Total synthesis of plakortone B.

Plakortone B is a naturally occurring bicyclic[3.3.0]furanolactone compound with attractive bioactivities. Although the relative configuration of plakortone B's central core had been established by NMR spectroscopic methods, the absolute configuration of its four stereocenters remained unknown. In the present paper, all four possible isomers of plakortone B were synthesized and one of these molecules was found to be identical with the natural plakortone B on the basis of (1)H, (13)C NMR spectra and specific rotation comparisons. Thus, the absolute configuration of the natural plakortone B was determined to be (3S,4S,6R,10R).

A glycoconjugate antigen based on the recognition motif of a broadly neutralizing human immunodeficiency virus antibody, 2G12, is immunogenic but elicits antibodies unable to bind to the self glycans of gp120.

The glycan shield of human immunodeficiency virus type 1 (HIV-1) gp120 contributes to viral evasion from humoral immune responses. However, the shield is recognized by the HIV-1 broadly neutralizing antibody (Ab), 2G12, at a relatively conserved cluster of oligomannose glycans. The discovery of 2G12 raises the possibility that a carbohydrate immunogen may be developed that could elicit 2G12-like neutralizing Abs and contribute to an AIDS vaccine. We have previously dissected the fine specificity of 2G12 and reported that the synthetic tetramannoside (Man(4)) that corresponds to the D1 arm of Man(9)GlcNAc(2) inhibits 2G12 binding to gp120 as efficiently as Man(9)GlcNAc(2) itself, indicating the potential use of Man(4) as a building block for creating immunogens. Here, we describe the development of neoglycoconjugates displaying variable copy numbers of Man(4) on bovine serum albumin (BSA) molecules by conjugation to Lys residues. The increased valency enhances the apparent affinity of 2G12 for Man(4) up to a limit which is achieved at approximately 10 copies per BSA molecule, beyond which no further enhancement is observed. Immunization of rabbits with BSA-(Man(4))(14) elicits significant serum Ab titers to Man(4). However, these Abs are unable to bind gp120. Further analysis reveals that the elicited Abs bind a variety of unbranched and, to a lesser extent, branched Man(9) derivatives but not natural N-linked oligomannose containing the chitobiose core. These results suggest that Abs can be readily elicited against the D1 arm; however, potential differences in the presentation of Man(4) on neoglycoconjugates, compared to glycoproteins, poses challenges for eliciting anti-mannose Abs capable of cross-reacting with gp120 and HIV-1.

Dissection of the carbohydrate specificity of the broadly neutralizing anti-HIV-1 antibody 2G12.

Human antibody 2G12 neutralizes a broad range of HIV-1 isolates. Hence, molecular characterization of its epitope, which corresponds to a conserved cluster of oligomannoses on the viral envelope glycoprotein gp120, is a high priority in HIV vaccine design. A prior crystal structure of 2G12 in complex with Man(9)GlcNAc(2) highlighted the central importance of the D1 arm in antibody binding. To characterize the specificity of 2G12 more precisely, we performed solution-phase ELISA, carbohydrate microarray analysis, and cocrystallized Fab 2G12 with four different oligomannose derivatives (Man(4), Man(5), Man(7), and Man(8)) that compete with gp120 for binding to 2G12. Our combined studies reveal that 2G12 is capable of binding both the D1 and D3 arms of the Man(9)GlcNAc(2) moiety, which would provide more flexibility to make the required multivalent interactions between the antibody and the gp120 oligomannose cluster than thought previously. These results have important consequences for the design of immunogens to elicit 2G12-like neutralizing antibodies as a component of an HIV vaccine.

Covalent display of oligosaccharide arrays in microtiter plates.

A covalent array for the display of complex oligosaccharides in microtiter plates has been developed. This strategy is conducive to the display of carbohydrates to proteins of interest such as lectins and antibodies, including the broadly neutralizing antibody 2G12 against HIV envelope oligomannose and can be cleaved from the surface for further characterization by mass spectrometry. The system was used to probe the multivalent interaction of 2G12 with an optimal epitope (Kd 0.1 muM).

Reactivity-based one-pot total synthesis of fucose GM1 oligosaccharide: a sialylated antigenic epitope of small-cell lung cancer.

The total synthesis of the sialic acid-containing antigenic epitope fucose GM(1) (Fuc-GM(1)) by an improved reactivity-based one-pot synthetic strategy is reported. Based on a thioglycoside reactivity database, three saccharide building blocks, 3, 4, and 5, were designed and prepared to incorporate a descending order of reactivity toward thiophilic activation. Using the reactivity-based one-pot synthetic method, the fully protected Fuc-GM(1) glycoside 2 was furnished in a facile manner, which was globally deprotected to give the Fuc-GM(1) glycoside 1. In addition, using the promoter system 1-(benzensulfinyl)piperidinetrifluoromethanesulfonic anhydride, the product yield was improved and the reaction time was reduced in comparison with the N-iodosuccinimidetrifluoromethanesulfonic acid- and dimethyl (thiomethyl) sulfonium trifluoromethanesulfonate-promoted systems.

An enantioselective synthetic pathway towards plakortones.

An enantioselective synthesis of functionalized bicyclic lactones 2, 3 and 4, core structures of plakortones, are described; the configurations of 2, 3 and 4 were confirmed by X-ray crystallographic analyses of their precursors 11, 19 and 24, respectively.