Fuminori Ohsawa

Replacing alkyl sulfonamide with aromatic sulfonamide in sulfonamide-type RXR agonists favors switch towards antagonist activity.

Retinoid X receptor (RXR) ligands are attractive candidates for clinical application because of their activity against tamoxifen-resistant breast cancer, taxol-resistant lung cancer, metabolic syndrome, and allergy. Though several RXR ligands, especially RXR antagonists, have been reported, the rational molecular design of such compounds is not well advanced. 4-[N-Methanesulfonyl-N-(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-2-naphthyl)amino]nicotinic acid (5a) is a moderately RXRalpha-preferential agonist, and we examined the feasibility of replacing the methyl group on the sulfonamide with a longer alkyl chain or an aromatic ring as an approach to produce new RXR antagonists. Several of the resulting benzenesulfonanilide-type compounds showed RXR antagonist activity. This design strategy should be a useful approach for addressing the lack of structure diversity of RXR antagonists.

Modification at the Lipophilic Domain of RXR Agonists Differentially Influences Activation of RXR Heterodimers.

RXR permissive heterodimers are reported to be activated differently depending upon the chemical structure of RXR agonists, but the relationship of agonist structure to differential heterodimer activation has not been explored in detail. In this study, we performed systematic conversion of the alkoxy side chain of 5a (6-[ethyl-(3-isopropoxy-4-isopropylphenyl)amino]nicotinic acid, NEt-3IP) and evaluated the RXR-, PPAR/RXR-, and LXR/RXR-agonist activities of the products. The cyclopropylmethoxy analogue (5c) showed similar RXR- and LXR/RXR-agonistic activities to the benzyloxy analogue (5i) and n-propoxy analogue (5k) but exhibited more potent PPAR/RXR-agonistic activity than 5i or 5k. Differential modulation of RXR heterodimer-activating ability by conversion of the alkoxy group located in the lipophilic domain of the RXR-agonist common structure is expected be a useful approach in the design of new RXR agonists for the treatment of hyperlipidemia or type 2 diabetes.

RXR Partial Agonist CBt-PMN Exerts Therapeutic Effects on Type 2 Diabetes without the Side Effects of RXR Full Agonists.

Treating insulin resistance and type 2 diabetes in rodents, currently known retinoid X receptor (RXR) agonists induce significant adverse effects. Here we introduce a novel RXR partial agonist CBt-PMN (11b), which shows a potent glucose-lowering effect and improvements of insulin secretion and glucose tolerance without the serious adverse effects caused by RXR full agonists. We suggest that RXR partial agonists may be a new class of antitype 2 diabetes drug candidates.

Mechanism of Retinoid X Receptor Partial Agonistic Action of 1-(3,5,5,8,8-Pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)-1H-benzotriazole-5-carboxylic Acid and Structural Development To Increase Potency

We have reported that retinoid X receptor (RXR) partial agonist 1-(3,5,5,8,8-pentamethyl-5,6,7,8-tetrahydro-2-naphthyl)-1H-benzotriazole-5-carboxylic acid (CBt-PMN, 4a) shows a significant antidiabetes effect in the KK-A(y) type 2 diabetes model mice, with reduced side effects compared to RXR full agonists. To elucidate the mechanism of the RXR partial agonist activity of 4a, we synthesized derivatives of 4a, evaluated their RXR agonist activity, and performed structure-activity relationship analysis. Reporter gene assay revealed that though 6b, which possesses an amino group at the 2-position of 5-carboxybenzimidazole, showed RXR full-agonist activity, compounds 6d and 6e, which possess an oxygen atom and a sulfur atom at the corresponding position, respectively, showed weak RXR agonist activity. On the other hand, 6c, which has a trifluoromethyl group at the corresponding position, acts as an RXR partial agonist, having similar Emax (67 ± 2%) and lower EC50 (15 ± 0 nM) compared to those of 4a (Emax = 75 ± 4%, EC50 = 143 ± 2 nM). A fluorescence polarization assay of cofactor recruitment confirmed that fluorescein-labeled D22 coactivator peptide was less efficiently recruited to RXR by 4a and 6c than by LGD1069 (1), a known RXR full agonist. Electrostatic potential field calculations and computational docking studies suggested that full agonists show an electrostatic attraction, which stabilizes the holo structure and favors coactivator recruitment, between the side chain at the benzimidazole 2-position and the α-carbonyl oxygen of asparagine-306 in helix 4 (H4) of the RXR receptor. However, RXR partial agonists 4a and 6c lack this interaction. Like 4a, 6c showed a significant antidiabetes effect in KK-A(y) type 2 diabetes model mice with reduced levels of the side effects associated with RXR full agonists. These findings should aid the design of new RXR partial agonists as antitype 2 diabetes drug candidates.

Identification of urine metabolites of TFAP, a cyclooxygenase-1 inhibitor.

Only a few COX-1-selective inhibitors are currently available, and the research on COX-1 selective inhibitors is not fully developed. The authors have produced several COX-1 selective inhibitors including N-(5-amino-2-pyridinyl)-4-trifluoromethylbenzamide: TFAP (3). Although 3 shows potent analgesic effect without gastric damage, the urine after administration of 3 becomes red-purple. Since the colored-urine should be avoided for clinical use, in this research we examined the cause of the colored-urine. UV-vis spectra and LC-MS/MS analyses of urine samples and metabolite candidates of 3 were performed to afford information that the main reason of the colored urine is a diaminopyridine (4), produced by metabolization of 3. This information is useful to design new COX-1 selective inhibitors without colored urine based on the chemical structure of 3.

Fluorescent retinoid X receptor ligands for fluorescence polarization assay.

Retinoid X receptor (RXR) agonists are candidate agents for the treatment of metabolic syndrome and type 2 diabetes via activation of peroxisome proliferator-activated receptor (PPAR)/RXR or liver X receptor (LXR)/RXR-heterodimers, which control lipid and glucose metabolism. Reporter gene assays or binding assays with radiolabeled compounds are available for RXR ligand screening, but are unsuitable for high-throughput screening. Therefore, as a first step towards stabilizing a fluorescence polarization (FP) assay system for high-throughput RXR ligand screening, we synthesized fluorescent RXR ligands by modification of the lipophilic domain of RXR ligands with a carbostyril fluorophore, and selected the fluorescent RXR agonist 6-[ethyl(1-isobutyl-2-oxo-4-trifluoromethyl-1,2-dihydroquinolin-7-yl)amino]nicotinic acid 8d for further characterization. Compound 8d showed FP in the presence of RXR and the FP was decreased in the presence of the RXR agonist LGD1069 (2). This compound should be a lead compound for use in high-throughput assay systems for screening RXR ligands.

Discovery of a Potent Retinoid X Receptor Antagonist Structurally Closely Related to RXR Agonist NEt-3IB

We discovered a potent retinoid X receptor (RXR) antagonist, 6-[N-ethyl-N-(5-isobutoxy-4-isopropyl-2-(E)-styrylphenyl)amino]nicotinic acid (13e), that is structurally closely related to the RXR full agonist 6-[N-ethyl-N-(3-isobutoxy-4-isopropylphenyl)amino]nicotinic acid (NEt-3IB) (4). Compound 13e was synthesized via a simple route from 11, a methyl ester precursor of 4. Because 11 possesses high electrophilic reactivity because of the amino and alkoxy groups, it was readily transformed to 12 by iodization, and the iodine atom of 12 was converted to a C-C or C-N bond by means of palladium-catalyzed reaction to afford 13. Transcriptional activation assay revealed that 13g (in which the iodine atom was replaced with an amino group) is a weak RXR agonist, while 13d (a phenyl group), 13e (a styryl group), and 13f (an anilino group) are RXR antagonists. Among them, 13e was found to be more potent than the known RXR antagonist PA452 (9).

Pharmacokinetic properties of newly synthesized retinoid X receptor agonists possessing a 6-[N-ethyl-N-(3-alkoxy-4-isopropylphenyl)amino]nicotinic acid skeleton in rats

Objective: The pharmacokinetic properties of three newly synthesized retinoid X receptor (RXR) agonists were evaluated in rats to elucidate the structural factor influencing their pharmacokinetic properties. Material and methods: Three RXR agonists possessing a common 6-[N-ethyl-N-(3-alkoxy-4-isopropylphenyl)amino]nicotinic acid skeleton and side chain structures that are slightly different from each other were prepared as we previously reported (Takamatsu et al., ChemMedChem, 2008; 3:780–787). The plasma concentration profiles of these compounds were evaluated following the intravenous and intra-intestinal administrations. Their hepatic metabolism was characterized using rat liver microsomes. Results: Based on the plasma concentration profile, NEt-3IP (3-isopropoxy) was shown to have a distribution volume of 4.53 L/kg, and to be cleared from the body with an elimination half-time of 0.95 h. The bioavailability of NEt-3IP is 16.4%, whereas those of the isobutoxy analog NEt-3IB and the cyclopropylmethoxy analog NEt-3cPM are 46.5% and 22.6%, respectively. Subsequently, in the experiments using rat liver microsomes, the Km and Vmax values of NEt-3IP were determined as 7.85 µM and 0.48 nmol/min/mg protein, respectively. This Km value is nearly the same as those of NEt-3IB and NEt-3cPM, but the Vmax value is noticeably smaller. Additionally, it was revealed that the CYP family mainly metabolizing NEt-3IP is different from those metabolizing the other analogs. Conclusion: Based on these findings, the pharmacokinetic properties of the compounds possessing this type of the skeleton seem to be largely influenced by a slight modification of the side chain structure.

Abstract 946: Discovery of a novel benzotriazole-type retinoid X receptor partial-agonist decreasing plasma glucose level in type-2 diabetes mellitus with decreased side effects

Insulin-independent hypoglycemic agents have attracted attention, because type 2 diabetes mellitus is reported to be associated with an increased risk of cancer due to mitogenic effect of insulin. Insulin-independent antihyperglycemic effect can be obtained by LXR activation. LXR functions as heterodimers with retinoid X receptors (RXRs). RXRs also work with PPARs, which are known as targets for improvement of insulin resistance. Since LXR/RXR and PPAR/RXR can be activated by RXR agonists alone, and we are interested in RXR agonists for new hyperglycemia agents. However, well-known RXR agonists (RXR full-agonists) show significant weight gain and triglyceride elevation. Since these side effects are caused by excess activation of RXR, we hypothesized that RXR partial-agonists might be effective to decrease plasma glucose level without these problems. As a result, CBt-PMN, which has a benzotriazole structure, was discovered as an RXR partial-agonist, and significantly decreases plasma glucose level in type 2 diabetes model mice without the side effects described above. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 946. doi:1538-7445.AM2012-946