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A next generation genetically attenuated Plasmodium falciparum parasite created by triple gene deletion

A next generation genetically attenuated Plasmodium falciparum parasite created by triple gene deletion
Author: 
Stefan H. Kappe, Ph.D.
Published: 
Sep 2014
Publisher: 
Mol Ther. Sep;22(9):1707-15

Mikolajczak SA1, Lakshmanan V1, Fishbaugher M1, Camargo N1, Harupa A1, Kaushansky A1, Douglass AN1, Baldwin M1, Healer J2, O'Neill M2, Phuong T2, Cowman A2, Kappe SH3.

Author information

  • 1Seattle Biomedical Research Institute (Seattle BioMed), Seattle, Washington, USA.
  • 2The Walter and Eliza Hall Institute of Medical Research, Victoria, Australia.
  • 31] Seattle Biomedical Research Institute (Seattle BioMed), Seattle, Washington, USA [2] Department of Global Health, University of Washington, Seattle, Washington, USA.

Abstract

Immunization with live-attenuated Plasmodium sporozoites completely protects against malaria infection. Genetic engineering offers a versatile platform to create live-attenuated sporozoite vaccine candidates. We previously generated a genetically attenuated parasite (GAP) by deleting the P52 and P36 genes in the NF54 wild-type (WT) strain of Plasmodium falciparum (Pf p52(-)/p36(-) GAP). Preclinical assessment of p52(-)/p36(-) GAP in a humanized mouse model indicated an early and severe liver stage growth defect. However, human exposure to >200 Pf p52(-)/p36(-) GAP-infected mosquito bites in a safety trial resulted in peripheral parasitemia in one of six volunteers, revealing that this GAP was incompletely attenuated. We have now created a triple gene deleted GAP by additionally removing the SAP1 gene (Pf p52(-)/p36(-)/sap1(-) GAP) and employed flippase (FLP)/flippase recognition target (FRT) recombination for drug selectable marker cassette removal. This next-generation GAP was indistinguishable from WT parasites in blood stage and mosquito stage development. Using an improved humanized mouse model transplanted with human hepatocytes and human red blood cells, we show that despite a high-dose sporozoite challenge, Pf p52(-)/p36(-)/sap1(-) GAP did not transition to blood stage infection and appeared to be completely attenuated. Thus, clinical testing of Pf p52(-)/p36(-)/sap1(-) GAP assessing safety, immunogenicity, and efficacy against sporozoite challenge is warranted.