Synthesis of α-l-rhamnosyl ceramide and evaluation of its binding with anti-rhamnose antibodies

Abstract

An α-L-rhamnosyl ceramide (1, α-L-RhaCer) has been prepared that was recognized by anti-L-rhamnose (anti-Rha) antibodies. During these studies we explored the use of an α-L-rhamnosyl thioglycoside and a trichloroacetimidate as a glycosyl donors. Subsequently, the acceptors desired for glycosylation, 3-O-benzoylazidosphingosine or 3-O-alloxycarbonylsphingosine, were prepared from D-xylose. The thioglycoside donor, 2,3,4-tri-O-acetyl-1-(4-tolyl)thio-α-L-rhamnopyranoside, and the trichloroacetimidate donor, 2,3,4-tri-O-acety-1-(2,2,2-trichloroethanimidate)-α-L-rhamnopyranoside, were synthesized in 50% and 78% yield overall, respectively. The synthesis of the glycosylation acceptor employed an addition-fragmentation olefination that was successfully carried out in 53% yield. With the successful synthesis of key intermediates, α-L-RhaCer (1) was prepared without any insurmountable obstacles. Anti-Rha antibodies were prepared in BALB/c mice by immunizing them with rhamnose-Ficoll with Sigma Adjuvant System (SAS) and the anti-L-Rha antibodies were isolated from the blood sera. Liposomes and EL4 tumor cells were used as model systems to demonstrate the ability of 1 to insert into a lipid bilayer. The interaction of the liposomes or the EL4 cells with α-L-RhaCer (1) and anti-Rha antibodies were investigated by fluorescence microscopy and flow cytometry, respectively, to confirm the ability of glycolipid 1 to be displayed on the tumor cell surface as well as the ability to be recognized by anti-Rha antibodies.

Synthesis of a Liposomal MUC1 Glycopeptide-Based Immunotherapeutic and Evaluation of the Effect of l-Rhamnose Targeting on Cellular Immune Responses

“…9, 50, 51 In fact, Rha is now one of the most effective natural antibody eliciting molecules known 52 and Rha has been conjugated to lipids to recruit anti-Rha antibodies to tumor cells. 53, 54 …”

Highly efficient chemoenzymatic synthesis and facile purification of α-Gal pentasaccharyl ceramide Galα3nLc4βCer

“…After removal of all benzoyl protecting groups by the Zemplén condition, several methods were tested to reduce the azido group while keep the alkene group intact. Methods using PPh 3 , 36 PMe 3 , 37 or Lindlar catalyst 38 led to poor yields (30–50%). The combination of 1,3-propanedithiol and triethylamine 39 were found to be the most efficient for selective reduction of the azido group to produce LacβSph ( 5 ) in an excellent yield (94%).…”

Why Do We Produce Anti-Gal

“…Galα1-3Glc and Galα1-3Gal epitopes were reported on both gram-positive and gram-negative bacteria (Han et al, 2012;Lüderitz et al, 1965). Additional examples of such antibodies in humans are anti-blood group A and B antibodies (Springer and Horton, 1969), natural antibody to N-glycolylneuraminic acid (called anti-Neu5Gc), which is produced in humans, and not in other Old World primates, or in nonprimate mammals (Higashi et al, 1977;Merrick et al, 1978;Zhu and Hurst, 2002;Padler-Karavani et al, 2008), and natural anti-rhamnose antibody (Chen et al, 2011;Sheridan et al, 2014;Long et al, 2014). As detailed in Chapter 3, the reason for the ability of anti-bacterial carbohydrate antibodies to bind mammalian carbohydrate antigens is that these antibodies are polyclonal, and different clones are capable of binding to various "facets" of a given carbohydrate antigen.…”