Targeting the Hypnozoites: Towards an in vitro System for Malaria Drug Discovery

Malaria still has a global incidence of about 219 million cases per year and the World Health Organization estimates that nearly half of the world’s population is at risk of the disease in 2017. The majority of the available antimalarials on the market exclusively target the asexual and proliferative stages of malaria parasites. However, to eliminate malaria, we will need drugs that prevent parasite transmission and eliminate the dormant liver stages found after infection with Plasmodium vivax (P. vivax) and Plasmodium ovale. These quiescent forms called hypnozoites can survive for months in the liver and reactivate into mature liver schizonts causing relapses in patients. In order to elucidate the mechanisms underlying liver stage dormancy and to accelerate drug discovery for P. vivax malaria, we developed an in vitro infection model using stem cell-derived hepatocytes that allows studies of the hypnozoites. Considering the difficulties related to P. vivax in vitro work, the simian parasite Plasmodium cynomolgi, which also produces hypnozoites, was chosen as a surrogate model. Hepatocyte-like cells generated from induced pluripotent stem cells (iPS) derived from Macaca fascicularis were successfully infected with Plasmodium cynomolgi sporozoites and both hypnozoites and liver schizonts were observed for 12 days post infection. This PhD thesis reports the development of a new iPS-based model that allows hypnozoite formation, overcomes the limitations of P. vivax and primary hepatocytes, and provides a promising and versatile model to investigate the dormant liver stage of malaria

Induced pluripotent stem cell-derived human macrophages as an infection model for Leishmania donovani.

The parasite Leishmania donovani is one of the species causing visceral leishmaniasis in humans, a deadly infection claiming up to 40,000 lives each year. The current drugs for leishmaniasis treatment have severe drawbacks and there is an urgent need to find new anti-leishmanial compounds. However, the search for drug candidates is complicated by the intracellular lifestyle of Leishmania. Here, we investigate the use of human induced pluripotent stem cell (iPS)-derived macrophages (iMACs) as host cells for L. donovani. iMACs obtained through embryoid body differentiation were infected with L. donovani promastigotes, and high-content imaging techniques were used to optimize the iMACs seeding density and multiplicity of infection, allowing us to reach infection rates up to 70% five days after infection. IC50 values obtained for miltefosine and amphotericin B using the infected iMACs or mouse peritoneal macrophages as host cells were comparable and in agreement with the literature, showing the potential of iMACs as an infection model for drug screening