Organocatalytic atroposelective construction of axially chiral arylquinones.

Atropisomeric biaryl motifs are ubiquitous in chiral catalysts and ligands. Numerous efficient strategies have been developed for the synthesis of axially chiral biaryls. In contrast, the asymmetric construction of o-quinone-aryl atropisomers has yet to be realized. Inspired by the rapid progress of the chemistry of biaryls, here we present our initial investigations about the atroposelective construction of axially chiral arylquinones by a bifunctional chiral phosphoric acid-catalyzed asymmetric conjugate addition and central-to-axial chirality conversion. With o-naphthoquinone as both the electrophile and the oxidant, three types of arylation counterparts, namely 2-naphthylamines, 2-naphthols and indoles, are utilized to assemble a series of atropisomeric scaffolds in good yields and excellent enantioselectivities. This approach not only expands the axially chiral library but also offers a route to a class of potential, chiral biomimetic catalysts.

Asymmetric phosphoric acid-catalyzed four-component Ugi reaction.

The Ugi reaction constructs α-acylaminoamide compounds by combining an aldehyde or ketone, an amine, a carboxylic acid, and an isocyanide in a single flask. Its appealing features include inherent atom and step economy together with the potential to generate products of broad structural diversity. However, control of the stereochemistry in this reaction has proven to be a formidable challenge. We describe an efficient enantioselective four-component Ugi reaction catalyzed by a chiral phosphoric acid derivative that delivers more than 80 α-acylaminoamides in good to excellent enantiomeric excess. Experimental and computational studies establish the reaction mechanism and origins of stereoselectivity.

Anti-inflammatory activity of cecropin-A2 from Musca domestica.

This study aimed to investigate the anti-inflammatory activity of Musca domestica cecropin-A2 (Mdc-A2) toward Staphylococcus aureus (S. aureus) to learn more about their immunological functions. RAW264.7 cells were transfected with recombinant lentiviruses introduce pLEX-Mdc-A2into the RAW264.7 cell line (RAW-Mdc-A2). The RAW264.7 cell line with empty pLEX (RAW-pLEX) was produced in the same manner as a negative control. Real-time quantitative reverse transcription PCR (RT-PCR) was performed to analyze the mRNA expression of TNF-a, IL-1ß, NFκB-1 and NFκB-2 in S. aureus-stimulated RAW-Mdc-A2 cells and RAW-pLEX cells in untreated cells and cells treated for 3 h, 6 h, 12 h and 24 h. RT-PCR was performed to analyze the mRNA expression of TNF-a, NFκB-1 and NFκB-2 stimulated by Lipoteichoic acid (LTA). Production of TNF-a was detected by enzyme-linked immunosorbent assay (ELISA). Colony counts were used to calculate the number of CFU per mL of cell culture supernatants. The results showed that compared to RAW-pLEX cells, stable transfection of Mdc-A2 in RAW264.7 cells stimulated by S. aureus significantly down-regulated the mRNA expression of TNF-a transcript variant 1 (TNF-a-tv-1) at 6 h and 12 h and the mRNA expression of TNF-a transcript variant 2 (TNF-a-tv-2) at 3 h, 6 h and 12 h. Compared to RAW-pLEX cells, stable transfection of Mdc-A2 in RAW264.7 cells stimulated by S. aureus significantly down-regulated the mRNA expression of IL-1ß-T at 3 h, 6 h and 12 h as well as the mRNA expression of IL-1ß at 3 h and 6 h. The expression and production of TNF-a and bacterial burden of cell culture supernatants were significantly down-regulated in RAW-Mdc-A2 cells stimulated by S. aureus, and the expression and production of TNF-a were significantly down-regulated in RAW-Mdc-A2 cells stimulated by LTA. Compared to RAW-pLEX cells, stable transfection of Mdc-A2 in RAW264.7 cells stimulated by S. aureus significantly down-regulated the mRNA expression of NFκB-1 at 3 h, 6 h and 12 h as well as the mRNA expression of NFκB-2 at 6 h. Additionally, stable transfection of Mdc-A2 in RAW264.7 cells stimulated by LTA significantly down-regulated the mRNA expression of NFκB-1. In conclusion, Mdc-A2 possesses potent anti-inflammatory activity and potent antimicrobial activity. Additionally, Mdc-A2 may interact with LTA and execute strong anti-inflammatory activity by blocking the activation of NF-κB signaling pathways.

Amphiregulin Antibody and Reduction of Axial Elongation in Experimental Myopia.

To examine the mechanism of ocular axial elongation in myopia, guinea pigs (age: 2-3weeks) which either underwent unilateral or bilateral lens-induced myopization (group 1) or which were primarily myopic at baseline (group 2) received unilateral intraocular injections of amphiregulin antibody (doses: 5, 10, or 15µg) three times in intervals of 9days. A third group of emmetropic guinea pigs got intraocular unilateral injections of amphiregulin (doses: 0.25, 0.50 or 1.00ng, respectively). In each group, the contralateral eyes received intraocular injections of Ringer's solution. In intra-animal inter-eye comparison and intra-eye follow-up comparison in groups 1 and 2, the study eyes as compared to the contralateral eyes showed a dose-dependent reduction in axial elongation. In group 3, study eyes and control eyes did not differ significantly in axial elongation. Immunohistochemistry revealed amphiregulin labelling at the retinal pigment epithelium in eyes with lens-induced myopization and Ringer's solution injection, but not in eyes with amphiregulin antibody injection. Intraocular injections of amphiregulin-antibody led to a reduction of lens-induced axial myopic elongation and of the physiological eye enlargement in young guinea pigs. In contrast, intraocularly injected amphiregulin in a dose of ≤1ng did not show a significant effect. Amphiregulin may be one of several essential molecular factors for axial elongation.

A small-molecule inhibitor suppresses the tumor-associated mitochondrial NAD(P)+-dependent malic enzyme (ME2) and induces cellular senescence.

Here, we found a natural compound, embonic acid (EA), that can specifically inhibit the enzymatic activity of mitochondrial NAD(P)+-dependent malic enzyme (m-NAD(P)-ME, ME2) either in vitro or in vivo. The in vitro IC50 value of EA for m-NAD(P)-ME was 1.4 ± 0.4 µM. Mutagenesis and binding studies revealed that the putative binding site of EA on m-NAD(P)-ME is located at the fumarate binding site or at the dimer interface near the site. Inhibition studies reveal that EA displayed a non-competitive inhibition pattern, which demonstrated that the binding site of EA was distinct from the active site of the enzyme. Therefore, EA is thought to be an allosteric inhibitor of m-NAD(P)-ME. Both EA treatment and knockdown of m-NAD(P)-ME by shRNA inhibited the growth of H1299 cancer cells. The protein expression and mRNA synthesis of m-NAD(P)-ME in H1299 cells were not influenced by EA, suggesting that the EA-inhibited H1299 cell growth occurs through the suppression of in vivo m-NAD(P)-ME activity EA treatment further induced the cellular senescence of H1299 cells. However, down-regulation of the enzyme-induced cellular senescence was not through p53. Therefore, the EA-evoked senescence of H1299 cells may occur directly through the inhibition of ME2 or a p53-independent pathway.

Structural characteristics of the nonallosteric human cytosolic malic enzyme.

Human cytosolic NADP(+)-dependent malic enzyme (c-NADP-ME) is neither a cooperative nor an allosteric enzyme, whereas mitochondrial NAD(P)(+)-dependent malic enzyme (m-NAD(P)-ME) is allosterically activated by fumarate. This study examines the molecular basis for the different allosteric properties and quaternary structural stability of m-NAD(P)-ME and c-NADP-ME. Multiple residues corresponding to the fumarate-binding site were mutated in human c-NADP-ME to correspond to those found in human m-NAD(P)-ME. Additionally, the crystal structure of the apo (ligand-free) human c-NADP-ME conformation was determined. Kinetic studies indicated no significant difference between the wild-type and mutant enzymes in Km,NADP, Km,malate, and kcat. A chimeric enzyme, [51-105]_c-NADP-ME, was designed to include the putative fumarate-binding site of m-NAD(P)-ME at the dimer interface of c-NADP-ME; however, this chimera remained nonallosteric. In addition to fumarate activation, the quaternary structural stability of c-NADP-ME and m-NAD(P)-ME is quite different; c-NADP-ME is a stable tetramer, whereas m-NAD(P)-ME exists in equilibrium between a dimer and a tetramer. The quaternary structures for the S57K/N59E/E73K/S102D and S57K/N59E/E73K/S102D/H74K/D78P/D80E/D87G mutants of c-NADP-ME are tetrameric, whereas the K57S/E59N/K73E/D102S m-NAD(P)-ME quadruple mutant is primarily monomeric with some dimer formation. These results strongly suggest that the structural features near the fumarate-binding site and the dimer interface are highly related to the quaternary structural stability of c-NADP-ME and m-NAD(P)-ME. In this study, we attempt to delineate the structural features governing the fumarate-induced allosteric activation of malic enzyme.