In vitro and in vivo antimalarial activity of amphiphilic naphthothiazolium salts with amine-bearing side chains.

Because of emerging resistance to existing drugs, new chemical classes of antimalarial drugs are urgently needed. We have rationally designed a library of compounds that were predicted to accumulate in the digestive vacuole and then decrystallize hemozoin by breaking the iron carboxylate bond in hemozoin. We report the synthesis of 16 naphthothiazolium salts with amine-bearing side chains and their activities against the erythrocytic stage of Plasmodium falciparum in vitro. KSWI-855, the compound with the highest efficacy against the asexual stages of P. falciparum in vitro, also had in vitro activity against P. falciparum gametocytes and in vivo activity against P. berghei in a murine malaria model

Protein O-Fucosyltransferase 2 Is Not Essential for Plasmodium berghei Development

Thrombospondin type I repeat (TSR) domains are commonly O-fucosylated by protein O-fucosyltransferase 2 (PoFUT2), and this modification is required for optimal folding and secretion of TSR-containing proteins. The human malaria parasite Plasmodium falciparum expresses proteins containing TSR domains, such as the thrombospondin-related anonymous protein (TRAP) and circumsporozoite surface protein (CSP), which are O-fucosylated. TRAP and CSP are present on the surface of sporozoites and play essential roles in mosquito and human host invasion processes during the transmission stages. Here, we have generated PoFUT2 null-mutant P. falciparum and Plasmodium berghei (rodent) malaria parasites and, by phenotyping them throughout their complete life cycle, we show that PoFUT2 disruption does not affect the growth through the mosquito stages for both species. However, contrary to what has been described previously by others, P. berghei PoFUT2 null mutant sporozoites showed no deleterious motility phenotypes and successfully established blood stage infection in mice. This unexpected result indicates that the importance of O-fucosylation of TSR domains may differ between human and RODENT malaria parasites; complicating our understanding of glycosylation modifications in malaria biology

Antimalarial Iron Chelator, FBS0701, Shows Asexual and Gametocyte Plasmodium falciparum Activity and Single Oral Dose Cure in a Murine Malaria Model

Iron chelators for the treatment of malaria have proven therapeutic activity in vitro and in vivo in both humans and mice, but their clinical use is limited by the unsuitable absorption and pharmacokinetic properties of the few available iron chelators. FBS0701, (S)3”-(HO)-desazadesferrithiocin-polyether [DADFT-PE], is an oral iron chelator currently in Phase 2 human studies for the treatment of transfusional iron overload. The drug has very favorable absorption and pharmacokinetic properties allowing for once-daily use to deplete circulating free iron with human plasma concentrations in the high µM range. Here we show that FBS0701 has inhibition concentration 50% (IC50) of 6 µM for Plasmodium falciparum in contrast to the IC50 for deferiprone and deferoxamine at 15 and 30 µM respectively. In combination, FBS0701 interfered with artemisinin parasite inhibition and was additive with chloroquine or quinine parasite inhibition. FBS0701 killed early stage P. falciparum gametocytes. In the P. berghei Thompson suppression test, a single dose of 100 mg/kg reduced day three parasitemia and prolonged survival, but did not cure mice. Treatment with a single oral dose of 100 mg/kg one day after infection with 10 million lethal P. yoelii 17XL cured all the mice. Pretreatment of mice with a single oral dose of FBS0701 seven days or one day before resulted in the cure of some mice. Plasma exposures and other pharmacokinetics parameters in mice of the 100 mg/kg dose are similar to a 3 mg/kg dose in humans. In conclusion, FBS0701 demonstrates a single oral dose cure of the lethal P. yoelii model. Significantly, this effect persists after the chelator has cleared from plasma. FBS0701 was demonstrated to remove labile iron from erythrocytes as well as enter erythrocytes to chelate iron. FBS0701 may find clinically utility as monotherapy, a malarial prophylactic or, more likely, in combination with other antimalarials

Clinically relevant atovaquone-resistant human malaria parasites fail to transmit by mosquito.

Long-acting injectable medications, such as atovaquone, offer the prospect of a "chemical vaccine" for malaria, combining drug efficacy with vaccine durability. However, selection and transmission of drug-resistant parasites is of concern. Laboratory studies have indicated that atovaquone resistance disadvantages parasites in mosquitoes, but lack of data on clinically relevant Plasmodium falciparum has hampered integration of these variable findings into drug development decisions. Here we generate atovaquone-resistant parasites that differ from wild type parent by only a Y268S mutation in cytochrome b, a modification associated with atovaquone treatment failure in humans. Relative to wild type, Y268S parasites evidence multiple defects, most marked in their development in mosquitoes, whether from Southeast Asia (Anopheles stephensi) or Africa (An. gambiae). Growth of asexual Y268S P. falciparum in human red cells is impaired, but parasite loss in the mosquito is progressive, from reduced gametocyte exflagellation, to smaller number and size of oocysts, and finally to absence of sporozoites. The Y268S mutant fails to transmit from mosquitoes to mice engrafted with human liver cells and erythrocytes. The severe-to-lethal fitness cost of clinically relevant atovaquone resistance to P. falciparum in the mosquito substantially lessens the likelihood of its transmission in the field

A G358S mutation in the Plasmodium falciparum Na⁺ pump PfATP4 confers clinically-relevant resistance to cipargamin

Diverse compounds target the Plasmodium falciparum Na(+) pump PfATP4, with cipargamin and (+)-SJ733 the most clinically-advanced. In a recent clinical trial for cipargamin, recrudescent parasites emerged, with most having a G358S mutation in PfATP4. Here, we show that PfATP4(G358S) parasites can withstand micromolar concentrations of cipargamin and (+)-SJ733, while remaining susceptible to antimalarials that do not target PfATP4. The G358S mutation in PfATP4, and the equivalent mutation in Toxoplasma gondii ATP4, decrease the sensitivity of ATP4 to inhibition by cipargamin and (+)-SJ733, thereby protecting parasites from disruption of Na(+) regulation. The G358S mutation reduces the affinity of PfATP4 for Na(+) and is associated with an increase in the parasite’s resting cytosolic [Na(+)]. However, no defect in parasite growth or transmissibility is observed. Our findings suggest that PfATP4 inhibitors in clinical development should be tested against PfATP4(G358S) parasites, and that their combination with unrelated antimalarials may mitigate against resistance development

Immune Regulation of Plasmodium Is Anopheles Species Specific and Infection Intensity Dependent

Malaria parasite ookinetes must traverse the vector mosquito midgut epithelium to transform into sporozoite-producing oocysts. The Anopheles innate immune system is a key regulator of this process, thereby determining vector competence and disease transmission. The role of Anopheles innate immunity factors as agonists or antagonists of malaria parasite infection has been previously determined using specific single Anopheles-Plasmodium species combinations. Here we show that the two C-type lectins CTL4 and CTLMA2 exert differential agonistic and antagonistic regulation of parasite killing in African and South American Anopheles species. The C-type lectins regulate both parasite melanization and lysis through independent mechanisms, and their implication in parasite melanization is dependent on infection intensity rather than mosquito-parasite species combination. We show that the leucine-rich repeat protein LRIM1 acts as an antagonist on the development of Plasmodium ookinetes and as a regulator of oocyst size and sporozoite production in the South American mosquito Anopheles albimanus. Our findings explain the rare observation of human Plasmodium falciparum melanization and define a key factor mediating the poor vector competence of Anopheles albimanus for Plasmodium berghei and Plasmodium falciparum