Emory University

Time From Onset to Diagnosis of Alpha-Gal Syndrome.

Alpha-Gal (Mammalian Meat) Allergy: Implications for Dermatologic Surgeons.

Unique repertoire of anti-carbohydrate antibodies in individual human serum.

Humoral immunity to pathogens and other environmental challenges is paramount to maintain normal health, and individuals lacking or unable to make antibodies are at risk. Recent studies indicate that many human protective antibodies are against carbohydrate antigens; however, little is known about repertoires and individual variation of anti-carbohydrate antibodies in healthy individuals. Here we analyzed anti-carbohydrate antibody repertoires (ACARs) of 105 healthy individual adult donors, aged 20-60 from different ethnic backgrounds to explore variations in antibodies, as defined by binding to glycan microarrays and by affinity purification. Using microarrays that contained > 1,000 glycans, including antigens from animal cells and microbes, we profiled the IgG and IgM ACARs from all donors. Each donor expressed many ACAs, but had a relatively unique ACAR, which included unanticipated antibodies to carbohydrate antigens not well studied, such as chitin oligosaccharides, Forssman-related antigens, globo-type antigens, and bacterial glycans. We also saw some expected antibodies to ABO(H) blood group and α-Gal-type antigens, although these also varied among individuals. Analysis suggests differences in ACARs are associated with ethnicity and age. Thus, each individual ACAR is relatively unique, suggesting that individualized information could be useful in precision medicine for predicting and monitoring immune health and resistance to disease.

Identification of the binding roles of terminal and internal glycan epitopes using enzymatically synthesized N-glycans containing tandem epitopes.

Glycans play diverse roles in a wide range of biological processes. Research on glycan-binding events is essential for learning their biological and pathological functions. However, the functions of terminal and internal glycan epitopes exhibited during binding with glycan-binding proteins (GBPs) and/or viruses need to be further identified. Therefore, a focused library of 36 biantennary asparagine (Asn)-linked glycans with some presenting tandem glycan epitopes was synthesized via a combined Core Isolation/Enzymatic Extension (CIEE) and one-pot multienzyme (OPME) synthetic strategy. These N-glycans include those containing a terminal sialyl N-acetyllactosamine (LacNAc), sialyl Lewis x (sLe) and Siaα2-8-Siaα2-3/6-R structures with N-acetylneuraminic acid (Neu5Ac) or N-glycolylneuraminic acid (Neu5Gc) sialic acid form, LacNAc, Lewis x (Le), α-Gal, and Galα1-3-Le; and tandem epitopes including α-Gal, Le, Galα1-3-Le, LacNAc, and sialyl LacNAc, presented with an internal sialyl LacNAc or 1-2 repeats of an internal LacNAc or Le component. They were synthesized in milligram-scale, purified to over 98% purity, and used to prepare a glycan microarray. Binding studies using selected plant lectins, antibodies, and viruses demonstrated, for the first time, that when interpreting the binding between glycans and GBPs/viruses, not only the structure of the terminal glycan epitopes, but also the internal epitopes and/or modifications of terminal epitopes needs to be taken into account.

Epidermal growth factor receptor inhibition augments the expression of MHC class I and II genes.

Diverse immune-related effects occur with the use of epidermal growth factor receptor inhibitors (EGFRI). In addition to the cutaneous inflammation induced by EGFRIs, these agents have been associated with the exacerbation of autoimmune skin disease and contact hypersensitivity, antiviral effects, and fatal alveolar damage in the setting of lung transplantation. Because EGFR ligands can modulate MHC class I (MHCI) and II (MHCII) molecule expression, we hypothesized that some of the immune-related effects of EGFRIs are due to direct effects on the expression of MHCI and/or MHCII molecules. Primary human keratinocytes and a malignant keratinocyte cell line (A431) were treated with EGFRIs alone or prior to IFN-γ, a potent inducer of MHCI and MHCII molecule expression. CIITA, MHCI, and MHCII RNA expression was measured using quantitative real-time reverse transcriptase PCR, and cell surface MHCI and MHCII protein expression was measured using flow cytometry. Skin biopsies from patients were analyzed for MHCI and MHCII protein expression before and during therapy with an EGFRI using immunohistochemistry. Both EGFR tyrosine kinase inhibitors and ligand-blocking antibodies (cetuximab) augmented the induction of MHCI and MHCII molecules by IFN-γ in primary and malignant human keratinocytes. Unexpectedly, the increase in MHCI protein expression did not require the presence of IFN-γ. Consistent with these in vitro findings, skin biopsies from cancer patients exhibited increased epidermal MHCI protein expression during therapy with an EGFRI as well as increases in MHCI and MHCII molecule RNA. These studies suggest that EGFRIs may influence immune/inflammatory responses by directly modulating MHC expression. Clin Cancer Res; 17(13); 4400-13. ©2011 AACR.

Vaccination-induced IgG response to Galalpha1-3GalNAc glycan epitopes in lambs protected against Haemonchus contortus challenge infection.

Lambs vaccinated with Haemonchus contortus excretory/secretory (ES) glycoproteins in combination with the adjuvant Alhydrogel are protected against H. contortus challenge infection. Using glycan micro-array analysis we showed that serum from such vaccinated lambs contains IgG antibodies that recognise the glycan antigen Galalpha1-3GalNAc-R and GalNAcbeta1-4(Fucalpha1-3)GlcNAc-R. Our studies revealed that H. contortus glycoproteins contain Galalpha1-3Gal-R as well as significant levels of Galalpha1-3GalNAc-R, which has not been previously reported. Extracts from H. contortus adult worms contain a galactosyltransferase acting on glycan substrates with a terminal GalNAc, indicating that the worms possess the enzymatic potential to synthesise terminal Gal-GalNAc moieties. These data illustrate that glycan micro-arrays constitute a promising technology for fast and specific analysis of serum anti-glycan antibodies in vaccination studies. In addition, this approach facilitates the discovery of novel, antigenic parasite glycan antigens that may have potential for developing glycoconjugate vaccines or utilization in diagnostics.

Phase II multicenter study of the epidermal growth factor receptor antibody cetuximab and cisplatin for recurrent and refractory squamous cell carcinoma of the head and neck.

This multicenter phase II study was undertaken to define the efficacy and safety of cetuximab, an antiepidermal growth factor receptor chimeric human and murine monoclonal antibody, administered with cisplatin to patients with refractory metastatic or recurrent squamous cell carcinoma of the head and neck (SCCHN). One hundred thirty-two patients were to receive two 3-week cycles with cisplatin/paclitaxel or cisplatin/fluorouracil. Patients (n = 30) with a complete or partial response continued standard therapy. Seventy-six patients with stable disease (SD; n = 51) or progressive disease (PD/1; n = 25) received combination therapy with cetuximab (400 mg/m2 intravenously on day 1, then 250 mg/m2/wk) and cisplatin (75 or 100 mg/m2 intravenously on day 1 every 3 weeks). The protocol was subsequently amended to enroll patients who had developed PD within 90 days after platinum-based therapy (PD/2; n = 54). Five patients (20%) in PD/1, three patients (6%) in PD/2, and nine patients (18%) with SD achieved an objective response. Median duration of response was 4.2, 4.1, and 7.4 months for the PD/1, PD/2, and SD groups, respectively, with median overall survival times of 6.1, 4.3, and 11.7 months. The most common toxicities were anemia, acne-like skin rash, leukopenia, fatigue and malaise, and nausea and vomiting. Seven patients (5%) developed a grade 3 or 4 hypersensitivity reaction to cetuximab. Cetuximab and cisplatin is an active regimen in refractory SCCHN. The relative contribution of cetuximab is better defined in a single-agent trial. Cetuximab did not exacerbate cisplatin toxicity but was associated with skin rash in a majority of patients and occasional serious allergic reactions. Further study of this compound is warranted.

Alpha-Gal Syndrome: A New Etiology for Periorbital Edema.

Alpha-gal syndrome is characterized by an IgE-mediated hypersensitivity response to the carbohydrate galactose-alpha-1,3-galactose. It can manifest as both an immediate drug allergy to pharmaceuticals containing alpha-gal as well as a delayed hypersensitivity response to the ingestion of mammalian meat products. In the United States, sensitization to alpha-gal is thought to occur from tick bites and can cause severe angioedema, most commonly periorbitally and periorally, with high rates of anaphylaxis. We present a unique case of chronic periorbital edema in a carnivorous individual after acquiring a new red meat allergy from Alpha-Gal Syndrome.