Regioisomeric SCFA Attachment to Hexosamines Separates Metabolic Flux from Cytotoxicity and MUC1 Suppression

Abstract

Chemical biology studies, exemplified by metabolic glycoengineering experiments that employ short chain fatty acid (SCFA)-hexosamine monosaccharide hybrid molecules, often suffer from off-target effects. Here we demonstrate that systematic structure-activity relationship (SAR) studies can deconvolute multiple biological activities of SCFA-hexosamine analogues by demonstrating that triacylated monosaccharides, including both n-butyrate- and acetate-modified ManNAc analogues, had dramatically different activities depending on whether the free hydroxyl group was at the C1 or C6 position. The C1-OH (hemiacetal) analogues enhanced growth inhibition in MDA-MB-231 human breast cancer cells and suppressed expression of MUC1, which are attractive properties for an anticancer agent. By contrast, C6-OH analogues supported high metabolic flux into the sialic acid pathway with negligible growth inhibition or toxicity, which are desirable properties for glycan labeling in healthy cells. Importantly, these SAR were general, applying to other hexosamines ( e.g., GlcNAc) and non-natural sugar "scaffolds" ( e.g., ManNLev). From a practical standpoint, the ability to separate toxicity from flux will facilitate the use of MOE analogues for cancer treatment and glycomics applications, respectively. Mechanistically, these findings overturn the premise that the bioactivities of SCFA-monosaccharide hybrid molecules result from their hydrolysis products ( e.g., n-butyrate, which acts as a histone deacetylase inhibitor, and ManNAc, which activates sialic acid biosynthesis); instead the SAR establish that inherent properties of partially acylated hexosamines supersede the cellular responses supported by either the acyl or monosaccharide moieties.

Metabolic Flux Increases Glycoprotein Sialylation: Implications for Cell Adhesion and Cancer Metastasis

“…CHO cells stably transfected with full-length E-selectin (CHO-E) or with phosphatidylinositol glycan-linked extracellular domain of P-selectin (CHO-P) were kindly donated by Affymax (Palo Alto, CA) and cultured as described previously (8). Synthesis of 1,3,4-O-Bu 3 ManNAc followed a previously reported procedure (9).…”

Ester Bonds in Prodrugs

“…An example is the area of "metabolic glycoengineering", where endogenous sugar-processing pathways are exploited to incorporate non-natural sugars into glycans for targeting, labeling, and other useful biological experiments (1). The paper by Yarema and co-workers on page 230 of this issue describes structureϪ activity relationship (SAR) studies on acylated carbohydrates for use in metabolic glycoengineering (2). The data indicate that the position of acyl groups affects strongly the biological activity of the carbohydrate.…”

“…The difference in activity between sugar derivatives 2 and 3 suggests that these synthetic ester bonds are not hydrolyzed immediately to yield free ManNAc. Instead, ester moieties must endure inside cells and influence biological activity (2).…”

Short-Chain Fatty Acid-Modified Hexosamine for Tissue-Engineering Osteoarthritic Cartilage

“…Both molecules were synthesized and characterized based on the previously reported methods. 12 3,4,6-O-Bu 3 GlcNAc was synthesized from 2-acetamido-1,3,4,6-tetra-O-butanoyl-2-deoxy-a,b-D-glucopyranose (1,3,4,6-O-Bu 4 GlcNAc) 13 using one-step selective deacylation reaction. Briefly, a mixture of butyric anhydride (15.6 mmol) and 4-(dimethylamino) pyridine (cat.)…”