Antiadhesive Receptor Mimetics for Cryptosporidiosis

Cryptosporidium parvum is a ubiquitous protozoan parasite that causes severe diarrheal disease in both humans and animals. This organism was recently responsible for the largest outbreak of waterborne disease in U. S. history. This massive outbreak in Milwaukee, Wisconsin in 1993 resulted in infection of more than 400,000 persons with mortality rates in immunocompromised individuals reaching nearly 70 percent. Cryptosporidium has become the most important contaminant found in drinking water and cryptosporidiosis is now recognized as both a significant emerging public health concern and a threat to United States water supplies. There are no current vaccines or clinically proven chemotherapies that reduce parasite infectivity, replication in host cells or the ability of C. parvum to destroy the host's intestinal lining and precipitate life-threatening fluid loss in the immunocompromised patient.
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The goal of this application is to identify specific mechanisms responsible for parasite attachment to enterocytes, and to explore the use of natural enterocyte receptor molecules or receptor mimetics as potential competitive inhibitors to ameliorate this severe diarrheal disease. There is currently almost no information describing enterocyte surface molecules used by the parasite for attachment. Recently, we have been able to establish an in vitro binding assay that measures in vivo-relevant sporozoite binding to enterocytes. Using this assay, we have collected preliminary data suggesting complex carbohydrate molecules on host cell-surface membranes and mucins are required for initial sporozoite recognition and attachment. In addition, we have developed a novel, in vivo xenograft animal model system that can be used to routinely evaluate therapeutic efficacy of these receptors or their carbohydrate derivatives.
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Accordingly, for this application we will focus on characterization of the structure and mechanism of function of these receptors as inhibitors of parasite infectivity and the feasibility of using these molecules or their active carbohydrate moieties as anti-adhesive mimetics for the treatment of cryptosporidiosis.
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We will accomplish these goals with the following specific aims: <ol> <li>To characterize biochemically a specific cell-surface membrane receptor(s) exploited by sporozoites and merozoites for binding to enterocytes,
<li>To define the mechanism of mucin-mediated inhibition of sporozoite attachment, and
<li>To quantify the in vivo ability of the mucins and enterocyte receptor(s) or their carbohydrate derivatives to inhibit C. parvum infection and resultant diarrheal disease.</ol>