Modification and Functionalization of the Guanidine Group by Tailor-made Precursors.

The guanidine group is one of the most important pharmacophoric groups in medicinal chemistry. The only amino acid carrying a guanidine group is arginine. In this article, an easy method for the modification of the guanidine group in peptidic ligands is provided, with an example of RGD-binding integrin ligands. It was recently demonstrated that the distinct modification of the guanidine group in these ligands allows for the selective modulation of the subtype (e.g., between the subtypes αv and α5). Moreover, a formerly unknown strategy for the functionalization via the guanidine group was demonstrated, and the synthetic approach is reviewed in this document. The modifications described here involve terminally (Nω) alkylated and acetylated guanidine groups. For the synthesis, tailor-made precursor molecules are synthesized, which are then subjected to a reaction with an orthogonally deprotected amine to transfer the pre-modified guanidine group. For the synthesis of alkylated guanidines, precursors based on N,N'-Di-Boc-1H-pyrazole-1-carboxamidine are used to synthesize acylated compounds, the precursor of choice being a correspondingly acylated derivative of N-Boc-S-methylisothiourea, which can be obtained in one- and two-step reactions.

Biomimetic synthesis of (+/-)-galanthamine and asymmetric synthesis of (-)-galanthamine using remote asymmetric induction.

(+/-)-Galanthamine (1) was synthesized in excellent yield by applying PIFA-mediated oxidative phenol coupling of N-(4-hydroxy)phenethyl-N-(3',4',5'-trialkoxy)benzyl formamide (15b) as a key step. Because of the symmetrical characteristics of the pyrogallol moiety in the substrate (15b), the phenol coupling resulted in a sole coupling product except for volatile components from the oxidizing agent. On the basis of the successful results of the above strategy, (-)-galanthamine (1) was synthesized by employing a novel remote asymmetric induction, where conformation of the seven-membered ring in the product of the phenol coupling was restricted by forming a fused-chiral imidazolidinone ring with D-phenylalanine on the benzylic C-N bond of the tri-O-alkylated gallyl amino moiety. The conformational restriction and successive debenzylation of the protected hydroxyl groups on the pyrogallol ring caused diastereoselective cyclization to yield a cyclic ether having the desired stereochemistry for the synthesis of (-)-1.

Design, synthesis, and neurochemical evaluation of 2-amino-5-(alkoxycarbonyl)-3,4,5,6-tetrahydropyridines and 2-amino-5-(alkoxycarbonyl)-1,4,5,6-tetrahydropyrimidines as M1 muscarinic receptor agonists.

Four regioisomers of 2-amino-(methoxycarbonyl)-3,4,5,6-tetrahydropyridine (2a-5a) were synthesized as the racemates to evaluate the utility of exocyclic amidines in the development of novel agonists for M1 muscarinic receptors. Of the four regioisomers, only racemic 2-amino-5-(methoxycarbonyl)-3,4,5,6-tetrahydropyridine (4a; CDD-0075-A) displayed high affinity (IC50 = 10 +/- 3.0 microM) and activity at muscarinic receptors coupled to PI metabolism in the rat cortex (260 +/- 4.5% stimulation above basal levels at 100 microM). A series of 2-amino-5-(alkoxycarbonyl)-3,4,5,6-tetrahydropyridines then was synthesized for further evaluation as M1 agonists. Only the propargyl derivative (4d) retained substantial agonist activity (120 +/- 14% at 100 microM) in this series. On the basis of the activity of the 5-(alkoxycarbonyl)-1,4,5,6- tetrahydropyrimidines (1a and 1d) and the 2-amino-5-(alkoxycarbonyl)-3,4,5,6-tetrahydropyridines, the corresponding cyclic guanidine derivatives were synthesized and tested. 2-Amino-5-(methoxycarbonyl)-1,4,5,6-tetrahydropyrimidine (7a) displayed a modest affinity for muscarinic receptors in the CNS (22 +/- 5.3 microM) and an ability to stimulate PI turnover in rat cerebral cortex (81 +/- 16% at 100 microM). The propargyl derivative (7d) also had modest binding affinity (31 +/- 15 microM) and high activity (150 +/- 8.5% at 100 microM), as expected based on the activity of propargyl esters of 1,4,5,6-tetrahydropyrimidine and 2-amino-3,4,5,6-tetrahydropyridine. Computational chemical studies revealed five distinct minimum-energy conformations for 1a, (R)-4a, and 7a, and three for 1d, (R)-4d, and 7d, each with a unique orientation of the ester moiety. Each of the five conformations for 1a could be superimposed upon a unique conformer of (R)-4a and 7a, suggesting that the compounds interact with muscarinic receptors in a similar fashion. Taken together, the data indicate the general utility of amidine systems as suitable replacements for the ammonium group of acetylcholine in developing ligands with activity at M1 muscarinic receptors in the central nervous system. Such compounds might be useful in the treatment of patients with Alzheimer's disease.

Central cholinergic agents. 6. Synthesis and evaluation of 3-[1-(phenylmethyl)-4-piperidinyl]-1-(2,3,4,5-tetrahydro-1H-1-benzazepi n-8-yl)-1-propanones and their analogs as central selective acetylcholinesterase inhibitors.

In an attempt to find central selective acetylcholinesterase (AChE) inhibitors, 3-[1-(phenylmethyl)-4-piperidinyl]-1-(2,3,4,5-tetrahydro-1H-1- benzazepin-8-yl)-1-propanones 9 and their analogs were designed on the basis of our working hypothesis of the enzyme's active site. These compounds were prepared by regioselective Friedel-Crafts acylation of 2,3,4,5-tetrahydro-1H-1-benzazepines and related nitrogen heterocycles as a key step. Most compounds showed potent inhibitory activities with IC50s in the 10-300 nM range. In order to estimate their central selectivities, we examined their effects on the apomorphine-induced circling behavior in rats with unilateral striatal lesions. Among compounds with potent AChE inhibition, 3-[1-(phenylmethyl)-4-piperidinyl]-1-(2,3,4,5-tetrahydro-1H-1- benzazepin-8-yl)-1-propanone fumarate (9a, TAK-147) (IC50 of AChE inhibition = 97.7 nM) inhibited the circling behavior at 3 mg/kg po, in which it had no significant effect on peripheral cholinergic effects. This demonstrates that 9a has favorable central selectivity. Furthermore, 9a significantly ameliorated diazepam-induced passive avoidance deficit at 1 mg/kg po. The benzazepine derivative 9a was selected as a candidate for clinical evaluation.

Synthesis and muscarinic properties of (1S*,3R*,5R*)-trimethyl(1-methyl-6-oxabicyclo[3.1.0]hex-3-yl)methyl ammonium iodide.

To acquire more information about the so-called "muscarinic subsite", compound 4 was synthesized and tested. The results show that in comparison with deoxamuscarine (23) the muscarinic potency of 4 on M2 and M3 subtypes is not significantly altered by the presence of an epoxidic function, which confirms the donor-acceptor hydrogen bonding character of this receptive site. Conversely, there is a negative influence on the transduction processes. In addition, a second hydroxylic function bound on the carbon carrying the terminal methyl of the fourth substituent on the nitrogen dramatically affects the muscarinic behavior; the resulting compounds (11-14) lack any agonist or antagonist activity.

Synthesis and muscarinic activity of some muscarone analogs.

The synthesis of some muscarone analogs has been accomplished and their muscarinic activity has been assayed in three in vitro functional tests. The results of such an investigation put in evidence the active role played by the methylene moiety, placed exocyclic to the pentaatomic ring, on the interaction of the ligands with the different muscarinic receptor subtypes. Worth noting is the ileum-atria selectivity displayed by methylene derivative 7.

Synthesis and muscarinic activity of the chiral forms of methylenemuscarones.

The synthesis of the two pairs of enantiomers of methylenemuscarones 3 and 4 has been accomplished by using (R)- and (S)-lactic esters as starting materials. Due to the existence of different muscarinic receptor subtypes, the compounds were examined for their ability to bind membranes from cerebral cortex (M1), heart (M2), and salivary glands (M3) and were assayed in "in vitro" functional tests as well. The results of such an investigation put in evidence that, in both binding and functional tests, (-)-3 (2S,5S) and (-)-4 (2R,5S) were the eutomers and shared the stereochemistry of the eutomer of muscarone and allomuscarone respectively. It is worth noting that the distomer of 3 and 4 behaves as a partial agonist in the cardiac tissue and as a full agonist in the other preparations. This peculiarity of the chiral forms of 3 and 4 could account for the remarkable selectivity for the ileum observed in the corresponding racemates.

Synthesis and pharmacological evaluation of a new class of 2-oxo-8-azaspiro (4,5)decan-1-ones as analogues of the muscarinic agonist RS-86.

A new series of 8-substituted-2-oxo-8-azaspiro (4,5)decan-1-ones has been synthesized and compounds tested for their cholinergic properties in comparison with the muscarinic agonist RS-86. Preliminary in vitro and in vivo pharmacological data indicate that none of them is provided with significant cholinergic effects either at central or peripheral level. A possible explanation for the lack of activity is given on the basis of conformational studies.

Selective signaling via unique M1 muscarinic agonists.

Rigid analogs of acetylcholine (ACh) were designed for selective actions at muscarinic receptor (mAChR) subtypes and distinct second messenger systems. AF102B, AF150, and AF151 are such rigid analogs of ACh. AF102B, AF150 and AF151 are centrally active M1 agonists. AF102B has a unique agonistic profile showing, inter alia: only part of the M1 electrophysiology of ACh and unusual binding parameters to mAChRs. AF150 and AF151 are more efficacious agonists than AF102B for M1 AChRS in rat cortex and in CHO cells stably transfected with the m1 AChR subtype. Notably, the selectivity of the new m1 agonists is reflected also by activation of select second messenger systems via distinct G-proteins. These compounds reflect a new pharmacological concept, tentatively defined as ligand-selective signaling. Thus, agonist/m1AChR complexes may activate different combinations of signaling pathways, depending on the ligand used. Rigid agonists may activate a limited repertoire of signaling systems. In various animal models for Alzheimer's disease (AD) the agonists AF102B, AF150 and AF151, exhibited positive effects on mnemomic processes and a wide safety margin. Such agonists, and especially AF102B, can be considered as a rational treatment strategy for AD.

Synthesis and structure-activity studies of a series of spirooxazolidine-2,4-diones: 4-oxa analogues of the muscarinic agonist 2-ethyl-8-methyl-2,8-diazaspiro[4.5]decane-1,3-dione.

A series of spirooxazolidine-2,4-dione derivatives related to the putative M1 agonist 2-ethyl-8-methyl-2,8-diazaspiro[4.5]decane-1,3-dione (RS86; 1) were synthesized. The compounds were evaluated as cholinergic agents in in vitro binding assays and in in vivo pharmacological tests including antiamnesic effects using scopolamine-treated mice, hypothermia, and salivation in mice. Four compounds (5a,c,f and 17a) exhibited affinity for cortical M1 receptors and reversed scopolamine-induced impairment of mouse passive avoidance tasks, as did 1. Among these compounds, only 5a exhibited M1-receptor stimulating activity in pithed rats. Structural requirements for muscarinic activity in this series of spirooxazolidine-2,4-dione derivatives were as strict as those reported for spirosuccinimide derivatives including 1. The antiamnesic dose of 3-ethyl-8-methyl-1-oxa-3,8-diazaspiro[4.5]decane-2,4-dione (5a) was 2 orders of magnitude lower than the doses inducing hypothermia and salivation, in contrast to 1 for which the former dose was only 5-10-fold lower than the latter. These results suggest that the 8-azaspiro[4.5]decane skeleton represents a useful template for designing new muscarinic agonists as antidementia drugs.

Design, synthesis, and neurochemical evaluation of 5-(3-alkyl-1,2,4- oxadiazol-5-yl)-1,4,5,6-tetrahydropyrimidines as M1 muscarinic receptor agonists.

A series of 5-(3-alkyl-1,2,4-oxadiazol-5-yl)-1,4,5,6-tetrahydropyrimidines+ ++ (7a-h) was synthesized for biological evaluation as selective agonists for M1 receptors coupled to phosphoinositide (PI) metabolism in the central nervous system. Each ligand bound with high affinity to muscarinic receptors from rat brain as measured by inhibition of [3H]-(R)-quinuclidinyl benzilate ([3H]-(R)-QNB) binding. 5-(3-Methyl-1,2,4-oxadiazol-5-yl)-1,4,5,6-tetrahydropyrimidine+ ++ trifluoroacetate (CDD-0098-J;7a) displayed high affinity (IC50 = 2.7 +/- 0.69 microM) and efficacy at muscarinic receptors coupled to PI metabolism in the rat cortex and hippocampus. Increasing the length of the alkyl substituent increased affinity for muscarinic receptors yet decreased activity in PI turnover assays. The hippocampal PI response of 7a was blocked by lower concentrations of pirenzepine (8) or by higher concentrations of either AF-DX 116 (9) or p-fluorohexahydrosiladifenidol (10), suggesting that at low concentrations 7a selectively stimulates PI turnover through M1 receptors.

Aminopyridazines--an alternative route to potent muscarinic agonists with no cholinergic syndrome.

In the search for central cholinergic agents which do not derive from already well-known agonists such as arecoline, pilocarpine or oxotremorine, we selected minaprine [3-(beta-morpholinoethylamino)-4-methyl-6-phenyl-pyridazine dihydrochloride] as a lead structure. Effectively, beside its antidepressive properties, minaprine shows a weak but highly selective affinity for hippocampic M1 receptors (IC50 = 17.10(-6) [3H]-pirenzepine). On the other hand minaprine has an excellent bioavailability and is well tolerated in human; particularly, minaprine does not induce any cholinergic syndrome. The synthesis of about 300 minaprine analogues was then undertaken using information resulting from a computer graphics analysis of published muscarinic ligands as well as classical structure-activity relationship considerations. These studies identified the features which are associated with high affinity for the muscarinic M1 receptors and led to the synthesis of ligands 5660 times more potent than minaprine. Taking into account factors such as the absence of a chiral center, the duration of activity, the metabolic stability led us to select compound SR 46559 A [3-(N-(2-diethylamino-2-methylpropyl)-6-phenyl-5-propylpyridazinamine sesquifumarate] for the industrial development. This compound is a potent orally active muscarinic agent, with no cholinergic symptom.

Solution structure and dynamics of cyclic and acyclic cholinergic agonists.

Two classes of nicotinic cholinergic agonists, which vary in flexibility and electronegativity, have been synthesized, and their structural and dynamic properties have been studied with nuclear magnetic resonance (NMR) spectroscopy. Although the compounds are chemically identical except for the presence or absence of one cyclicizing C--C bond, single channel recording and radioligand binding studies have shown that the cyclic compounds are considerably more potent than the acyclic derivatives (McGroddy, K.A., A.A. Carter, M.M. Tubbert, and R.E. Oswald. 1993. Biophys. J. 64:325-338). Using one- and two-dimensional NMR spectroscopy, we have shown that these molecules exist in two distinct stable conformers, which differ in the orientation of the amide bond. The cyclic 1,1-dimethyl-4-trifluoroacetyl-piperazinium iodide and its trifluoromethyl derivative compounds are symmetric, and the two conformers are of equal energy. The acyclic N,N,N,N'-tetramethyl-N'-acetylethylene-diamine iodide (TED) and its trifluoromethyl derivative derivatives, however, populate two energetically unequal solution conformations. Using variable temperature NMR spectroscopy on these molecules and their uncharged precursors, we have characterized the energetics of amide bond isomerization and have distinguished steric and electrostatic contributions to the equilibrium between the two conformers. The more populated TED conformer has the amide methyl group trans to the carbonyl oxygen, and it is stabilized by an electrostatic attraction between the partially negative carbonyl oxygen and the positively charged quaternary amine nitrogen. As discussed in the accompanying paper (McGroddy, K.A., A.A. Carter, M.M. Tubbert, and R.E. Oswald. 1993. Biophys. J. 64:325-338), the differences in the stable solution structures of the TED derivatives and their interconversion kinetics may be of biological significance.

[The synthesis and bioactivities of cholinergic tropane alkaloids].

Ten analogs of baogongteng A, the natural cholinergic tropane alkaloid isolated from Erycibe obtusifolia Benth, were designed and synthesized. The tropane skeleton was kept unchanged, while the substituting groups on N and C3 were modified. In myotic experiments in rabbits 3-paramethyl benzoyloxy-6-acetoxy nortropane(4), 3-propionyloxy-6-acetoxy nortropane(5), and 3-isobutyryloxy-6-acetoxy nortropane(6) showed cholinergic activities.

Synthesis and SAR of bulky 1-azabicyclo[2.2.1]-3-one oximes as muscarinic receptor subtype selective agonists.

The synthesis of a series of potent and efficacious 1-azabicyclo[2.2.1]heptan-3-one oxime muscarinic agonists is described. The oximes have extended appendages designed to span the cavity defined by the seven transmembrane helices of the muscarinic receptor. Some members of the series are selective for receptors of the m1 subtype. One such oxime, 31, shows affinity and functional selectivity for m1 over m2, m3, and m4 muscarinic receptor types.

Synthesis of 2-(3-substituted-1,2,4-oxadiazol-5-yl)-8-methyl-8-azabicyclo [3.2.1]octanes and 2 alpha-(3-substituted-1,2,4-oxadiazol-5-yl)-8-methyl-8- azabicyclo[3.2.1]oct-2-enes as potential muscarinic agonists.

Radioligand binding affinities of seven muscarinic receptor ligands which possess an oxadiazole ring side chain have been determined in rat heart, rat brain, and m1- or m3-transfected CHO cell membrane preparations to determine the selectivity for subtypes of muscarinic receptor. The ratios of binding constants in brain membranes were measured as an indicator of potential agonist activity against [3H]QNB and [3H]Oxo-M. These muscarinic ligands did not discriminate the subtypes of muscarinic receptors. Six muscarinic ligands which have a 3-amino- or 3-methyl-1,2,4-oxadiazol-5-yl groups attached to the 8-methyl-8-azabicyclo[3.2.1]oct-2-ene or 8-methyl-8-azabicyclo[3.2.1]octane head group show binding constants between 2.04 x 10(-6) and 1.79 x 10(-5) M in rat heart, rat brain, and m1- or m3-transfected CHO cell membrane preparations. 1-Methyl-2-[3-amino-1,2,4-oxadiazol-5-yl]piperidine shows low binding constants of approximately 10(-4) M in rat heart and rat brain. (1R,5S)-2-[3-Amino-1,2,4-oxadiazol-5-yl]-8-methyl-8-azabicyclo- [3.2.1]oct-2-ene [(1R,5S)-17] was the most active compound.

Novel functional M1 selective muscarinic agonists. 2. Synthesis and structure-activity relationships of 3-pyrazinyl-1,2,5,6-tetrahydro-1-methylpyridines. Construction of a molecular model for the M1 pharmacophore.

A series of 3-(3-substituted-pyrazinyl)-1,2,5,6-tetrahydro-1-methylpyridines were synthesized and found to have high affinity for central muscarinic receptors. The ability of some of these compounds to inhibit the electrically stimulated twitch of the guinea pig vas deferens indicated that the compounds were M1 agonists. M1 agonist activity was related to the length of the side chain attached to the pyrazine ring, with maximal activity being obtained with the hexyloxy side chain. The (hexyloxy)pyrazine 3f lacked M2 agonist activity as it failed to affect the guinea pig atria and was also relatively devoid of M3 agonist activity as determined by its lack of tremorogenic and sialogogic effects in mice. A comparison of the M1 agonist efficacy of these pyrazines and related 1,2,5-thiadiazoles and 1,2,5-oxadiazoles suggested that M1 efficacy was related to the magnitude of electrostatic potential located over the nitrogens of the respective heterocycles. The heteroatom directly attached to the 3 position of the pyrazine or 1,2,5-thiadiazole heterocycle markedly influenced the M1 efficacy of the compounds by determining the energetically favorably conformers for rotation about the bond connecting the tetrahydropyridyl ring and the heterocycle. A three-dimensional model for the M1-activating pharmacophore was proposed based on computational studies and the model of the muscarinic pharmacophore proposed by Schulman.

Indonesian medicinal plants. I. Chemical structures of calotroposides A and B, two new oxypregnane-oligoglycosides from the root of Calotropis gigantea (Asclepiadaceae).

Two new oxypregnane-oligoglycosides named calotroposides A (1) and B (2) have been isolated from the root of Calotropis gigantea (Asclepiadaceae), an Indonesian medicinal plant, and their chemical structures have been elucidated by chemical and spectroscopic methods as 12-O-benzoyllineolon 3-O-beta-D-cymaropyranosyl(1----4)-beta-D-oleandropyranosyl( 1----4)- beta-D-oleandropyranosyl(1----4)-beta-D-cymaropyranosyl(1--- -4)-beta-D- cymaropyranoside and 12-O-benzoyldeacetylmetaplexigenin 3-O-beta-D-cymaropyranosyl(1---4)-beta-D-oleandropyranosyl(- ---4)- beta-D-oleandropyranosyl(1----4)-beta-D-cymaropyranosyl(1--- -4)- beta-D-cymaropyranoside, respectively.

Stereoisomerism and muscarinic receptor agonists: synthesis and effects of the stereoisomers of 3-[5-(3-amino-1,2,4-oxadiazol)yl]-1- azabicyclo[2.2.1]heptane.

The preparation and the biological activities of the four stereoisomers of 3-[5-(3-amino-1,2,4-oxadiazol)yl]-1-azabicyclo[2.2.1]heptane are described. The most potent stereoisomer, 3a, has the 3R,4R configuration, and in vitro activities in (pD2(% efficacy): ileum 8.8 (87%), hippocampus 9.8 (116%) and ganglion 10.2 (36%)). 3b (3S,4S) was weaker (ileum 8.1 (121%), hippocampus 8.5 (107%), ganglion 9.0 (63%)). The other two stereoisomers, 4a (3S,4R; ileum 7.1 (108%), hippocampus 8.2 (116%), ganglion 7.3 (31%)) and 4b (3R,4S; ileum 7.0 (100%), hippocampus 7.0 (120%), ganglion 7.2 (67%)) are of comparable activity, with an analogous profile to that of the more potent stereoisomers. Thus, compounds 3a and 4a, possessing the 4R stereochemistry, showed selectivity for the hippocampus over the ileum. Compound 3a was, however, more potent in the ganglion than in the hippocampus. All four stereoisomers were full agonists in the hippocampus, indicating M1 activity; however, they were partial agonists in the depolarisation of the rat superior cervical ganglion, another M1-mediated response. This may be due to M2-mediated hyperpolarization. With 3a (0.01 mg/kg i.p.), expression of c-fos mRNA was observed in the hypothalamus and in brain areas involved in sensory processing; these effects were totally blocked by pretreatment with 2 mg/kg scopolamine. In particular, activation of the superior colliculus is consistent with potent M2 activity.