The new antiestrogen RU 16117, at doses of 8 or 24 mcg daily, had been shown to completely prevent the development of rat mammary cancer when given from the day after 7,12-dimethylbenz(a)anthracene (DMBA) administration. This study was undertaken to investigate the effect of this compound on the growth of DMBA-induced tumors which had already developed in Sprague-Dawley rats. The effect was compared with that of castration. Levels of receptors for 17beta-estradiol (E2), progesterone, and prolactin (PRL) were correlated with the response. At about 3 months after DMBA administration animals with palpable tumors were selected. The rats were then treated daily for 4 weeks with .1, .5, 2.5, or 12.5 mcg E2 or with 2, 8, or 24 mcg RU 16117 injected in .1 ml of 1% gelatin in .9% NaCl. Controls were injected with the vehicle alone. For comparison, a group of rats were ovariectomized. After 4 weeks' treatment rats were killed, blood collected, and a cytosol was prepared from tumor tissues. Binding assays and radioimmunoassays were done. 8 and 24 mcg doses of RU 16117 led to 45 and 65% inhibition of tumor number, respectively, and tumor size was markedly reduced. Lower doses had less effect. Ovariectomy had an effect similar to that of 24 mcg RU 16117. E2 doses did not change the number or size of tumors. Decreased levels of receptors for E2, progesterone, and PRL were found in the tumors remaining after ovariectomy. The 24 mcg dose of RU 16117 had a similar effect on levels of E2 and PRL receptors. It was considered likely that RU 16117 exerts its inhibitory activity at both the hypothalamic-pituitary and tumor levels.