staining index (SI = 0-9) based on the staining intensity and the number of stained cells was calculated. DNA extracted from paraffin wax-embedded tissues was subjected to PCR-restriction fragment length polymorphism analysis (PCR-RFLP) for determination of the PROGINS DNA sequence alterations and the +331G/A-promoter polymorphism. Low PR expression (SI < 1.0) was observed in 7% of primary tumours derived from controls, 25% of primary tumours from patients with recurrence, and 38% of recurrent tumours. The expression of PR was significantly lower in primary tumours from patients with recurrence (SI = 4.0 +/- 0.5) than in the tumours in the control group (SI = 5.6 +/- 0.5) (T-test for paired analysis, p < 0.05). The PROGINS and +331G/A-promoter polymorphism were not related to age at diagnosis, tumour grade or myometrial invasion. The +331G/A-promoter polymorphism was present in 14% of primary tumours from patients without recurrence, compared with 17% of patients with recurrence. The PROGINS polymorphism was observed in 16% of primary tumours from patients without, and in 34% of patients with, recurrence (OR 2.6; 95% CI: 0.9-7.6). Most interestingly, patients who carried the PROGINS variant and in whom a PR-expressing tumour was diagnosed were at significantly enhanced risk of relapse (OR 4.7; 95% CI: 1.3-17.1). In conclusion, low PR expression tended to be associated with recurrent disease, and PR expression in tumours from patients carrying the PROGINS allele was predictive of the risk of recurrence.

Defective mismatch repair and the development of recurrent endometrial carcinoma.

OBJECTIVE: To investigate whether defective DNA mismatch repair (MMR) defines a subgroup at risk for recurrence in sporadic endometrial carcinoma patients. METHODS: Primary tumors from 44 patients with recurrent stage I endometrial carcinoma were compared after matching, with tumors of 44 patients being free of recurrence for minimal 3 years. Paraffin-embedded primary tumors (n = 88) and recurrent tumors (n = 32) were subjected to immunohistochemical analysis for hMSH2 and hMLH1 expression. Subsequently, a staining index (SI = 0-9) was calculated based on staining intensity and quantity. DNA was extracted from paraffin-embedded tissues, and promoter methylation of hMLH1 was determined by nested methylation-specific PCR (MSP). Microsatellite instability (MSI) was assessed by BAT-26 or BAT-25. RESULTS: Low hMSH2 expression was observed in 2% of primary tumors of control patients without recurrence, 14% of primary tumors of patients with recurrence, and 0% of recurrent tumors. Low hMLH1 expression was observed in 32%, 19%, and 22%, respectively. hMLH1 gene promoter methylation was detected in 50%, 47%, and 32%, and MSI was found in 16%, 14%, and 30%, respectively. No significant differences were found between primary tumors of patients with and without recurrence with respect to hMSH2 and hMLH1 expression, hMLH1 promoter methylation, and MSI. When primary and recurrent tumors were compared, there was an increased correlation of hMLH1 methylation with low hMLH1 expression and MSI in recurrent tumors. CONCLUSION: MSI, hMLH1 promoter methylation, and the expression of hMLH1 and hMSH2 are not predictive for the development of recurrent stage I endometrial carcinoma. In the progression of tumor, "de novo" hMLH1 methylation rarely occurs, instead there is further derailment of the MMR pathway in affected tumors.