Leishmaniasis is an infectious vector-borne disease caused by a number of different species of the protozoan parasite Leishmania. There are different forms of leishmaniasis, the most common being cutaneous leishmaniasis (CL), which is associated with large, open skin ulcers that may heal over many months, but can lead to complications due to secondary infections. CL affects 12 million people worldwide, particularly in Northern Africa, the Middle East, Asia, and in Latin America. Mostly due to military deployment to endemic regions (e.g., Afghanistan and Iraq), CL is also on the rise in the U.S. Proper diagnostic tools (PCR) are often not available in developing countries, and the presentation of CL can be easily confused with other skin conditions. Even though a number of drugs ? most of which are highly toxic? are available to treat CL, there is no perfect drug with equal efficacy for all CL-causing Leishmania species. For example, ketoconazole is effective for CL caused by L. major, but it is almost entirely ineffective against L. tropica. Thus, there is a need not only for reliable diagnostic tools for the accurate diagnosis of CL, but also for the differentiation between CL infections by different Leishmania species, which is important to determine drug treatment regimens. Leishmania spp. express unusual glycans on their cell surfaces that are foreign to humans, which may elicit a strong antibody (IgG) response. Specifically, it is known that the glycoinositol phospholipids (GIPLs) of L. major contain unusual ?-galactopyranosyl (?Gal) and ?-galactofuranosyl (?Galf)-residues. Likewise, L. tropica expresses lipophosphoglycans (LPGs) that contain ?-arabinose (?Ara), suggesting that L. major and L. tropica could be serologically distinguishable. Our preliminary data show that the reactivities of sera from CL patients against certain synthetic ?Gal and/or ?Galf epitopes differ depending on the Leishmania species that causes the underlying CL infection. However, some antibody cross-reactivities are also observed. Whitesides and others have demonstrated that the density at which glycan antigens are presented has a major influence on antibody recognition and specificity. We hypothesize that ?Gal- and ?Galf-containing glycans that exist in the GIPLs of L. major, and ?Ara- containing glycans that exist in the LPGs of L. tropica, cause a strong IgG response in CL patients. We further hypothesize that varying the densities at which certain immunodominant glycotopes are displayed will identify conditions at which these glycans show large differentials in enzyme-linked immunosorbent assay (CL-ELISA) between CL patients and the control group, and between CL patients with L. major and L. tropica infections.
The specific aims are to synthesize ?Gal-, ?Galf-, or ?Ara containing partial structures of the cell surface glycans of L. major and L. tropica, and to identify those that are strongly recognized by antibodies in the sera of CL patients that are infected with L. major or L. tropica using CL-ELISA. The most promising glycans will be subjected to CL-ELISA at different glycan densities. The goal is to identify conditions under which a glycan shows large differentials between CL patients and the control group, and also between CL patients with L. major or L. tropica infection. Can the newly discovered CL biomarkers indeed achieve a differential diagnosis between L. major and L. tropica infections? Their specificity will be put to a test by screening the individual sera of a large cohort of CL patients with an active L. major or L. tropica infection by CL-ELISA. !
The proposed work aims at discovering glycan biomarkers for the disease cutaneous leishmaniasis, which is caused by a number of parasites of the genus Leishmania and causes large skin ulcers that heal over months and sometimes years. Biomarkers are needed for a proper diagnosis of the disease, which can easily be confused with other skin conditions. Even though several drugs are available to treat this disease, there is no drug that is equally effective for all Leishmania species. Therefore, a differential diagnosis is needed to determine drug treatment regimens. The glycan biomarkers to be discovered are partial structures of cell surface glycans of two different Leishmania species (L. major and L. tropica) that may elicit antibodies in patients. Optimized immunoassays may be developed to differentiate between infections with L. major and L. tropica.
