Alexey Stepanov

Ubiquitin-independent proteosomal degradation of myelin basic protein contributes to development of neurodegenerative autoimmunity

Recent findings indicate that the ubiquitin‐proteasome system is involved in the pathogenesis of cancer as well as autoimmune and several neurodegenerative diseases, and is thus a target for novel therapeutics. One disease that is related to aberrant protein degradation is multiple sclerosis, an autoimmune disorder involving the processing and presentation of myelin autoantigens that leads to the destruction of axons. Here, we show that brain‐derived proteasomes from SJL mice with experimental autoimmune encephalomyelitis (EAE) in an ubiquitin‐independent manner generate significantly increased amounts of myelin basic protein peptides that induces cytotoxic lymphocytes to target mature oligodendrocytes ex vivo. Ten times enhanced release of immunogenic peptides by cerebral proteasomes from EAE‐SJL mice is caused by a dramatic shift in the balance between constitutive and β1ihigh immunoproteasomes in the CNS of SJL mice with EAE. We found that during EAE, β1i is increased in resident CNS cells, whereas β5i is imported by infiltrating lymphocytes through the blood‐brain barrier. Peptidyl epoxyketone specifically inhibits brain‐derived β1ihigh immunoproteasomes in vitro (kobs/[I] = 240 M‐1s‐1), and at a dose of 0.5 mg/kg, it ameliorates ongoing EAE in vivo. Therefore, our findings provide novel insights into myelin metabolism in pathophysiologic conditions and reveal that the β1i subunit of the immunoproteasome is a potential target to treat autoimmune neurologic diseases.—Belogurov Jr., A., Kuzina, E., Kudriaeva, A., Kononikhin, A., Kovalchuk, S., Surina, Y., Smirnov, I., Lomakin, Y., Bacheva, A., Stepanov, A., Karpova, Y., Lyupina, Y., Kharybin, O., Melamed, D., Ponomarenko, N., Sharova, N., Nikolaev, E., Gabibov, A. Ubiquitin‐independent proteosomal degradation of myelin basic protein contributes to development of neurodegenerative autoimmunity. FASEB J. 29, 1901‐1913 (2015). www.fasebj.org

Voltammetric study of metal ions binding by biindolizine heterocyclophanes and their acyclic analogues

The binding of ions Li+, Na+, K+, (group I), Mg2+, Al3+, Ga3+ (group II), Ca2+, Pb2+ (group III) ions, Ba2+ and paraquat by heterocyclophanes containing biindolizine and quinoxaline fragments connected by 3,6,9-trioxaundecane and 5,8,11,14,17-pentaoxageneicosane spacers, and also their acyclic analogues, in the acetonitrile-0.1 M Bu4NBF4 is studied by cyclic voltammetry. A conclusion is drawn that the ions of the group I are not bound by these compounds; the paraquat is not bound by heterocyclophane with the 5,8,11,14,17-pentaoxageneicosan spacers. For ions of the group II, reversible redox-switchable binding by the macrocycles with the 3,6,9-trioxaundecane and 5,8,11,14,17-pentaoxageneicosan spacers is observed: the initial compounds show the binding; their radical cations and dications do not. The binding of the ions of the group III and Ba2+ is determined by the macrocycles’ size. In particular, these ions are bound not only by the heterocyclophane with 3,6,9-trioxaundecane spacers but also by its radical cation or dication. The binding results in the corresponding dication stabilization. The heterocyclophane with the 5,8,11,14,17-pentaoxageneicosan spacers demonstrates the redox-switchable binding of Ca2+ and Pb2+ ions; no effect of Ba2+ ions on the cyclic voltammograms of this heterocyclophane was observed. In the ternary system “heterocyclophane with 3,6,9-trioxaundecane spacers + ions of the group II (Al3+, Ga3+) + ions of the group III (Ca2+, Pb2+)” either primary binding of the group III ion Pb2+ or concurrent binding of the ions of the group II and the group III, with the system’s reversible redox-switching from one metal complex to another, was observed.

Electricoswitchable bonding of metal ions and complexes by calixarenes

The results of authors on designing electroswitchable supramolecular systems based on calix[4]arenes, calix[4]resorcinols, and transition metal ions and complexes are summarized. We consider systems in which the switching is performed owing to electrochemical reactions both of groups grafted to the macrocycle and bound substrates. Examples of electrochemically switchable luminescence are given.

Electrochemical properties of n-sulfonatothiacalyx[4]arene complexes with Fe3+ and [Co(dipy)3]3+ ions

The electrochemical properties of n-sulfonatothiacalyx[4]arene (H4XNa4) complexes with [Co(dipy)3]3+ and Fe3+ ions were studied by means of cyclic voltammetry in aqueous solution at pH 2.5. The observed single-electron reduction of [Co(dipy)3]3+ bound extraspherically to the upper rim and Fe3+ ion bound intraspherically to the lower rim of n-sulfonatothiacalyx[4]arene in binary [Co(dipy)3]3+ · H3X5−, H3X5− · Fe3+, and ternary [Co(dipy)3]3+ · H3X5− · Fe3+ heterometal complexes was more difficult than in the free state. The reversible single-electron transfer to the metal ion results in lower binding energy ([Co(dipy)3]3+, ΔΔG0 = 3.9 kJ/mol) or in full fast dissociation of the complex (Fe3+). The ternary complex in the solution forms the aggregates, in which inner encapsulated Fe(III) and Co(III) ions are not reduced on the electrode. Their quantitative reduction takes place by the relay mechanism of intra- and intermolecular electron transfer through electrochemically generated [Co(dipy)3]2+ outer ions.

Tuning the non-covalent confinement of Gd(III) complexes in silica nanoparticles for high T1-weighted MR imaging capability

The present work introduces deliberate synthesis of Gd(III)-doped silica nanoparticles with high relaxivity at magnetic field strengths below 1.5T. Modified microemulsion water-in-oil procedure was used in order to achieve superficial localization of Gd(III) complexes within 40-55nm sized silica spheres. The relaxivities of the prepared nanoparticles were measured at 0.47, 1.41 and 1.5T with the use of both NMR analyzer and whole body NMR scanner. Longitudinal relaxivities of the obtained silica nanoparticles reveal significant dependence on the confinement mode, changing from 4.1 to 49.6mM-1s-1 at 0.47T when the localization of Gd(III) complexes changes from core to superficial zones of the silica spheres. The results highlight predominant contribution of the complexes located close to silica/water interface to the relaxivity of the nanoparticles. Low effect of blood proteins on the relaxivity in the aqueous colloids of the nanoparticles was exemplified by serum bovine albumin. T1- weighted MRI data indicate that the nanoparticles provide strong positive contrast at 1.5T, which along with low cytotoxicity effect make a good basis for their application as contrast agents.

Nanoscale hydrophilic colloids with high relaxivity and low cytotoxicity based on Gd(III) complexes with Keplerate polyanions

The work introduces for the first time Mo-132-type Keplerates (Kp) and Gd3+ ions as a basis for the one-pot synthesis of aqueous colloids with high longitudinal relaxivity (r1 = 96.4 mM−1 s−1 at 20 MHz). The hydrophilic coating of small (3–5 nm) Gdx(Kp)y-based cores provides their colloid stabilization, protection from degradation and low cytotoxicity.

Administration of Myelin Basic Protein Peptides Encapsulated in Mannosylated Liposomes Normalizes Level of Serum TNF-α and IL-2 and Chemoattractants CCL2 and CCL4 in Multiple Sclerosis Patients

We have previously shown that immunodominant MBP peptides encapsulated in mannosylated liposomes (Xemys) effectively suppressed experimental allergic encephalomyelitis (EAE). Within the frames of the successfully completed phase I clinical trial, we investigated changes in the serum cytokine profile after Xemys administration in MS patients. We observed a statistically significant decrease of MCP-1/CCL2, MIP-1β/CCL4, IL-7, and IL-2 at the time of study completion. In contrast, the serum levels of TNF-α were remarkably elevated. Our data suggest that the administration of Xemys leads to a normalization of cytokine status in MS patients to values commonly reported for healthy subjects. These data are an important contribution for the upcoming Xemys clinical trials.

Electroswitchable binding of [Co(dipy)3]3+ and [Fe(dipy)3]2+n-sulfonato(thia)calix[4]arenas

The methods of cyclic voltammetry, electrolysis, and spectrophotometry were used to study electrochemical properties of (TCAS + Fe3+ + dipy), (CCAS + Fe3+ + dipy), and (CCAS + Fe3+ + [Co(dipy)3]3+) triple systems (where TCAS is n-sulfonatothiacalix[4]arene, CCAS is tetracarboxylate n-sulfonatocalix[4]arene, and dipy = α,α′-dipyridyl) in an aqueous solution. One-electron reduction of Fe(III) in the (TCAS + Fe3+ + dipy) system at pH 2.5 results in electroswitching of iron ions from the lower TCAS ring to the upper ([Fe(dipy)3]2+). Reverse electrochemical switching of the system is impossible due to mediator ([Fe(dipy)3]2+/3+) oxidation of TCAS. Reverse electroswitching of Fe(III) ions from unbound to bound state as ([Fe(dipy)3]2+) with CCAS has been revealed in the system (CCAS + Fe3+ + dipy) (pH 1.7) upon single-electron transfer, whereas reversible electroswitching by the upper rim of CCAS from one complex ion ([Co(dipy)3]3+) to another ([Fe(dipy)3]2+) has been demonstrated in the system ([Co(dipy)3]3+ + CCAS + Fe3+ upon double-electron transfer. In all systems, electric switching was accompanied by synchronous color switching.

Colloid Properties of Paramagnetic Nanoparticles Based on Novel Thiacalix[4]arenes and Gd(III) Ions

The present work introduces colloid behaviour of core-shell nanoparticles, where hard paramagnetic cores are coated by hydrophilic polyelectrolyte-based shells. Paramagnetic cores are built from water insoluble Gd(III) complexes with alkyl-malonate derivatives of thiacalix[4]arenes adopting 1,3-alternate conformation. Polystyrenesulfonate (PSS) and polyethyleneimine (PEI) are used to deposit one (PSS) or two (PSS-PEI) polyelectrolyte layers onto the cores. Small angle light scattering was used to reveal both size and aggregation behaviour of the core-shell colloids in aqueous solutions.