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In an effort to obtain more information on the structure-activity relationship among the 7 alpha-(phenylthio)androstenedione inhibitors of the enzyme aromatase, a series of compounds containing both electron-donating and electron-withdrawing ring substituents was synthesized and tested for aromatase inhibitory activity. No linear correlation between substituent electronic effects and inhibitory activity was observed. The halogen-containing compounds, particularly 8, appeared to be quite potent inhibitors. The 125I analogue of 8 was synthesized in order to evaluate the possibility of side-chain elimination under the assay conditions. Approximately 90% of [125I]-8 remained intact for up to 1 h under assay conditions.

A new class of compounds, polyiodinated sterol esters, has recently been synthesized and found to have characteristics suggesting potential value as site-specific hepatic computed tomographic (CT) contrast agents. A prototype compound, cholesteryl iopanoate, was administered intravenously to rabbits in doses of 65 mg I/kg. The compound was found to significantly enhance hepatic parenchyma to a maximal level of 65 Hounsfield units (HU) above base-line values without observable splenic enhancement. A subsequent study comparing the administered dose with the change in CT attenuation values of liver demonstrated that near-maximum enhancement reaching 22 HU was achieved at a dose of 30 mg I/kg within 24 h after injection. The enhancement characteristics suggest the mechanism of hepatic accumulation may not be mediated solely by the reticuloendothelial system. Administration of cholesteryl iopanoate (30 mg I/kg) to rabbits with hepatic tumors (V x 2 adenocarcinoma) resulted in the CT imaging of tumors as small as 2 mm in diameter. Further investigation of this new class of lipid-soluble contrast agents seems warranted.

Analogues of diacylglycerol containing a 3-(3-amino-2,4,6-triiodophenyl)-2-ethylpropanoyl or 3-(3-amino-2,4,6-triiodophenyl)propanoyl group in the 2-position (1a and 1b, respectively) were synthesized and shown to compete with [3H]phorbol dibutyrate [( 3H]PDBu) for binding in a crude rat brain preparation. Phorbol diesters have been shown to bind specifically to protein kinase C and the PDBu receptor has been copurified with protein kinase C activity. The four diastereomers of 1a (1c-f) were synthesized from chiral starting material and studied in the same assay. The affinities for the [3H]PDBu binding site of 1a, 1b, and two isomers of 1a with naturally occurring L configuration were comparable to that of 1-oleoyl-2-acetyl-rac-glycerol (OAG), but the D isomers of 1a were essentially inactive. The chirality of the side chain did not influence the binding affinity. Activation of protein kinase C by 1a, 1c, and 1e demonstrated the same stereochemical requirements, but none were as active as OAG. For the 1,3-isomers 2, 2a, and 2b, the competitive binding studies gave different results. The racemic mixture and the D isomer, 2b, were able to compete for binding, but the L isomer, 2a, did not compete. These studies demonstrate that diacylglycerol binding to and activation of protein kinase C is stereospecific for the glycerol backbone, but not the side chain. Furthermore, the D-1,3-isomer must exist in a conformation such that the acyl and hydroxyl oxygens assume a spatial relationship similar to that in the L-1,2-isomers.

The activity of 33 steroids, injected intramuscularly, was determined in a castrate rat assay utilizing the responses of the seminal vesicle glands and ventral lobe of the prostate glands as independent parameters of androgenicity and the response of the levator ani muscle as an index of anabolic activity. Testosterone propionate was the reference standard. None of the compounds possessed high androgenic or anabolic activity. The myotrophic and average androgenic relative potencies of the 8 most active analogues ranged from 30 to 89% and 3.1 to 39%, respectively. The most active anabolic compound was the 1-dehydro-3-keto derivative of 17α-methyl-5α-androstan-17β-ol while the widest separation between anabolic and androgenic activities, myotrophic to androgenic ratio of 11.3, was obtained with the 2-dehydro-2-formyl derivative. The effect of various modifications of the A-ring on biological activity are outlined in detail.

19-Iodocholesterol- 125 I was synthesized for study as a possible agent for photoscanning the adrenal gland and associated tumors. In contrast to previous radioiodinated steroids, it was found to be much less prone to rapid in vivo deiodination. Preliminary tissue distribution studies and biochemical analyses have revealed a marked similarity in the behavior of the radioiodinated derivative with the natural steroid. The concentration of radioactivity in the adrenal cortex of dogs at 48 hours was found to greatly exceed that found in other organs.

SummaryA new inhibitor of cholesterol synthesis, SC-11952, closely related structurally to cholesterol, has been characterized in terms of its effects on acetate and meva-lonate metabolism in vitro. The compound did not reduce acetate conversion to CO2, fatty acids, or keto-acids but did inhibit incorporation of this substrate into cholesterol. Inhibition of mevalonate conversion to cholesterol was less than when acetate was the substrate. The similarity of the effects of SC-11952 to those produced by cholesterol feeding is discussed.

Previous studies with progesterone-4-14C suggested that a radioiodinated analog might be useful for photoscanning the adrenal gland and associated tumors. 21-Iodoprogesterone was prepared by conversion of deoxycorticosterone to its mesylate and subsequent displacement with iodide. Depending on the conditions for mesylation, 21-chloroprogesterone could also be isolated. 21-Iodopregnenolone acetate was obtained in a similar manner from the 21-bromo analog which in turn was prepared in good yield from the enol acetate of pregnenolone acetate. The radioiodinated steroids were prepared by isotope exchange with sodium iodide-125 in refluxing acetone. Tissue distribution studies showed that the radioactivity concentrated mainly in the thyroid gland presumably as a result of rapid in vivo deiodination.

Synthesis of esters of 1- and 3-hydroxyquinolizidine as well as of the parent aminoalcohols is described, and preliminary screening suggests that antispasmodic activity is enhanced by the beta -arrangement. Suitable esters of all three aminoalcohols have also been synthesized to determine the degree of local anesthetic activity associated with them. A significant degree of inhibition of spontaneous motor activity in rats was noted with the trimethoxybenzoates of 1- and 3-hydroxyquinolizidine.

Three radioiodinated phenytoin analogs were synthesized, and tissue distribution studies were conducted in rats. Except for a greater retention of radioactivity following administration of the radioiodinated derivatives, the tissue distribution patterns were qualitatively similar to those found for 14C phenytoin. In nearly all cases, the adrenals, heart, kidneys, and liver displayed the greatest capacity to retain radioactivity. The high uptake of radioactivity observed for the thyroid was attributed to in vivo metabolic deiodination of the radioiodinated derivatives.

A number of A or B ring modified 20- and 22-azacholesterol analogues (1 and 2, respectively) were synthesized in an attempt to ascertain the structural requirements for inhibition of the cholesterol side-chain cleavage reaction in bovine adrenocortical mitochondrial acetone powder preparations. The inhibition studies of these analogues revealed that (1) the 3-methyl ethers were as active as the parent compounds and that (2) reduction of the delta 5 double bond greatly lessened the inhibitory activity. These studies demonstrated a crucial role of the delta 5 double bond for inhibitory activity, while a free hydroxyl group at C-3 is not essential for this action. Furthermore, as in the parent compounds, 22-azacholesterol analogues were more potent than their 20-azacholesterol counterparts.

2-selena-A-nor-5α-androstan-17β-ol was studied in vitro and in vivo in the rat prostate gland. The data demonstrates the ability of this compound to selectively complex with the specific receptors of 5α-dihydrotestosfcerone (5α-DHT) in the cytosol and to be retained in the nuclei in an unaltered form. Studies with selenium-75 labeled material suggests that the uptake and localization is similar to endogenous 5α-dihydrotestosterone.

Alkylated derivatives of 17-acetoxyprogesterone were prepared in order to test the hypothesis that bulky groups in certain positions of the steroid molecule have the effect of transforming progestogens into antiprogestogens. These groups might exert binding influence outside the area occupied by progesterone itself. The compounds were tested for competitive affinity against tritiated progesterone and receptor from rabbit uterus cytosol. The low affinity of all derivatives makes it unlikely that they would be active as antiprogestational agents.

Despite the paucity of literature dealing with the effect of plasma lipoproteins upon drug disposition, evidence is accumulating that shows that these macromolecular complexes can play important roles in the absorption and transport of lipid-soluble agents. Moreover, preliminary studies have demonstrated that radiotracers can be directed to specific tissues by prior incorporation into the hydrophobic core of specific lipoproteins. Although these studies offer encouragement for the possible use of lipoproteins in the site-specific delivery of radiopharmaceuticals that are used in tracer doses, the use of lipoproteins in the transport of drugs at pharmacological concentrations represents a much greater challenge. Nothing is known at this time about the saturation kinetics of drug incorporation into lipoproteins or partially delipidated lipoproteins. Nor is there any assurance that drug-laden lipoproteins will participate in receptor-mediated uptake processes similar to native lipoproteins. Moreover, receptor-mediated uptake of lipoproteins is a saturable process and may not permit attainment of sufficient drug concentrations within cells. It could be argued, however, that receptor-mediated uptake could be enhanced by prior treatment with hypocholesterolemic drugs as has been shown for cholestyramine. In any event, the possible use of lipoproteins for the site-specific delivery of intravenously administered radiodiagnostics or highly potent drugs (e.g., anticancer agents) appears promising. Only the results of ongoing studies will determine the practicality of this approach.

Despite the paucity of literature dealing with the effect of plasma lipoproteins upon drug disposition, evidence is accumulating that shows that these macromolecular complexes can play important roles in the absorption and transport of lipid-soluble agents. Moreover, preliminary studies have demonstrated that radiotracers can be directed to specific tissues by prior incorporation into the hydrophobic core of specific lipoproteins. Although these studies offer encouragement for the possible use of lipoproteins in the site-specific delivery of radiopharmaceuticals that are used in tracer doses, the use of lipoproteins in the transport of drugs at pharmacological concentrations represents a much greater challenge. Nothing is known at this time about the saturation kinetics of drug incorporation into lipoproteins or partially delipidated lipoproteins. Nor is there any assurance that drug-laden lipoproteins will participate in receptor-mediated uptake processes similar to native lipoproteins. Moreover, receptor-mediated uptake of lipoproteins is a saturable process and may not permit attainment of sufficient drug concentrations within cells. It could be argued, however, that receptor-mediated uptake could be enhanced by prior treatment with hypocholesterolemic drugs as has been shown for cholestyramine. In any event, the possible use of lipoproteins for the site-specific delivery of intravenously administered radiodiagnostics or highly potent drugs (e.g., anticancer agents) appears promising. Only the results of ongoing studies will determine the practicality of this approach.

A no-carrier added synthesis of [11C]thymidine from cyclotron produced [11C]methyl iodide is presented. The bis-silylated derivative of the lithium salt of 5-bromo-2'-deoxyuridine is treated with [11C]methyl iodide in THF at -50 degrees C giving 20-30 mCi [11C]thymidine in a 40-70% radiochemical yield. Purification is performed by reversed phase chromatography on a C18 column. The whole procedure takes 25 min and the specific activity is about 20-30 mCi/mumol at the end of the synthesis.

A series of substituted 5alpha-androstan-17beta-ols was synthesized and evaluated for their potential use in the development of a prostate imaging agent. The ability of the synthesized compounds to compete with [3H]-5alpha-dihydrotestosterone for rat prostate androgen receptor protein served as the screening assay. For 3-substituted derivatives, the order of binding to the androgen receptor protein was =O greater than -OH greater than H approximately or equal to F. 3beta-Fluoro-5alpha-androstan-17beta-ol was found to have approximately 5% the androgenic activity of testosterone propionate in the castrated rat. The low biological activity for the 3beta-fluoro derivatives, coupled with the synthetic obstacles associated with introducing fluorine-18, has led us to search for more suitable halo steroids as potential radiodiagnostics.

Derivatives of 19- azaandrostenedione (10 beta-amino-4- estrene -3,17-dione, 2) and 19-amino-4-androstene-3,17-dione (3) were synthesized as potential inhibitors of aromtase (estrogen synthetase). Compound 2 and its derivatives were synthesized from 3,17-dioxo-4-androsten-19-oic acid (5) via a Curtius rearrangement. Derivatives of 3 were synthesized from the intermediate 3,17-bis(ethylenedioxy)-5-androsten-19-al oxime (14), which was reduced to the corresponding amine (16) with Raney nickel. However, attempts to synthesize the parent compound, 3, from 16 by several different methods were unsuccessful. The compounds obtained were tested for inhibitory activity in the tritiated water assay for aromatase, with human placental microsomes as the enzyme source and [1-3H]-androst-4-ene-3,17-dione (83% 3H 1 beta) as the substrate. All of the compounds caused less than 20% inhibition of enzyme activity when tested at one and five times the substrate concentration (0.25 microM, 1.25 microM) and were poorer inhibitors than two known inhibitors, 7 alpha-[(p-aminophenyl)thio] androstenedione (7- APTA ) and 4-hydroxy-4-androstene-3,17-dione (4-OHA).

SUMMARY Radiolabeled 19-iodocholesterol is widely used to obtain images of human adrenals. We have shown in rats and mice that 85-95% of the radiolabel is present in the esterified form. Using rat adrenal cytosolic cholesterol esterase, the kinetic parameters K~t and V were determined for commercially available cholesteryl [l-14C]-oleate (!) and [125I]-19-iodocholesteryl oleate (It). The Kra and V were found to be 16.2/zM and 602 pmol/min/mg prot. respectively for I compared to 76.2 ttM and 37.6 pmol/min/mg prot. for II. Since V/Ku is 37.3 ml/min/mg prot. for the normal ester and 0.49ml/min/mg prot. for the iodinated analog, it appears that the normal substrate is 76 times more specific than II. In addition, direct competition experiments were run in which 67/~M I was used as substrate. The addition of 65 and 325/tM 19-iodocholesteryl oleate caused 20 and 49% inhibition, respectively. On the basis of these studies, it is not surprising that 19-iodocholesterol would accumulate in the adrenals in an esterified form and thus provide an effective agent for adrenal visualization.

Analogues of dioctanoylglycerol (diC8) and 1-oleoyl-2-acetylglycerol (OAG) containing an iodoaryl group have been synthesized and shown to compete with [3H]phorbol dibutyrate [( ([3H]PDBu) for binding to protein kinase C in a crude rat brain preparation. Phorbol diesters have been shown to bind specifically to protein kinase C and the PDBu receptor has been copurified with protein kinase C activity. All three diacylglycerol analogues were comparable to OAG in binding affinity. In an assay of protein kinase C activation, the diC8 analogue was more active than the OAG analogues, thus demonstrating greater structural specificity under the conditions of this assay.

Durch Oxidation des Hydroxy-androstendions (I) erhalt man die Carbonsaure (II), die uber (III) und (IV) zu den Verbindungen (V) bzw. (VI) fuhrt.

Reports of rapid accumulation of 3H- or 14 C-labelled hexamethonium in cartilaginous tissues after intravenous administration to rats and mice and of binding of hexamethonium to chondroitin sulfate in vitro prompted synthesis of radioiodinated analogs of hexamethonium. Tissue distribution studies in rats performed at 0.5 and 2 h after intravenous administration of 14 C-hexamethonium confirmed the propensity of this drug to accumulate in cartilaginous tissues. Tissue distribution studies were performed in both normal and chondrosarcoma tumor-bearing rats at 0.5 and 2 h after intravenous administration of the radioiodinated analogs. The radioiodinated analogs showed profiles of distribution of radioactivity comparable to that of hexamethonium. Levels of uptake were highest in kidney and urine at all time periods studied. High levels of activity were seen in trachea, intervertebral discs and, when present, tumor. A rapid dispersal of a relatively low level of radioactivity was seen in most other tissues and was followed by rapid clearance in all tissues except meninges which continued to accumulate radioactivity for as long as 2 h. A chondrosarcoma tumor was successfully imaged in a rat 15 min following intravenous administration of a radioiodinated analog of hexamethonium.

A series of substituted 5alpha-androstan-17beta-ols was synthesized and evaluated for their potential use in the development of a prostate imaging agent. The ability of the synthesized compounds to compete with [3H]-5alpha-dihydrotestosterone for rat prostate androgen receptor protein served as the screening assay. For 3-substituted derivatives, the order of binding to the androgen receptor protein was =O greater than -OH greater than H approximately or equal to F. 3beta-Fluoro-5alpha-androstan-17beta-ol was found to have approximately 5% the androgenic activity of testosterone propionate in the castrated rat. The low biological activity for the 3beta-fluoro derivatives, coupled with the synthetic obstacles associated with introducing fluorine-18, has led us to search for more suitable halo steroids as potential radiodiagnostics.

A series of five 1-(arylsulfonyl)-3,5-dialkyl-s-triazine-2,4,6-(1H,3H,5H)-triones was synthesized by the base-catalyzed reaction of arylsulfonamides with alkyl isocyanates. The compounds were tested for apomorphine antagonism in mice, diuretic activity in fasted rats, antiviral activity in mice, antiviral activity in infected cell cultures, CNS stimulant activity in mice, and hypoglycemic activity in rats. The only biological activity exhibited by these compounds was a slight tendency to lower blood sugar levels in glucose-primed, fasted rats. This activity may be attributable to their structural similarity to the sulfonylurea hypoglycemics.

Radioiodinated cholesteryl oleate (125I-CO) was found to associate rapidly with plasma lipoproteins following intravenous administration to rats. The high-density lipoprotein (HDL) fraction was observed to contain the highest amount of radioiodinated ester. Isolation and purification of this HDL fraction (125I-CO-HDL) and subsequent administration to rats demonstrated a plasma clearance similar to that previously observed for HDL labeled by direct iodination. Moreover, the concentration of radioactivity appearing in the adrenal cortex and ovary 0.5 h after intravenous administration of 125I-CO-HDL was greater than that observed after administration of 125I-CO, and the uptake of radioactivity by these tissues was considerably greater in hypolipidemic rats. These findings are consistent with existing knowledge relating to the metabolic fate of HDL and radioiodinated cholesterol derivatives in the rat, and suggest that radioiodinated cholesteryl esters may become useful probes for labeling lipoproteins.

Durch Umsetzung von Androstendion (I) mit den Thiolen (II) werden die Thio-substituierten Steroide (III) erhalten; das Iod-substituierte Derivat (IV) entsteht aus der entsprechenden Aminoverbindung (III) uber das Diazoniumsalz.

A radioiodinated analogue of a naturally occurring alkyl lysophospholipid (ALP) was synthesized for evaluation as a potential tumor-localizing imaging agent. rac-1-(12-(m-Iodophenyl)dodecyl)-2-methylglycero-3-phosphocholine (ET-12IP-OMe, 14) was radiolabeled with iodine-125 via an isotope-exchange procedure. Tissue distribution studies with ({sup 125}I)ET-12IP-OMe in tumor-bearing rats revealed an immediate tumor uptake of radioactivity. Although radioactivity was also present in nontarget tissues at this time, clearance of tracer from the tumor was much slower and thus provided a suitable tumor to nontarget tissue ratio at 24 h. As a result of this selective accumulation, it was possible to clearly delineate the tumor with gamma-camera scintigraphy.

The synthesis and preliminary biodistribution data for a series of sterol-like esters of iopanoic acid having potential value as liver-specific CT contrast agents are described. Structural modification of the sterol portion of the iopanoate ester afforded a group of compounds that displayed tissue specificity similar to cholesteryl iopanoate, the prototype ester of this series, but were rapidly cleared from the target tissues after hydrolysis. From the biodistribution data, the most promising of these agents, pregnenolone iopanoate (PI), was evaluated by CT in rabbits receiving a radiologic dose equivalent to 30 mg I/kg. The hepatic parenchyma was enhanced within 2 h of infusion to a maximal level of 31 HU above precontrast values. Hepatic CT attenuation returned to normal within 24 h. However, CT performed after PI infusion into Vx2 tumor-bearing rabbits failed to provide superior images compared with those acquired following bolus administration of urographic contrast.

SummaryA series of five 17-iminoste-roids was tested in combination with testosterone propionate in castrated male rats. All of these compounds inhibited the response to testosterone propionate in the seminal vesicles and ventral prostate glands. There was no indication of inhibition of pituitary gonadotropin secretion in spayed female rats treated with several of these compounds. While inhibition of the action of exogenous testosterone propionate was obtained, no effect on endogenous androgen was observed in intact males at the dose used.