Project Grant R01AI173171

EXTENT, DYNAMICS AND MECHANISMS OF PLASMODIUM VIVAX IMMUNE EVASION CAUSED BY PVDBP GENE AMPLIFICATION - ELIMINATION OF PLASMODIUM VIVAX (PV) MALARIA PARASITES WOULD GREATLY BENEFIT FROM A BLOOD-STAGE VACCINE. PVDBP IS A PARASITE LIGAND INVOLVED IN ERYTHROCYTE INVASION THROUGH THE INTERACTION WITH ITS HUMAN RECEPTOR, THE DUFFY PROTEIN. THIS INTERACTION IS CRITICAL FOR THE PARASITE'S ENTRY MAKING PVDBP THE MOST ADVANCED CANDIDATE FOR A BLOOD-STAGE VACCINE WITH PHASE II CLINICAL TRIALS UNDERGOING. RECENT WORK HAS IDENTIFIED AND CHARACTERIZED HUMAN MONOCLONAL ANTIBODIES (HUMABS) THAT ALLOW STRAIN-TRANSCENDING NEUTRALIZATION OF PARASITES REGARDLESS OF THEIR PVDBP SEQUENCE DIVERSITY. HOWEVER, WE HAVE DEMONSTRATED THAT PV COLLECTED IN CAMBODIA WITH MULTIPLE COPIES OF THE PVDBP GENE WERE ABLE TO OVERCOME IN VITRO NEUTRALIZATION BY THESE HUMABS. THESE OBSERVATIONS PROVIDED THE FIRST EVIDENCE FOR AN EVOLUTIONARY ADVANTAGE FOR PVDBP AMPLIFICATION, WIDESPREAD IN PV POPULATIONS, AND CREATED A NEW PARADIGM IN WHICH TO CONSIDER PATHOGEN IMMUNE EVASION MECHANISMS. THESE RESULTS RAISE THE CONCERN THAT IMPLEMENTATION OF A PVDBP VACCINE MAY SELECT FOR MULTI-PVDBP COPY PARASITES. THE OVERALL GOAL OF THIS PROPOSAL IS PRECISELY TO DETERMINE IF PVDBP AMPLIFICATION WILL LIKELY COMPROMISE A PVDBP VACCINE STRATEGY. THE FIRST SPECIFIC AIM (SA) IS TO DETERMINE TO WHAT EXTENT MULTI-PVDBP COPY PARASITES GENETICALLY DISTANT FROM CAMBODIAN ISOLATES RESPOND TO ANTI-PVDBP HUMABS AND TO EVALUATE IF PVDBP AMPLIFICATION IS ASSOCIATED TO DUFFY POLYMORPHISMS IN HUMAN POPULATIONS. BY EVALUATING THE IN VITRO NEUTRALIZATION BY ANTI-PVDBP HUMABS OF SINGLE AND MULTI-PVDBP COPY PARASITES FROM ETHIOPIA, WE WILL BE ABLE TO EVALUATE THE EXTENT OF THE IMMUNE EVASION PHENOTYPE CONFERRED BY PVDBP AMPLIFICATION DESCRIBED WITH CAMBODIAN PV. BY (I) ASSOCIATING IN VITRO INVASION RATES WITH THE FULL-LENGTH DUFFY SEQUENCES OF INVADED ERYTHROCYTES, AND (II) PROSPECTIVELY TESTING FOR ASSOCIATION BETWEEN PVDBP COPY NUMBER AND HUMAN DUFFY SEQUENCES IN PARTICIPANTS ENROLLED IN LONGITUDINAL COHORTS IN CAMBODIA AND IN ETHIOPIA, WE WILL BE ABLE TO DETERMINE THE RELATION BETWEEN PVDBP AMPLIFICATION AND DUFFY HUMAN POLYMORPHISM. OUR SECOND SA WILL BE TO EVALUATE THE WITHIN-HOSTS AND WITHIN-POPULATION DYNAMICS OF PVDBP AMPLIFICATION OVER TIME. THROUGH THE ANALYSIS OF THE SEROLOGICAL DYNAMICS OF OUR LONGITUDINAL COHORTS' PARTICIPANTS, THE MEASURE OF PV INFECTIONS AND THE PVDBP COPY NUMBER OF INFECTING PARASITES, WE WILL BE ABLE TO TEST IF THE GENE AMPLIFICATION IS SELECTED IN VIVO BY THE IMMUNE STATUS OF HUMAN HOSTS AND HOW IT CORRELATES WITH CHANGES IN PV PREVALENCE IN THE POPULATION. IN VITRO EXPERIMENTAL EVOLUTION OF PK LINES WILL PROVIDE COMPLEMENTARY EVIDENCE FOR SELECTION OF PVDBP AMPLIFICATION BY ANTI-PVDBP HUMABS. OUR THIRD SA WILL BE TO DECIPHER THE MOLECULAR MECHANISMS ENABLING MULTI-COPY PARASITES TO EVADE ANTI-PVDBP HUMABS' NEUTRALIZATION. WE WILL SPECIFICALLY TEST IF IMMUNE EVASION RESULTS FROM INCREASED PROTEIN QUANTITY PRODUCED BY MULTI-COPY PARASITES AND/OR FROM EPITOPE VARIATIONS THROUGH MULTIPLE, DIFFERENT ALLELES AND VARIANTS PRESENT SIMULTANEOUSLY IN A GIVEN PARASITE. THROUGH A COMBINATION OF PHENOTYPING AND GENOMIC APPROACHES, OUR RESULTS WILL PROVIDE INVALUABLE DATA TO INFORM ON STRATEGIES TO OVERCOME THIS IMMUNE EVASION IN THE CONTEXT OF VACCINE DEVELOPMENT. .