Montoya AL, Carvajal EG, Ortega-Rodriguez U, Estevao IL, Ashmus RA, et al.
Molecules (Basel, Switzerland) ā¢
Chagas disease (CD) is caused by the parasite and affects 6-7 million people worldwide. The diagnosis is still challenging, due to extensive parasite diversity encompassing seven genotypes (TcI-VI and Tcbat) with diverse ecoepidemiological, biological, and pathological traits. Chemotherapeutic intervention is usually effective but associated with severe adverse events. The development of safer, more effective therapies is hampered by the lack of biomarker(s) (BMKs) for the early assessment of therapeutic outcomes. The mammal-dwelling trypomastigote parasite stage expresses glycosylphosphatidylinositol-anchored mucins (tGPI-MUC), whose -glycans are mostly branched with terminal, nonreducing Ī±-galactopyranosyl (Ī±-Gal) glycotopes. These are absent in humans, and thus highly immunogenic and inducers of specific CD anti-Ī±-Gal antibodies. In search for Ī±-Gal-based BMKs, here we describe the synthesis of neoglycoprotein NGP11b, comprised of a carrier protein decorated with the branched trisaccharide GalĪ±(1,2)[GalĪ±(1,6)]GalĪ². By chemiluminescent immunoassay using sera/plasma from chronic CD (CCD) patients from Venezuela and Mexico and healthy controls, NGP11b exhibited sensitivity and specificity similar to that of tGPI-MUC from genotype TcI, predominant in those countries. Preliminary evaluation of CCD patients subjected to chemotherapy showed a significant reduction in anti-Ī±-Gal antibody reactivity to NGP11b. Our data indicated that NGP11b is a potential BMK for diagnosis and treatment assessment in CCD patients.
Montoya AL, Gil ER, Heydemann EL, Estevao IL, Luna BE, et al.
Molecules (Basel, Switzerland) ā¢
Chagas disease (CD) can be accurately diagnosed by detecting 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-Ī±-galactopyranosyl (Ī±-Gal) antibodies in chronic CD (CCD) patients elicited by Ī±-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 Ī²-galactofuranosyl (Ī²-Gal) residues, which are equally foreign to humans and, therefore, highly immunogenic. Here we report that CCD patients' sera react specifically with synthetic Ī²-Gal-containing glycans. We took a reversed immunoglycomics approach that entailed: (a) Synthesis of GIPL-derived GalĪ²1,3ManĪ±-(CH)SH (glycan ) and GalĪ²1,3ManĪ±1,2-[GalĪ²1,3]ManĪ±-(CH)SH (glycan ); and (b) preparation of neoglycoproteins and , and their evaluation in a chemiluminescent immunoassay. Receiver-operating characteristic analysis revealed that can distinguish CCD sera from sera of healthy individuals with 85.3% sensitivity and 100% specificity. This suggests that GalĪ²1,3ManĪ±1,2-[GalĪ²1,3]ManĪ± is an immunodominant glycotope and that could potentially be used as a novel CCD biomarker.
Ortega-Rodriguez U, Portillo S, Ashmus RA, Duran JA, Schocker NS, et al.
Methods in molecular biology (Clifton, N.J.) ā¢
Chagas disease (ChD), caused by the protozoan parasite Trypanosoma cruzi, affects millions of people worldwide. Chemotherapy is restricted to two drugs, which are partially effective and may cause severe side effects, leading to cessation of treatment in a significant number of patients. Currently, there are no biomarkers to assess therapeutic efficacy of these drugs in the chronic stage. Moreover, no preventive or therapeutic vaccines are available. In this chapter, we describe the purification of Trypanosoma cruzi trypomastigote-derived glycosylphosphatidylinositol (GPI)-anchored mucins (tGPI-mucins) for their use as antigens for the reliable primary or confirmatory diagnosis and as prognostic biomarkers for early assessment of cure following ChD chemotherapy. We also describe, as an example, the synthesis of a potential tGPI-mucin-derived Ī±-Gal-terminating glycan and its coupling to a carrier protein for use as diagnostic and prognostic biomarker in ChD.
Pathobiology : journal of immunopathology, molecular and cellular biology ā¢
This study describes the presence of alpha-galactosyl epitopes on 12 cervical biopsy samples with features of human papillomavirus infection (HPV) and different stages of cervical intraepithelial neoplasia. An avidin-biotin-peroxidase assay with a monoclonal antibody recognizing gal(alpha 1-->3)gal residues was strongly positive in 5 of 12 cases. None of the controls stained (p = 0.02). Immunostaining was intense in the areas with the highest viral load (koilocytes and keratinocytes) and absent in malignant foci. Immunostaining was also absent in normal exo- and endocervical epithelium of 12 controls with no features of HPV infection. A faint background staining in cases and controls was evident, but similar. These initial findings suggest that alpha-galactosyl epitopes are expressed in cervical squamous cells infected with HPV, turning them vulnerable to lysis by natural anti-alpha-galactosyl antibodies.
