Specific Recognition of β-Galactofuranose-Containing Glycans of Synthetic Neoglycoproteins by Sera of Chronic Chagas Disease Patients.

Alba L Montoya,Maria Paola Zago,Emily L Heydemann,Belkisyolé Alarcón De Noya,Eileni R Gil,Igor L Estevao,Igor C Almeida,Katja Michael,Bianca E Luna,Oscar Noya,Sohan R Jankuru,Cameron C Ellis

doi:10.3390/molecules27020411

Abstract

Chagas disease (CD) can be accurately diagnosed by detecting Trypanosoma cruzi in patients’ blood using polymerase chain reaction (PCR). However, parasite-derived biomarkers are of great interest for the serological diagnosis and early evaluation of chemotherapeutic efficacy when PCR may fail, owing to a blood parasite load below the method’s limit of detection. Previously, we focused on the detection of specific anti-α-galactopyranosyl (α-Gal) antibodies in chronic CD (CCD) patients elicited by α-Gal glycotopes copiously expressed on insect-derived and mammal-dwelling infective parasite stages. Nevertheless, these stages also abundantly express cell surface glycosylphosphatidylinositol (GPI)-anchored glycoproteins and glycoinositolphospholipids (GIPLs) bearing nonreducing terminal β-galactofuranosyl (β-Galf) residues, which are equally foreign to humans and, therefore, highly immunogenic. Here we report that CCD patients’ sera react specifically with synthetic β-Galf-containing glycans. We took a reversed immunoglycomics approach that entailed: (a) Synthesis of T. cruzi GIPL-derived Galfβ1,3Manpα-(CH2)3SH (glycan G29SH) and Galfβ1,3Manpα1,2-[Galfβ1,3]Manpα-(CH2)3SH (glycan G32SH); and (b) preparation of neoglycoproteins NGP29b and NGP32b, and their evaluation in a chemiluminescent immunoassay. Receiver-operating characteristic analysis revealed that NGP32b can distinguish CCD sera from sera of healthy individuals with 85.3% sensitivity and 100% specificity. This suggests that Galfβ1,3Manpα1,2-[Galfβ1,3]Manpα is an immunodominant glycotope and that NGP32b could potentially be used as a novel CCD biomarker.

Highlights

The protozoan parasite Trypanosoma cruzi is the causative agent of Chagas disease (CD), which is endemic throughout Latin America, where millions of people are affected
A negative polymerase chain reaction (PCR) test does not necessarily mean cure, as the concentration of circulating parasites may have decreased below the limit of detection (LOD) of the assay, and/or parasites may still reside in the tissues [5]
Using a reversed immunoglycomics approach, a combination of the chemical synthesis of suspected T. cruzi-derived β-Galf -containing glycotopes and their immunological evaluation with sera of chronic CD (CCD) patients led to the discovery of two novel BMKs for CD (i.e., NGP29b and NGP32b)

Summary

Introduction

The protozoan parasite Trypanosoma cruzi is the causative agent of Chagas disease (CD), which is endemic throughout Latin America, where millions of people are affected. CD can be accurately diagnosed by detecting parasite DNA in patient’s blood using polymerase chain reaction (PCR) [4]. A negative PCR test does not necessarily mean cure, as the concentration of circulating parasites may have decreased below the limit of detection (LOD) of the assay, and/or parasites may still reside in the tissues [5]. Detection of anti-parasitic antibodies (Abs) in chronic CD (CCD) may be more suitable for evaluating treatment efficacy because the continuous stimulation of the immune system even by very few bloodstream or tissue-residing parasites, or circulating T. cruzi-derived antigens results in high levels of Ab titers [6]

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Results

Conclusion