-glycolyl groups of nonhuman chondroitin sulfates survive in ancient fossils.

Biosynthesis of the common mammalian sialic acid -glycolylneuraminic acid (Neu5Gc) was lost during human evolution due to inactivation of the gene, possibly expediting divergence of the lineage, due to a partial fertility barrier. Neu5Gc catabolism generates -glycolylhexosamines, which are potential precursors for glycoconjugate biosynthesis. We carried out metabolic labeling experiments and studies of mice with human-like Neu5Gc deficiency to show that Neu5Gc degradation is the metabolic source of UDP-GlcNGc and UDP-GalNGc and the latter allows an unexpectedly selective incorporation of -glycolyl groups into chondroitin sulfate (CS) over other potential glycoconjugate products.

Novel anti-Sialyl-Tn monoclonal antibodies and antibody-drug conjugates demonstrate tumor specificity and anti-tumor activity.

Targeted therapeutics that can differentiate between normal and malignant tumor cells represent the ideal standard for the development of a successful anti-cancer strategy. The Sialyl-Thomsen-nouveau antigen (STn or Sialyl-Tn, also known as CD175s) is rarely seen in normal adult tissues, but it is abundantly expressed in many types of human epithelial cancers. We have identified novel antibodies that specifically target with high affinity the STn glycan independent of its carrier protein, affording the potential to recognize a wider array of cancer-specific sialylated proteins.

Production platforms for biotherapeutic glycoproteins. Occurrence, impact, and challenges of non-human sialylation.

One of the fastest growing fields in the pharmaceutical industry is the market for therapeutic glycoproteins. Today, these molecules play a major role in the treatment of various diseases, and include several protein classes, i.e.

Sexual selection by female immunity against paternal antigens can fix loss of function alleles.

Humans lack the common mammalian cell surface molecule N-glycolylneuraminic acid (Neu5Gc) due to a CMAH gene inactivation, which occurred approximately three million years ago. Modern humans produce antibodies specific for Neu5Gc. We hypothesized that anti-Neu5Gc antibodies could enter the female reproductive tract and target Neu5Gc-positive sperm or fetal tissues, reducing reproductive compatibility.

Human xeno-autoantibodies against a non-human sialic acid serve as novel serum biomarkers and immunotherapeutics in cancer.

Human carcinomas can metabolically incorporate and present the dietary non-human sialic acid Neu5Gc, which differs from the human sialic acid N-acetylneuraminic acid (Neu5Ac) by 1 oxygen atom. Tumor-associated Neu5Gc can interact with low levels of circulating anti-Neu5Gc antibodies, thereby facilitating tumor progression via chronic inflammation in a human-like Neu5Gc-deficient mouse model. Here we show that human anti-Neu5Gc antibodies can be affinity-purified in substantial amounts from clinically approved intravenous IgG (IVIG) and used at higher concentrations to suppress growth of the same Neu5Gc-expressing tumors.

Implications of the presence of N-glycolylneuraminic acid in recombinant therapeutic glycoproteins.

Recombinant glycoprotein therapeutics produced in nonhuman mammalian cell lines and/or with animal serum are often modified with the nonhuman sialic acid N-glycolylneuraminic acid (Neu5Gc; refs. 1,2). This documented contamination has generally been ignored in drug development because healthy individuals were not thought to react to Neu5Gc (ref.

Novel mechanism for the generation of human xeno-autoantibodies against the nonhuman sialic acid N-glycolylneuraminic acid.

The nonhuman sialic acid N-glycolylneuraminic acid (Neu5Gc) is metabolically incorporated into human tissues from certain mammalian-derived foods, and this occurs in the face of an anti-Neu5Gc "xeno-autoantibody" response. Given evidence that this process contributes to chronic inflammation in some diseases, it is important to understand when and how these antibodies are generated in humans. We show here that human anti-Neu5Gc antibodies appear during infancy and correlate with weaning and exposure to dietary Neu5Gc.

Sensitive and specific detection of the non-human sialic Acid N-glycolylneuraminic acid in human tissues and biotherapeutic products.

Background: Humans are genetically defective in synthesizing the common mammalian sialic acid N-glycolylneuraminic acid (Neu5Gc), but can metabolically incorporate it from dietary sources (particularly red meat and milk) into glycoproteins and glycolipids of human tumors, fetuses and some normal tissues. Metabolic incorporation of Neu5Gc from animal-derived cells and medium components also results in variable contamination of molecules and cells intended for human therapies. These Neu5Gc-incorporation phenomena are practically significant, because normal humans can have high levels of circulating anti-Neu5Gc antibodies.

Evidence for dynamic interplay of different oligomeric states of UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase by biophysical methods.

The bifunctional enzyme UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE) is a key enzyme for the biosynthesis of sialic acids, the terminal sugars of glycoconjugates associated with a variety of physiological and pathological processes such as cell adhesion, development, inflammation and cancer. In this study, we characterized rat GNE by different biophysical methods, analytical ultracentrifugation, dynamic light-scattering and size-exclusion chromatography, all revealing the native hydrodynamic behavior and molar mass of the protein. We show that GNE is able to reversibly self-associate into different oligomeric states including monomers, dimers and tetramers.

Influence of UDP-GlcNAc 2-epimerase/ManNAc kinase mutant proteins on hereditary inclusion body myopathy.

Hereditary inclusion body myopathy (HIBM), a neuromuscular disorder, is caused by mutations in UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE), the key enzyme of sialic acid biosynthesis. To date, more than 40 different mutations in the GNE gene have been reported to cause the disease. Ten of them, representing mutations in both functional domains of GNE, were recombinantly expressed in insect cells (Sf9).

UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase, functionally expressed in and purified from Escherichia coli, yeast, and insect cells.

UDP-GlcNAc 2-epimerase/ManNAc kinase is the key enzyme of sialic acid biosynthesis in mammals. Its functional expression is a prerequisite for early embryogenesis and for the synthesis of several cell recognition motifs in adult organism. This bifunctional enzyme is involved in the development of different diseases like sialuria or hereditary inclusion body myopathy.

Identification of human L-fucose kinase amino acid sequence.

Fucose is a major component of complex carbohydrates. L-Fucose kinase (fucokinase) takes part in the salvage pathway for reutilization of fucose from the degradation of oligosaccharides. The amino acid sequence of human fucokinase was derived from a cDNA encoding a protein of hitherto unidentified function.