One of the most consistent epidemiological associations between diet and human disease risk is the impact of consuming red meat and processed meat products. In recent years, the health concerns ...surrounding red meat and processed meat have gained worldwide attention. The fact that humans have lost the ability to synthesize N‐glycolylneuraminic acid (Neu5Gc) makes red meat and processed meat products the most important source of exogenous Neu5Gc for humans. As our research of Neu5Gc has increased, it has been discovered that Neu5Gc in red meat and processed meat is a key factor in many major diseases. Given the objective evidence of the harmful risk caused by Neu5Gc in red meat and processed meat to human health, there is a need for heightened attention in the field of food. This updated review has several Neu5Gc aspects given including biosynthetic pathway of Neu5Gc and its accumulation in the human body, the distribution of Neu5Gc in food, the methods for detecting Neu5Gc, and most importantly, a systematic review of the existing methods for reducing the content of Neu5Gc in red meat and processed meat. It also provides some insights into the current status and future directions in this area.
The emerging field of biotherapeutics provides successful treatments for various diseases, yet immunogenicity and limited efficacy remain major concerns for many products. Glycosylation is a key ...factor determining the pharmacological properties of biotherapeutics, including their stability, solubility, bioavailability, pharmacokinetics, and immunogenicity. Hence, an increased attention is directed at optimizing the glycosylation properties of biotherapeutics. Currently, most biotherapeutics are produced in non-human mammalian cells in light of their ability to produce human-like glycosylation. However, most mammals produce the sialic acid
-glycolylneuraminic acid (Neu5Gc), while humans cannot due to a specific genetic defect. Humans consume Neu5Gc in their diet from mammalian derived foods (red meat and dairy) and produce polyclonal antibodies against diverse Neu5Gc-glycans. Moreover, Neu5Gc can metabolically incorporate into human cells and become presented on surface or secreted glycans, glycoproteins, and glycolipids. Several studies in mice suggested that the combination of Neu5Gc-containing epitopes and anti-Neu5Gc antibodies could contribute to exacerbation of chronic inflammation-mediated diseases (e.g., cancer, cardiovascular diseases, and autoimmunity). This could potentially become complicated with exposure to Neu5Gc-containing biotherapeutics, bio-devices or xenografts. Indeed, Neu5Gc can be found on various approved and marketed biotherapeutics. Here, we provide a perspective review on the possible consequences of Neu5Gc glycosylation of therapeutic protein drugs due to the limited published evidence of Neu5Gc glycosylation on marketed biotherapeutics and studies on their putative effects on immunogenicity, drug efficacy, and safety.
This study compared the concentrations, types and distributions of sialic acid (SA) in human milk at different stages of the postnatal period with those in a range of infant formulas. Breast milk ...from mothers of healthy, full-term and exclusively breastfed infants was collected on the 2nd (
246), 7th (
135), 30th (
85) and 90th (
48) day after birth. The SA profiles of human milk, including their distribution, were analysed and compared with twenty-four different infant formulas. Outcome of this observational study was the result of natural exposure. Only SA of type Neu5Ac was detected in human milk. Total SA concentrations were highest in colostrum and reduced significantly over the next 3 months. Approximately 68·7–76·1 % of all SA in human milk were bound to oligosaccharides. Two types of SA, Neu5Ac and Neu5Gc, have been detected in infant formulas. Most SA was present in infant formulas combined with protein. Breastfed infants could receive more SA than formula-fed infants with the same energy intake. Overall, human milk is a preferable source of SA than infant formulas in terms of total SA content, dynamics, distribution and type. These SA profiles in the natural state are worth to be considered by the production of formulas because they may have a great effect on infant nutrition and development.
Entry of influenza A viruses (IAVs) into host cells is initiated by binding to sialic acids (Sias), their primary host cell receptor, followed by endocytosis and membrane fusion to release the viral ...genome into the cytoplasm of the host cell. Host tropism is affected by these entry processes, with a primary factor being receptor specificity. Sias exist in several different chemical forms, including the hydroxylated N-glycolylneuraminic acid (Neu5Gc), which is found in many hosts; however, it has not been clear how modified Sias affect viral binding and entry. Neu5Gc is commonly found in many natural influenza hosts, including pigs and horses, but not in humans or ferrets. Here, we engineered HEK293 cells to express the hydoxylase gene (CMAH) that converts Neu5Ac to Neu5Gc, or knocked out the Sia-CMP transport gene (SLC35A1), resulting in cells that express 95% Neu5Gc or minimal level of Sias, respectively. H3N2 (X-31) showed significantly reduced infectivity in Neu5Gc-rich cells compared to wild-type HEK293 (>95% Neu5Ac). To determine the effects on binding and fusion, we generated supported lipid bilayers (SLBs) derived from the plasma membranes of these cells and carried out single particle microscopy. H3N2 (X-31) exhibited decreased binding to Neu5Gc-containing SLBs, but no significant difference in H3N2 (X-31)'s fusion kinetics to either SLB type, suggesting that reduced receptor binding does not affect subsequent membrane fusion. This finding suggests that for this virus to adapt to host cells rich in Neu5Gc, only receptor affinity changes are required without further adaptation of virus fusion machinery.
Influenza A virus (IAV) infections continue to threaten human health, causing over 300,000 deaths yearly. IAV infection is initiated by the binding of influenza glycoprotein hemagglutinin (HA) to host cell sialic acids (Sias) and the subsequent viral-host membrane fusion. Generally, human IAVs preferentially bind to the Sia N-acetylneuraminic acid (Neu5Ac). Yet, other mammalian hosts, including pigs, express diverse nonhuman Sias, including N-glycolylneuraminic acid (Neu5Gc). The role of Neu5Gc in human IAV infections in those hosts is not well-understood, and the variant form may play a role in incidents of cross-species transmission and emergence of new epidemic variants. Therefore, it is important to investigate how human IAVs interact with Neu5Ac and Neu5Gc. Here, we use membrane platforms that mimic the host cell surface to examine receptor binding and membrane fusion events of human IAV H3N2. Our findings improve the understanding of viral entry mechanisms that can affect host tropism and virus evolution.
N-glycolylneuraminic acid (Neu5Gc), a sialic acid predominantly found in the non-neurohumoral fluids of hind-mouthed animals, is incapable of synthesizing Neu5Gc due to a deletion in the CMAH exon of ...the gene encoding human CMP-Neu5Gc hydroxylase. But consumption of animal-derived foods that contain Neu5Gc, such as red meat, can instigate an immune response in humans, as Neu5Gc is recognized as a foreign substance by the human immune system. This recognition leads to the production of anti-Neu5Gc antibodies, subsequently resulting in chronic inflammation. When Neu5Gc is consumed excessively or frequently, it may contribute to the development of heart disease and cancer. This makes Neu5Gc, an endogenous pathogenic factor derived from red meat, a new hot topic in red meat safety research. In this study, aptamers obtained by the magnetic bead SELEX technique were subjected to homology and secondary structure prediction analysis as well as affinity determination. The result indicated that the aptamer 2B.N2A9 exhibited a robust binding affinity, with an affinity constant (Ka) of 1.87 × 10
L/mol. This aptamer demonstrated optimal binding specificity within a pH range of 5.4 to 7.4. Molecular docking analysis further revealed that aptamer 2B.N2A9 formed stable binding interactions with the target Neu5Gc at specific sites, namely G-14, C-15, G-13, G-58, G-60, and C-59. An Enzyme-Linked Oligonucleotide Sorbent Assay (ELOSA) methodology was established to detect the endogenous pathogenic factor Neu5Gc present in red meat. This method demonstrated a limit of detection (LOD) of 0.71 ng/mL, along with an average recovery rate of 92.23%. The aptamer obtained in this study exhibited favorable binding properties to Neu5Gc. The assay was relatively convenient and demonstrated good sensitivity. Further investigation into the distribution of Neu5Gc in various red meats is of public health significance and scientific potential. A practical detection method should be provided to guide red meat diets and ensure the nutrition and safety of meat products.
Seventy to ninety percentage of preformed xenoreactive antibodies in human serum bind to the galactose-α(1,3)-galactose Gal epitope, and the creation of Gal knockout (KO) pigs has eliminated ...hyperacute rejection as a barrier to xenotransplantation. Now other glycan antigens are barriers to move ahead with xenotransplantation, and the N-glycolyl neuraminic acid, Neu5Gc (or Hanganutziu-Deicher antigen), is also a major pig xenoantigen. Humans have anti-Neu5Gc antibodies. Several data indicate a strong immunogenicity of Neu5Gc in humans that may contribute to an important part in antibody-dependent injury to pig xenografts. Pig islets express Neu5Gc, which reacted with diet-derived human antibodies and mice deleted for Neu5Gc reject pancreatic islets from wild-type counterpart. However, Neu5Gc positive heart were not rejected in Neu5Gc KO mice indicating that the role of Neu5Gc-specific antibodies has to be nuanced and depend of the graft situation parameters (organ/tissue, recipient, implication of other glycan antigens). Recently generated Gal/Neu5Gc KO pigs eliminate the expression of Gal and Neu5Gc, and improve the crossmatch of humans with the pig. This review summarizes the current and recent experimental and (pre)clinical data on the Neu5Gc immunogenicity and emphasize of the potential impact of anti-Neu5Gc antibodies in limiting xenotransplantation in humans.
Normal human tissues do not express glycans terminating with the sialic acid N-glycolylneuraminic acid (Neu5Gc), yet Neu5Gc-containing glycans have been consistently found in human tumor tissues, ...cells and secretions and have been proposed as a cancer biomarker. We engineered a Neu5Gc-specific lectin called SubB2M, and previously reported elevated Neu5Gc biomarkers in serum from ovarian cancer patients using a Surface Plasmon Resonance (SPR)-based assay. Here we report an optimized SubB2M SPR-based assay and use this new assay to analyse sera from breast cancer patients for Neu5Gc levels.
To enhance specificity of our SPR-based assay, we included a non-sialic acid binding version of SubB, SubB
, to control for any non-specific binding to SubB2M, which improved discrimination of cancer-free controls from early-stage ovarian cancer. We analysed 96 serum samples from breast cancer patients at all stages of disease compared to 22 cancer-free controls using our optimized SubB2M-
-SPR assay. We also analysed a collection of serum samples collected at 6 monthly intervals from breast cancer patients at high risk for disease recurrence or spread.
Analysis of sera from breast cancer cases revealed significantly elevated levels of Neu5Gc biomarkers at all stages of breast cancer. We show that Neu5Gc serum biomarker levels can discriminate breast cancer patients from cancer-free individuals with 98.96% sensitivity and 100% specificity. Analysis of serum collected prospectively, post-diagnosis, from breast cancer patients at high risk for disease recurrence showed a trend for a decrease in Neu5Gc levels immediately following treatment for those in remission.
Neu5Gc serum biomarkers are a promising new tool for early detection and disease monitoring for breast cancer that may complement current imaging- and biopsy-based approaches.
Background
Clinical xenotransplantation is not possible because humans possess antibodies that recognize antigens on the surface of pig cells. Galα‐1,3‐Gal (Gal) and N‐glycolylneuraminic acid ...(Neu5Gc) are two known xenoantigens.
Methods
We report the homozygous disruption of the α1, 3‐galactosyltransferase (GGTA1) and the cytidine monophosphate‐N‐acetylneuraminic acid hydroxylase (CMAH) genes in liver‐derived female pig cells using zinc‐finger nucleases (ZFNs). Somatic cell nuclear transfer (SCNT) was used to produce healthy cloned piglets from the genetically modified liver cells. Antibody‐binding and antibody‐mediated complement‐dependent cytotoxicity assays were used to examine the immunoreactivity of pig cells deficient in Neu5Gc and Gal.
Results
This approach enabled rapid production of a pig strain deficient in multiple genes without extensive breeding protocols. Immune recognition studies showed that pigs lacking both CMAH and GGTA1 gene activities reduce the humoral barrier to xenotransplantation, further than pigs lacking only GGTA1.
Conclusions
This technology will accelerate the development of pigs for xenotransplantation research.
Human heterophile antibodies that agglutinate animal erythrocytes are known to detect the nonhuman sialic acid N-glycolylneuraminic acid (Neu5Gc). This monosaccharide cannot by itself fill the ...binding site (paratope) of an antibody and can also be modified and presented in various linkages, on diverse underlying glycans. Thus, we hypothesized that the human anti-Neu5Gc antibody response is diverse and polyclonal. Here, we use a novel set of natural and chemoenzymatically synthesized glycans to show that normal humans have an abundant and diverse spectrum of such anti-Neu5Gc antibodies, directed against a variety of Neu5Gc-containing epitopes. High sensitivity and specificity assays were achieved by using N-acetylneuraminic acid (Neu5Ac)-containing probes (differing from Neu5Gc by one less oxygen atom) as optimal background controls. The commonest anti-Neu5Gc antibodies are of the IgG class. Moreover, the range of reactivity and Ig classes of antibodies vary greatly amongst normal humans, with some individuals having remarkably large amounts, even surpassing levels of some well-known natural blood group and xenoreactive antibodies. We purified these anti-Neu5Gc antibodies from individual human sera using a newly developed affinity method and showed that they bind to wild-type but not Neu5Gc-deficient mouse tissues. Moreover, they bind back to human carcinomas that have accumulated Neu5Gc in vivo. As dietary Neu5Gc is primarily found in red meat and milk products, we suggest that this ongoing antigen-antibody reaction may generate chronic inflammation, possibly contributing to the high frequency of diet-related carcinomas and other diseases in humans.
Tick bite induced α‐gal syndrome (AGS) following consumption of mammalian meat is a recently described intriguing disease occurring worldwide. Here we argue that AGS and delayed allergy in general is ...an adaptive defence method against cancer. Our hypothesis synthesizes two lines of supporting evidence. First, allergy has been shown to have direct anti‐cancer effects with unknown mechanism. Second, eating processed meat was shown to be linked to developing cancer. Humans lost their genes encoding molecules α‐gal 30 MYA and Neu5Gc 2 MYA, the latter co‐occurring with the start of using fire. These molecules are acquired from external sources, as tick bite for α‐gal and mammalian meat for Neu5Gc, the latter accumulating in tumors. The resulting specific delayed allergic response is a molecular adaptation to fight cancer. By further testing and applying our hypothesis, new avenues in cancer research and therapy will open that might save lives and decrease human suffering.
Tick bite induced α‐gal syndrome (AGS) following consumption of mammalian meat is a recently described intriguing disease occurring worldwide. Here, we argue that AGS and delayed allergy in general is an adaptive defence method against cancer. Our hypothesis might open new avenues in cancer research and therapy.
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