We created neutral antimalarial prodrugs that deliver bisthiazolium compounds with antimalarial activity in the nanomolar range. These drugs primarily affect early intraerythrocytic stages through rapid, nonreversible cytotoxicity. The compounds are suitable for both parenteral and oral use and plasma promotes rapid conversion of the prodrug into the drug. We demonstrate that very low doses offer protection in a murine model of malaria. The drugs show great potential for curing high parasitemia with short-course treatments. Oral administration of the TE3 prodrug completely cures Plasmodium cynomolgi infection in rhesus monkeys. The drugs specifically accumulate inside infected erythrocytes, block phosphatidylcholine biosynthesis, and interact with hemozoin. To our knowledge, this class of compounds represents one of the most potent antimalarials tested to date. These unique properties signal a promising future for this class of antimalarial.M alaria is a public health problem affecting Ͼ40% of the world's population. It causes up to 2 million deaths, mostly young children, in Africa, and Ͼ300 million clinical cases each year (1), with major consequent impact on economic productivity and livelihood (2). Currently, there are no licensed vaccines and the spread of drug-resistant parasites and insecticideresistant mosquitoes is an increasing problem. New antimalarial compounds, particularly those based on compounds structurally unrelated to existing antimalarial drugs with new mechanisms of action, are urgently needed (3-5).We have developed classes of antimalarial drugs targeting membrane biogenesis during intraerythrocytic Plasmodium falciparum development. We have focused on mono-and bisquaternary ammonium compounds for their potent antimalarial activity in vitro and in vivo (6, 7). These compounds mimic choline structure; they potently inhibit the low-affinity choline carrier related to phospholipid biosynthesis in eukaryotic cells and the high-affinity carrier involved in biosynthesis of the neurotransmitter acetylcholine in the CNS (8, 9). These compounds have exceptional in vitro and in vivo antimalarial properties devoid of mutagenic activity (10 -12).G25 [1,16-hexadecamethylenebis(N-methylpyrrolidinium) dibromide] and other drugs in this class possess a permanently charged cationic group (7, 13) that is essential for activity but detrimental to oral absorption. This action prejudiced development for the clinic. Oral administration is essential for treatment in dispensaries in endemic countries and for prophylactic or curative treatment for travelers. A targeting carrier system is unsuitable because, in the absence of carrier-mediated specific processes, quaternary ammonium compounds are intrinsically incapable of crossing bilayered cellular membranes. Designing drugs that mask the ionizable groups was therefore the most attractive solution. This was the rationale behind development of a chemically modified form of drug that is converted into an active ionized form by enzymes present in plasma. Here, we repo...

Erythrocytes infected with Plasmodium falciparum or Plasmodium knowlesi efficiently incorporated radioactive serine into phosphatidylserine (PtdSer), phosphatidylethanolamine (PtdEtn) and phosphatidylcholine (PtdCho). Serine was also metabolized into ethanolamine (Etn) and phosphorylethanolamine (P-Etn) via direct serine decarboxylation ; this is a major phenomenon since together these metabolites represent 60 % of total radioactive water-soluble metabolites. They were identified by reverse-phase HPLC and two TLC-type analyses and confirmed by alkaline phosphatase treatment, which depleted the radioactive P-Etn peak completely with a concomitant increase in that of Etn. In the presence of 5 µM labelled serine, radioactivity appeared in Etn and P-Etn after a 25 min lag period, and isotopic equilibrium was reached at 40 and 95 min respectively. There was a similar lag period for PtdEtn formation, which accumulated steadily for at least 180 min. Incorporation of serine into phospholipids and water-soluble metabolites increased in the presence of up to 500 µM external serine. An apparent plateau was then reached for all metabolites except intracellular serine and Etn. Exogenous Etn (at 20 µM) induced a concomitant dramatic decrease in serine incorporation into P-Etn and all phospholipids, but not

A key mechanism underlying physiological angiogenesis of the human endometrium is its ability to regenerate the vascular capillary network and to perform vascular remodeling (i.e., development of spiral arteries). Vascular endothelial growth factor (VEGF) is associated with angiogenesis and capillary permeability in this tissue. VEGF is expressed as several spliced variants, its main human isoforms contain 121 and 165 aa; 17␤-estradiol (E2) increases endometrial VEGF, possibly in all isoforms. Here we show that progesterone (P) selectively increases the expression of the VEGF189 (V189) isoform in the human uterus. V189 is identified in the conditioned medium of stromal cells treated with E2 ؉ P; its presence in this in vitro model of decidual stromal cells is detected after 6 -8 days, using ELISA, and after 8 -10 days, using Western blot analysis with different antibodies, including one specific for V189. The secretion pattern of V189 parallels that of the decidual protein IGFBP-1. V189 is secreted as a native isoform, as compared with the migration of recombinant V189 by SDS͞PAGE. In situ hybridization and immunocytochemistry, performed on the same biopsies, suggest that decidual cells express V189 during the midlate secretory phase of the menstrual cycle and early gestation. Finally, using an in vivo permeability assay, we show that native V189 increases capillary permeability. These observations demonstrate that P regulates V189 expression in decidual cells, which could have important implications for understanding uterine vascular remodeling and implantation, and may be relevant in a range of disease states such as edema and irregular bleeding.A s a tissue that exhibits rapid cyclic changes (1), the human endometrium is a good model for the study of physiological angiogenesis, which is under the control of 17␤ estradiol (E 2 ), progesterone (P), and vascular endothelial growth factor (VEGF) (2-7).VEGF is a protein with angiogenic activity and is a potent stimulator of microvascular permeability (8, 9); it plays an important role in physiological and pathological neovascularization, via its receptors Flt-1͞VEGFR1 and Flk-1͞VEGFR2 (10-13). Molecular cloning of the cDNA for VEGF revealed that differential exon splicing generates several isoforms containing 121, 165, 189, and 206 aa (V 121, V 165, V 189, and V 206 ) (14, 15). In most systems V 121 and V 165 are the major species expressed, whereas V 189 is only minimally present and V 206 is limited to embryonic tissue (9, 15). The different isoforms appear to have similar functions, but differ in their binding affinity for extracellular matrix (ECM). In contrast to VEGF 121 , which is secreted and found freely soluble in the culture medium, the bioavailability of VEGF 165 and to a greater extent V 189 appears to be regulated by binding to heparan sulfate proteoglycans (HSPG) in the ECM (9, 16). Bound VEGF isoforms could provide a reserve of growth factors available after cleavage by heparinase or urokinase-type plasminogen activator (uPA) (17, 18).Previous stud...