Total Synthesis of a Conjugation-Ready Tetrasaccharide Repeating Unit of Vibrio cholerae O:3 O-antigen Polysaccharide

Abstract

Herein, we report the first total synthesis of the tetrasaccharide repeating unit of Vibrio cholerae O:3 O-antigen polysaccharide. The highly complex tetrasaccharide contains rare amino sugars such as D-bacillosamine and L-fucosamine, highly labile sugar ascarylose, and higher carbon sugar D-D-heptose. Stereoselective glycosylation of the notoriously reactive ascarylose with D-D-heptose, poor nucleophilicity of the axial C4-OH of Lfucosamine, and amide coupling are the key challenges encountered in the total synthesis, which was completed via a longest linear sequence of 23 steps in 4.2% overall yield.Vibrio cholerae is a Gram-negative, facultative anaerobic, and comma-shaped bacteria, commonly found in contaminated water and food. According to the WHO fact sheet, Vibrio cholerae is a highly contagious pathogen, causing over 1 million cases of diarrhea and a significant number of deaths globally every year. 1 The world has faced seven massive cholerae pandemics, epidemics, endemics, and seasonal or sporadic outbreaks. V. cholerae can survive in adverse aquatic environments for prolonged periods, which makes the bacteria highly virulent. 2 This high-risk bacterium can cause watery diarrhea associated with dehydration, vomiting, unconsciousness, and death within a few hours if left untreated. Furthermore, epidemiological study proves that V. cholerae shows antimicrobial resistance toward first-and second-line antibiotics. 3 The prophylaxis strategies are therefore required to curb the disease. There are a few oral cholera vaccines (OCVs) approved by WHO. However, degradation of the antigen exposed to acidic pH in the stomach and short-lived immune protection leaves room for further development for a multicomponent glycoconjugate vaccine against all O-antigen serogroups of V. cholerae. 4 There are more than 200 serogroups based on O-antigen of the lipopolysaccharide present on the bacterial cell surface. 2 Recently, strain O:3 was detected in the archived clinical isolates obtained from cholera outbreaks that took place in southern Ghana during 2012− 2015. 3 Surface exposed carbohydrates are antigenic materials and are key components of a glycoconjugate vaccine. 5 Also, they are known to be safe and create a long-lasting T-cell-dependent memory, upon protein conjugation. 6 The bacterial cell surface carries glycoconjugates and oligosaccharides containing rare deoxy amino sugars that are absent in human cells. 7 This critical difference can be exploited to develop carbohydrate-

Total Synthesis of the Conjugation-Ready Tetrasaccharide Repeating Unit of Shewanella japonica Type Strain KMM 3299T

“…For the ensuing glycosylation with bacillosamine acceptor 7 , since a distal C4 participating group is absent in l -fucosamine, we thought of exploring the solvent participation effect using Et 2 O. According to that, disaccharide thioglycoside donor 6 on glycosylation using NIS and TMSOTf as a promoter in dry CH 2 Cl 2 :Et 2 O (1:1) with acceptor 7 , at −40 to 0 °C afforded the corresponding trisaccharide, which upon benzylidene hydrolysis under acidic medium using p -TSA and CH 3 OH:CH 2 Cl 2 (4:1) furnished trisaccharide diol 16 in 78% yield over two steps. The characteristic NMR signals 5.29 (s, 1H, H-1″), 5.19 (s, 1H, H-1′), and 4.87 (d, J = 10.0 Hz, 1H, H-1) and 13 C NMR δ 99.4 (C-1′), 98.8 (C-1), and 98.7 (C-1″) ppm confirmed the formation of the α -linked trisaccharide diol 16 , exclusively.…”