Inhibition by stabilization: targeting the Plasmodium falciparum aldolase-TRAP complex.
BACKGROUND
Emerging resistance of the malaria parasite Plasmodium to current therapies underscores the critical importance of exploring novel strategies for disease eradication. Plasmodium species are obligate intracellular protozoan parasites. They rely on an unusual form of substrate-dependent motility for their migration on and across host-cell membranes and for host cell invasion. This peculiar motility mechanism is driven by the 'glideosome', an actin-myosin associated, macromolecular complex anchored to the inner membrane complex of the parasite. Myosin A, actin, aldolase, and thrombospondin-related anonymous protein (TRAP) constitute the molecular core of the glideosome in the sporozoite, the mosquito stage that brings the infection into mammals.


METHODS
Virtual library screening of a large compound library against the PfAldolase-TRAP complex was used to identify candidate compounds that stabilize and prevent the disassembly of the glideosome. The mechanism of these compounds was confirmed by biochemical, biophysical and parasitological methods.


RESULTS
A novel inhibitory effect on the parasite was achieved by stabilizing a protein-protein interaction within the glideosome components. Compound 24 disrupts the gliding and invasive capabilities of Plasmodium parasites in in vitro parasite assays. A high-resolution, ternary X-ray crystal structure of PfAldolase-TRAP in complex with compound 24 confirms the mode of interaction and serves as a platform for future ligand optimization.


CONCLUSIONS
This proof-of-concept study presents a novel approach to anti-malarial drug discovery and design. By strengthening a protein-protein interaction within the parasite, an avenue towards inhibiting a previously "undruggable" target is revealed and the motility motor responsible for successful invasion of host cells is rendered inactive. This study provides new insights into the malaria parasite cell invasion machinery and convincingly demonstrates that liver cell invasion is dramatically reduced by 95 % in the presence of the small molecule stabilizer compound 24.
Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, USA. maureen.nemetski@gmail.com.|Department of Pediatrics, Phyllis and David Komansky Center for Children's Health, New York-Presbyterian Hospital-Weill Cornell Medical College, New York, USA. maureen.nemetski@gmail.com.
Sondra Maureen
SM
Nemetski
Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, USA. Timothy.Cardozo@nyumc.org.|Institute for Systems Genetics, New York University School of Medicine, New York, USA. Timothy.Cardozo@nyumc.org.
Timothy J
TJ
Cardozo
Department of Molecular Microbiology and Immunology, Johns Hopkins University Bloomberg School of Public Health, Baltimore, USA. gbosch@jhu.edu.|Johns Hopkins Malaria Research Institute (JHMRI), Baltimore, USA. gbosch@jhu.edu.
Gundula
G
Bosch
Department of Biochemistry and Molecular Biology, Johns Hopkins University Bloomberg School of Public Health, Baltimore, USA. ryan.weltzer@gmail.com.|Johns Hopkins Malaria Research Institute (JHMRI), Baltimore, USA. ryan.weltzer@gmail.com.
Ryan
R
Weltzer
Department of Biochemistry and Molecular Biology, Johns Hopkins University Bloomberg School of Public Health, Baltimore, USA. kevinomalley87@gmail.com.|Johns Hopkins Malaria Research Institute (JHMRI), Baltimore, USA. kevinomalley87@gmail.com.
Kevin
K
O'Malley
Department of Medical Parasitology, New York University School of Medicine, New York, USA. ijeoma_ejigiri@yahoo.com.
Ijeoma
I
Ejigiri
Michael Heidelberg Division of Pathology of Infectious Diseases, Department of Pathology, New York University School of Medicine, New York, USA. kumar_arun03@yahoo.com.|Department of Animal Sciences, School of Life Sciences, University of Hyderabad, Hyderabad, 500046, India. kumar_arun03@yahoo.com.
Kota Arun
KA
Kumar
Instituto de Investigaciones Biotecnológicas-Instituto Tecnológico de Chascomús (IIB-INTECH), Universidad Nacional de General San Martín-CONICET, 1650, San Martín, Buenos Aires, Argentina. cbuscaglia89@gmail.com.
Carlos A
CA
Buscaglia
Michael Heidelberg Division of Pathology of Infectious Diseases, Department of Pathology, New York University School of Medicine, New York, USA. Victor.Nussenzweig@nyumc.org.
Victor
V
Nussenzweig
Department of Molecular Microbiology and Immunology, Johns Hopkins University Bloomberg School of Public Health, Baltimore, USA. psinnis1@jhu.edu.|Department of Medical Parasitology, New York University School of Medicine, New York, USA. psinnis1@jhu.edu.|Johns Hopkins Malaria Research Institute (JHMRI), Baltimore, USA. psinnis1@jhu.edu.
Photini
P
Sinnis
Department of Molecular Microbiology and Immunology, Johns Hopkins University Bloomberg School of Public Health, Baltimore, USA. lelena1@jhu.edu.|Johns Hopkins Malaria Research Institute (JHMRI), Baltimore, USA. lelena1@jhu.edu.
Jelena
J
Levitskaya
Department of Biochemistry and Molecular Biology, Johns Hopkins University Bloomberg School of Public Health, Baltimore, USA. jbosch@jhu.edu.|Johns Hopkins Malaria Research Institute (JHMRI), Baltimore, USA. jbosch@jhu.edu.
Jürgen
J
Bosch
D000818:Animals; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D018360:Crystallography, X-Ray; D005634:Fructose-Bisphosphate Aldolase / Q000737:chemistry / Q000378:metabolism*; D022781:Hepatocytes / Q000187:drug effects; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D062105:Molecular Docking Simulation; D046912:Multiprotein Complexes / Q000737:chemistry* / Q000187:drug effects; D010963:Plasmodium falciparum / Q000737:chemistry; D055550:Protein Stability / Q000187:drug effects; D015800:Protozoan Proteins / Q000737:chemistry / Q000378:metabolism*; D011817:Rabbits; D054852:Small Molecule Libraries / Q000737:chemistry / Q000494:pharmacology / Q000633:toxicity; D020349:Surface Plasmon Resonance
D008565:Membrane Proteins; D046912:Multiprotein Complexes; D015800:Protozoan Proteins; D054852:Small Molecule Libraries; C069428:thrombospondin-related adhesive protein, protozoan; D005634:Fructose-Bisphosphate Aldolase