An Oxidation Study of Phthalimide-Derived Hydroxylactams.

A systematic study of the oxidation of 3-hydroxy-2-substituted isoindolin-1-ones (hydroxylactams) and their conversion to the corresponding phthalimides was undertaken using three oxidants. Of special interest was the introduction of nickel peroxide (NiO2) as an oxidation system for hydroxylactams and comparison of its performance with the commonly used pyridinium chlorochromate (PCC) and iodoxybenzoic acid (IBX) reagents. Using a range of hydroxylactams, optimal conversions of these substrates to the corresponding imides was achieved with 50 equivalents of freshly prepared NiO2 in refluxing toluene over 5-32 h reaction times. By comparison, oxidations of the same substrates using PCC/silica gel (three equivalents) and IBX (three equivalents) required oxidation times of 1-3 h for full conversion but required lengthier purification. While nominal amounts (~25 mg) of substrate hydroxylactams were used to ascertain conversion, scale-up procedures using all three methods gave good to excellent isolated yields of imides.

Chemoenzymatic route to stereodefined 2-(azidophenyl)oxazolines for click chemistry.

Aryl-substituted esters of a racemic diprotected 2-azido-1-alkanol were submitted to the Staudinger/aza-Wittig reaction in order to assess scope and establish conditions for their cyclization to the corresponding 2,4,5-trisubstituted oxazolines. Following the cyclization study, the (2R,3R)-antipode of the azidoalkanol was obtained in high ee by incubation of the corresponding racemic azidoacetate with pig liver esterase (PLE). The p-nitrobenzoate of the enantioenriched 2-azido-1-alcohol was cyclized by the Staudinger/aza-Wittig to give the corresponding (4R,5R)-disubstituted-2-(4-nitrophenyl) oxazoline. Selective reduction of the nitrophenyloxazoline to the corresponding aminophenyloxazoline using aluminum amalgam followed by direct azidation of the 2-(4-aminophenyl) moiety provided the corresponding (4R,5R)-2-(4-azidophenyl) oxazoline derivative. The azidophenyl oxazoline was reacted with a proven click partner 4-ethynylfluorobenzene under copper/sodium ascorbate mediation to provide the click triazole product in high yield.

Applicability of aluminum amalgam to the reduction of arylnitro groups.

An array of arylnitro compounds with various functionality were treated with freshly-prepared aluminum amalgam in THF/water solution and resulted in the corresponding arylamines. The Al(Hg)-mediated reductions are relatively rapid with consumption of the amalgam and disappearance of starting material occurring over 20-30 minutes. The workup of the reductions involves only removal of the insoluble by-products by filtration followed by concentration. Only in some cases is chromatography required to secure the pure product. The desired arylamines are furnished in quantities of 25-100 mg, which in some cases, could be taken on to the next reaction without further purification. Reductions of 4-nitrobenzyl derivatives of carbohydrates or nucleosides were selective in affording the corresponding 4-aminobenzyl products. To show applicability in click chemistry, selected aminobenzyl products are directly azidated to yield products that were then used in click reactions to afford the corresponding 1,2,3-triazoles.

In Vitro and In Vivo Activity of Peptidomimetic Compounds That Target the Periodontal Pathogen Porphyromonas gingivalis.

The interaction of the periodontal pathogen Porphyromonas gingivalis with oral streptococci is important for initial colonization of the oral cavity by P. gingivalis and is mediated by a discrete motif of the streptococcal antigen I/II protein. A synthetic peptide encompassing this motif functions as a potent inhibitor of P. gingivalis adherence, but the use of peptides as topically applied therapeutic agents in the oral cavity has limitations arising from the relatively high cost of peptide synthesis and their susceptibility to degradation by proteases expressed by oral organisms. In this study, we demonstrate the in vitro and in vivo activity of five small-molecule mimetic compounds of the streptococcal peptide. Using a three-species biofilm model, all five compounds were shown to effectively inhibit the incorporation of P. gingivalis into in vitro biofilms and exhibited 50% inhibitory concentrations (IC50s) of 10 to 20 µM. Four of the five compounds also significantly reduced maxillary alveolar bone resorption induced by P. gingivalis infection in a mouse model of periodontitis. All of the compounds were nontoxic toward a human telomerase immortalized gingival keratinocyte cell line. Three compounds exhibited slight toxicity against the murine macrophage J774A.1 cell line at the highest concentration tested. Compound PCP-III-201 was nontoxic to both cell lines and the most potent inhibitor of P. gingivalis virulence and thus may represent a novel potential therapeutic agent that targets P. gingivalis by preventing its colonization of the oral cavity.

Antiangiogenic Activity and Chemical Derivatization of the Neurotoxic Acetogenin Annonacin Isolated from Asimina triloba.

Annonacin (1) was isolated from the North American pawpaw ( Asimina triloba), as reported earlier from these laboratories. Natural 1 was submitted to the rat aortic ring bioassay for evaluation of antiangiogenic activity and was found to inhibit microvessel growth (IC50 value of 3 µM). 4,10,15,20-Tetraazido derivatives of 1 were prepared by permesylation followed by azide displacement or by iodination followed by azide displacement. The tetraazide derived from mesylation/azidation was antiangiogenic, while that derived from iodination/azidation exhibited no appreciable activity. The membrane permeability of natural 1 was evaluated using the parallel artificial membrane permeability assay and was found to be marginally permeable as compared to several clinically relevant compounds.

Alternate pathway for the click reaction of 2-(2-azidophenyl)-4,5-diaryloxazoles.

The CuSO4/ascorbate-mediated 'click' reaction of 2-(2-azidophenyl)-4,5-diaryloxazoles and arylacetylenes proceeded through an alternate pathway whereby reduction of the azide predominated over formation of the 1,2,3-triazole-forming cycloaddition. The unimolecular product, 2-(2-aminophenyl)-4,5-diphenyloxazole, was isolated which appears to be a formal reduction of the arylazide to the corresponding arylamine. A series of oxazoles which possessed various substituents (F, Cl, Br, OCH3) on the 4,5-diaryl rings and having the 2-azido group on the 2-oxazolylphenyl position were submitted to the same 'click' conditions and gave the corresponding arylamine products (73-99%). The reaction appears to be specific toward the ortho-azido substitution of the polycyclic system, as the corresponding azidomethyl-substituted phenyl oxazoles do not give the 'reduction' products but gave the expected click products with the acetylenic co-reactants.

The intramolecular click reaction using 'carbocontiguous' precursors.

The synthesis and utilization of all carbon-chain 'carbocontiguous' azidoalkynyl precursors for an intramolecular click reaction is described. The substrates contain both azidoalkyl and ethynylmethyl groups which are conjoined by a 2-(phenylsulfonylmethyl)-4,5-diphenyloxazole lynchpin and are suitably disposed for ring closure. On promotion by copper salts, a number of cyclic click products having the 1,4-disubstituted endo-fused triazole component and the 4,5-diphenyloxazole component are obtained. In one case, removal of the phenylsulfonylmethyl group from the substrate prior to cyclization gave the 1,5-disubstituted exo-fused triazole. The utilization of CuSO4/sodium ascorbate system appears to be the optimal conditions for closure/cyclization and afforded the cyclized products in yields of 84-95%.

m-Chloroperbenzoic acid-oxchromium (VI)-mediated cleavage of 2,4,5-trisubstituted oxazoles.

An array of 2-substituted-4,5-diphenyloxazoles were found to be cleaved to triacylamines and diacylamines (imides) using a reagent system composed of 3-chloroperbenzoic acid (MCPBA) and 2,2'-bipyridinium chlorochromate (BPCC). The 2-alkyl-4,5-diphenyloxazoles give imides (38-60%) as the predominant cleavage product while the 2-aryl-4,5-diphenyloxazoles give triacylamines (44-71%). Two mechanisms involving intermediates such as cyclic endoperoxides or oxachromacycles were proposed. An application of the oxidative cleavage to the multi-step synthesis of (±)-phoracantholide I seco acid is detailed.

Selective alkylation/oxidation of N-substituted isoindolinone derivatives: synthesis of N-phthaloylated natural and unnatural α-amino acid analogues.

The interchangeability of the isoindolinone group as a nitrogen protecting group for amino acid intermediates is demonstrated by the preparation of several natural and unnatural α-amino acid derivatives using a two-carbon N-isoindolinone (phthalimidine) scaffold. Using a selective benzylic oxidation, the N-isoindolinone group is then converted to the N-phthaloyl group for convenient removal (65-98%). For preparation of the isoindolinone products which were to be the substrates for benzylic oxidation, a range of side chains were installed on the isoindolinone-protected glycine equivalent on deprotonation to demonstrate the utility of the N-protected isoindolinone synthon (51-93%). While the ensuing benzylic oxidation is employed successfully for converting the N-isoindolinone group to the N-phthaloyl group in simple substrates, substrates bearing unsaturated or electron-rich side chains respond poorly to the oxidation.

Synthesis of Extended Oxazoles III: Reactions of 2-(Phenylsulfonyl)methyl-4,5-diaryloxazoles.

2-((Phenylsulfonyl)methyl)-4,5-diphenyloxazole is a useful scaffold for synthetic elaboration at the 2-methylene position thereby affording extended oxazoles. The corresponding α-sulfonyl anion reacts smoothly with diverse alkyl halides giving monoalkylated (47-90%), dialkylated (50-97%), and cyclic (59-93%) products. The reductive desulfonylation of the monoalkylated and selected dialkylated products was optimized with a magnesium/mercuric chloride reagent system and afforded desulfonylated products in the range of 66-97%. The anti-inflammatory Oxaprozin was prepared using the α-sulfonyl carbanion strategy along with optimized desulfonylation.

1,2,3-Triazole-based inhibitors of Porphyromonas gingivalis adherence to oral streptococci and biofilm formation.

The development and use of small-molecule inhibitors of the adherence of Porphyromonas gingivalis to oral streptococci represents a potential therapy for the treatment of periodontal disease as these organisms work in tandem to colonize the oral cavity. Earlier work from these laboratories demonstrated that a small synthetic peptide was an effective inhibitor of the interaction between P. gingivalis and Streptococcus gordonii and that a small-molecule peptidomimetic would provide a more stable, less expensive and more effective inhibitor. An array of 2-(azidomethyl)- and 2-(azidophenyl)-4,5-diaryloxazoles having a full range of hydrophobic groups were prepared and reacted with substituted arylacetylenes to afford the corresponding 'click' products. The title compounds were evaluated for their ability to inhibit P. gingivalis' adherence to oral streptococci and several were found to be inhibitory in the range of (IC50) 5.3-67µM.

Synthesis of extended oxazoles II: Reaction manifold of 2-(halomethyl)-4,5-diaryloxazoles.

2-(Halomethyl)-4,5-diphenyloxazoles are effective, reactive scaffolds which can be utilized for synthetic elaboration at the 2-position. Through substitution reactions, the chloromethyl analogue is used to prepare a number of 2-alkylamino-, 2-alkylthio- and 2-alkoxy-(methyl) oxazoles. The 2-bromomethyl analogue offers a more reactive alternative to the chloromethyl compounds and is useful in the C-alkylation of a stabilized (malonate) carbanion as exemplified by a concise synthesis of Oxaprozin.

Oxazoles for click chemistry II: synthesis of extended heterocyclic scaffolds.

New routes to 2, 4, 5-trisubstituted oxazoles were established whereby the substitution pattern was established by the structure of the starting nonsymmetrical acyloins. 2-Chloromethyl-4, 5-disubstituted oxazoles were prepared by refinements of an earlier described process whereby chloroacetyl esters of symmetrical and non-symmetrical acyloins were cyclized using an ammonium acetate/acetic acid protocol. After substitution is effected, the azide moiety is then installed by substitution under mild conditions. While dibrominated and iodinated phenyloxazoles are required for further synthetic elaboration, the cyclization reaction was found to be very sensitive to the relative positions of the halogens in the starting materials.

Preparation of azidoaryl- and azidoalkyloxazoles for click chemistry.

A series of azidoaryl- and azidoalkyl(diphenyl)oxazole scaffolds were warranted for biofilm inhibition studies. Cyclization of azidoaryl- or azidoalkyl esters of benzoin with ammonium acetate in acetic acid gives 2-azidoaryl- or 2-azidoalkyl-4,5-diphenyloxazoles. The azidoaryl esters are prepared from the corresponding azidocarboxylic acids/acid chlorides while the azidoalkyl esters are prepared from the corresponding haloalkyl esters.

Synthetically useful Brønsted acid-promoted arylbenzyl ether --> o-benzylphenol rearrangements.

Camphorsulfonic acid in warm fluorobenzene facilitates the ortho rearrangement of (alkoxy-substituted) benzyl ethers of 1-(O-methyl)-2-nitroresorcinols to the corresponding o-(alkoxy-substituted) arylmethylnitrophenols. The substrate phenolic ethers are prepared by ultrasound-promoted arylmethylation of the appropriate 1-alkoxy-substituted 2-nitroresorcinol.

'Second-generation' 1,2,3-triazole-based inhibitors of Porphyromonas gingivalis adherence to oral streptococci and biofilm formation.

Several 'second-generation' click inhibitors of the multi-species biofilm propagated by the adherence of the oral pathogen Porphyromonas gingivalis to Streptococcus gordonii were synthesized and evaluated. The design of the structures was based on the results obtained with the first-generation diphenyloxazole 'click' inhibitors which bear suitable hydrophobic and polar groups within a dual scaffold molecule bearing a 1,2,3-triazole spacer. The structures of the synthetic targets reported herein now consist of a triazolyl(phenylsulfonylmethyl) and a triazolyl(phenylsulfinylmethyl) spacer which joins a 4,5-diphenyloxazole with both phenyl rings bearing lipophilic substituents. The triazolyl "linker" group is formed by a click reaction between the 4-azido(phenylsulfonyl/sulfinylmethyl) oxazoles and acetylenic components having aryl groups bearing hydrophobic substituents. The 1,3,5-trisubstituted-2,4,6-triazine scaffold of the most active click compounds were modeled after the structural motif termed the VXXLL nuclear receptor (NR) box. When substituted at the 3- and 5-positions with 2- and 4-fluorophenylamino and N,N-diethylamino units, the candidates bearing the 1,3,5-trisubstituted-2,4,6-triazine scaffold formed a substantial subset of the second-generation click candidates. Four of the click products, compounds 95, 111, 115 and 122 showed inhibition of the adherence of P. gingivalis to S. gordonii with an IC50 range of 2.3-4.3 µM and only 111 exhibited cytotoxic activity against telomerase immortalized gingival keratinocytes at 60 µM. These results suggest that compounds 95, 115, 122, and possibly 111 represent the most suitable compounds to evaluate for activity in vivo.

Efficient preparation and processing of the 4-methoxybenzyl (PMB) group for phenolic protection using ultrasound.

Power ultrasound efficiently facilitates the rapid preparation and reaction of 4-methoxybenzyl chloride (PMB-Cl) 1 in providing protected phenolic ether intermediates for organic synthesis. Using two-phase systems in both the ultrasound-promoted preparation and reactions of PMB-Cl, typical runs produce PMB-protected products within 15 min. When compared with nonsonicated control reactions, the results demonstrate clear advantage in terms of efficiency when the protocol is applied to the mild and selective protection of various multisubstituted phenols including sensitive phenolic aldehydes.

Unnatural Amino Acid Derivatives through Click Chemistry: Synthesis of Triazolylalanine Analogues.

A novel tert-butyl 2-(1-oxoisolndolin-2-yl)acetate derivative is selectively alkylated with propargyl bromide in the presence of lithium hexamethyldisilazide. After removal of the tert-butyl protecting group, the resulting N-isoindolinyl (ethynylalanine) derivative is reacted with a series of azides under 'click conditions'. The click reactions afford an array of N-isoindolinyl-1,2,3-triazolylalanine derivatives as the free carboxylic adds. Following esterification, the N-isoindolinone protecting group is then transformed into the more easily removable phthaloyl group by selective oxidation at the benzylic position.

Antiangiogenic activity of N-substituted and tetrafluorinated thalidomide analogues.

Inhibition of angiogenesis is currently perceived as one of the promising strategies in the treatment of cancer. The antiangiogenic property of thalidomide has inspired a second wave of research on this teratogenic drug. Previous studies from our group and others demonstrated that metabolites of thalidomide are responsible for the drug's pharmacological actions. On the basis of the structures of these metabolites, we synthesized 118 thalidomide analogues. Preliminary screening selected 7 of these 118 analogues for more extensive testing in the current study. In the rat aortic ring assay, all 4 analogues in the N-substituted class and 2 of the 3 analogues in the tetrafluorinated class significantly inhibited microvessel outgrowth at 12.5-200 microM. Thalidomide failed to block angiogenesis at similar concentrations. Subsequently, the effects of these analogues on human umbilical vein endothelial cell proliferation and tube formation were determined. Those analogues showing antiangiogenicity in the rat aortic ring assay also demonstrated antiproliferative action in human umbilical vein endothelial cells. Cell proliferation was not affected by thalidomide. Interestingly, all 7 analogues as well as thalidomide suppressed tube formation. Two tetrafluorinated analogues consistently showed the highest potency and efficacy in all three assays. The in vivo toxicity of representative analogues from each class was also evaluated. Taken together, our results support the further development and evaluation of novel thalidomide analogues as antiangiogenic agents.

Thalidomide metabolites and analogues. 3. Synthesis and antiangiogenic activity of the teratogenic and TNFalpha-modulatory thalidomide analogue 2-(2,6-dioxopiperidine-3-yl)phthalimidine.

Versatile synthesis of the teratogenic, TNFalpha-modulatory, and antiangiogenic thalidomide analogue 2-(2,6-dioxopiperidine-3-yl)phthalimidine (1) and its direct antiangiogenic properties are described. With thalidomide or thalidomide derivatives as precursors, the synthesis involved either carbonyl reduction/thiation-desulfurization or carbonyl reduction/acyliminium ion reduction protocols. Compared to earlier studies with thalidomide, which was only active with microsomal treatment, 1 exhibited marginal inhibitory activity in the rat aortic ring assay, thereby demonstrating the requirement for metabolic activation.