The human malaria parasite Plasmodium falciparum imports and catabolizes massive amounts of host erythrocyte cytosol (mainly hemoglobin) during its asexual replication cycle. The mechanism and regulation of hemoglobin uptake, which occurs through a specialized protozoan structure termed the cytostome, is poorly understood. Here, we present new data which, together with prior published work, strongly suggests that the anti-malarial action of mefloquine (MQ) is closely associated with the inhibition of hemoglobin endocytosis. In this proposal, we will test the hypothesis that MQ binds to a protein target that plays a key role in the parasite?s endocytosis of host erythrocyte hemoglobin.
In Aim 1, we will synthesize a library of photoaffinity probes that are based on the MQ pharmacophore. Probe design will focus on the incorporation of ?all-in-one? moieties that enable the introduction of photoactivatable and affinity groups with minimal structural perturbation. Phenotypic and parasite viability assays will be employed to characterize probe potency and to verify retention of MQ?s mode of action.
In Aim 2, these probes will be used to irreversibly label the protein target of MQ. Specificity of labeling will be assessed by competition with unmodified MQ and the characterization of purified enantiomers of selected probes. Putative targets will be affinity purified, identified by mass spectrometry, and validated using a suite of complementary approaches. These studies hold great promise to address the long-standing questions of the molecular mechanisms of hemoglobin endocytosis and of the mode of action of MQ, thereby identifying a potentially high-value anti-malarial target.

Public Health Relevance

Althoughmefloquinehasbeenusedclinicallyfordecades,themodeof actionofthisanti-malarialdrughasnotbeenconclusivelyestablished.This projectseekstotestthehypothesisthatmefloquineactsbyblockingthemalaria parasite?suptakeofhosterythrocytehemoglobin.Thiswillbeaccomplishedby generatingalibraryofnovel,mefloquine-basedaffinityphotoprobesthatwillbe employedtoidentifytheproteintargetofmefloquineinparasitizederythrocytes.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Exploratory/Developmental Grants (R21)
Project #
5R21AI149190-02
Application #
10089405
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
O'Neil, Michael T
Project Start
2020-02-01
Project End
2022-01-31
Budget Start
2021-02-01
Budget End
2022-01-31
Support Year
2
Fiscal Year
2021
Total Cost
Indirect Cost
Name
Virginia Polytechnic Institute and State University
Department
Biochemistry
Type
Earth Sciences/Resources
DUNS #
003137015
City
Blacksburg
State
VA
Country
United States
Zip Code
24061