The lysosomal acidic protease cathepsin D, a recognized independent predictor of prognosis in human breast cancer, has not been studied widely in patients with endometrial adenocarcinoma. Cathepsin D levels (52-kD precursor plus 48-kD intermediate and 34/14-kD mature form) were measured in tumor cytosols from 26 hysterectomy specimens by immunoradiometric assay. Significant correlation between cathepsin D levels and tumor differentiation was noted with linear increase in cathepsin D from 8 pmol/mg (standard error of the mean [SEM], 1.73 pmol/mg) for Grade I tumors to 28 pmol/mg (SEM, 3.91 pmol/mg) for Grade III tumors. A group of four papillary serous carcinomas showed relatively high cathepsin D levels reaching 39 pmol/mg. A significant stepwise increase in cathepsin D levels was associated with increased depth of myometrial invasion. Noninvasive tumors averaged 7 pmol/mg (SEM, 4.0 pmol/mg); intramural tumors averaged 15 pmol/mg (SEM, 2.45 pmol/mg); and transmural invasive tumors averaged 30 pmol/mg (SEM, 3.72 pmol/mg). There was no significant correlation of cathepsin D levels with age, estrogen/progesterone receptor hormone status, clinical stage, and lymph node metastasis. Cathepsin D levels correlate significantly with tumor differentiation and myometrial invasiveness and may show promise as a clinically useful adjunct to prognosis assessment and the planning of therapy for patients with endometrial adenocarcinoma.