Long chain analogs of physostigmine as potential drugs for Alzheimer's disease: new insights into the mechanism of action in the inhibition of acetylcholinesterase.

Heptylphysostigmine is in advanced clinical trial as a drug for Alzheimer's disease. 8-Morpholinooctylphysostigmine and 8-(cis-2,6-dimethylmorpholino)octylphysostigmine are currently undergoing pre-clinical evaluation. The mechanism of action of these compounds in the inhibition of acetylcholinesterase has been investigated. All the examined compounds display non competitive-like kinetics of inhibition. There are no reversible components in the observed inhibition: the whole inhibitory effect is due to the time-dependent pseudo-irreversible carbamylation of the active site. Yet the observed time course of the inhibition does not match a simple second order kinetics. An influence of the quaternary structure of the enzyme on the more complex kinetics of carbamylation is hypothesized. Reactivation experiments on the inhibited enzyme show long lasting inhibitory effects for these compounds. The higher duration of the anticholinesterase effect of the morpholino derivatives compared to heptylphysostigmine should provide the basis for their higher therapeutic potential.

Synthesis and antibacterial activity of some ester, amides, and hydrazides of 3-carboxyrifamycin S. Relationship between structure and activity of ansamycins.

Esters, amides, and hydrazides of 3-carboxyrifamycin S were synthesized by oxidizing the cyanohydrin of 3-formylrifamycin SV to 3-(cyanocarbonyl)rifamycin S, followed by treatment with alcohols, amines and hydrazines. The in vitro microbiological activity of the derivatives was quite low, especially toward Gram-negative bacteria. This poor activity was not shown to be due to the inadequate inhibiting action on the bacterial DNA-dependent RNA polymerase but to the poor penetration of the compounds through the bacterial cell wall. The microbiological activity was correlated to the chemical properties of the substituent on C3.

Potentiation of cADPR-induced Ca(2+)-release by methylxanthine analogues.

Caffeine and other methylxanthines are known to induce Ca(2+)-release from intracellular stores via the ryanodine receptor. In the present work, a range of caffeine analogues, in which methyl groups at the 1 and 7 positions were replaced with alkyl chains containing different functional groups (oxo, hydroxyl, propargyl, ester, and acids), were synthesized. These compounds were then screened for their ability to potentiate Ca(2+)-release induced by cADPR (an endogenous modulator of ryanodine receptors) in sea urchin egg homogenates. Two of the synthesized methylxanthines, 1, 3-dimethyl-7-(7-hydroxyoctyl)xanthine (37) and 3-methyl-7-(7-oxooctyl)-1-propargylxanthine (66), were shown to be more potent than caffeine in potentiating cADPR-induced Ca(2+)-release, while 1,3-dimethyl-7-(5-ethylcarboxypentyl)xanthine (14) was shown to be more efficacious. The development of new methylxanthine analogues may lead to a better understanding of ryanodine receptor function and could possibly provide novel therapeutic agents.

Synthesis and antibacterial activity of some derivatives of tolypomycinone. Relationship between structure and activity in ansamycins.

3-Aminotolypomycinoes and 3,16-diamino-16,17-dihydrotolypomycinones are formed by the addition of primary and secondary amines to tolypomycinone, obtained by mild hydrolysis of the antibiotic tolypomycin Y.3-Amino-16,17-dihydrotolypomycinones are formed by the addition of primary and secondary amines to 16,17-dihydrotolypomycinone. In vitro microbiological tests showed high antibacterial activity in compounds obtained by the addition of primary amines, which must be unbranched in the alpha position to the nitrogen atom to position 3 of the naphthoquinone ring. The relationship between structure and activity is described, and evidence is presented that hydrogen bonding between the amino NH bonded to C3 and the amide CO of tolypomycinone is very important for biological activity.

Synthesis of 2-amino-5-sulfanyl-1,3,4-thiadiazole derivatives and evaluation of their antidepressant and anxiolytic activity.

Recently a series of 2-amino-5-sulfanyl-1,3,4-thiadiazole derivatives bearing different substituents were synthesized and screened pharmacologically in order to evaluate their central nervous system activity. The purpose of this study was to evaluate the effects of the title compounds on CNS activity by varying the substituents in the thiadiazole moiety. It was found that some of these compounds possess marked antidepressant and anxiolytic properties comparable in efficiency to the reference drugs Imipramine and Diazepam. The most potent compound 3k was further investigated to complete its pharmacological profile with respect to undesired side effects. Behavioral results showed that 3k is a very promising compound, characterized by a mixed antidepressant-anxiolytic activity accompanied by a therapeutic dose range that is essentially 2 orders of magnitude less than that at which side effects such as sedation and amnesia are evident.

4-Deoxypyrido[1',2':1,2]imidazo[5,4-c]rifamycin SV derivatives. A new series of semisynthetic rifamycins with high antibacterial activity and low gastroenteric absorption.

A series of 4-deoxypyrido[1',2':1,2]imidazo[5,4-c]rifamycin SV derivatives (6-11) were prepared that demonstrated high antibacterial activity suitable for an intestinal disinfectant. These compounds are zwitterionic in nature and are poorly absorbed through the gastroenteric tract but maintain the ability to cross the bacterial cell wall. X-ray crystallographic data are presented to demonstrate the zwitterionic nature of these compounds. The structure-activity relationship of this novel series of antibiotics is discussed and the derivative with the highest ratio between subcutaneous and oral activity (6) was selected for clinical development. At the outset of this work several 3-(quaternary ammonium bromides) (1-5) were prepared and tested for antibacterial activity. These compounds were demonstrated to be too polar to even cross the bacterial cell wall but led to the synthesis of 6-11.

Acetazolamide-like carbonic anhydrase inhibitors with topical ocular hypotensive activity.

New carbonic anhydrase (EC 4.2.1.1) inhibitors were synthesized as potential drugs for the topical treatment of glaucoma. They were obtained by substituting the acetyl group of acetazolamide and methazolamide with bicarboxylic acids of different chain length (C4-C6). The terminal carboxyl was either kept free or esterified with alcohols of different size (C1-C12). A gamma-aminovaleric derivative was also prepared. All compounds proved active as carbonic anhydrase inhibitors in vitro, with an average IC50 of about 0.5 microM. Some proved also to be topically active in vivo in lowering the artificially elevated intraocular pressure in rabbits. The most active compound, carrying a succinic acid side chain, is the most soluble in aqueous buffers. Its duration of action is about 8 h and it is under evaluation as a topical antiglaucoma drug. It is hypothesized that the duration of action could be longer in compounds having both the same high water solubility and partition coefficient.

Inflammatory molecule release by beta-amyloid-treated T98G astrocytoma cells: role of prostaglandins and modulation by paracetamol.

Deposition of beta-amyloid in the brain triggers an inflammatory response which accompanies the neuropathologic events of Alzheimer's disease and contributes to the destruction of brain tissue. The present study shows that beta-amyloid can stimulate human astrocytoma cells (T98G) to secrete the proinflammatory factors interleukin-6 and prostaglandins. Furthermore, prostaglandins can stimulate T98G to secrete interleukin-6, which in turn triggers the formation of additional prostaglandins. Prostaglandins are, therefore, a key element in the induction and maintenance of a state of chronic inflammation in the brain which may exacerbate the fundamental pathology in Alzheimer patients. Paracetamol (0.01-1000 microM), an unusual analgesic/antipyretic drug which acts preferentially by reducing prostaglandin production within the central nervous system, and indomethacin (0.001-10 microM) caused a clear dose-dependent reduction of prostaglandin E2 production by stimulated T98G cells whereas interleukin-6 release was not affected. These data provide further evidence of the involvement of non-steroidal anti-inflammatory drugs in the inflammatory processes that can be generated by glial cells in intact brain.

New analogs of physostigmine: alternative drugs for Alzheimer's disease?

The synthesis of a series of physostigmine analogs, in which the methylcarbamyl group has been substituted with monoalkylcarbamyl, dimethyl- and diethylcarbamyl groups, is reported. These compounds were prepared with the aim of investigating their possible therapeutic effects in the treatment of Alzheimer's type dementia. The new analogs of physostigmine are inhibitors of acetylcholinesterase from Electroforus electricus, with a value of the reactivation constant, k3 smaller than the one of physostigmine. The percentage of anticholinesterase activity in vitro and in vivo, the acute toxicity and some behavioural effects were also evaluated for selected derivatives. The reactivation constant, in vitro, supports the view that the derivatives described would be more suitable for therapeutic use than physostigmine.

A long-lasting cholinesterase inhibitor affecting neural and behavioral processes.

A series of analogues of physostigmine were prepared with the aim of investigating their inhibitory effects on acetylcholinesterase in the treatment of Alzheimer's disease. One of the isomers prepared was evaluated for its anticholinesterase activity in vivo, acute toxicity, and some behavioral effects. This compound was a competitive inhibitor of the enzyme and was found to antagonize the stimulating effect produced by scopolamine on locomotor activity and to facilitate memory consolidation.

Synthesis of a carbocyclic sialic acid analogue for the inhibition of influenza virus neuraminidase.

The influenza virus neuraminidase (NA) is essential for viral infection and offers a potential target for antiviral drug development. We prepared a carbocyclic sialic acid analogue, potentially able to inhibit NA. Its structure is an analogue of the transition-state of the reaction catalysed by NA. As starting material, quinic acid was selected owing to its ready availability and its stereochemical feature suitable for the target structure. The quinic acid was first converted in the shikimic acid; then two of the three hydroxyl functions of this product were selectively functionalised to obtain the target molecule (3R,4S,5R)-4-acetamido-3-guanidino-5-hydroxycyclohex-1-ene-1-carboxylic acid.

Synthesis of some carbamates of myo-inositol.

The syntheses are described of the 1-O-carbamoyl (11), 1-O-carbamoyl-2-O-stearoyl (10), 1-O-(acetylcarbamoyl)-2-O-stearoyl (12), 1-O-(heptylcarbamoyl) (13), 2-O-(heptylcarbamoyl) (14) 1,2-di-O-(heptylcarbamoyl) (15), and 1-O-(octadecylcarbamoyl) (16) derivatives of myo-inositol. None of these compounds had significant activity against phospholipase C.

The synthesis of 4-deoxypyrido[1',2'-1,2]imidazo[5,4-c]rifamycin SV derivatives.

Two series of new semisynthetic rifamycin SV derivatives have been prepared. One of them bears a quaternary ammonium salt at C3 (1-5), and the other a pyridoimidazo system condensed at C3 and C4 (6-11). While compounds 1-5 had poor antibacterial activity in vitro, compounds 6-11 were found to be highly active in vitro but poorly absorbed in vivo. They could thus have potential as agents in the therapy of intestinal infections. The synthesis and the 1H NMR structure determination of these new compounds are reported.

Chemical modifications of the aliphatic bridge of ansamycins. Synthesis and activity of 25-deacetoxy-25-epi-hydroxyrifamycin S.

Rifamycins are supposed to bind to, and inhibit the bacterial DNA-dependent RNA polymerase (DDRP) by the formation of hydrogen bonds through O (1), O (2), O (9), O (10). Therefore, with the aim of increasing the intrinsic activity of rifamycin S (1), the 25-deacetoxy-25-epi-hydroxyrifamycin S (8), was synthesized, which displays an additional hydroxyl available for the inhibiting interaction with the bacterial enzyme. The configuration and conformation of the new compound were as expected, but the biological evaluation did not confirm the hypothesis.

Chemical modifications of the aliphatic bridge of ansamycins. 3. Synthesis and activity of 21-epi-rifamycin S.

Rifamycins inhibit bacterial DNA-dependent RNA polymerase through the formation of non-covalent bonds by the oxygenated groups at C(1), C(8), C(21), and C(23). These must be unhindered and underivatized, with the antibiotic in a proper overall molecular conformation. The present study shows that contrary to previous conclusions the availability of the hydroxyl group at C(21) is not as important as that of the other three groups. In support of this is the observation that 21-epi-rifamycin S is partially active, both on the isolated DNA-dependent RNA polymerase and on some Gram-positive bacterial strains.

X-ray crystal structure of 4-deoxy-3'-bromopyrido[1',2'-1,2]imidazo[5,4-c]rifamycin S.

This paper reports the determination of the X-ray molecular structure of 4-deoxy-3'-bromopyrido[1',2'-1,2]imidazo[5,4-c]rifamycin S, carried out in order to unequivocally define the general structure of a new series of rifamycin SV derivatives, which are potent antibacterial agents, and are not absorbed at the gastroenteric level. They have been prepared by Alfa Farmaceutici, Bologna, by condensing 2-aminopyridine derivatives to 3-bromorifamycin S. The solid state X-ray study has confirmed the structure proposed on the basis of 1H NMR studies in solution. It has also shown that the newly formed pyridoimidazo system is in a mesomeric betaine form, the pyrido nitrogen being positively charged and the imidazo nitrogen being negatively charged. This feature is believed responsible for the pharmacokinetic behavior of these new drugs, one of which, denoted either as rifamycin L 105 or rifaximin, is actually under clinical trial as a topical intestinal disinfectant.

Metabolites of microorganisms. 229. Absolute configuration of naphthomycin A determined by X-ray analysis and chemical degradation.

The relative and absolute configurations of naphthomycin A were elucidated by an X-ray structural analysis of a methylation product, 25-O-methylnaphthomycin A iminomethyl ether. The absolute configuration was confirmed by degradation (O3, NaBH4) to (S)-butane-1,2,4-triol.

A new simple method for determining the kinetic constants of inhibited acetylcholinesterase.

The authors, with a spectrophotometric procedure, have evaluated the kinetic constants of physostigmine, a cholinesterase inhibitor which assumes an important function when the degeneration of cholinergic projections is observed. This approach overcomes some of the difficulties encountered in measuring the binding and the carbamylation rate constants. A kinetic treatment, consistent with the experimental results observed in the enzyme-inhibitor system in the presence of substrate, is developed.

Acetylcholinesterase inhibition by eserine: rate constants of reaction. Part II.

The inhibition of eel acetylcholinesterase by physostigmine at 20 degrees and 25 degrees C have been investigated. In our evaluation the unimolecular reactivation rate constant, k3, the carbamylation rate constant, k2, and the binding constant, Ka, are the first simultaneously determined. The mechanism of this reaction is discussed.

Aromatic sialic acid analogues as potential inhibitors of influenza virus neuraminidase.

The influenza virus neuraminidase (NA) is an enzyme essential for viral infection and offers a potential target for antiviral drug development. We aimed our research at the synthesis of non-carbohydrate molecules able to inhibit NA as transition-state analogues. Aromatic sialic acid analogues (compound 5 and compound 10) were synthesised in good yields starting from commercially available benzoic acids using a suitable synthetic strategy.