Retinoic acid receptors (RAR), thyroid hormone receptors (TR), peroxisome proliferator activated receptors (PPARs) and the orphan receptor, LXR, bind preferentially to DNA as heterodimers with a common partner, retinoid X receptor (RXR), to regulate transcription. We investigated whether RXR-selective agonists replicate the activity of ligands for several of these receptors? We demonstrate here that RXR-selective ligands (referred to as rexinoids) function as RXR heterodimer-selective agonists, activating RXR: PPARgamma and RXR:LXR dimers but not RXR:RAR or RXR:TR heterodimers. Because PPARgamma is a target for antidiabetic agents, we investigated whether RXR ligands could alter insulin and glucose signalling. In mouse models of noninsulin-dependent diabetes mellitus (NIDDM) and obesity, RXR agonists function as insulin sensitizers and can decrease hyperglycaemia, hypertriglyceridaemia and hyperinsulinaemia. This antidiabetic activity can be further enhanced by combination treatment with PPARgamma agonists, such as thiazolidinediones. These data suggest that the RXR:PPARgamma heterodimer is a single-function complex serving as a molecular target for treatment of insulin resistance. Activation of the RXR:PPARgamma dimer with rexinoids may provide a new and effective treatment for NIDDM.

Structural modifications of cellular macromolecules by chemical carcinogens may represent early and requisite events in neoplastic transformation (1, 2). Through interactions of this nature, qualitative changes could be induced in informational macromolecules such as DNA and RNA, and these lesions could provide a molecular basis for alteration of gene expression in carcinogenesis. Identification of the products of these reactions (herein referred to as adducts) is essential in order to: (i) gain insights into mechanisms of carcinogen activation; (ii) determine the reactive centers in these macromolecules; (iii) follow the kinetics of appearance and disappearance of adducts in the cell; and (iv) relate specific patterns of macromolecule modification with the ultimate development of tumors in target organs of susceptible species.Aflatoxin B1 (AFB1) is a very potent liver carcinogen in several animal species (3), and epidemiologic evidence indicates that it is also an important factor in the etiology of human liver cancer in certain sections of the world (4). AFB1 binds covalently to cellular macromolecules, including DNA, in mvo (5-7) and in vitro after metabolic activation (8-10). The relationship of this type of interaction to its mechanism of action has been emphasized (11). Strong indirect evidence has indicated the production of AFB1-2,3-oxide as a major activated metabolite responsible for macromolecular binding in vitro and in vivo (5-7, 9, 12), but structures of specific adducts formed with nucleic acids or proteins have not been determined. The purpose of the research reported here was to determine the structure of the major adduct formed with DNA by AFB1 activated metabolically in vitro. The results indicate that approximately 90% of the binding in vitro can be attributed to a single adduct, which was isolated in sufficient quantity for structural analysis and identified as 2,3-dihydro-2-(N7-guanyl)-3-hydroxyaflatoxin B1 (structure I).,H0 Ho (c) H3C (c) Hk I MATERIALS AND METHODS Liver microsomes used for metabolic activation of AFB1 were prepared from phenobarbital-treated male Fischer rats (13) by the procedure of Kinoshita et al. (14). The incubation mixture (400 ml) for the binding of AFB1 to DNA included Tris-HCl (pH 7.5,45 mM), MgCl2 (3 mM), glucose-6-phosphate (5 mM), NADP (0.8 mM, Sigma Chemical Co.), glucose-6-phosphate dehydrogenase (0.4 unit/ml, Sigma Chemical Co.), approximately 1 mg of microsomal protein per ml, calf thymus DNA (20 A260 units/ml or a total of 340 mg; type I, Sigma Chemical Co.), AFB1 [224 ,uM added Abbreviations: AFB1, aflatoxin B1; I, 2,3-dihydro-2-(N7-guanyl)-3-hydroxyaflatoxin Bj; II, 2,3-dihydro-3-hydroxy-2-(4-nitrobenzoxy)-aflatoxin B1; HPLC, high-pressure liquid chromatography; NMR, nuclear magnetic resonance; FD, field-desorption mass spectrometry; EI, electron-impact mass spectrometry.

Abnormally high rates of fatty acid oxidation and low rates of glucose oxidation are important contributors to the severity of ischemic heart disease. Malonyl coenzyme A (CoA) regulates fatty acid oxidation by inhibiting mitochondrial uptake of fatty acids. Malonyl CoA decarboxylase (MCD) is involved in the decarboxylation of malonyl CoA to acetyl CoA. Therefore, inhibition of MCD may decrease fatty acid oxidation and protect the ischemic heart, secondary to increasing malonyl CoA levels. Ex vivo working rat hearts aerobically perfused in the presence of newly developed MCD inhibitors showed an increase in malonyl CoA levels, which was accompanied by both a significant decrease in fatty acid oxidation rates and an increase in glucose oxidation rates compared with controls. Using a model of demand-induced ischemia in pigs, MCD inhibition significantly increased glucose oxidation rates and reduced lactate production compared with vehicle-treated hearts, which was accompanied by a significant increase in cardiac work compared with controls. In a more severe rat heart global ischemia/reperfusion model, glucose oxidation was significantly increased and cardiac function was significantly improved during reperfusion in hearts treated with the MCD inhibitor compared with controls. Together, our data show that MCD inhibitors, which increase myocardial malonyl CoA levels, decrease fatty acid oxidation and accelerate glucose oxidation in both ex vivo rat hearts and in vivo pig hearts. This switch in energy substrate preference improves cardiac function during and after ischemia, suggesting that pharmacological inhibition of MCD may be a novel approach to treating ischemic heart disease.

Retinoid X receptor (RXR) plays a central role in the regulation of many intracellular receptor signalling pathways and can mediate ligand-dependent transcription, acting as a homodimer or as a heterodimer. Here we identify an antagonist towards RXR homodimers which also functions as an agonist when RXR is paired as a heterodimer to specific partners, including peroxisome proliferator-activated receptor and retinoic acid receptor. This dimer-selective ligand confers differential interactions on the transcription machinery: the antagonist promotes association with TAF110 (TATA-binding protein (TBP)-associated factor 110) and the co-repressor SMRT, but not with TBP, and these properties are distinct from pure RXR agonists. This unique class of RXR ligands will provide a means to control distinct target genes at the level of transcription and allow the development of retinoids with a new pharmacological action.

These novel VDR modulators may have potential as therapeutics for cancer, leukemia and psoriasis with less calcium mobilization side effects than are associated with secosteroidal 1,25(OH)2D3 analogs.

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The literature route to racemic cis-and trans-3-substituted prolines starting from acetamidomalonate and a,@-unsaturated aldehydes has been refined and applied to the synthesis of 3-phenyl-and 3-n-propylproline.A key improvement in the sequence was the acid-catalyzed silane reduction of the initial hydroxylactam Michael adduct 2, which allowed subsequent transformations to proceed cleanly and in high yield. Both the Nand 3-substitutents were found to have an effect on selectivity in the saponification of trans-3-substituted proline esters in the presence of the corresponding cis esters. The trans isomers in each series were resolved via the diastereomeric (S)-a-methylbenzylamides, and the absolute configurations of the resulting pure optical isomers were assigned.Proline is a well-known means of inducing conformational constraints into peptides.l Because of motional restrictions inherent to the pyrrolidine ring, the presence of a proline residue greatly reduces the available conformational space of a peptide and gives rise to conformers separated by relatively high interconversion barriem2 Thus, information may be obtained about the bioactive conformation of a peptide, and the biological potency may be increased by incorporation of a proline residue. However, when substitution of an amino acid iesidue with proline leads to a reduction in biological activity, the question arises whether the loss is due to conformational or steric considerations or to loss of a favorable interaction associated with the side chain of the original amino acid residue. In this vein, we have examined the incorporation of 3-substituted proline derivatives into peptides, where the 3-substituent corresponds to the substituent on the @-carbon of standard amino acids. For example, 3-substituted prolines la-c are conformationally constrained analogues of phenylalanine, norleucine, and aspartate, respectively.In this paper, we describe our work on the synthesis and enantiomeric resolution of derivatives of 3-phenyl-and 3-n-propylproline. A stereospecific synthesis of N-Boctrans-3-n-propyl-~-proline, 1 b, from 4-hydroxy-~-proline was recently developed in our l a b~r a t o r y ,~ but the route was not applicable to the synthesis of la and was not amenable to large-scale work. We were attracted to the schemes outlined by Cox et d.,4 Mauger et al.? and Sarges (1) (a) Momany, F. A; Chuman, H. Methods Enzymol. 1986, 124, 3. (b) Marshall, G. R. In Chemical Recognition in Biological Systems; Creighton, A. M., Turner, S., EMS.; The Chemical Society: London, 1982; p 279. (c) Arison, B. H.; Hirschmann, R.; Veber, D. F. Bioorg. Chem. 1978, 7, 447. (2) (a) Holldsi, M.; Radica, L.; Wieland, T. Znt. J. Peptide Protein Res. 1977, 10, 286. (b) Delaney, N. G.; Madison, V. Int. J. Peptide Protein Res. 1982, 19, 543, and references therein. (3) Holladay, M. W.; May, C. S.; Arnold, W. A., unpublished results. lb: mp 86-90 'C; [ a ]~*~ = -40.6' (c 1, CHCl,). Optical purity of this product was assessed as >98% based on HPLC analysis of derived amethylbenzylamides. ...