The alpha-gal story: Lessons learned from connecting the dots
Authors: John W. Steinke, Thomas A.E. Platts-Mills, Scott P. Commins
Journal: Journal of Allergy and Clinical Immunology (2015)

Anaphylaxis is a severe allergic reaction that can be rapidly progressing and fatal, and therefore establishing its cause is pivotal to long-term risk management. Our recent work has identified a novel IgE antibody response to a mammalian oligosaccharide epitope, galactose-alpha-1,3-galactose (alpha-gal). IgE to alpha-gal has been associated with 2 distinct forms of anaphylaxis: (1) immediate-onset anaphylaxis during first exposure to intravenous cetuximab and (2) delayed-onset anaphylaxis 3 to 6 hours after ingestion of mammalian food products (eg, beef and pork). Results of our studies and those of others strongly suggest that tick bites are a cause, if not the only significant cause, of IgE antibody responses to alpha-gal in the southern, eastern, and central United States; Europe; Australia; and parts of Asia. Typical immune responses to carbohydrates are considered to be T-cell independent, whereas IgE antibody production is thought to involve sequential class-switching that requires input from T cells. Therefore, establishing the mechanism of the specific IgE antibody response to alpha-gal will be an important aspect to address as this area of research continues. Anaphylaxis is a severe allergic reaction that can be rapidly progressing and fatal, and therefore establishing its cause is pivotal to long-term risk management. Our recent work has identified a novel IgE antibody response to a mammalian oligosaccharide epitope, galactose-alpha-1,3-galactose (alpha-gal). IgE to alpha-gal has been associated with 2 distinct forms of anaphylaxis: (1) immediate-onset anaphylaxis during first exposure to intravenous cetuximab and (2) delayed-onset anaphylaxis 3 to 6 hours after ingestion of mammalian food products (eg, beef and pork). Results of our studies and those of others strongly suggest that tick bites are a cause, if not the only significant cause, of IgE antibody responses to alpha-gal in the southern, eastern, and central United States; Europe; Australia; and parts of Asia. Typical immune responses to carbohydrates are considered to be T-cell independent, whereas IgE antibody production is thought to involve sequential class-switching that requires input from T cells. Therefore, establishing the mechanism of the specific IgE antibody response to alpha-gal will be an important aspect to address as this area of research continues. Information For Category 1 CME CreditCredit can now be obtained, free for a limited time, by reading the review articles in this issue. Please note the following instructions.Method of Physician Participation in Learning Process: The core material for these activities can be read in this issue of the Journal or online at the JACI Web site: www.jacionline.org. The accompanying tests may only be submitted online at www.jacionline.org. Fax or other copies will not be accepted.Date of Original Release: March 2015. Credit may be obtained for these courses until February 29, 2016.Copyright Statement: Copyright © 2015-2016. All rights reserved.Overall Purpose/Goal: To provide excellent reviews on key aspects of allergic disease to those who research, treat, or manage allergic disease.Target Audience: Physicians and researchers within the field of allergic disease.Accreditation/Provider Statements and Credit Designation: The American Academy of Allergy, Asthma & Immunology (AAAAI) is accredited by the Accreditation Council for Continuing Medical Education (ACCME) to provide continuing medical education for physicians. The AAAAI designates this journal-based CME activity for a maximum of 1 AMA PRA Category 1 Credit™. Physicians should claim only the credit commensurate with the extent of their participation in the activity.List of Design Committee Members: John W. Steinke, PhD, Thomas A. E. Platts-Mills, MD, PhD, FRS, and Scott P. Commins, MD, PhDDisclosure of Significant Relationships with Relevant Commercial Companies/Organizations: J. W. Steinke has received research support from Genentech (HSR 17123), has received a lecture honorarium from Northwestern, and receives royalties for human mast cell line developed. T. A. E. Platts-Mills has received research support from the National Institutes of Health (NIH; AI-20565)/National Institute of Allergy and Infectious Diseases (NIAID; AI-085190), has received travel support from Phadia/Thermo Fisher, and receives royalties from IBT/Viracor. S. P. Commins has received research support from the NIH/NIAID (AI-20565; AI-085190; grant K08), has received consultancy fees from Sanofi, and receives royalties from UpToDate.Activity Objectives1.To describe the history and presenting symptoms of galactose-alpha-1,3-galactose (alpha-gal).2.To describe the pathophysiology of the IgE-mediated response to alpha-gal in human subjects.3.To list the diagnostic approach to alpha-gal.Recognition of Commercial Support: This CME activity has not received external commercial support.List of CME Exam Authors: Jill Hanson, MD, Alison Humphrey, MD, David Jara, MD, Neha Patel, MD, and Paul Dowling, MDDisclosure of Significant Relationships with Relevant Commercial Companies/Organizations: The exam authors disclosed no relevant financial relationships. Credit can now be obtained, free for a limited time, by reading the review articles in this issue. Please note the following instructions. Method of Physician Participation in Learning Process: The core material for these activities can be read in this issue of the Journal or online at the JACI Web site: www.jacionline.org. The accompanying tests may only be submitted online at www.jacionline.org. Fax or other copies will not be accepted. Date of Original Release: March 2015. Credit may be obtained for these courses until February 29, 2016. Copyright Statement: Copyright © 2015-2016. All rights reserved. Overall Purpose/Goal: To provide excellent reviews on key aspects of allergic disease to those who research, treat, or manage allergic disease. Target Audience: Physicians and researchers within the field of allergic disease. Accreditation/Provider Statements and Credit Designation: The American Academy of Allergy, Asthma & Immunology (AAAAI) is accredited by the Accreditation Council for Continuing Medical Education (ACCME) to provide continuing medical education for physicians. The AAAAI designates this journal-based CME activity for a maximum of 1 AMA PRA Category 1 Credit™. Physicians should claim only the credit commensurate with the extent of their participation in the activity. List of Design Committee Members: John W. Steinke, PhD, Thomas A. E. Platts-Mills, MD, PhD, FRS, and Scott P. Commins, MD, PhD Disclosure of Significant Relationships with Relevant Commercial Companies/Organizations: J. W. Steinke has received research support from Genentech (HSR 17123), has received a lecture honorarium from Northwestern, and receives royalties for human mast cell line developed. T. A. E. Platts-Mills has received research support from the National Institutes of Health (NIH; AI-20565)/National Institute of Allergy and Infectious Diseases (NIAID; AI-085190), has received travel support from Phadia/Thermo Fisher, and receives royalties from IBT/Viracor. S. P. Commins has received research support from the NIH/NIAID (AI-20565; AI-085190; grant K08), has received consultancy fees from Sanofi, and receives royalties from UpToDate. Activity Objectives1.To describe the history and presenting symptoms of galactose-alpha-1,3-galactose (alpha-gal).2.To describe the pathophysiology of the IgE-mediated response to alpha-gal in human subjects.3.To list the diagnostic approach to alpha-gal. Recognition of Commercial Support: This CME activity has not received external commercial support. List of CME Exam Authors: Jill Hanson, MD, Alison Humphrey, MD, David Jara, MD, Neha Patel, MD, and Paul Dowling, MD Disclosure of Significant Relationships with Relevant Commercial Companies/Organizations: The exam authors disclosed no relevant financial relationships. Discuss this article on the JACI Journal Club blog: www.jaci-online.blogspot.com. Hypersensitivity in the allergic setting refers to immune reactions stimulated by soluble antigens that can be rapidly progressing and, in the case of anaphylaxis, are occasionally fatal. Because the number of known exposures associated with anaphylaxis is limited, identification of novel causative agents is important in facilitating both education and other allergen-specific approaches that are crucial to long-term risk management. Within the last 10 years, several seemingly separate observations were recognized as related, all of which resulted from the development of antibodies to a carbohydrate moiety on proteins in which exposure differed from airborne allergens but that were nevertheless capable of producing anaphylactic and hypersensitivity reactions. Our recent work has identified these responses as being due to a novel IgE antibody directed against a mammalian oligosaccharide epitope, galactose-alpha-1,3-galactose (alpha-gal).1Chung C.H. Mirakhur B. Chan E. Le Q.T. Berlin J. Morse M. et al.Cetuximab-induced anaphylaxis and IgE specific for galactose-alpha-1,3-galactose.N Engl J Med. 2008; 358: 1109-1117Crossref PubMed Scopus (1180) Google Scholar This review will present the history and biology of alpha-gal and discuss the evidence that the IgE response to alpha-gal is different from typical IgE responses directed toward protein allergens. In 2004, ImClone and Bristol-Myers Squibb were investigating an mAb (cetuximab) specific for the epidermal growth factor receptor in clinical trials for the treatment of metastatic colorectal cancer. Early in those studies, it became clear that the antibody was causing hypersensitivity reactions; however, these reactions were occurring primarily in a group of southern US states (Table I). These reactions to cetuximab developed rapidly, and symptoms often peaked within 20 minutes after or during the first infusion of the antibody and occasionally proved fatal.1Chung C.H. Mirakhur B. Chan E. Le Q.T. Berlin J. Morse M. et al.Cetuximab-induced anaphylaxis and IgE specific for galactose-alpha-1,3-galactose.N Engl J Med. 2008; 358: 1109-1117Crossref PubMed Scopus (1180) Google Scholar, 2O'Neil B.H. Allen R. Spigel D.R. Stinchcombe T.E. Moore D.T. Berlin J.D. et al.High incidence of cetuximab-related infusion reactions in Tennessee and North Carolina and the association with atopic history.J Clin Oncol. 2007; 25: 3644-3648Crossref PubMed Scopus (283) Google Scholar, 3Maier S. Chung C.H. Morse M. Platts-Mills T. Townes L. Mukhopadhyay P. et al.A retrospective analysis of cross-reacting cetuximab IgE antibody and its association with severe infusion reactions.Cancer Med. 2015; 4: 36-42Crossref PubMed Scopus (31) Google Scholar Because of delays in marketing, it was not until 2006 that the true severity of the reactions became obvious.2O'Neil B.H. Allen R. Spigel D.R. Stinchcombe T.E. Moore D.T. Berlin J.D. et al.High incidence of cetuximab-related infusion reactions in Tennessee and North Carolina and the association with atopic history.J Clin Oncol. 2007; 25: 3644-3648Crossref PubMed Scopus (283) Google ScholarTable ITime course of the alpha-gal storyYear: Events leading to our understanding of red meat allergy∼2000: At least 2 groups reported cases of meat allergy that started after tick bites.2003: IgE to cat allergens is common in an African village but not related to symptoms.2005: There are reports of hypersensitivity reactions to first infusion of cetuximab in clinical trials.2007: Severe reactions to cetuximab are common in Tennessee, North Carolina, Arkansas, Missouri, and Virginia.2007: Two cases in Virginia of adult-onset delayed anaphylaxis occurring 3 to 6 hours after eating beef are reported.2008: Alpha-gal is identified as the epitope on cetuximab.2009: Twenty-four cases of delayed anaphylaxis to red meat are found in the United States.Multiple cases of meat allergy after tick bites are reported in Sydney, Australia.2010: There is a range of evidence that ticks are responsible for the IgE response in the United States.2011: There is extensive evidence that the IgE response is not related to asthma, despite cross-reactions with dog and cat.2014: Open challenge tests confirm the delay in reactions to red meat. Open table in a new tab At this time, our group began preliminary experiments examining the IgE response to this molecule. Dr Hatley, who was working in Bentonville, Arkansas, convinced our group to develop a new version of the IgE fluorometric enzyme immunoassay or CAP assay to cetuximab using the streptavidin technique. In this assay streptavidin is coupled to the solid phase of the CAP to provide a matrix for the binding of biotinylated novel or purified allergens.4Erwin E.A. Custis N.J. Satinover S.M. Perzanowski M.S. Woodfolk J.A. Crane J. et al.Quantitative measurement of IgE antibodies to purified allergens using streptavidin linked to a high-capacity solid phase.J Allergy Clin Immunol. 2005; 115: 1029-1035Abstract Full Text Full Text PDF PubMed Scopus (89) Google Scholar We were subsequently asked to investigate the reactions to cetuximab, in part because we had already developed the IgE assay to cetuximab. In collaboration with Dr Chung from Nashville, Dr Mirakhur from Bristol-Myers Squibb, and Dr Hicklin from ImClone, we demonstrated that the patients who had reactions to cetuximab also had IgE antibodies specific for this molecule before they started treatment.1Chung C.H. Mirakhur B. Chan E. Le Q.T. Berlin J. Morse M. et al.Cetuximab-induced anaphylaxis and IgE specific for galactose-alpha-1,3-galactose.N Engl J Med. 2008; 358: 1109-1117Crossref PubMed Scopus (1180) Google Scholar The question remained as to what epitope the IgE antibody was recognizing on the cetuximab molecule. Early work by Karl Landsteiner discovered that all human subjects had antibodies to a blood group "B-like" oligosaccharide found on nonprimate red blood cells.5Landsteiner K. The specificity of serological reactions. Charles C Thomas, Baltimore (MD)1936Google Scholar That antigen was subsequently identified as alpha-gal, which represents a major transplantation barrier between primates and other mammals.6Milland J. Sandrin M.S. ABO blood group and related antigens, natural antibodies and transplantation.Tissue Antigens. 2006; 68: 459-466Crossref PubMed Scopus (73) Google Scholar, 7Koike C. Uddin M. Wildman D.E. Gray E.A. Trucco M. Starzl T.E. et al.Functionally important glycosyltransferase gain and loss during catarrhine primate emergence.Proc Natl Acad Sci U S A. 2007; 104: 559-564Crossref PubMed Scopus (66) Google Scholar, 8Macher B.A. Galili U. The Galalpha1,3Galbeta1,4GlcNAc-R (alpha-Gal) epitope: a carbohydrate of unique evolution and clinical relevance.Biochim Biophys Acta. 2008; 1780: 75-88Crossref PubMed Scopus (323) Google Scholar Antibodies against alpha-gal are present in all nonimmunocompromised human subjects, and some early studies suggested that the IgG antibodies against alpha-gal constituted about 1% of circulating immunoglobulins in human subjects, apes, and Old World monkeys.9Galili U. Rachmilewitz E.A. Peleg A. Flechner I. A unique natural human IgG antibody with anti-alpha-galactosyl specificity.J Exp Med. 1984; 160: 1519-1531Crossref PubMed Scopus (582) Google Scholar Recent work in our laboratory with specific assays for IgG antibodies suggests that the percentages are not this high. As discussed below, the fact that all nonprimate mammals, including mice, can make oligosaccharides that are foreign to human subjects is an important component of our story. Glycosylation of proteins is a posttranscriptional modification that can play key roles in many processes, including protein folding, protein stability, intracellular trafficking, and cellular adhesion, as reviewed by Hurtado-Guerrero and Davies.10Hurtado-Guerrero R. Davies G.J. Recent structural and mechanistic insights into post-translational enzymatic glycosylation.Curr Opin Chem Biol. 2012; 16: 479-487Crossref PubMed Scopus (25) Google Scholar Characterization of cetuximab glycosylation, as measured based on peak area on time-of-flight mass spectrometric spectra, revealed 21 distinct oligosaccharide structures, of which approximately 30% have 1 or more alpha-1,3–linked galactosyl residues.11Qian J. Liu T. Yang L. Daus A. Crowley R. Zhou Q. Structural characterization of N-linked oligosaccharides on monoclonal antibody cetuximab by the combination of orthogonal matrix-assisted laser desorption/ionization hybrid quadrupole-quadrupole time-of-flight tandem mass spectrometry and sequential enzymatic digestion.Anal Biochem. 2007; 364: 8-18Crossref PubMed Scopus (242) Google Scholar Analysis of the IgE antibodies to cetuximab demonstrated that these antibodies were specific for the oligosaccharide residues on the heavy chain of the Fab portion of the mAb. From the known glycosylation of the molecule at amino acids 88 and 299 (Fig 1),11Qian J. Liu T. Yang L. Daus A. Crowley R. Zhou Q. Structural characterization of N-linked oligosaccharides on monoclonal antibody cetuximab by the combination of orthogonal matrix-assisted laser desorption/ionization hybrid quadrupole-quadrupole time-of-flight tandem mass spectrometry and sequential enzymatic digestion.Anal Biochem. 2007; 364: 8-18Crossref PubMed Scopus (242) Google Scholar alpha-gal was identified as the relevant epitope. Of the total alpha-gal in cetuximab, most of it is located in the Fab domain (Fab domain: 990 nmol alpha-gal/μmol IgG vs Fc domain: 140 nmol alpha-gal/μmol IgG).11Qian J. Liu T. Yang L. Daus A. Crowley R. Zhou Q. Structural characterization of N-linked oligosaccharides on monoclonal antibody cetuximab by the combination of orthogonal matrix-assisted laser desorption/ionization hybrid quadrupole-quadrupole time-of-flight tandem mass spectrometry and sequential enzymatic digestion.Anal Biochem. 2007; 364: 8-18Crossref PubMed Scopus (242) Google Scholar Recent mass spectrometric analysis indicates that glycosylation of cetuximab might be more complex than previously thought, containing both dianternary and trianternary structures.12Ayoub D. Jabs W. Resemann A. Evers W. Evans C. Main L. et al.Correct primary structure assessment and extensive glyco-profiling of cetuximab by a combination of intact, middle-up, middle-down and bottom-up ESI and MALDI mass spectrometry techniques.mAbs. 2013; 5: 699-710Crossref PubMed Scopus (150) Google Scholar Synthesis of alpha-gal requires the gene encoding alpha-1,3-galactosyltransferase. In human subjects and higher primates this gene is not functional, and therefore these species cannot produce alpha-gal, which in turn makes it possible for these animals to initially make IgG and IgM antibodies directed toward this oligosaccharide.7Koike C. Uddin M. Wildman D.E. Gray E.A. Trucco M. Starzl T.E. et al.Functionally important glycosyltransferase gain and loss during catarrhine primate emergence.Proc Natl Acad Sci U S A. 2007; 104: 559-564Crossref PubMed Scopus (66) Google Scholar, 13Galili U. The alpha-gal epitope and the anti-Gal antibody in xenotransplantation and in cancer immunotherapy.Immunol Cell Biol. 2005; 83: 674-686Crossref PubMed Scopus (280) Google Scholar How IgE to alpha-gal gets made and the nature of the IgE response will be considered later. Of considerable importance to the development of biologics, in particular mAbs, is the observation that murine cell lines, such as NS0 and Sp2/0, can synthesize galactose in an alpha-1,3 linkage such that alpha-gal is present on the molecules. Sp2/0 was the cell line used to produce cetuximab. In those subjects with IgE to alpha-gal (≥0.35 IU/mL), reactions are likely to occur directed against this mAb.3Maier S. Chung C.H. Morse M. Platts-Mills T. Townes L. Mukhopadhyay P. et al.A retrospective analysis of cross-reacting cetuximab IgE antibody and its association with severe infusion reactions.Cancer Med. 2015; 4: 36-42Crossref PubMed Scopus (31) Google Scholar During this same time period (2006-2008), we evaluated a number of patients, most of whom spent a significant amount of time outdoors, who had presented with episodes of generalized urticaria, angioedema, or recurrent anaphylaxis. The importance of the time spent outdoors was not clear at that time. There was no obvious immediate cause for the symptoms, but in several cases the patients reported that they believed the reactions might be due to consumption of meat 3 to 5 hours earlier. Skin prick tests were performed with commercial extracts of beef, pork, or lamb and produced small wheals only 2 to 4 mm in diameter that often would be interpreted as negative results. However, given the compelling history described by the patients, we extended our analysis to intradermal skin testing with commercial meat extracts or skin prick tests with fresh meat extracts, both of which demonstrated strong positive results.14Commins S.P. Satinover S.M. Hosen J. Mozena J. Borish L. Lewis B.D. et al.Delayed anaphylaxis, angioedema, or urticaria after consumption of red meat in patients with IgE antibodies specific for galactose-alpha-1,3-galactose.J Allergy Clin Immunol. 2009; 123: 426-433Abstract Full Text Full Text PDF PubMed Scopus (514) Google Scholar These results were confirmed with blood tests for specific IgE antibody to red meats.14Commins S.P. Satinover S.M. Hosen J. Mozena J. Borish L. Lewis B.D. et al.Delayed anaphylaxis, angioedema, or urticaria after consumption of red meat in patients with IgE antibodies specific for galactose-alpha-1,3-galactose.J Allergy Clin Immunol. 2009; 123: 426-433Abstract Full Text Full Text PDF PubMed Scopus (514) Google Scholar Although not published, similar sensitivity to red meat had been previously noted in Georgia. Starting in 1989, Mrs Sandra Latimer, together with Dr Antony Deutsch from Athens, Georgia, collected 10 cases of delayed reactions to mammalian meat and made a connection with the occurrence of tick bites several weeks or months before the first episode of hives or anaphylaxis. They presented these findings to the Georgia Allergy Society and to the US Centers for Disease Control and Prevention in 1991, but no additional reports or statements were issued by either of these organizations. The characteristics of red meat allergy are different from typical allergic reactions. Common complaints include both gastrointestinal symptoms and urticaria, but unlike most allergic reactions, patients do not have any symptoms for at least 2 hours after eating red meat, whereas many reactions are delayed for 3 to 5 hours or even longer. Nonetheless, symptoms can be severe or even life-threatening. Many of the patients described nausea, diarrhea, or indigestion before a reaction; however, the most common symptom reported was itching. The presence of symptoms before a severe reaction is common but not a requirement. Many patients do not have any symptoms, and symptoms do not occur with every exposure to red meat even in those who have had them previously. All of the patients had consumed red meat without complications for many years before the onset of the syndrome. Although some patients had a prior history of allergy, most of them had no previous allergic symptoms, and thus an atopic disposition does not appear to predispose patients to this kind of IgE response. Three observations led us to investigate whether IgE antibodies to alpha-gal were present in the sera of adult patients reporting reactions to beef. Alpha-gal is known to be present on both tissues and meat from nonprimate mammals,15Thall A. Galili U. Distribution of Gal alpha 1–3Gal beta 1–4GlcNAc residues on secreted mammalian glycoproteins (thyroglobulin, fibrinogen, and immunoglobulin G) as measured by a sensitive solid-phase radioimmunoassay.Biochemistry. 1990; 29: 3959-3965Crossref PubMed Scopus (114) Google Scholar the antibodies causing reactions to cetuximab were directed against alpha-gal, and the geographic distribution of the reactions to cetuximab overlapped the same geographic area where the red meat–induced reactions were occurring. Not surprisingly, the patients' sera had positive results for IgE to beef, pork, lamb, cat, and dog but not to nonmammalian meat, such as turkey, fish, or chicken.14Commins S.P. Satinover S.M. Hosen J. Mozena J. Borish L. Lewis B.D. et al.Delayed anaphylaxis, angioedema, or urticaria after consumption of red meat in patients with IgE antibodies specific for galactose-alpha-1,3-galactose.J Allergy Clin Immunol. 2009; 123: 426-433Abstract Full Text Full Text PDF PubMed Scopus (514) Google Scholar, 16Kennedy J.L. Stallings A.P. Platts-Mills T.A. Oliveira W.M. Workman L. James H.R. et al.Galactose-alpha-1,3-galactose and delayed anaphylaxis, angioedema, and urticaria in children.Pediatrics. 2013; 131: e1545-e1552Crossref PubMed Scopus (100) Google Scholar The presence of alpha-gal was confirmed by using 2 different absorption assays, one with alpha-gal on human serum albumin and the other with mammalian or beef thyroglobulin, which is heavily decorated with alpha-gal. The glycosylated antigens were bound to sepharose beads.14Commins S.P. Satinover S.M. Hosen J. Mozena J. Borish L. Lewis B.D. et al.Delayed anaphylaxis, angioedema, or urticaria after consumption of red meat in patients with IgE antibodies specific for galactose-alpha-1,3-galactose.J Allergy Clin Immunol. 2009; 123: 426-433Abstract Full Text Full Text PDF PubMed Scopus (514) Google Scholar In each case, levels of the specific IgE binding to beef, pork, lamb, cat, and dog were reduced by greater than 75%. More recently, evidence has been obtained from a study examining beef extracts using 2-diemnsional gel electrophoresis. The authors demonstrated 7 alpha-gal–containing IgE-binding proteins, 4 of which survived heating the beef extract.17Apostolovic D. Tran T. Hamsten C. Starkhammar M. Cirkovic Velickovic T. van Hage M. Immunoproteomics of processed beef proteins reveal novel galactose-alpha-1,3-galactose containing allergens.Allergy. 2014; 69: 1308-1315Crossref PubMed Scopus (54) Google Scholar How alpha-gal is structurally expressed on red meat remains unclear. Also unclear is whether differences in the structure exist and whether these differences affect IgE binding. The terminal carbohydrate residue on red meat is likely alpha-gal based on the binding of IgE from sera of patients with red meat allergy to cetuximab, which, as discussed, also has terminal alpha-gal residues. However, one can envision a difference in carbohydrate structure, such that only a single exposed alpha-gal–binding site is present in the oligosaccharide chain on meat, contrasting the 2 found predominately on cetuximab. The majority of alpha-gal found on cetuximab has a dianternary structure (Fig 1). The structure on meat has not been determined. Whether having 2 alpha-gal residues on the terminus of the carbohydrate structure has an effect on the strength of IgE binding is unknown. In 2008, as the specificity of the IgE antibodies to alpha-gal that caused reactions to cetuximab became clearer, the number of reports describing delayed reactions to red meat was also increasing. A relationship between mammalian meat allergy and tick bites had already been suggested in Australia18Van Nunen S.A. O'Connor K.S. Clarke L.R. Boyle R.X. Fernando S.L. An association between tick bite reactions and red meat allergy in humans.Med J Aust. 2009; 190: 510-511PubMed Google Scholar; however, the role of alpha-gal was not known, and the tick connection was not yet obvious in the United States. What caught our attention was that both cetuximab reactions and delayed reactions to red meat were being reported from the same region of the country, a group of southeastern states. However, it was not clear why these cases were geographically localized, and the only area that was comparable was the maximum incidence of Rocky Mountain spotted fever (RMSF). At this time, red meat allergy developed in 3 members of our group, and each one distinctly remembered being bitten by ticks weeks or months before the development of symptoms. Sera from these persons that had been obtained before the tick bite were compared with sera collected after the bite, and it was found that serum levels of IgE to alpha-gal had increased dramatically (4- to 10-fold). Following up on this connection, we started to ask patients about tick bites and rapidly became aware that most of those with delayed anaphylaxis had experienced recent bites from adult or larval ticks. Examination of US Centers for Disease Control and Prevention maps of the distribution of the tick Amblyomma americanum (lone star tick) revealed an overlap with the region of both cetuximab sensitivity and red meat allergy. Additional indications that tick bites are involved in the development of specific IgE to alpha-gal include histories of bites that have itched for 2 or more weeks, a significant correlation between IgE antibodies to alpha-gal and IgE to lone star tick (Fig 2), and prospective data on the increase in IgE levels to alpha-gal after known lone star tick bites.19Commins S.P. James H.R. Kelly L.A. Pochan S.L. Workman L.J. Perzanowski M.S. et al.The relevance of tick bites to the production of IgE antibodies to the mammalian oligosaccharide galactose-alpha-1,3-galactose.J Allergy Clin Immunol. 2011; 127: 1286-1293.e6Abstract Full Text Full Text PDF PubMed Scopus (437) Google Scholar Allergy to red meat is now being reported in other countries, but the ticks giving rise to this response are not the same species as in the United States. In Europe Ixodes ricinus has been implicated, whereas in Australia the relevant tick is Ixodes holocyclus.18Van Nunen S.A. O'Connor K.S. Clarke L.R. Boyle R.X. Fernando S.L. An association between tick bite reactions and red meat allergy in humans.Med J Aust. 2009; 190: 510-511PubMed Google Scholar, 20Hamsten C. Tran