Project Grant F31AI200042

EVALUATING NEUTRALIZING IMMUNE RESPONSES TO THE ESSENTIAL INVASION PROTEIN RIPR ACROSS HUMAN PLASMODIUM SPECIES - PROJECT SUMMARY (REVISED) MALARIA IS A MOSQUITO-BORNE DISEASE THAT CONTINUES TO BE A SIGNIFICANT GLOBAL HEALTH CHALLENGE, POSING RISKS TO US TRAVELERS AND THE US MILITARY. WHILE VECTOR CONTROL MEASURES AND TREATMENT HAVE LED TO GLOBAL PROGRESS IN CONTROLLING MALARIA CASES AND DEATHS IN RECENT DECADES, THIS PROGRESS HAS STALLED SINCE 2015. CURRENT MALARIA VACCINES (RTS,S/AS01 AND R21/MATRIX-M) HAVE SHORT-LIVED PROTECTION, LIMITED EFFICACY AGAINST VARIOUS STRAINS, AND SPECIES-SPECIFICITY. AS A RESULT, DEVELOPING NEXT-GENERATION VACCINES TARGETING BLOOD-STAGE (MEROZOITE) ANTIGENS WHICH WILL REDUCE CLINICAL SYMPTOMS RESULTING FROM PLASMODIUM PARASITE REPLICATION IN RED BLOOD CELLS (RBCS) IS A PRIORITY. MY PROJECT FOCUSES ON RIPR, A PLASMODIUM PROTEIN ESSENTIAL FOR RBC INVASION THAT IS CURRENTLY BEING EVALUATED AS A BLOOD-STAGE VACCINE CANDIDATE (R78C) IN PHASE 1A CLINICAL TRIALS. MY CENTRAL HYPOTHESIS IS THAT RIPR WILL SERVE AS A TARGET OF NEUTRALIZING ANTIBODIES ACROSS HUMAN PLASMODIUM FALCIPARUM STRAINS AND WILL SERVE AS A CROSS-SPECIES BLOOD-STAGE VACCINE CANDIDATE. THE FIRST GOAL OF THE PROJECT IS TO DETERMINE THE NEUTRALIZING AND SYNERGISTIC ACTIVITY OF RIPR VACCINE (R78C)-INDUCED ANTIBODIES AGAINST GENETIC VARIANTS OF P. FALCIPARUM FROM A REGION OF HIGH MALARIA TRANSMISSION. I WILL PAIR PHENOTYPIC DATA FROM FIELD-BASED EX VIVO NEUTRALIZATION ASSAYS WITH GENOMIC AND PROTEIN STRUCTURE ANALYSIS OF IDENTIFIED RIPR SINGLE NUCLEOTIDE POLYMORPHISMS (SNPS) TO CHARACTERIZE THE FUNCTIONAL EFFECTS OF MUTATIONS OF INTEREST (EG: IMMUNE EVASION). FOR OTHER HUMAN PLASMODIUM SPECIES, WE LACK A CLEAR UNDERSTANDING OF THE FUNCTIONAL ROLE OF RIPR IN RBC INVASION. THEREFORE, THE SECOND GOAL OF THE PROJECT IS TO IDENTIFY THE NEUTRALIZABLE EPITOPES OF RIPR IN THE FIVE NON-FALCIPARUM HUMAN PLASMODIUM SPECIES. AS P. KNOWLESI IS THE ONLY NON-FALCIPARUM PLASMODIUM SPECIES THAT CAN BE CULTURED LONG-TERM IN VITRO, I WILL USE IT AS A SYSTEM TO IDENTIFY NEUTRALIZING DOMAINS OF RIPR IN THESE SPECIES USING PHENOTYPIC ASSAYS AND PROTEIN BIOCHEMISTRY (SURFACE PLASMON RESONANCE, SPR). I WILL THEN QUANTIFY THE CROSS-SPECIES NEUTRALIZING ACTIVITY OF P. FALCIPARUM RIPR (PFRIPR)-SPECIFIC ANTIBODIES TARGETING THESE SPECIFIC DOMAINS USING TWO COMPLEMENTARY METHODS: CRISPR/CAS9-GENERATED CHIMERIC P. KNOWLESI PARASITES AND EX VIVO CLINICAL ISOLATES OF P. MALARIAE, P. OVALE CURTISI, AND P. OVALE WALLIKERI. IN SUMMARY, MY PROPOSED PROJECT WILL SHED LIGHT ON A HIGHLY CONSERVED RED BLOOD CELL INVASION PATHWAY IN HUMAN PLASMODIUM SPECIES AND EVALUATE ITS POTENTIAL AS A CROSS-SPECIES VACCINE CANDIDATE. THROUGH MY PROPOSED TRAINING ACTIVITIES AT YALE AND COLLABORATING INSTITUTIONS, I AIM TO DEVELOP TECHNICAL SKILLS IN FIELD-BASED EX VIVO ASSAYS, PROTEIN BIOCHEMISTRY, CRISPR/CAS9-BASED GENOME EDITING OF PLASMODIUM PARASITES, AND METHODS DEVELOPMENT IN RESOURCE-LIMITED SETTINGS. I ALSO AIM TO DEVELOP A STRONG CONCEPTUAL BACKGROUND IN IMMUNOLOGY, PARASITOLOGY, AND EARLY-STAGE VACCINE DEVELOPMENT AND IMPROVE MY SCIENTIFIC COMMUNICATION AND MENTORSHIP SKILLS, WITH THE GOAL OF PURSUING A CAREER AS AN INDEPENDENT INVESTIGATOR IN TRANSLATIONAL TROPICAL INFECTIOUS DISEASE RESEARCH.