High throughput screening for anti-adhesion drugs to treat severe malaria

Seattle Biomedical Research Institute, Seattle WA

Investigators

Linked publications & trials

Abstract

DESCRIPTION (provided by applicant): The long-term aim of this research is to develop a high throughput screening (HTS) platform for drugs that specifically prevent and/or reverse adhesion of P. falciparum parasite-infected erythrocytes (PE) to the host endothelium and red blood cells. PE adhesion involves a large family (~60 members) of variant surface- expressed erythrocyte membrane proteins called PfEMP1. Specific adhesion of these extremely complex multidomain PfEMP1 proteins to host cell receptors mediates severe complications in malaria (SM) including cerebral malaria, placental malaria, and respiratory distress, but only a few PfEMP1s are involved in SM. Currently available drugs fail in 10-15 % of SM cases. Specific treatment with drugs reversing and preventing PE adhesion may significantly improve disease outcome and save lives in severe malaria syndromes. Anti- adhesion drugs (AAD) may be particularly useful because they 1) will stimulate de-sequestration of PE which would lead to decreases in microvasculature obstruction and help to re-establish the microvascular blood flow;2) will mediate their effect without penetrating parasites and thus have more immediate pharmacological activity;3) will make parasites vulnerable to clearance in the spleen and, possibly, by other innate and adaptive immune mechanisms. These properties also make such drugs good candidates for adjunctive therapy. During our R21-funded work, we constructed functional genome-wide PfEMP1 domain arrays and successfully used them to identify domains that bind specifically to two host cell receptors (ICAM1 and CD36). We further demonstrated that antibodies that block adhesion of a PfEMP1 domain to the ICAM1 receptor can be detected and quantified using this platform. To establish the basis for AAD HTS, we propose the following specific aims: 1) Identify PfEMP1 domains that mediate binding to a further nine endothelial cell receptors, implicated in PE adhesion, using our multiplexed assay based on BioPlex micro-bead technology. We predict that the systematic HT analysis of functional PfEMP1 domains will reveal new domain-receptor pairs that contribute to PE adhesion. We will verify that domains identified in vitro are also functional in the live PE. 2) Validate the PfEMP1 domain array for HTS of molecules that inhibit specific domain-host receptor adhesion. A small molecule (EGCG) will be used as a positive control for inhibition of the ICAM1 receptor binding. With the other receptors we will test for inhibition of domain binding using: a) several macromolecules previously shown to inhibit PE-receptor binding;b) unlabeled receptors;c) antibodies against receptors and against receptor- binding PfEMP1 domains;d) immune sera collected from patients living in malaria endemic areas. All positive hits from HTS will be tested for inhibition/reversal of binding of live PE to corresponding receptors. Finally, we will screen a commercially available small-molecule medium-size library as a proof-of-principle. These experiments will validate our platform for HTS of AAD for the treatment of severe malaria, and will support further development of this platform for HTS of molecules that target other protein-protein interactions of PE. PUBLIC HEALTH RELEVANCE: The long-term aim of this work is to develop a high throughput screening (HTS) platform for drugs that specifically prevent adhesion of malaria parasites to the host endothelial and red blood cells. The key player involved in parasite adhesion is a family (~60 members) of clonally variant surface-expressed Erythrocyte Membrane Protein 1 (PfEMP1). Specific treatment with drugs reversing and preventing parasite adhesion may significantly improve disease outcome and save many lives in severe malaria syndromes. Suggested experiments will develop and validate our functional PfEMP1 domain array platform for HTS for malaria antiadhesion compounds.