In Vitro and In Vivo Effects of Soluble, Monovalent Globotriose on Bacterial Attachment and Colonization

Abstract

Epithelial cells lining the urinary tract are rich in globo series glycolipids, structurally defined by a Gal␣1,4Gal motif in the oligosaccharide moiety of this glycolipid family. This Gal␣1,4Gal motif is the attachment target for the P-fimbrial adhesin of uropathogenic Escherichia coli. We investigated the ability of a trisaccharide analog of this core motif, globotriose (Gal␣1,4Gal␤1,4Glc), to interfere with uropathogen attachment and colonization in vitro and in vivo. We assessed the ability of globotriose to inhibit and reverse the binding and agglutination of a P-fimbriated strain of E. coli (JR1) using human erythrocytes and immortalized human colonic epithelial cells as targets. Globotriose (5 mg/ml) completely inhibited and reversed cell agglutination and caused a 10-to 100-fold reduction in JR1 binding to target cells, as determined by flow cytometry. In preparation for an in vivo efficacy study, we investigated the distribution and pharmacokinetics of globotriose in the BALB/c mouse. Globotriose was administered via the tail vein, targeting an instantaneous plasma concentration of 5 mg/ml, and in a different experiment, animals were gavaged at 10 times the intravenous (i.v.) dose. Globotriose was rapidly cleared from plasma (half-life [t 1/2 ], 6 min) and slowly excreted via the kidney (t 1/2 , 4 h). Urine levels of >5 mg/ml were maintained from 4 to 12 h after the i.v. bolus dose, which resulted in a 1-log reduction in established bladder colonization by JR1. These results suggest that free, soluble globotriose is a feasible alternative therapy for urinary tract infections.Glycolipids and other cell surface glycoconjugates play important roles in tissue development, cell trafficking and sorting, and cell-cell recognition. Independently of their inherent biological functions, cell surface glycolipids are used as targets by bacteria, viruses, and toxins to recognize and adhere to cells. Several groups have described and proposed the use of oligosaccharides to prevent or reverse pathological events occurring at the cell surface (7,14,15). This oligosaccharide receptor decoy concept is based on a rich body of experimental evidence demonstrating the importance of cell surface oligosaccharides in health and disease. Microbial lectins have been shown to have low affinities for their monovalent carbohydrate binding targets (4, 5), and carbohydrate targets of microbial lectins are normally present in a multivalent fashion on mammalian cell surfaces. The microbial lectins themselves are often displayed as multivalent complexes, or, as is the case with many microbial toxins, each contains a multimeric carbohydrate binding subunit. This multivalent presentation of binding targets and microbial lectins drives the equilibrium strongly in favor of the formation of target-ligand pairs. For these reasons, multivalent glycoconjugates have been favored for development as antiadhesive therapeutic candidates, despite the difficulties associated with the synthesis and the poor bioavailability of multivalent preparations.Re...

2′-fucosyllactose: an abundant, genetically determined soluble glycan present in human milk

“…123,127 There are limited data on the absorption, distribution, metabolism, and excretion of non-digestible soluble glycans, although a published study demonstrates that a trisaccharide is quickly cleared from the plasma into a depot compartment and that the bulk is cleared precisely through the urine. 128 This was true for intravenous and oral administration of the synthetic soluble glycan globotriose. The prebiotic effect attributed to milk oligosaccharides is consistent with the fact that these glycans can be found in the feces of breastfed infants, thus demonstrating that they are not digestible in vivo.…”

“…These researchers designed a set of in vivo studies to investigate the fate of milk oligosaccharides by providing a lactating mother with a bolus of 13 C‐Gal and measuring the appearance of 13 C‐ labeled HMO in the urine of her breastfed infant . There are limited data on the absorption, distribution, metabolism, and excretion of non‐digestible soluble glycans, although a published study demonstrates that a trisaccharide is quickly cleared from the plasma into a depot compartment and that the bulk is cleared precisely through the urine . This was true for intravenous and oral administration of the synthetic soluble glycan globotriose.…”

TLR4 Facilitates Translocation of Bacteria across Renal Collecting Duct Cells

“…Preincubation of the cells with the type 1 fimbriae receptor analogue D-mannose 10 induced significant inhibition of invasion by CFT073, HT7, and HT91 (45.8 -55.4%; Figure 1C). Globotriose, shown to reverse binding of P fimbriated UPEC to target cells, 29,30 also inhibited in an additive manner with mannose the ability of HT7, CFT073, and HT91 (data not shown) to invade cells ( Figure 1D). …”

Urinary Tract Infections and the Mucosal Immune System

“…Soluble Galα1-4Galβ-containing oligosaccharides present receptor epitopes to P fimbriae and act as soluble inhibitors of attachment to human uroepithelial cells Svanborg Eden et al, 1983;Kihlberg et al, 1989;Leach et al, 2005). Therapeutic efficacy has also been demonstrated in vivo .…”