The putative presence of a mutation in codon 12 of the K-ras gene was investigated in the endometrium of tamoxifen (TAM) and toremifene (TOR)-treated breast cancer patients. DNA was extracted from fresh cytologic samples of the endometrium in 86 TAM and 21 TOR-treated breast cancer patients. Mutations were detected by enriched PCR and an enzyme-linked mini-sequence assay (ELMA). K-ras mutation was found in 35 TAM-treated endometrial samples, and in only one TOR-treated endometrium (Po0.003). In 24 premenopausal patients, K-ras mutation was found in seven (43.8%) of 16 patients with less than 47 months of TAM treatment, while none was found in eight patients with more than 48 months of TAM treatment (Po0.03). In 62 postmenopausal-amenorrheic patients, K-ras mutation was found in three (15.8%) of 19 patients with less than 23 months of TAM treatment, while it was found in 16 (61.5%) of 26 patients with 24 -47 months of TAM treatment and nine (52.9%) of 17 patients with more than 48 months of TAM treatment (P ¼ 0.002). The presence of K-ras mutation is significantly influenced by the duration of TAM treatment and menstrual status of the patients. TOR may have a lower potential genotoxicity than TAM.

Proliferation and apoptosis of the endometrium throughout the menstrual cycle were evaluated. Sections from 30 premenopausal women were examined using antibodies of c-jun protein, c-fos protein, estrogen receptors alpha and beta (ER-α and ER-β), progesterone receptor (PR), and Ki-67. Apoptotic cells were identified using a modified terminal deoxynucleotidyl-transferase-mediated biotinylated deoxyuridine triphosphate nick-end labeling (TUNEL) method. The cyclic changes of c-jun protein, c-fos protein, ER-α, PR, and Ki-67 were shown in glandular epithelial cells. Although the stromal expression of ER-α decreased during the secretory phase, a high stromal expression of c-jun protein and PR was still observed during the late secretory phase. The expression of ER-β appeared lower as compared with that of ER-α, without a cyclic change. The apoptotic index was significantly elevated in the glandular epithelial cells of the late secretory phase, whereas a few apoptotic cells were detected in the stromal cells at any stage of the cycle. The cyclic change of c-jun protein probably plays an important role in proliferation and apoptosis of glandular epithelial cells. The persistent stromal expression of c-jun protein and PR is thus considered to prevent stromal cells from entering into apoptosis during the late secretory phase.

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