3-Aryl-5-hydroxy-5-vinyl-2-isoxazolines were synthesized by reacting aryl nitrile oxides with the lithium enolate of methyl vinyl ketone (MVK) at-78 °C. Fair to good yields are obtained in the case of aryl nitrile oxides bearing electron-withdrawing groups on the aryl moiety or less bulky groups. Conversely, lower yields or no reaction was observed in the presence of hindered aryl nitrile oxides. Such a behavior was confirmed by ab initio calculations of the activation energies for three reactions. A number of 3-aryl-5-vinylisoxazoles were quantitatively obtained by dehydration/aromatization of the corresponding 5-hydroxy-2-isoxazolines under acidic conditions. The side-chain elaboration is reported as a synthetic utility of some vinylisoxazoles and vinylisoxazolines

A novel procedure for the synthesis of N-unsubstituted β-enaminothioesters, ( Z)- S-phenyl3-amino-3-arylprop-2-enethioates, from 3-arylisoxazoles or 3-aryl-5-phenylthio-2-isoxazolinesis described and a plausible mechanism for the reaction is proposed. Someexamples of conversion of one β-enamino thioester [( Z)- S-phenyl3-amino-3-phenylprop-2-enethioate] into N-unsubstituted β-enaminoacylderivatives (β-enamino esters, β-enamino amides, and β-ketoamides) are also reported.

The invention claims the identification of beta3-adrenoceptor inverse agonists, the synthetic strategy for their preparation and their use as a drug

Optically active molecules are important building blocks for the synthesis of many chemicals and biologically active compounds. Among the known catalysts, isolated enzymes have some advantages over conventional methods in the asymmetric synthesis, such as chemo-, regio-, and stereo-selectivity, together with very mild reaction conditions [1]. The asymmetric synthesis accomplished by using whole-cells has also further advantages [2, 3] because all the necessary cofactors and all required substances for their regeneration are present in their natural environment, thus making the catalytic system more efficient [4]. Many research groups have focused their attention on looking for non-conventional yeasts, to study in comparison to the deeply investigated Saccharomyces cerevisiae [5]. For several years our interests focused on using non-conventional yeasts to prepare new EPCs: among these, thermotolerant Kluyveromyces marxianus CBS 6556, not widely investigated in asymmetric synthesis, was preliminarly and successfully used by us for the stereoselective bioreduction of prostereogenic keto-esters to prepare optically active building-blocks in the synthesis of pharmacologically active compounds [6, 7]. These studies allowed also the isolation of an unkown ADH from this yeast [8], able to mediate the highly stereoselective bioreduction of prostereogenic 3-oxo esters. Herein, we report the continuation of such studies, in which the Kluyveromyces marxianus CBS 6556 is used in the bioreduction of various prochiral ketones, with the aim to deepen its substrate specificity, turnover rate, regio- chemo- and enantioselectivity.

Inverse agonists are useful active ingredient of drugs clinically used to treat diseases mainly involving receptors endowed with nonendogenous agonist induced activity (constitutive or basal activity). SP-1e and SP-1g are the first two potent and highly selective β3-adrenoceptor inverse agonists [EC50=181 nM (IA=- 64%) and 136 nM (IA=-73%), respectively], which their peculiar activity seems due to the absolute configurations of the two stereogenic centres present in each molecule. Rat proximal colon motility measurements allowed their further pharmacological characterization and pA2 values determination by Schild analysis (7.89 and 8.16, respectively). The purpose of our work is a further characterization of our novel β3-adrenoceptor agonists (SP-1a-d, SP-1f,1h) and inverse agonists (SP-1e and SP1g) on rat proximal colon motility and a confirmation of their inverse agonist nature in a more complex system like the functional test on rat proximal colon. Male Wistar rats segment of the proximal colon were placed in organ baths containing Krebs solution. Muscle tension was recorded isotonically. Cumulative β3-AR agonists doses experiments were performed for each test compound: isoprenaline, BRL37344, SP-1a-d, SP-1f and SP-1h were dissolved in Krebs. The EC50 values of each agonists and pA2 of inverse agonists were determined. SP-1a-d, SP-1f and SP-1h in rat colon have a muscle relaxing effect thus confirming their partial agonist activity found in CHO-K1 cell line. SP-1e and SP-1g behaved as antagonists with pA2 values of 7.89 and 8.16, espectively. In conclusion, experiments carried out by using isolated rat proximal colon allowed us to determine the pA2 values of the two β3-AR inverse agonists and add knowledge on the behavior of a novel set of compounds and their possible value as agents useful whenever is necessary to also control the colon motility.

The synthetic utility of enolates in the heterocycles synthesis constitutes the main topic of this review. Appropriate enolates, in a number of synthetic approaches, are easily formed in situ from a variety of carbonyl compounds (aldehydes, ketones, amides, etc.) and used for the ring construction of (poly)substituted heterocycles. Often, the synthesis of five-membered rings such as isoxazoles, triazoles, pyrazoles, etc. has been performed by using these intermediates through a two-step or even by one pot procedures. The selectivity and the versatility of enolates intermediates will be discussed, with particular attention to those methodologies useful for preparation of heterocycle containing pharmacologically active molecules

A novel set of 1,4-diaryl-1,2,3-triazoles were projected as a tool to study the effect of both the heteroaromatic triazole as a core ring and a variety of chemical groups with different electronic features, size and shape on the catalytic activity of the two COX isoenzymes. The new triazoles were synthesized in fair to good yields and then evaluated for their inhibitory activity towards COXs arachidonic acid conversion catalysis. Their COXs selectivity was also measured. A predictive pharmacometric Volsurf plus model, experimentally confirmed by the percentage (%) of COXs inhibition at the concentration of 50 μM and IC50 values of the tested compounds, was built by using a number of isoxazoles of known COXs inhibitory activity as a training set. It was found that two compounds {4-(5-methyl-4-phenyl-1H-1,2,3-triazol-1-yl)benzenamine (18) and 4-[1-(4-methoxyphenyl)-5-methyl-1H-1,2,3-triazole-4-yl]benzenamine (19)} bearing an amino group (NH2) are potent and selective COX-1 inhibitors (IC50 Combining double low line 15 and 3 μM, respectively) and that the presence of a methylsulfamoyl group (SO2CH3) is not a rule to have a Coxib. In fact, 4-(4-methoxyphenyl)-5-methyl-1-[4-(methylsulfonyl)phenyl]-1H-1,2,3-triazole (23) has COX-1 IC50 Combining double low line 23 μM and was found inactive towards COX-2.

COX-1 isoenzyme has been recently reconsidered as therapeutic target, due to its crucial role exerted in a variety of pathological conditions, such as atherosclerosis, endothelial dysfunction, neuroinflammation, pain processing, pre-term labor and some type of cancers. Hence, highly selective COX-1 inhibitors might be particularly relevant for the treatment of several diseases [1]. 3-(5-Chlorofuran-2-yl)-5-methyl-4- phenylisoxazole, P6, a highly selective COX-1 inhibitor, recently uncovered by us[2], has been chosen as !lead compound" for structure-activity relationship studies [3].They assessed that the presence of the P6-furanyl group is crucial for COX-1 inhibitory potency and selectivity, as it is important the substituent size (bromine, chlorine or methyl group) on that furanyl. In addition, the replacement of a methyl by CF3-group at isoxazole C5 and the introduction of a substituent on the phenyl bonded to the isoxazole C4 still provide selective COX-1 inhibitors. Among the diarylheterocycle class of COX-1 inhibitors, the most studied COX-1 inhibitor is the SC-560 that has a pyrazole as a core ring instead of an isoxazole [4]. Thus, to identify the P6 and SC-560 common (if any) structural and/or electronic determinants responsible of the selective COX-1 inhibition, a series of new pyrazole analogues of P6 have been prepared by substituting the P6-isoxazole core ring with a pyrazole. The results of this investigation will be presented

The use of microorganism growing cells is a well recognized methodology in biocatalyzed organic reactions. A non-conventional thermotolerant Kluyveromyces marxianus yeast strain was used for the bioreduction of different arylketones. Differently substituted ketones were converted into the corresponding (S)-alcohols with up to 96% enantiomeric excess under very mild reaction conditions. Kluyveromyces marxianus represents a promising biocatalyst for the production of optically active 1-arylethanols.

INTRODUCTION: Stimulation of the β(3)-adrenoceptor (β(3)-AR) is thought to be a valuable approach for the treatment of obesity, type 2 diabetes, heart failure, frequent urination, preterm labor, anxiety and depression. Therefore, the β(3)-AR is recognized as an attractive target for drug discovery. Simultaneous activation of the β(1)- and β(2)-AR can cause undesirable side effects such as increased heart rate and muscle tremors. Consequently, much effort has been directed towards the design and development of selective β(3)-AR agonists through original synthetic chemistry, extensive in vitro tests and detailed preclinical investigations to various phases of clinical trials. AREAS COVERED: SciFinder Scholar, PubMed, ISI web of Knowledge(SM), Espacenet, ClinicalTrials and Google have been used as the main sources for retrieving literature and patents filed since the discovery of β(3)-AR through to June 2010. This review discusses the enormous efforts made by private and public research laboratories to uncover β(3)-AR ligands and to prove their usefulness as drugs. EXPERT OPINION: Remarkable knowledge has been gained about the physio-pathological role of the β(3)-AR to date. Many highly potent and selective β(3)-AR ligands (agonists, antagonists and inverse agonists) have been discovered; however, further investigations are still needed to identify novel compounds acting as β(3)-AR ligands in order to adequately treat the diseases in which β(3)-AR is involved.

The present invention relates to novel pyrazoles which are potent and selective inhibitors of cyclooxygenase-1 (COX-1) and to their radiolabeled derivatives thereof which are both useful as theranostics of a number of pathologies.

COX-1 plays a previously unrecognized part in the neuroinflammation. Genetic ablation or pharmacological inhibition of COX-1 activity attenuates the inflammatory response and neuronal loss. In this context, the effects of selective COX-1 inhibitors (P6, P10, SC-560, aspirin) and coxibs (celecoxib and etoricoxib) on LPS-stimulated microglial cell function (a worldwide accepted neuroinflammation model) were investigated, and the effects on COX-1/COX-2, cPGES mRNA and iNOS expression, PGE2 and NO production and NF-kB activation by IkBalpha phosphorylation were evaluated. The total suppression of the expressionof both COX-1 and COX-2 by their respective selective inhibitors occurred. NF-kB remained almost completely inactive in the presence of coxibs, as expected, and totally inactive in the presence of P6. P6 also markedly counteracted LPS enhancing cPGES mRNA expression and PGE2 production. Since COX-1 is predominantly localized in microglia, its high selective inhibition rather than COX-2 (by coxibs) is more likely to reduce neuroinflammation and has been further investigated as a potential therapeutic approach and prevention in neurodegenerative diseases with a marked inflammatory component.

Cyclooxygenase-1 (COX-1), but not COX-2, is expressed at high levels in the early stages of human epithelial ovarian cancer where it seems to play a key role in cancer onset and progression. As a consequence, COX-1 is an ideal biomarker for early ovarian cancer detection. A series of novel fluorinated COX-1-targeted imaging agents derived from P6 was developed by using a highly selective COX-1 inhibitor as a lead compound. Among these new compounds, designed by structural modification of P6, 3- (5-chlorofuran-2-yl)-5-(fluoromethyl)-4-phenylisoxazole ([18/19F]-P6) is the most promising derivative [IC50 ¼ 2.0 mM (purified oCOX-1) and 1.37 mM (hOVCAR-3 cell COX-1)]. Its tosylate precursor was also prepared and, a method for radio[18F]chemistry was developed and optimized. The radiochemistry was carried out using a carrier-free K18F/Kryptofix 2.2.2 complex, that afforded [18F]-P6 in good radiochemical yield (18%) and high purity (>95%). In vivo PET/CT imaging data showed that the radiotracer [18F]-P6 was selectively taken up by COX-1-expressing ovarian carcinoma (OVCAR 3) tumor xenografts as compared with the normal leg muscle. Our results suggest that [18F]-P6 might be an useful radiotracer in preclinical and clinical settings for in vivo PET-CT imaging of tissues that express elevated levels of COX-1.

3-(5-Chlorofuran-2-yl)-5-methyl-4-phenylisoxazole (P6), a known selective cyclooxygenase-1 (COX-1) inhibitor, was used to design a new series of 3,4-diarylisoxazoles in order to improve its biochemical COX-1 selectivity and antiplatelet efficacy. Structure−activity relationships were studied using human whole blood assays for COX-1 and COX-2 inhibition in vitro, and results showed that the simultaneous presence of 5-methyl (or -CF3), 4-phenyl, and 5-chloro(-bromo or -methyl)furan-2-yl groups on the isoxazole core was essential for their selectivity toward COX-1. 3g, 3s, 3d were potent and selective COX-1 inhibitors that affected platelet aggregation in vitro through the inhibition of COX-1-dependent thromboxane (TX) A2. Moreover, we characterized their kinetics of COX-1 inhibition. 3g, 3s, and 3d were more potent inhibitors of platelet COX-1 and aggregation than P6 (named 6) for their tighter binding to the enzyme. The pharmacological results were supported by docking simulations. The oral administration of 3d to mice translated into preferential inhibition of platelet-derived TXA2 over protective vascular-derived prostacyclin (PGI2).

La presente invenzione ha per oggetto un nuovo procedimento per la sintesi di alcoli chirali mediante l’impiego di Lactobacillus reuteri. L’invenzione ha anche per oggetto l’uso di tale microorganismo per la sintesi di detti alcoli chirali.

Kluyveromyces marxianus CBS 6556 growing cell versatility in the enantioselective reduction of ketone-carbonyl functionality to the corresponding alcohols was exploited. In particular, methyl ketones were reduced to (S)-alcohols with ee up to 96%. Longer chain alkyl ketones afford, in the same experimental condition, (R)-alcohols with ee up to 84%. Interestingly, carbon-carbon double and triple bond of ketones can also be reduced in the presence of Kluyveromyces marxianus CBS 6556 yeast. Endocyclic carbonyl functionality, such as 2-tetralone carbonyl, is also quantitatively reduced to its (S)-alcohol with ee = 76%.

Since cyclooxygenase (COX) isozymes discovery, many papers and reviews have been published to describe the structural bases of COX inhibition, and to debate on the therapeutic and adverse effects of worldwide clinically used nonsteroidal anti-inflammatory drugs (NSAIDs), included COX-2 selective inhibitors (well known as Coxibs). COX-2 inhibition has been widely investigated, whereas the role of COX-1 in human pathophysiology is mostly not yet well ascertained. As time goes on, the cliché that the constitutively expressed isoform COX-1 is only involved in normal physiological functions, such as platelet aggregation, gastric mucosa protection and renal electrolyte homeostasis is going to be shattered. Low-dose aspirin, behaving as a preferential inhibitor of platelet COX-1, allowed to enlighten the role exerted by this isoenzyme in many mammalian cell types. This review would elucidate the most recent findings on selective COX-1 inhibition and their relevance to human pathology such as cancer, neuro-inflammation, cardioprotection, fever and pain. It would also focus on the design and development of new highly selective COX-1 inhibitors, useful tools in pharmacological studies aimed at gaining a deeper insight of the role of COX-1 in human health and disease. Among the traditional NSAIDs, other then aspirin and indomethacin, only few examples of selective COX-1 inhibitors (SC-560, FR122047, mofezolac, P6 and TFAP) have been so far identified. This review has also the scope to stimulate the development of novel drugs, which activity is COX-1 mediated.

Background/Aims: Gastrointestinal damage (GD) is commonly associated with the inhibition of cyclooxygenase (COX)-1, one of the two known COXs, by traditional non-steroidal anti-inflammatory drugs. More recent evidences have proven that GD is caused by the simultaneous inhibition of the two COXs. This study was designed to evaluate the effect of the selective COX-1 inhibition on gastric integrity. Methods: GD was evaluated in male CD1 mice. Drugs were administered by gastric gavage at a dose of 50 mg/kg (injection volume of 100 μl). Control mice received an equal volume of the vehicle (10% ethanol). Each mouse, in groups of at least 6 mice, received one dose/day for 5 days. Results: In Western blot analysis, COX-1 expression levels were found to be significantly reduced in mice treated with 3-(5-chlorofuran-2-yl)-5-methyl-4-phenylisoxazole (P6) in comparison to mice pretreated with aspirin (ASA), which exhibited higher levels of COX-1, thus confirming the high selectivity of P6 towards COX-1 enzyme inhibition. Mucosal sections obtained from ASA-treated mice showed breaks in the epithelial barrier and a marked alteration of foveolae and gastric glands, whereas stomachs isolated from mice sacrificed after 5 days of chronic administration of P6 (at a dose of up to 50 mg/kg/day) showed sporadic transient mucosal hyperemia and did not seem to display any significant gastric damage. Conclusions: The selective COX-1 inhibition by P6 does not cause gastric damage in mice but preserves mucosal integrity.

Microglia, the immune cells of the brain, are one of the key mediators of neuroinflammation. Microglial activation leads these cells to produce various proinflammatory and neurotoxic substances. Several in vitro and in vivo studies have demonstrated that a decrease in the levels of pro-inflammatory mediators in microglia can attenuate the severity of neuro-degenerative diseases, including AD, PD, ALS, MS and Huntington’s disease. Thus, regulation of excessive microglial activation should be of therapeutic value especially in neuro-degenerative disorders. Cyclooxygenase(COX) is a key enzyme in the conversion of arachidonic acid into prostaglandins and other lipid mediators. It is widely accepted that this enzyme play a pivotal role in the neuro-inflammatory process exacerbation. Two COX isoforms are known: COX-1, constitutively expressed in most tissues, classically considered as the isoform primarily responsible for maintaining the homeostasis by mediating physiological responses, and COX-2, which represents the inducible form, mainly activated in response to inflammatory stimuli. In this respect, recent studies have also indicated a previously unrecognized pro-inflammatory role of COX-1 in the pathophysiology of acute and chronic neurological disorders. Consequently, it was reconsidered the potential beneficial effect of COX-1 inhibition in the treatment of neuroinflammation [1]. These findings prompted us to investigate the behaviour of two selective COX-1 inhibitors such as P6 (COX-1 IC50= 0.5 mM and COX-2 IC50 >100 mM) and P10 (COX-1 IC50= 0.09 mM and COX-2 IC50 = 2.49 mM) [2] in an in vitro experimental model of inflammation, represented by mouse N13 microglial cells activated by lipopolysaccharide (LPS) [3]. LPS is a component of the outer cell wall of gramnegative bacteria and is well known as an inducer of inflammatory responses. New selective COX-1 inhibitors are here proposed also as useful tools in pharmacological studies to investigate the role of COX-1 isoenzyme in neuroinflammation or more in general in all the investigations in which only COX-1 needs to be selectively inhibited. The results of this study as well as their rationale will be presented and discussed. References: 1. Perrone, M.G.; Scilimati, A.; Simone, L., Vitale, P. Curr. Med. Chem.2010,17, 3769-805. 2. Di Nunno. L., Vitale, P., Scilimati, A., Tacconelli, S.; Patignani, P. J. Med. Chem. 2004, 47, 4881-90. 3. Calvello, R.; Panaro, M.A.; Carbone, M.L.; Cianciulli, A.; Perrone, M.G.; Vitale, P.; Malerba, P.; Scilimati, A. Biochem. Pharmacol.,2011, submitted for publication.

3- and 4-N,N-Dialkylaminobenzonitriles and 4-chloro-(N,N-dialkyl)benzamidines were isolated by reacting 4-chlorobenzonitrile with hindered lithium amides under thermodynamic (0 °C) and kinetic control conditions (-78 °C), respectively. As previously reported, a benzyne mechanism seems to be confirmed since N,N-dialkylaminobenzonitriles are formed. Only benzamidines were isolated in fair to high yields at both 0 °C and -78 °C with non-hindered lithium amides. Exploitation and mechanistic rationale of the reaction of different halobenzonitriles are also reported

isoxazole derivatives development as cyclooxygenase inhibitors

La presente invenzione si riferisce ai derivati ​​di isossazolo, in particolare agli inibitori dei derivati ​​del diarilisoxazolo della cicloossigenasi (COX), in particolare la cicloossigenasi-1 (COX-1), alle loro composizioni farmaceutiche, al processo per la loro preparazione e al loro uso per la chemioprevenzione e trattamento dell'infiammatorio sindromi e nella prevenzione e trattamento dei carcinomi, in particolare dei carcinomi intestinali, ovarici e cutanei, nel trattamento delle sindromi dolorose, in particolare dopo l'intervento chirurgico, e nel campo cardiovascolare come antitrombotici / vasoprotettivi / cardioprotettivi.

La presente invenzione riguarda composti eterociclici di formula generale (II) che sono ligandi del recettore adrenergico beta-3 e il loro uso in terapia.

La presente invenzione si riferisce a nuovi eterocicli che sono inibitori potenti e selettivi della cicloossigenasi-1 (COX-1) e ai loro derivati ​​radiomarcati che sono entrambi utili come teranostici di un certo numero di patologie.

L'invenzione riguarda una nuova classe di composti di formula (I) indirizzati a COX-1. L'invenzione riguarda anche l'uso di alcuni di tali composti come uno strumento per studiare la struttura e la funzione dell'enzima, nel trattamento mirato alla COX-1 o al rilevamento di COX-1 in disturbi o malattie correlate come il cancro e la neuroinfiammazione, in particolare in neurologico (es. disturbi dello spettro autistico) e malattie neurodegenerative (es Malattie di Alzheimer, Malattia di Parkinson, sclerosi laterale amiotrofica (SLA), sclerosi multipla (SM), trauma cranico (TBI), demenza da HIV e malattie da prioni) e nel tumore ginecologico (es. Cancro ovarico), tumore del collo e della testa e ematologico tumori (ad esempio mieloma multiplo) e nella rilevazione di COX-1 in "in vitro" (cellule e tessuti) e in "in vivo".

Formulazione farmaceutica comprendente 3,4-diidrossi-6- [18F] -fluoro-L-fenilalanina e almeno un agente tampone in un veicolo acquoso, in cui la formulazione ha un valore di pH compreso tra 4,0 e 5,5, preferibilmente tra 4,5 e 5,0, più preferibilmente circa 5 e usi corrispondenti nei metodi di imaging diagnostico.

The present invention relates to novel heterocycles which are potent and selective inhibitors of cyclooxygenase-1 (COX-1) and to their radiolabeled derivatives thereof which are both useful as theranostics of a number of pathologies.