Aziridine synthesis by coupling amines and alkenes via an electrogenerated dication.

Aziridines-three-membered nitrogen-containing cyclic molecules-are important synthetic targets. Their substantial ring strain and resultant proclivity towards ring-opening reactions makes them versatile precursors of diverse amine products1-3, and, in some cases, the aziridine functional group itself imbues important biological (for example, anti-tumour) activity4-6. Transformation of ubiquitous alkenes into aziridines is an attractive synthetic strategy, but is typically accomplished using electrophilic nitrogen sources rather than widely available amine nucleophiles. Here we show that unactivated alkenes can be electrochemically transformed into a metastable, dicationic intermediate that undergoes aziridination with primary amines under basic conditions. This new approach expands the scope of readily accessible N-alkyl aziridine products relative to those obtained through existing state-of-the-art methods. A key strategic advantage of this approach is that oxidative alkene activation is decoupled from the aziridination step, enabling a wide range of commercially available but oxidatively sensitive7 amines to act as coupling partners for this strain-inducing transformation. More broadly, our work lays the foundations for a diverse array of difunctionalization reactions using this dication pool approach.

Aza-Prilezhaev Aziridination-Enabled Multidimensional Analysis of Isomeric Lipids via High-Resolution U-Shaped Mobility Analyzer-Mass Spectrometry.

Unsaturated lipids constitute a significant portion of the lipidome, serving as players of multifaceted functions involving cellular signaling, membrane structure, and bioenergetics. While derivatization-assisted liquid chromatography tandem mass spectrometry (LC-MS/MS) remains the gold standard technique in lipidome, it mainly faces challenges in efficiently labeling the carbon-carbon double bond (C═C) and differentiating isomeric lipids in full dimension. This presents a need for new orthogonal methodologies. Herein, a metal- and additive-free aza-Prilezhaev aziridination (APA)-enabled ion mobility mass spectrometric method is developed for probing multiple levels of unsaturated lipid isomerization with high sensitivity. Both unsaturated polar and nonpolar lipids can be efficiently labeled in the form of N-H aziridine without significant side reactions. The signal intensity can be increased by up to 3 orders of magnitude, achieving the nM detection limit. Abundant site-specific fragmentation ions indicate C═C location and sn-position in MS/MS spectra. Better yet, a stable monoaziridination product is dominant, simplifying the spectrum for lipids with multiple double bonds. Coupled with a U-shaped mobility analyzer, identification of geometric isomers and separation of different lipid classes can be achieved. Additionally, a unique pseudo MS3 mode with UMA-QTOF MS boosts the sensitivity for generating diagnostic fragments. Overall, the current method provides a comprehensive solution for deep-profiling lipidomics, which is valuable for lipid marker discovery in disease monitoring and diagnosis.

Recent Developments in the Biosynthesis of Aziridines.

Only 0.016 % of all known natural products contain an aziridine ring, but this unique structural feature imparts high reactivity and cytotoxicity to the compounds in which it is found. Until 2021, no naturally occurring aziridine-forming enzymes had been identified. Since 2021, the biosynthetic enzymes for ~10 % of known aziridine containing natural products have been identified and characterized. This article describes the recent advances in our understanding of enzyme-catalyzed aziridine formation in the context of historical methods for aziridine formation through synthetic chemistry.

Total Synthesis and Biological Profiling of Putative (±)-Marinoaziridine B and (±)-N-Methyl Marinoaziridine A.

The total synthesis of two new marine natural products, (±)-marinoaziridine B 7 and (±)-N-methyl marinoaziridine A 8, was accomplished. The (±)-marinoaziridine 7 was prepared in a six-step linear sequence with a 2% overall yield. The key steps in our strategy were the preparation of the chiral epoxide (±)-5 using the Johnson Corey Chaykovsky reaction, followed by the ring-opening reaction and the Staudinger reaction. The N,N-dimethylation of compound (±)-7 gives (±)-N-methyl marinoaziridine A 8. The NMR spectra of synthetized (±)-marinoaziridine B 7 and isolated natural product did not match. The compounds are biologically characterized using relevant in silico, in vitro and in vivo methods. In silico ADMET and bioactivity profiling predicted toxic and neuromodulatory effects. In vitro screening by MTT assay on three cell lines (MCF-7, H-460, HEK293T) showed that both compounds exhibited moderate to strong antiproliferative and cytotoxic effects. Antimicrobial tests on bacterial cultures of Escherichia coli and Staphylococcus aureus demonstrated the dose-dependent inhibition of the growth of both bacteria. In vivo toxicological tests were performed on zebrafish Danio rerio and showed a significant reduction of zebrafish mortality due to N-methylation in (±)-8.

Characterization of AST-001 non-clinical pharmacokinetics: A novel selective AKR1C3-activated prodrug in mice, rats, and cynomolgus monkeys.

AST-001 is a chemically synthesized inactive nitrogen mustard prodrug that is selectively cleaved to a cytotoxic aziridine (AST-2660) via aldo-keto reductase family 1 member C3 (AKR1C3). The purpose of this study was to investigate the pharmacokinetics and tissue distribution of the prodrug, AST-001, and its active metabolite, AST-2660, in mice, rats, and monkeys. After single and once daily intravenous bolus doses of 1.5, 4.5, and 13.5 mg/kg AST-001 to Sprague-Dawley rats and once daily 1 h intravenous infusions of 0.5, 1.5, and 4.5 mg/kg AST-001 to cynomolgus monkeys, AST-001 exhibited dose-dependent pharmacokinetics and reached peak plasma levels at the end of the infusion. No significant accumulation and gender differences were observed after 7 days of repeated dosing. In rats, the half-life of AST-001 was dose independent and ranged from 4.89 to 5.75 h. In cynomolgus monkeys, the half-life of AST-001 was from 1.66 to 5.56 h and increased with dose. In tissue distribution studies conducted in Sprague-Dawley rats and in liver cancer PDX models in female athymic nude mice implanted with LI6643 or LI6280 HepG2-GFP tumor fragments, AST-001 was extensively distributed to selected tissues. Following a single intravenous dose, AST-001 was not excreted primarily as the prodrug, AST-001 or the metabolite AST-2660 in the urine, feces, and bile. A comprehensive analysis of the preclinical data and inter-species allometric scaling were used to estimate the pharmacokinetic parameters of AST-001 in humans and led to the recommendation of a starting dose of 5 mg/m2 in the first-in-human dose escalation study.

Structural Evaluation of a Nitroreductase Engineered for Improved Activation of the 5-Nitroimidazole PET Probe SN33623.

Bacterial nitroreductase enzymes capable of activating imaging probes and prodrugs are valuable tools for gene-directed enzyme prodrug therapies and targeted cell ablation models. We recently engineered a nitroreductase (E. coli NfsB F70A/F108Y) for the substantially enhanced reduction of the 5-nitroimidazole PET-capable probe, SN33623, which permits the theranostic imaging of vectors labeled with oxygen-insensitive bacterial nitroreductases. This mutant enzyme also shows improved activation of the DNA-alkylation prodrugs CB1954 and metronidazole. To elucidate the mechanism behind these enhancements, we resolved the crystal structure of the mutant enzyme to 1.98 Å and compared it to the wild-type enzyme. Structural analysis revealed an expanded substrate access channel and new hydrogen bonding interactions. Additionally, computational modeling of SN33623, CB1954, and metronidazole binding in the active sites of both the mutant and wild-type enzymes revealed key differences in substrate orientations and interactions, with improvements in activity being mirrored by reduced distances between the N5-H of isoalloxazine and the substrate nitro group oxygen in the mutant models. These findings deepen our understanding of nitroreductase substrate specificity and catalytic mechanisms and have potential implications for developing more effective theranostic imaging strategies in cancer treatment.

A Study on the Biological Activity of Optically Pure Aziridine Phosphines and Phosphine Oxides.

A series of optically pure aziridine phosphines and their corresponding phosphine oxides were synthesized through established chemical methodologies. The compounds were systematically investigated for their biological properties. Notably, all synthesized compounds demonstrated moderate antibacterial activity only against the reference strain of Staphylococcus aureus. However, compounds 5 and 7 exhibited noteworthy cell viability inhibition of human cervical epithelioid carcinoma HeLa cells and endometrial adenocarcinoma Ishikawa cells. Further studies of these compounds revealed additional biological effects, including disruption of the cell membrane in high concentrations, cell cycle arrest in the S phase, and the induction of reactive oxygen species (ROS). Comparative analysis of the two classes of chiral organophosphorus derivatives of aziridines indicated that chiral phosphine oxides displayed significantly higher biological activity. Consequently, these findings suggest that chiral phosphine oxides may be potential candidates for the development of anticancer drugs. In light of the significant interest in preparations whose structure is based on a three-membered aziridine ring in terms of potential anticancer therapy, this research fits into the current research trend and should constitute a valuable addition to the current state of knowledge and the existing library of aziridine derivatives with anticancer properties.

Lactic Acid Bacteria-Derived Postbiotics as Adjunctive Agents in Breast Cancer Treatment to Boost the Antineoplastic Effect of a Conventional Therapeutic Comprising Tamoxifen and a New Drug Candidate: An Aziridine-Hydrazide Hydrazone Derivative.

Breast cancer is associated with high mortality and morbidity rates. As about 20-30% of patients exhibiting ER-positive phenotype are resistant to hormonal treatment with the standard drug tamoxifen, finding new therapies is a necessity. Postbiotics, metabolites, and macromolecules isolated from probiotic bacteria cultures have been proven to have sufficient bioactivity to exert prohealth and anticancer effects, making them viable adjunctive agents for the treatment of various neoplasms, including breast cancer. In the current study, postbiotics derived from L. plantarum and L. rhamnosus cultures were assessed on an in vitro breast cancer model as potential adjunctive agents to therapy utilizing tamoxifen and a candidate aziridine-hydrazide hydrazone derivative drug. Cell viability and cell death processes, including apoptosis, were analyzed for neoplastic MCF-7 cells treated with postbiotics and synthetic compounds. Cell cycle progression and proliferation were analyzed by PI-based flow cytometry and Ki-67 immunostaining. Postbiotics decreased viability and triggered apoptosis in MCF-7, modestly affecting the cell cycle and showing a lack of negative impact on normal cell viability. Moreover, they enhanced the cytotoxic effect of tamoxifen and the new candidate drug toward MCF-7, accelerating apoptosis and the inhibition of proliferation. This illustrates postbiotics' potential as natural adjunctive agents supporting anticancer therapy based on synthetic drugs.

Mechanistic Studies of Aziridine Formation Catalyzed by Mononuclear Non-Heme Iron Enzymes.

Aziridines are compounds with a nitrogen-containing three-membered ring. When it is incorporated into natural products, the reactivity of the strained ring often drives the biological activities of aziridines. Despite its importance, the enzymes and biosynthetic strategies deployed to install this reactive moiety remain understudied. Herein, we report the use of in silico methods to identify enzymes with potential aziridine-installing (aziridinase) functionality. To validate candidates, we reconstitute enzymatic activity in vitro and demonstrate that an iron(IV)-oxo species initiates aziridine ring closure by the C-H bond cleavage. Furthermore, we divert the reaction pathway from aziridination to hydroxylation using mechanistic probes. This observation, isotope tracing experiments using H218O and 18O2, and quantitative product analysis, provide evidence for the polar capture of a carbocation species by the amine in the pathway to aziridine installation.

Protonated Porphyrins: Bifunctional Catalysts for the Metal-Free Synthesis of N-Alkyl-Oxazolidinones.

The protonation of commercially available porphyrin ligands yields a class of bifunctional catalysts able to promote the synthesis of N-alkyl oxazolidinones by CO2 cycloaddition to corresponding aziridines. The catalytic system does not require the presence of any Lewis base or additive, and shows interesting features both in terms of cost effectiveness and eco-compatibility. The metal-free methodology is active even with a low catalytic loading of 1 % mol, and the chemical stability of the protonated porphyrin allowed it to be recycled three times without any decrease in performance. In addition, a DFT study was performed in order to suggest how a simple protonated porphyrin can mediate CO2 cycloaddition to aziridines to yield oxazolidinones.

Ring Opening of Aziridines by Pendant Silanols Allows for Stereospecific Preparations of 1'-Amino-tetrahydrofurans.

We have developed a highly stereospecific cyclization of aziridine silanols into 1'-amino-tetrahydrofurans. Our protocol of stirring a substrate with 10 mol % Sc (OTf)3 and 1 equivalent of NaHCO3 in CH2Cl2 is mild and compatible with a range of activating aziridine N-substituents (including tosylates, mesylates, and carbamates) and functional groups on the alkyl chains (including substituted aryl rings, alkyl bromides, and alkyl ethers). In all cases examined, trans di-substituted aziridine silanols give products with an erythro configuration; conversely, cis di-substituted aziridine silanols give products with a threo configuration. While literature syntheses of 1'-amino-tetrahydrofurans exist, only one example, contemporaneous with our work, uses a similar cyclization for their construction. Control experiments demonstrate that, for this transformation, the silanol is not particularly privileged, and a variety of protecting groups on the alcohol (including other silicon protecting groups, benzyl ethers, and MOM ethers) are compatible with product formation.

Recent advances in the accessibility, synthetic utility, and biological applications of aziridines.

Compounds featuring aziridine moieties are widely known and extensively reported in the literature. Due to their great potential from both synthetic and pharmacological points of view, many researchers have focused their efforts on the development of new methodologies for the preparation and transformation of these interesting compounds. Over the years, more and more ways to obtain molecules bearing these three-membered functional groups, which are challenging due to their inherent reactivity, have been described. Among them, several are more sustainable. In this review, we report the recent advances in the biological and chemical evolution of aziridine derivatives, in particular, the variety of methodologies described for the synthesis of aziridines and their chemical transformations leading to the formation of interesting derivatives, such as 4-7 membered heterocycles of pharmaceutical interest due to their promising biological activities.

Catalytic enantioselective desymmetrization of meso-aziridines.

As a type of readily available small strained-ring heterocycle, meso-aziridines may undergo catalytic desymmetrizing transformations to empower the rapid construction of diverse nitrogen-containing structures bearing contiguous stereocenters, which have great relevance in natural product synthesis, drug development and the design and synthesis of chiral catalysts/ligands for asymmetric catalysis. This review outlines the advances achieved in the catalytic asymmetric desymmetrization of meso aziridines and highlights some promising avenues for further work in this realm.

Aromatic sulphonamides of aziridine-2-carboxylic acid derivatives as novel PDIA1 and PDIA3 inhibitors.

In this study, we report a series of newly synthesised sulphonamides of aziridine-2-carboxylic acid (Az-COOH) ester and amide analogues as potent protein disulphide isomerase (PDI, EC 5.3.4.1) inhibitors. The inhibitory activity on PDI was determined against recombinant human PDIA1 and PDIA3 proteins using an insulin reduction assay. These compounds in low micromolar to low nanomolar concentrations showed the effective in vitro inhibitory properties of PDIA1 with weaker effects on PDIA3. Complexes of 15N- and 15N,13C- uniformly labelled recombinant human PDIA1a with two PDIA1 inhibitors were produced and investigated by a protein nuclear magnetic resonance (NMR) spectroscopy. It was found that both C53 and C56 of the PDIA1 enzyme were involved in covalent binding. Finally, in a range of pharmacological studies, we demonstrated that investigated compounds displayed anti-cancer and anti-thrombotic activity. These findings demonstrate that sulphonamides of Az-COOH derivatives are promising candidates for the development of novel anti-cancer and anti-thrombotic agents.

Systematic identification of engineered methionines and oxaziridines for efficient, stable, and site-specific antibody bioconjugation.

The field of chemical modification of proteins has been dominated by random modification of lysines or more site-specific labeling of cysteines, each with attendant challenges. Recently, we have developed oxaziridine chemistry for highly selective modification of methionine called redox-activated chemical tagging (ReACT) but have not broadly tested the molecular parameters for efficient and stable protein modification. Here we systematically scanned methionines throughout one of the most popular antibody scaffolds, trastuzumab, used for antibody engineering and drug conjugation. We tested the expression, reactivities, and stabilities of 123 single engineered methionines distributed over the surface of the antibody when reacted with oxaziridine. We found uniformly high expression for these mutants and excellent reaction efficiencies with a panel of oxaziridines. Remarkably, the stability to hydrolysis of the sulfimide varied more than 10-fold depending on temperature and the site of the engineered methionine. Interestingly, the most stable and reactive sites were those that were partially buried, presumably because of their reduced access to water. There was also a 10-fold variation in stability depending on the nature of the oxaziridine, which was determined to be inversely correlated with the electrophilic nature of the sulfimide. Importantly, the stabilities of the best analogs were sufficient to support their use as antibody drug conjugates and potent in a breast cancer mouse xenograft model over a month. These studies provide key parameters for broad application of ReACT for efficient, stable, and site-specific antibody and protein bioconjugation to native or engineered methionines.

Sulfenate Anion Catalyzed Diastereoselective Synthesis of Aziridines.

Aziridines are highly valued synthetic targets in organic and medicinal chemistry. The organocatalytic synthesis of such structures with broad substrate scope and good diastereoselectivity, however, is rare. Herein, we report a broadly applicable and diastereoselective synthetic method for the synthesis of trans-aziridines from imines and benzylic or alkyl halides utilizing sulfenate anions (PhSO- ) as the catalyst. Substrates bearing heterocyclic aromatic groups, alkyl, and electron-rich and electron-poor aryl groups were shown to be compatible with this method (33 examples), giving good yields and high diastereoselectivities (trans : cis >20 : 1). Further functionalization of aziridines containing cyclopropyl or cyclobutyl groups was achieved through ring-opening reactions, with a cyclobutyl-substituted norephedrine derivative obtained through a four-step synthesis. We offer a mechanistic proposal involving reversible addition of the deprotonated benzyl sulfoxide to the imine to explain the high trans-diastereoselectivity.

Ni/Photoredox-Catalyzed C(sp3)-C(sp3) Coupling between Aziridines and Acetals as Alcohol-Derived Alkyl Radical Precursors.

Aziridines are readily available C(sp3) precursors that afford valuable ß-functionalized amines upon ring opening. In this article, we report a Ni/photoredox methodology for C(sp3)-C(sp3) cross-coupling between aziridines and methyl/1°/2° aliphatic alcohols activated as benzaldehyde dialkyl acetals. Orthogonal activation modes of each alkyl coupling partner facilitate cross-selectivity in the C(sp3)-C(sp3) bond-forming reaction: the benzaldehyde dialkyl acetal is activated via hydrogen atom abstraction and ß-scission via a bromine radical (generated in situ from single-electron oxidation of bromide), whereas the aziridine is activated at the Ni center via reduction. We demonstrate that an Ni(II) azametallacycle, conventionally proposed in aziridine cross-coupling, is not an intermediate in the productive cross-coupling. Rather, stoichiometric organometallic and linear free energy relationship studies indicate that aziridine activation proceeds via Ni(I) oxidative addition, a previously unexplored elementary step.

Biocatalytic One-Carbon Ring Expansion of Aziridines to Azetidines via a Highly Enantioselective [1,2]-Stevens Rearrangement.

We report enantioselective one-carbon ring expansion of aziridines to make azetidines as a new-to-nature activity of engineered "carbene transferase" enzymes. A laboratory-evolved variant of cytochrome P450BM3, P411-AzetS, not only exerts unparalleled stereocontrol (99:1 er) over a [1,2]-Stevens rearrangement but also overrides the inherent reactivity of aziridinium ylides, cheletropic extrusion of olefins, to perform a [1,2]-Stevens rearrangement. By controlling the fate of the highly reactive aziridinium ylide intermediates, these evolvable biocatalysts promote a transformation which cannot currently be performed using other catalyst classes.

Aza-Quasi-Favorskii Reaction: Construction of Highly Substituted Aziridines through a Concerted Multibond Rearrangement Process.

A new molecular rearrangement, the aza-Quasi-Favorskii rearrangement, has been developed for the construction of highly substituted aziridines. Electron-deficient O-sulfonyl oximes react readily with α,α-disubstituted acetophenone-derived enolates to furnish highly substituted aziridines via this unprecedented domino process. In-depth computational studies reveal an asynchronous yet concerted nitrenoid-type rearrangement pathway.

Molecular Basis for Enzymatic Aziridine Formation via Sulfate Elimination.

Natural products containing an aziridine ring, such as mitomycin C and azinomycin B, exhibit antitumor activities by alkylating DNA via their aziridine rings; however, the biosynthetic mechanisms underlying the formation of these rings have not yet been elucidated. We herein investigated the biosynthesis of vazabitide A, the structure of which is similar to that of azinomycin B, and demonstrated that Vzb10/11, with no similarities to known enzymes, catalyzed the formation of the aziridine ring via sulfate elimination. To elucidate the detailed reaction mechanism, crystallization of Vzb10/11 and the homologous enzyme, AziU3/U2, in the biosynthesis of azinomycin B was attempted, and the structure of AziU3/U2, which had a new protein fold overall, was successfully determined. The structural analysis revealed that these enzymes adjusted the dihedral angle between the amino group and the adjacent sulfate group of the substrate to almost 180° and enhanced the nucleophilicity of the C6-amino group temporarily, facilitating the SN2-like reaction to form the aziridine ring. The present study reports for the first time the molecular basis for aziridine ring formation.