Membrane receptors for luteinizing hormone-releasing hormone (LH-RH), somatostatin, and prolactin (PRL) were investigated in the Dunning R-3327H rat prostate adenocarcinoma specimens after in vivo treatment with microcapsules of the agonist [D-TrpWLH-RH (Kd = 0.72 nM; B, = 161 fmol/mg) were also detected. Treatment with either analog reduced Bx by 50%, but a much greater reduction of PRL binding capacity was revealed after in vitro dissociation of the bound endogenous PRL by MgC12. The dramatic fall in the total number of PRL receptors after combination treatment with both analogs could be partially responsible for the decrease in the weight and volume of prostate tumors. The findings support the concept that analogs of LH-RH and somatostatin can inhibit tumors directly through their own respective receptors. One of several mechanisms of the antineoplastic activity of these analogs could be the elimination of tumor growth-promoting effect of PRL by the reduction of the total number of PRL receptors.The primary treatment of advanced prostate carcinomas is based on the elimination of testicular androgen secretion by hormonal manipulation (1-3). A new endocrine therapy for prostate cancer makes use of superactive agonists of luteinizing hormone-releasing hormone (LH-RH) that after chronic administration produces an inhibition of circulating testosterone (2-5). The development of once-a-month injectable microcapsules of an analog of LH-RH with Dtryptophan in position 6 ([D-Trp6]LH-RH), in a biodegradable polymer, provides'a practical, convenient, and efficacious way of administration (3-6).Other hormones in addition to testosterone could influence the growth of prostatic carcinoma. Prolactin (PRL) could be involved in prostate cancer as a cofactor (4, 5). It was demonstrated that 'PRL potentiates the effects of testosterone on the prostate (7,8), and PRL receptors are found in normal and cancerous prostate tissue (9,10 890The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

A sensitive micromethod for the determination of free and total prolactin receptors in normal or malignant tissues has been developed. Positive and negative quality controls are incorporated in the procedure. Either whole tissue or the pellet fraction remaining from tissue that had undergone processing for estrogen receptors can be used. Crude microsomal and plasma membrane fractions obtained by homogenization and differential centrifugation are incubated with labeled prolactin in the presence or absence of increasing amounts of unlabeled hormone. The labeled ligand is prepared by a stoichiometric iodination procedure in which one atom of iodine-125 is incorporated into one molecule of the hormone, resulting in an intact labeled prolactin with a high specific activity of 170-186 muCi/micrograms (1 Ci = 37 GBq). Human prolactin labeled by this procedure has much greater specific binding capacity to various rat tissues than does iodinated rat prolactin. This technique permits an accurate measurement of prolactin receptors in as little as 50 micrograms of membrane protein. Highest levels of free and total prolactin receptors were found in the liver of 60-day-old female rats that served as a positive control. Liver of immature 21-day-old male rats, devoid of prolactin receptors, was used as a negative control. The amount of detectable free receptors was dependent on the level of circulating plasma prolactin. In 3-day postpartum lactating rats with high prolactin levels in plasma, all prolactin receptors in the mammary glands were found to be occupied, and no free receptors could be detected. When these receptors were desaturated from the endogenous prolactin by exposure to 3 M MgCl2, one class of receptors in a high quantity (1.75 nmol/mg of protein) and with a moderate affinity (Kd = 6.41 X 10(-9) M) was detected. A similar type of receptor was found in the mammary glands of rats at midpregnancy and of cycling adult female rats. In malignant rat mammary tissue, however, fewer receptors (27 pmol/mg of protein) but with a very high affinity (Kd = 6.8 X 10(-14) M) were detected. Normal ventral and dorsolateral rat prostate contained two classes of prolactin receptors (Kd = 3.46 X 10(-10) M and 1.93 X 10(-8) M). In the cancerous rat prostate, however, only one of these two classes of receptors was detected, and the number was smaller.

Using a sensitive micromethod for the determination of prolactin (PRL) binding sites based on 125I-human PRL ligand, PRL receptor levels in specimens of benign prostatic hyperplasia (BPH) and human prostate cancer were estimated by the one-point assay system. Ten of 19 BPH specimens (53%), showed significant PRL binding, four being in the 9-12 fmol/mg range. All ten of these cases had an histological diagnosis of nodular glandular hyperplasia. Of ten adenocarcinomas examined, four samples (40%) exhibited positive PRL binding, the highest receptor levels being 10.2 fmol/mg protein. To characterize the receptors from BPH membranes, samples were then separately pooled according to the results obtained in one-point assays. In the PRL-negative pool no displacement could be detected. In the PRL-positive pool, the Scatchard analysis revealed one class of receptors with an average affinity Kd = 1.1 X 10(-9) M and capacity Bmax = 287 fmol/mg protein. In the prostate cancer specimens, luteinizing hormone-releasing hormone receptors with a high affinity and a low capacity were also found. The results indicate the presence of prolactin receptors in prostate cancer and in BPH. The clinical implications of such findings are not clear, but it is possible that a certain proportion of BPH and prostate cancers might be in part PRL dependent. Further studies are necessary to ascertain this hypothesis in an attempt to improve the treatment of BPH and prostate cancer.

Nine new analogues of acetyl-CCK-heptapeptide (Ac-Tyr(SO3H)2-Met3-Gly4-Trp5-Met6-Asp7-Phe8-NH2 ) were synthesized by solid-phase methodology. In a first series, the Asp7 residue was replaced by hydroxy amino acid sulfate esters. In another series, Gly4 was substituted by D-Ala, while Trp5 and Met6 were replaced by their D enantiomer. The introduction of the sulfate ester was performed with a new, mild, crystalline, and stable reagent, pyridinium acetyl sulfate. Each analogue that contained Tyr(SO3H)2 and a hydroxy amino acid sulfate ester [Ser(SO3H), Thr(SO3H), or Hyp(SO3H)] in position 7 proved to be more potent (1.9, 1.7, and 3.0 times, respectively) than CCK-8 in vitro (isolated gallbladder strips). While devoid of gastrin-like activity in vivo, these analogues had potent anticonvulsive activity. The analogues containing a D-amino acid residue were less potent than the parent compound in vitro. The D-Ala4 replacement, however, yielded a compound that was 40% as potent as CCK-8 in the in vitro test but showed prolonged duration of action on sphincter Oddi. While the 7-substituted Ac-CCK heptapeptides are among the most potent CCK analogues reported so far, the D-Ala4 replacement resulted, for the first time, in prolonged activity in vivo.

The effects of intracerebroventricular administration of an 80 pmole dose of cholecystokinin octapeptide sulfate ester (CCK-8-SE) were tested on the dopamine (DA), norepinephrine (NE) and serotonin (5-HT) turnovers of the hypothalamus, mesencephalon, amygdala, septum, striatum and cerebral cortex in rats. CCK-8-SE in an 80 pmole dose decreased the DA turnovers of the hypothalamus, mesencephalon, amygdala and septum, while it increased that of the striatum. The NE turnovers were increased in the hypothalamus and amygdala, but decreased in the striatum. The 5-HT turnover decreased only in the hypothalamus.