Miller MJ, Lee P, Lee BG, Flegel WA, West-Mitchell K, et al.
Transfusion •
The U.S. Centers for Disease Control and Prevention (CDC) has reported increasing rates of alpha-gal syndrome, an allergic response after meat ingestion (AGS). AGS has been associated with prior exposure to tick bites or other biologics characterized by a life-threatening immunoglobulin E (IgE)-mediated hypersensitivity to galactose-alpha-1,3-galactose (alpha-gal) an oligosaccharide structurally similar to the group B antigen on red blood cells (RBC) found in most non-primate mammalian meat and products derived from these mammals. In 2023, Transfusion reported 3 group O recipients of group B plasma in the Washington, D.C. metropolitan area with no history of meat allergy who had anaphylactic transfusion reactions compatible with AGS. We investigated allergic reactions in 2 additional patients who received ABO minor-incompatible blood products at 2 hospitals in the D.C. area during fall 2023. For both patients, a medical chart review was performed and IgE levels to alpha-gal were measured. The first patient, a 64-year-old, O-positive patient status post heart transplant with no known allergies, was admitted with acute COVID-19 induced antibody-mediated transplant rejection and placed on extracorporeal membrane oxygenation (ECMO). While undergoing plasma exchange (PLEX) (50% albumin/50% fresh frozen plasma (FFP)), the patient tolerated 2 units of group O FFP and 1 unit of group A FFP before becoming hemodynamically unstable during transfusion of 1 unit of B-positive FFP. PLEX was stopped. The patient later died of sepsis from underlying causes. The second patient, a 57-year-old O-positive man with a history of melanoma and neuro fibromatosis type 1, was undergoing an abdominal resection including transfusion of 3 units of O-positive RBC when he suffered hypotension and ventricular tachycardia requiring intraoperative code after receiving 2 units of group B FFP. Hiveswere noted after resuscitation. The patient had a history of tick bites but no known allergies. He is alive 5 months after the possible allergic event. Both patients had full transfusion reaction evaluations and immunology testing results above the positive cutoff for anti-alpha-gal IgE. Two patients with O-positive blood and no known allergies experience danaphyl axis after transfusion with group B FFP. The symptoms cannot definitively be imputed to an allergic transfusion reaction, but the presence of IgE against alpha-gal supports an association. Medicating patients with antihistamines and IV steroids pre-transfusion may prevent allergic reactions. Restricting group B plasma-containing products (plasma, platelets, cryoprecipitate) for patients who experience AGS-like symptoms may be considered.
Biochemical and biophysical research communications •
The N-linked oligosaccharides of cobra venom factor (CVF) contain unique terminal alpha-galactosylated Lewis X structures. We have previously shown that CVF immobilized on nylon membranes binds naturally occurring human anti-alpha-Gal antibody. The present study shows that soluble CVF can effectively inhibit the binding of anti-alpha-Gal antibody to CVF-coated microtiter plates, indicating that the terminal alpha-galactosyl residues of the functionally active CVF are accessible to anti-alpha-Gal antibody binding. Modification of the terminal galactosyl residues of CVF by treatment with galactose oxidase and in situ derivatization of the generated aldehyde groups with hydrazides abolished the human anti-alpha-Gal antibody immunoreactivity without affecting the complement-activating activity.
Cobra venom factor (CVF) is the nontoxic C-activating glycoprotein in cobra venom. It is a structural and functional analogue of complement component C3. Previous work has established that the major oligosaccharide chain of CVF is a symmetric fucosylated biantennary complex-type N-linked chain containing an alpha-galactosylated Le(x) antigenic epitope, Gal alpha 1-3Gal beta 1-4 (Fuc alpha 1-3) GlcNAc beta 1, a novel carbohydrate structure. We show that naturally occurring anti-alpha-Gal Ab present in normal human serum binds to CVF. In view of a possible human application of CVF, we studied the effect of this anti-alpha-Gal Ab on CVF function as well as the effect of oligosaccharide modification or removal on CVF activity and immunoreactivity with the anti-alpha-Gal Ab. The immunoreactivity of CVF with the anti-alpha-Gal Ab is abolished upon de-alpha-galactosylation or complete deglycosylation. De-alpha-galactosylated CVF and deglycosylated CVF were found to be equally active in forming a stable C3/C5 convertase with human factor B and in decomplementing human serum indicating that the oligosaccharide chains of CVF are not required for its C-activating function. Consistent with the unaltered functional activity, deglycosylation of the molecule caused only minor changes in secondary structure as judged by far UV circular dichroism analysis. However, the oligosaccharide chains appear to contribute to the thermal stability of CVF, because deglycosylation caused the molecule to be more sensitive to temperature. Inasmuch as rCVF produced in mammalian cells can be expected to contain sialylated oligosaccharide chains, we also prepared sialylated CVF by enzymatic sialylation of de-alpha-galactosylated and de-alpha-fucosylated CVF with 2,6-sialyltransferase. It was found that the secondary structure and the activity of sialylated CVF were unchanged compared to native CVF.
A high molecular weight, heavily glycosylated protein fraction was isolated from cobra venom. It consists of mucin-like glycoproteins (designated as cobra venom mucin) in noncovalent association with several lower molecular weight proteins and glycoproteins. The mucin was purified by CsBr density gradient centrifugation under dissociative conditions. The purified venom mucin comprised about 85% carbohydrate and 15% protein and was rich in Thr, Ser, Pro, Gly, Glu, Asp, and Ala. The mucin was resolved into two or more distinct classes of mucin-like glycoproteins which differ in their amino acid compositions and/or carbohydrate content. Unlike other mucins, cobra venom mucin does not form highly viscous solutions. It appears to keep several venom proteins and glycoproteins soluble by noncovalent interactions. Cobra venom mucin contains both O- and N-linked oligosaccharides; 1 N-linked chain for every 8-10 O-linked oligosaccharides. The O-linked chains are novel structures with high molar proportions of fucose, galactose, and N-acetylglucosamine relative to N-acetylgalactosamine; they have a very low sialic acid content and lack sulfate esters. The majority of the O-linked oligosaccharides are unusually large and contain 15 to as many as 50 sugar residues. The O-linked oligosaccharides are poly-N-acetyllactosaminyl chains consisting of -3Gal beta 1-4GlcNAc beta 1- and -3Gal beta 1-4(Fuc alpha 1-3)GlcNAc beta 1- repeats and thus they contain inner Le(X) antigenic determinants. These oligosaccharides terminate with novel alpha-galactosylated Le(X) and Le(a) epitopes. Due to the abundance of terminal alpha-galactosyl residues, cobra venom mucin reacts with anti-alpha-Gal antibodies that are normally present in human serum.
