ABSTRACT: Optimization of Lead BKIs for Cryptosporidiosis Therapy
The broad, long-term objective of this research is to develop a therapeutic for treatment of Cryptosporidium
infection. Cryptosporidium infection causes persistent diarrhea (cryptosporidiosis) that is associated with
increased morbidity and mortality in children and immunocompromised individuals. Cryptosporidium is one of
the most important pathogens leading to poor outcomes in <2-yo malnourished children in resource poor
countries, including >3-fold mortality and strong associations with stunting and impaired neurological
development. The only available therapeutic, nitazoxanide, does not work in immunocompromised individuals
and has <30% efficacy in malnourished children. New therapeutics for Cryptosporidium are badly needed. We
have been developing bumped-kinase inhibitors (BKIs) that selectively target Cryptosporidium calcium-
dependent protein kinase 1 (CDPK1) as therapeutics for cryptosporidiosis. In our program, our leads have
performed very well, demonstrating efficacy at low oral dosages in mouse, calf, and piglet models of C. parvum
and C. hominis, while retaining almost all the favorable safety aspects consistent with a late lead. However,
safety issues with BKI leads have stopped us from developing current late leads. We now understand the
safety issues associated with late BKI leads and have found these safety issues are not inextricably associated
with the structure-activity relationship (SAR) of BKIs’ efficacy against cryptosporidiosis. In this proposal, we will
use the efficacy and safety SAR to help drive medicinal chemistry towards a late pre-clinical lead with safety,
pharmacokinetic, and efficacy properties that can be developed for treatment in the target population of <2-yo
malnourished children and immunocompromised individuals. The Aims are: AIM 1) Establish late leads for
cryptosporidiosis therapy, including Subaims 1A) In vitro efficacy and safety testing, 1B) IFN- ¿ -KO mouse
nLuc-C. parvum efficacy testing, 1C) Mouse multidosing, 7d safety testing, 1D) Determine metabolites of BKIs,
and, 1E) Design and synthesize new BKIs; AIM 2) Test novel late leads in calf clinical model for
cryptosporidiosis; and, AIM 3) Assess advanced late leads for late safety testing, including subaims 3A) Bone
safety testing, 3B) Pregnancy/developmental/fetal safety testing; 3C) Rat and dog cardiovascular (CV) safety
testing, and, 3D) Pre-GLP safety and polymorph testing. At the end of the grant period, we expect to have a
preclinical lead and at least one back up molecule that is ready to move into GLP safety testing, pre-GMP
manufacturing scale up, and IND filing to move towards human trials for a BKI for therapy of cryptosporidiosis.