Synthesis and Functionalization of Azetidine-Containing Small Macrocyclic Peptides.

Cyclic peptides are increasingly important structures in drugs but their development can be impeded by difficulties associated with their synthesis. Here, we introduce the 3-aminoazetidine (3-AAz) subunit as a new turn-inducing element for the efficient synthesis of small head-to-tail cyclic peptides. Greatly improved cyclizations of tetra-, penta- and hexapeptides (28 examples) under standard reaction conditions are achieved by introduction of this element within the linear peptide precursor. Post-cyclization deprotection of the amino acid side chains with strong acid is realized without degradation of the strained four-membered azetidine. A special feature of this chemistry is that further late-stage modification of the resultant macrocyclic peptides can be achieved via the 3-AAz unit. This is done by: (i) chemoselective deprotection and substitution at the azetidine nitrogen, or by (ii) a click-based approach employing a 2-propynyl carbamate on the azetidine nitrogen. In this way, a range of dye and biotin tagged macrocycles are readily produced. Structural insights gained by XRD analysis of a cyclic tetrapeptide indicate that the azetidine ring encourages access to the less stable, all-trans conformation. Moreover, introduction of a 3-AAz into a representative cyclohexapeptide improves stability towards proteases compared to the homodetic macrocycle.

Stereoselective synthesis of (R)- and (S)-1,2-diazetidine-3-carboxylic acid derivatives for peptidomimetics.

The stereoselective synthesis of both enantiomers of N-protected 1,2-diazetidine-3-carboxylic acid (aAze) from homochiral glycidol is described. Orthogonal protection of this novel cyclic α-hydrazino acid allows for selective functionalisation at either Nγ or Nδ. This novel peptidomimetic building block was incorporated into the pseudotripeptides Gly-γaAze-Ala and Gly-δaAze-Ala.

Synthesis of Arylidene-ß-lactams via exo-Selective Matsuda-Heck Arylation of Methylene-ß-lactams.

exo-Methylene-ß-lactams were synthesized in two steps from commercially available 3-bromo-2-(bromomethyl)propionic acid and reacted with arene diazonium salts in a Heck-type arylation in the presence of catalytic amounts of Pd(OAc)2 under ligand-free conditions. The products, arylidene-ß-lactams, were obtained in high yields as single isomers. The ß-hydride elimination step of the Pd-catalyzed coupling reaction proceeds with high exo-regioselectivity and E-stereoselectivity. With aryl iodides, triflates, or bromides, the coupling products were isolated only in low yields, due to extensive decomposition of the starting material at elevated temperatures. This underlines that arene diazonium salts can be superior arylating reagents in Heck-type reactions and yield coupling products in synthetically useful yields and selectivities when conventional conditions fail.

Development of oxetane modified building blocks for peptide synthesis.

The synthesis and use of oxetane modified dipeptide building blocks in solution and solid-phase peptide synthesis (SPPS) is reported. The preparation of building blocks containing non-glycine residues at the N-terminus in a stereochemically controlled manner is challenging. Here, a practical 4-step route to such building blocks is demonstrated, through the synthesis of dipeptides containing contiguous alanine residues. The incorporation of these new derivatives at specific sites along the backbone of an alanine-rich peptide sequence containing eighteen amino acids is demonstrated via solid-phase peptide synthesis. Additionally, new methods to enable the incorporation of all 20 of the proteinogenic amino acids into such dipeptide building blocks are reported through modifications of the synthetic route (for Cys and Met) and by changes to the protecting group strategy (for His, Ser and Thr).

Impact of oxetane incorporation on the structure and stability of alpha-helical peptides.

Peptide-based drugs combine advantages of larger biological therapeutics with those of small molecule drugs, but they generally display poor permeability and metabolic stability. Recently, we introduced a new type of peptide bond isostere, in which the backbone carbonyl is replaced with a 3-amino oxetane heterocycle, into short linear peptides with the aim of improving their therapeutic potential. In this study, we have explored the impact of oxetane modification on α-helical peptides to establish whether or not this modification is tolerated in this biologically important structural motif. The oxetane modification was introduced at two positions in a well-characterised helical peptide sequence, and circular dichroism and NMR spectroscopy were used to measure the resulting secondary structure content under different experimental conditions. Our data demonstrated that introduction of an oxetane into the peptide backbone results in a significant loss of helicity, regardless of where in the sequence the modification is placed. The molecular determinants of this destabilisation were then explored using steered molecular dynamics simulations, a computational method analogous to single molecule spectroscopy. Our simulations indicated that oxetane modification introduces a kink in the helical axis, alters the dihedral angles of residues up to three positions away from the modification, and disrupts the (i, i + 4) hydrogen bonding pattern characteristic of α-helices in favour of new, short-range hydrogen bonds. The detailed structural understanding provided in this work can direct future design of chemically modified peptides.

Iterative Arylation of Itaconimides with Diazonium Salts through Electrophilic Palladium Catalysis: Divergent ß-H-Elimination Pathways in Repetitive Matsuda-Heck Reactions.

N-Arylitaconimides, accessible from maleic anhydride, anilines, and paraformaldehyde, react with arene diazonium salts in Pd-catalyzed Matsuda-Heck arylation to the pharmacologically relevant E-configured 3-arylmethylidene pyrrolidine-2,5-diones (also known as arylmethylidene succinimides) through exo-selective ß-H-elimination. The coupling proceeds at ambient temperature with the simple and easy-to-handle precatalyst Pd-II-acetate under ligand- and base-free conditions. Notable features are high isolated yields, regio- and stereoselectivities, and short reaction times. In a comparative investigation, aryl iodides, bromides, and triflates were shown to be inferior coupling reagents in this reaction. The 3-arylmethylidene pyrrolidine-2,5-diones undergo second Matsuda-Heck coupling, which proceeds via endo-selective ß-H-elimination to give diarylmethyl-substituted maleimides as coupling products. These products can also be accessed in one flask by sequential addition of different arene diazonium salts to the starting itaconimide. The potential of 3-arylmethylidene succinimides as photoswitches was tested. Upon irradiation of the E-isomer at 300 nm, partial isomerization to the Z-isomer (E/Z = 65:35 in the photostationary state) was observed. The isomerization was found to be nearly completely reversible by irradiating the mixture at 400 nm.

Synthesis of 4,5-Diazaspiro[2.3]hexanes and 1,2-Diazaspiro[3.3]heptanes as Hexahydropyridazine Analogues.

4,5-Diazaspiro[2.3]hexanes are made by dihalocarbene addition across the exocyclic double bond of readily accessible 3-alkylidene-1,2-diazetidines. Using difluorocarbene, generated from TMSCF3/NaI, these spirocycles were produced in yields up to 97% by stereospecific addition across the alkene. Lower yields (up to 64%) were observed using more reactive dichlorocarbene, due to competitive insertion of the carbene into the N-N bond. Larger 1,2-diazaspiro[3.3]heptanes are produced by [2 + 2] cycloaddition of 3-alkylidene-1,2-diazetidines with tetracyanoethylene (TCNE) in up to 99% yield.

Asymmetric Synthesis of 2-Substituted Azetidin-3-ones via Metalated SAMP/RAMP Hydrazones.

2-Substituted azetidin-3-ones can be prepared in good yields and enantioselectivities (up to 85% ee) by a one-pot procedure involving the metalation of the SAMP/RAMP hydrazones of N-Boc-azetidin-3-one, reaction with a wide range of electrophiles, including alkyl, allyl, and benzyl halides and carbonyl compounds, followed by hydrolysis using oxalic acid.

Nitrogen Stereodynamics and Complexation Phenomena as Key Factors in the Deprotonative Dynamic Resolution of Alkylideneaziridines: A Spectroscopic and Computational Study.

The present work is aimed at shedding light on the origin of the stereoselectivity observed in the reactions of chiral heterosubstituted organolithiums, generated by lithiation of alkylideneaziridines. Factors such as the nitrogen inversion barrier, the stereochemistry at the nitrogen atom, the substitution pattern of the alkylideneaziridines, and the reaction conditions are taken into consideration. The interplay between nitrogen stereodynamics and complexation phenomena seems to be crucial in determining the stereochemical outcome of the lithiation/trapping sequence. The findings were rationalized by a synergistic use of NMR experiments, run on the lithiated intermediates, alongside computational data. It has been demonstrated that, in such systems, the stereochemistry-determining step is the deprotonation reaction, and a model based on a deprotonative dynamic resolution has been proposed. Such findings could find application in dynamic systems other than aziridines.

Ring closing metathesis reactions of α-methylene-ß-lactams: application to the synthesis of a simplified phyllostictine analogue with herbicidal activity.

Ring closing metathesis (RCM) reactions of α-methylene-ß-lactams are used to construct strained 11- and 12-membered macrocycles that mimic key structural elements of phyllostictine A. The highest yield and stereoselectivity was achieved making 12-membered macrocycle Z-19 with use of a p-methoxyphenyl group on the lactam nitrogen. Interestingly, substrate concentration had an important influence on the stereochemical course of the reaction. A simplified analogue produced using this approach displays phytotoxic activity against Chlamydomonas reinhardtii suggesting that the α-methylene-ß-lactam subunit is responsible, at least in part, for the herbicidal activity of phyllostictine A.

Stereocontrolled lithiation/trapping of chiral 2-alkylideneaziridines: investigation into the role of the aziridine nitrogen stereodynamics.

The origin of the stereoselectivity in the lithiation/trapping of 2-alkylideneaziridines bearing a chiral group as the nitrogen substituent was investigated. Optimal reaction conditions were discovered by in situ FT-IR monitoring. In addition, it has been found that the solvent and the alkene substitution pattern are important factors able to impart a switch in stereoselectivity. While lithiation of the alkylideneaziridine ring flanked by either a fully substituted or a Z-configured alkene pendant occurs stereoselectively in THF, in contrast unsubstituted 2-methyleneaziridine undergoes lithiation in toluene with the opposite sense of stereoinduction. Lithiation experiments, on deuterium labelled 2-alkylideneaziridines, confirmed the configurational stability of the lithiated intermediates. A model based on complexation and proximity effects was proposed to rationalize the reactivity. This model assumes that slowly equilibrating N-invertomers undergo deprotonation (lithiation) at different rates and that the stereochemical outcome is established during the deprotonation step.

Palladium-catalyzed multicomponent synthesis of 2-aryl-2-imidazolines from aryl halides and diamines.

An efficient palladium-catalyzed three-component reaction that combines aryl halides, isocyanides, and diamines provides access to 2-aryl-2-imidazolines in yields up to 96%. Through variation of the diamine component, the reaction can be extended to the synthesis of 2-aryl-1H-benzimidazoles and 2-aryl-1,4,5,6-tetrahydropyrimidines.

Asymmetric synthesis of 2-substituted oxetan-3-ones via metalated SAMP/RAMP hydrazones.

2-Substituted oxetan-3-ones can be prepared in good yields and enantioselectivities (up to 84% ee) by the metalation of the SAMP/RAMP hydrazones of oxetan-3-one, followed by reaction with a range of electrophiles that include alkyl, allyl, and benzyl halides. Additionally, both chiral 2,2- and 2,4-disubstituted oxetan-3-ones can be made in high ee (86-90%) by repetition of this lithiation/alkylation sequence under appropriately controlled conditions. Hydrolysis of the resultant hydrazones with aqueous oxalic acid provides the 2-substituted oxetan-3-ones without detectable racemization.

Lewis acid promoted intramolecular (3 + 2) 'cycloadditions' of methyleneaziridines with alkene and alkyne acceptors.

2-Methyleneaziridines can be tethered to a variety of alkene or alkyne acceptors through the saturated carbon of the heterocyclic ring by application of a simple lithiation/alkylation sequence (8 examples, 31-59%). Treatment of the resultant alkene bearing substrates with BF(3)·OEt(2) leads to cis-octahydrocyclopenta[c]pyrroles in which up to four contiguous stereocentres are created in a diastereocontrolled manner after reductive work-up. Using an alkyne based substrate, a 2,4,5,6-tetrahydrocyclopenta[c]pyrrole is produced by rapid tautomerisation of the initially formed bisenamine. Evidence that these (3 + 2) 'cycloadditions' proceed in a stepwise manner via a 2-aminoallyl cation is presented.

Selective aldehyde reductions in neutral water catalysed by encapsulation in a supramolecular cage.

The enhancement of reactivity inside supramolecular coordination cages has many analogies to the mode of action of enzymes, and continues to inspire the design of new catalysts for a range of reactions. However, despite being a near-ubiquitous class of reactions in organic chemistry, enhancement of the reduction of carbonyls to their corresponding alcohols remains very much underexplored in supramolecular coordination cages. Herein, we show that encapsulation of small aromatic aldehydes inside a supramolecular coordination cage allows the reduction of these aldehydes with the mild reducing agent sodium cyanoborohydride to proceed with high selectivity (ketones and esters are not reduced) and in good yields. In the absence of the cage, low pH conditions are essential for any appreciable conversion of the aldehydes to the alcohols. In contrast, the specific microenvironment inside the cage allows this reaction to proceed in bulk solution that is pH-neutral, or even basic. We propose that the cage acts to stabilise the protonated oxocarbenium ion reaction intermediates (enhancing aldehyde reactivity) whilst simultaneously favouring the encapsulation and reduction of smaller aldehydes (which fit more easily inside the cage). Such dual action (enhancement of reactivity and size-selectivity) is reminiscent of the mode of operation of natural enzymes and highlights the tremendous promise of cage architectures as selective catalysts.

Readily accessible sp3-rich cyclic hydrazine frameworks exploiting nitrogen fluxionality.

Increased molecular complexity correlates with improved chances of success in the drug development process. Here, a strategy for the creation of sp3-rich, non-planar heterocyclic scaffolds suitable for drug discovery is described that obviates the need to generate multiple stereogenic centers with independent control. Asymmetric transfer hydrogenation using a tethered Ru-catalyst is used to efficiently produce a range of enantiopure cyclic hydrazine building blocks (up to 99% ee). Iterative C-N functionalization at the two nitrogen atoms of these compounds produces novel hydrazine and hydrazide based chemical libraries. Wide chemical diversification is possible through variation in the hydrazine structure, use of different functionalization chemistries and coupling partners, and controlled engagement of each nitrogen of the hydrazine in turn. Principal Moment of Inertia (PMI) analysis of this small hydrazine library reveals excellent shape diversity and three-dimensionality. NMR and crystallographic studies confirm these frameworks prefer to orient their substituents in three-dimensional space under the control of a single stereogenic center through exploitation of the fluxional behavior of the two nitrogen atoms.

Four-component reaction for the preparation of alpha-amino phosphonates from methyleneaziridines.

Alpha-amino phosphonates can be rapidly assembled in moderate to good yields (42-65%) via a "one-pot" process that brings together four components through the construction of three new intermolecular bonds.

Towards a better understanding of the electrosynthesis of 2,5-dicarboxy-2,5-dihydrofurans: structure, mechanism and influence over stereochemistry.

2,5-Dicarboxy-2,5-dihydrofurans are key constituents of a number of natural products and have roles as intermediates in the formation of other such compounds of interest. Typically, these species are synthesized using toxic Pb(IV) salts. Electrochemical syntheses of 2,5-diacetoxy-2,5-dihydrofuran that do not require the use of lead have been reported, but a general lack of experimental detail has prevented these procedures from being more widely adopted. Moreover, no electrochemical study has yet reported the ratio of cis and trans isomers produced. Herein, we compare the chemical, lead-based route to 2,5-diacetoxy-2,5-dihydrofuran with a fully described electrosynthesis method. In doing so, we have discovered that the cis and trans isomers of this compound were previously incorrectly assigned in the literature, an error that we correct by obtaining the crystal structure of cis-2,5-diacetoxy-2,5-dihydrofuran. This allows the ratios of the isomers as prepared by the chemical (2 : 1 cis : trans) and electrochemical (7 : 5 cis : trans) methods to be obtained. Through experimental and computational insights, we propose a mechanism for the electrochemical synthesis of 2,5-dicarboxy-2,5-dihydrofurans and go some way towards validating this mechanism by synthesizing 2,5-dibutoxy-2,5-dihydrofuran electrochemically for the first time. We hope that these findings will provide some greater clarity to the literature surrounding the electrosynthesis and potential applications of 2,5-dicarboxy-2,5-dihydrofurans.

Macrocyclisation of small peptides enabled by oxetane incorporation.

Cyclic peptides are an important source of new drugs but are challenging to produce synthetically. We show that head-to-tail peptide macrocyclisations are greatly improved, as measured by isolated yields, reaction rates and product distribution, by substitution of one of the backbone amide C[double bond, length as m-dash]O bonds with an oxetane ring. The cyclisation precursors are easily made by standard solution- or solid-phase peptide synthesis techniques. Macrocyclisations across a range of challenging ring sizes (tetra-, penta- and hexapeptides) are enabled by incorporation of this turn-inducing element. Oxetane incorporation is shown to be superior to other established amino acid modifications such as N-methylation. The positional dependence of the modification on cyclisation efficiency is mapped using a cyclic peptide of sequence LAGAY. We provide the first direct experimental evidence that oxetane modification induces a turn in linear peptide backbones, through the observation of d NN (i, i + 2) and d αN (i, i + 2) NOEs, which offers an explanation for these improvements. For cyclic peptide, cLAGAY, a combination of NMR derived distance restraints and molecular dynamics simulations are used to show that this modification alters the backbone conformation in proximity to the oxetane, with the flexibility of the ring reduced and a new intramolecular H-bond established. Finally, we incorporated an oxetane into a cyclic pentapeptide inhibitor of Aminopeptidase N, a transmembrane metalloprotease overexpressed on the surface of cancer cells. The inhibitor, cCNGRC, displayed similar IC50 values in the presence or absence of an oxetane at the glycine residue, indicating that bioactivity is fully retained upon amide C[double bond, length as m-dash]O bond replacement.

Rapid synthesis of 1,3,4,4-tetrasubstituted beta-lactams from methyleneaziridines using a four-component reaction.

2-Methyleneaziridines can be transformed into a variety of 1,3,4,4-tetrasubstituted beta-lactams in moderate to good yields (46-63%) via a "one-pot" process that brings together four components with the formation of three new intermolecular carbon-carbon bonds.