Metal-organic coordination polymers for delivery of immunomodulatory agents, and infectious disease and cancer vaccines.

Metal-organic coordination polymers (CPs) are a broad class of materials that include metal-organic frameworks (MOFs). CPs are highly ordered crystalline materials that are composed of metal ions (or metal ion clusters) and multidentate organic ligands that serve as linkers. One-, two-, and three-dimensional CPs can be formed, with 2D and 3D structures referred to as MOFs. CPs have gained a lot of attention due to attractive structural features like structure versatility and tunability, and well-defined pores that enable the encapsulation of cargo. Further, CPs show a lot of promise for drug delivery applications, but only a very limited number of CPs are currently being evaluated in clinical trials. In this review, we outlined features that are desired for CP-based drug delivery platform, and briefly described most relevant characterization techniques. We highlighted some of the recent efforts directed toward developing CP-based drug delivery platforms with the emphasis on vaccines against cancer, infectious diseases, and viruses. We hope this review will be a helpful guide for those interested in the design and evaluation of CP-based immunological drug delivery platforms. This article is categorized under: Therapeutic Approaches and Drug Discovery > Nanomedicine for Infectious Disease Therapeutic Approaches and Drug Discovery > Nanomedicine for Oncologic Disease.

Vinyl Sulfone-functionalized Acetalated Dextran Microparticles as a Subunit Broadly Acting Influenza Vaccine.

Influenza is a global health concern with millions of infections occurring yearly. Seasonal flu vaccines are one way to combat this virus; however, they are poorly protective against influenza as the virus is constantly mutating, particularly at the immunodominant hemagglutinin (HA) head group. A more broadly acting approach involves Computationally Optimized Broadly Reactive Antigen (COBRA). COBRA HA generates a broad immune response that is capable of protecting against mutating strains. Unfortunately, protein-based vaccines are often weekly immunogenic, so to help boost the immune response, we employed the use of acetalated dextran (Ace-DEX) microparticles (MPs) two ways: one to conjugate COBRA HA to the surface and a second to encapsulate cGAMP. To conjugate the COBRA HA to the surface of the Ace-DEX MPs, a poly(L-lactide)-polyethylene glycol co-polymer with a vinyl sulfone terminal group (PLLA-PEG-VS) was used. MPs encapsulating the STING agonist cGAMP were co-delivered with the antigen to form a broadly active influenza vaccine. This vaccine approach was evaluated in vivo with a prime-boost-boost vaccination schedule and illustrated generation of a humoral and cellular response that could protect against a lethal challenge of A/California/07/2009 in BALB/c mice.

Metal-Organic Coordination Polymer for Delivery of a Subunit Broadly Acting Influenza Vaccine.

A zinc-carnosine (ZnCar) metal-organic coordination polymer was fabricated in biologically relevant N-(2-hydroxyethyl)piperazine-N'-ethanesulfonic acid (HEPES) buffer for use as a vaccine platform. In vitro, ZnCar exhibited significantly less cytotoxicity than a well-established zeolitic imidazolate framework (ZIF-8). Adsorption of CpG on the ZnCar surface resulted in enhanced innate immune activation compared to soluble CpG. The model antigen ovalbumin (OVA) was encapsulated in ZnCar and exhibited acid-sensitive release in vitro. When injected intramuscularly on days 0 and 21 in C57BL/6 mice, OVA-specific serum total IgG and IgG1 were significantly greater in all groups with ZnCar and antigen compared to soluble controls. Th1-skewed IgG2c antibodies were significantly greater in OVA and CpG groups delivered with ZnCar for all time points, regardless of whether the antigen and adjuvant were co-formulated in one material or co-delivered in separate materials. When broadly acting Computationally Optimized Broadly Reactive Antigen (COBRA) P1 influenza hemagglutinin (HA) was ligated to ZnCar via its His-tag, significantly greater antibody levels were observed at all time points compared to soluble antigen and CpG. ZnCar-formulated antigen elicited increased peptide presentation to B3Z T cells in vitro and production of IL-2 after ex vivo antigen recall of splenocytes isolated from vaccinated mice. Overall, this work displays the formation of a zinc-carnosine metal-organic coordination polymer that can be applied as a platform for recombinant protein-based vaccines.

Dexamethasone and Fumaric Acid Ester Conjugate Synergistically Inhibits Inflammation and NF-κB in Macrophages.

Macrophage-mediated inflammation drives autoimmune and chronic inflammatory diseases. Treatment with anti-inflammatory agents can be an effective strategy to reduce this inflammation; however, high concentrations of these agents can have immune-dampening and other serious side effects. Synergistic combination of anti-inflammatory agents can mitigate dosing by requiring less drug. Multiple anti-inflammatory agents were evaluated in combination for synergistic inhibition of macrophage inflammation. The most potent synergy was observed between dexamethasone (DXM) and fumaric acid esters (e.g., monomethyl fumarate (MMF)). Furthermore, this combination was found to synergistically inhibit inflammatory nuclear factor κB (NF-κB) transcription factor activity. The optimal ratio for synergy was determined to be 1:1, and DXM and MMF were conjugated by esterification at this molar ratio. The DXM-MMF conjugate displayed improved inhibition of inflammation over the unconjugated combination in both murine and human macrophages. In the treatment of human donor monocyte-derived macrophages, the combination of DXM and MMF significantly inhibited inflammatory gene expression downstream of NF-κB and overall performed better than either agent alone. Further, the DXM-MMF conjugate significantly inhibited expression of NOD-, LRR-, and pyrin domain-containing protein 3 (NLRP3) inflammasome-associated genes. The potent anti-inflammatory activity of the DXM-MMF conjugate in human macrophages indicates that it may have benefits in the treatment of autoimmune and inflammatory diseases.

Biochemical Characterization of the Two-Component Flavin-Dependent Monooxygenase Involved in Valanimycin Biosynthesis.

The flavin reductase (FRED) and isobutylamine N-hydroxylase (IBAH) from Streptomyces viridifaciens constitute a two-component, flavin-dependent monooxygenase system that catalyzes the first step in valanimycin biosynthesis. FRED is an oxidoreductase that provides the reduced flavin to IBAH, which then catalyzes the hydroxylation of isobutylamine (IBA) to isobutylhydroxylamine (IBHA). In this work, we used several complementary methods to investigate FAD binding, steady-state and rapid reaction kinetics, and enzyme-enzyme interactions in the FRED:IBAH system. The affinity of FRED for FADox is higher than its affinity for FADred, consistent with its function as a flavin reductase. Conversely, IBAH binds FADred more tightly than FADox, consistent with its role as a monooxygenase. FRED exhibits a strong preference (28-fold) for NADPH over NADH as the electron source for FAD reduction. Isothermal titration calorimetry was used to study the association of FRED and IBAH. In the presence of FAD, either oxidized or reduced, FRED and IBAH associate with a dissociation constant of 7-8 µM. No interaction was observed in the absence of FAD. These results are consistent with the formation of a protein-protein complex for direct transfer of reduced flavin from the reductase to the monooxygenase in this two-component system.

Author Correction: Identification of Aspergillus fumigatus UDP-Galactopyranose Mutase Inhibitors.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

Identification of Aspergillus fumigatus UDP-Galactopyranose Mutase Inhibitors.

Aspergillus fumigatus is an opportunistic human pathogen responsible for deadly, invasive infections in immunocompromised patients. The A. fumigatus cell wall is a complex network of polysaccharides among them galactofuran, which is absent in humans. UDP-galactopyranose mutase (UGM) catalyzes the conversion of UDP-galactofuranose (UDP-Galf) to UDP-galactopyranose (UDP-Galp) and is an important virulence factor. UGM is a flavin-dependent enzyme that requires the reduced flavin for activity; flavin reduction is achieved by reaction with NADPH. The aim of this work was to discover inhibitors of UGM by targeting the NADPH binding site using an ADP-TAMRA probe in a high-throughput screening assay. The flavonoids (2S)-hesperetin and (2S)-naringenin were validated as competitive inhibitors of UGM against NADPH with Ki values of 6 µM and 74 µM, respectively. To gain insight into the active chemical substituents involved in the inhibition of UGM, several derivatives of these inhibitors were studied. The results show that the hydroxyl groups of (2S)-hesperetin are important for inhibition, in particular the phenyl-chroman moiety. Congo red susceptibility assay and growth temperature effects showed that these compounds affected cell wall biosynthesis in A. fumigatus. This work is the first report of inhibition studies on UGM from eukaryotic human pathogens.

Identification of eukaryotic UDP-galactopyranose mutase inhibitors using the ThermoFAD assay.

Aspergillus fumigatus is a human pathogen responsible for deadly infections in immune-compromised patients. A potential strategy for treating A. fumigatus infections is by targeting the biosynthesis of cell wall components, such as galactofuranase, which is absent in humans. Galactofuranose biosynthesis is initiated by the flavoenzyme UDP-galactopyranose mutase (UGM), which converts UDP-galactopyranose (UDP-Galp) to UDP-galactofuranose (UDP-Galf). UGM requires the reduced form of the flavin for activity, which is obtained by reacting with NADPH. We aimed to identify inhibitors of UGM by screening a kinase inhibitor library using ThermoFAD, a flavin fluorescence thermal shift assay. The screening assay identified flavopiridol as a compound that increased the melting temperature of A. fumigatus UGM. Further characterization showed that flavopiridol is a non-competitive inhibitor of UGM and docking studies suggest that it binds in the active site. This compound does not inhibit the prokaryotic UGM from Mycobacteria tuberculosis.

Cold denaturation induces inversion of dipole and spin transfer in chiral peptide monolayers.

Chirality-induced spin selectivity is a recently-discovered effect, which results in spin selectivity for electrons transmitted through chiral peptide monolayers. Here, we use this spin selectivity to probe the organization of self-assembled α-helix peptide monolayers and examine the relation between structural and spin transfer phenomena. We show that the α-helix structure of oligopeptides based on alanine and aminoisobutyric acid is transformed to a more linear one upon cooling. This process is similar to the known cold denaturation in peptides, but here the self-assembled monolayer plays the role of the solvent. The structural change results in a flip in the direction of the electrical dipole moment of the adsorbed molecules. The dipole flip is accompanied by a concomitant change in the spin that is preferred in electron transfer through the molecules, observed via a new solid-state hybrid organic-inorganic device that is based on the Hall effect, but operates with no external magnetic field or magnetic material.

A versatile water-soluble chelating and radical scavenging platform.

The phenol-diamide compound, 5-(tert-butyl)-2-hydroxy-N1,N3-bis(2-hydroxyethyl)isophthalamide (), is water-soluble, non-cytotoxic, and capable of both, trapping ROS species and chelating Cu(ii) and Fe(iii) ions; these combined properties confer a protective effect against ROS induced cell death.

Nickel(III) complexes of di-amidato-di-phenolato ligands: effect of H-bonding.

The one-electron oxidation of the dianionic diamido-diphenoxo Ni(II) complexes involving H-bonding (1(2-)), or not (2(2-)), yields the corresponding Ni(III) species; the formation, stability and electronic structures of which are affected by the H-bonding interactions.

A novel di-compartmental bis-(2-hydroxyisophtalamide) macrocyclic ligand and its mononuclear Cu(II) and Ni(II) complexes.

The synthesis and characterisation of the new di-compartmental bis-(2-hydroxyisophtalamide) macrocyclic pro-ligand, LH(6), which comprises two phenol-diamide units linked by ethylene bridges, is herein reported, together with its corresponding di-phenolate salt, [NBu(4)](2)[LH(4)]. The three macrocyclic compounds, [LH(4)(OMe)(2)] (protected ligand), LH(6) and [LH(4)][NBu(4)](2) were fully characterised including X-ray crystallography for [LH(4)(OMe)(2)] and [NBu(4)](2)[LH(4)]. The results of solid-state and solution studies have indicated that the macrocycle can adopt specific conformations, which are influenced by H-bonding interactions as well as the deviation of the amide carbonyl relative to the phenol plane. LH(6) reacts with M(II)(acetate)(2)·(H(2)O)(6) (M = Ni, Cu) in a 1 : 1 ratio in the presence of 4 eq of [NBu(4)](OH) in methanol to afford the dianionic [M(LH(2))](2-) complexes, (2-) and (2-), respectively. The X-ray crystallography, EPR, NMR and UV-vis spectroscopic data, combined with DFT calculations, indicate that (2-) and (2-) are unique unsymmetrical square planar mononuclear complexes that are intramolecularly H-bonded. Thus, one macrocyclic compartment contains a M(II)-N(2)O(2) centre resulting from the tetra-anionic di-phenolato di-amidato ligation; the other compartment possesses two protonated amide N-H groups that are H-bonded the coordinated phenolate O atoms. This represents a unique example in which a phenolate is both coordinated and intramolecularly H-bonded. This H-bonding appears unusually strong as revealed by N(H/D) exchange experiments; and may be responsible for the stability of the mononuclear complex, and the difficulty in isolating the corresponding dinuclear complex [M(2)(L)](2-).