The protozoan parasite Cryptosporidium has caused massive waterborne outbreaks in the U.S, including the 1993 Milwaukee outbreak, where ~400,000 individuals contracted disease, with an economic cost of $31.7 million in medical expenses and another $64.6 million in productivity losses. Highly concentrated Cryptosporidium samples can be prepared with modest effort and simple equipment, and the water supply is easily accessed, leading to its classification as a category B bio-warfare agent. Immunocompromised patients, pregnant women and the elderly are at risk of serious disease;infection can be chronic and fatal in AIDS patients. The tools to respond to such an incident are woefully inadequate: no vaccine or effective drug treatment is currently available. We have discovered that this eukaryotic pathogen has obtained numerous genes by horizontal transfer from bacteria. The enzymes encoded by these bacterial genes provide highly divergent targets for the design of parasite-specific drugs. One such enzyme is IMP dehydrogenase (IMPDH), which catalyzes a key step in guanine nucleotide biosynthesis. Using the RO1 funding mechanism, we have validated IMPDH as a target for parasite treatment and identified ten Principle Hits that selectively inhibit the parasite enzyme. Here we propose a medicinal chemistry program to optimize these compounds and identify a drug candidate with antiparasitic activity in a mouse model of infection: Aim 1: Assess principal hits and select four lead compounds for optimization. Aim 2: Optimize four Lead Series through a medicinal chemistry program. Aim 3: Characterize Advanced Leads in an animal model of Cryptosporidium infection and chose a drug candidate. Following selection of the Drug Candidate, and in work not included in this proposal, we will execute a Pre- Clinical Development Plan leading to submission of an IND application to the FDA. If successful, the work described in this proposal will develop an urgently needed drug for the management of cryptosporidiosis in epidemic outbreaks and AIDS patients that will be invaluable in the event of a bioterrorist attack.