A B S T R A C T The failure of certain adrenal tumors to respond to ACTH was investigated in vivo by administration of corticotropin-( 1-24) -tetracosapeptide (ACTH1-2) and dexamethasone and in vitro by studying the binding properties of ACTH1-2 and prostaglandin E1 (PGE1) and their effect on adenylate cyclase activity of the tumors' crude membranes; in addition, in five cases the stimulation of cortisol production in isolated adrenal cells by both hormones and dibutyryl cyclic adenosine 3',5'-monophosphate (cAMP) was also studied. The results obtained in 13 hormone-producing tumors of the human adrenal cortex, i.e. 10 carcinomas and 3 adenomas, were compared with those found in normal human adrenal glands.According to the adenylate cyclase responses to ACTH1-and PGE1, the tumors fall into different categories. In the first group are six tumors in which the adenylate cyclase was stimulated by both ACTHl-2X and PGE1; in addition specific binding could be demonstrated for the two hormones in all six. The binding affinity for 'I-ACTH1-2 was found to be about 10 times higher than that for 'I-ACTHu-24. In the one tumor in which the experiment was performed, bound 'I-ACTH1-X was displaced by ACTH1-1o. These results are similar to the ones found in normal human adrenal preparations. For two tumors of the group in which ACTH did not increase steroidogenesis in vivo, the biochemical abnormality might be located beyond cAMP formation.A preliminary report of this work was presented at the 55th Meeting of the Endocrine