Manufacture of aseptic, purified, cryopreserved Plasmodium vivax sporozoites

ABSTRACT Plasmodium vivax (Pv) malaria is second to P. falciparum (Pf) malaria with regard to global prevalence and morbidity. Unlike Pf, control and elimination of Pv is complicated by frequent relapses – reactivation of dormant parasitic forms called hypnozoites that persist in the liver for months to more than a year, that are not eliminated by standard treatment, and that cause recurrent blood stage infections. Only primaquine and tafenoquine target hypnozoites, drugs potentially causing life threatening acute hemolytic anemia in those with G6PD deficiency which affects 8% of people in malaria-endemic regions. Efforts to develop better drugs or a vaccine are hampered by the inability to propagate blood stages of Pv in vitro. Therefore, generating Pv- infected mosquitoes for controlled human malaria infection (CHMI) to assess drugs, vaccines, and diagnostics is reliant on feeding mosquitoes on fresh, Pv-infected blood from Pv patients and is rarely done due to complex logistics. Absence of a readily available Pv CHMI of consistent quality hampers the development and testing of interventions against Pv malaria compared to Pf. In our phase II grant, we produced Pv sporozoites (SPZ) of the Chesson strain by feeding Pv-infected blood from specific pathogen free (SPF) Saimiri boliviensis (Sb) monkeys to aseptic Anopheles stephensi mosquitoes, and demonstrated that PvSPZ were highly infectious to FRG mice with humanized livers. In compliance with GMPs we manufactured 1) a master cell bank (MCB) of Pv (Chesson) asexual erythrocytic stage parasites in SPF Sb blood, and 2) one lot of aseptic, purified, vialed cryopreserved PvSPZ, Sanaria® PvSPZ Challenge (Chesson). We submitted a pre-IND package to the FDA detailing chemistry, manufacturing and controls and a proposal for a clinical trial and received positive feedback and recommendations. In this competitive renewal, we plan to complete all quality control (QC) assays to release the MCB and the cGMP PvSPZ Challenge lot, compile and submit an IND to the FDA that includes a protocol for a CHMI study to assess the safety and infectivity of PvSPZ Challenge (Chesson). We also propose manufacturing and releasing additional PvSPZ Challenge for future use in CHMI and as a potent immunogen in the vaccination approach called PvSPZ-CVac (PvSPZ administered with an antimalarial drug). Finally, to obviate the long-term need for producing and maintaining SPF Sb, we propose optimizing production of aseptic, purified PvSPZ by feeding on blood from volunteers undergoing CHMI by injection of Pv-infected erythrocytes. In summary, PfSPZ Challenge has revolutionized CHMI studies for Pf malaria, especially in the 6 African countries where it has been used, and brought Sanaria significant income. PvSPZ Challenge should have equivalent impact, providing malaria investigators with a heretofore non-existent, transformative CHMI model to assess efficacy of drugs, vaccines, and diagnostics for Pv malaria. It will be a safe quality-controlled reagent with minimal variability in potency, logistically simple to use, and available to any research center in the world and the foundation for establishing a highly protective approach to vaccinating against Pv malaria.