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Pyrazoleamide compounds are potent antimalarials that target Na+ homeostasis in intraerythrocytic Plasmodium falciparum.

Vaidya AB, Morrisey JM, Zhang Z, Das S, Daly TM, Otto TD, Spillman NJ, Wyvratt M, Siegl P, Marfurt J, Wirjanata G, Sebayang BF, Price RN, Chatterjee A, Nagle A, Stasiak M, Charman SA, Angulo-Barturen I, Ferrer S, Belén Jiménez-Díaz M, Martínez MS, Gamo FJ, Avery VM, Ruecker A, Delves M, Kirk K, Berriman M, Kortagere S, Burrows J, Fan E, Bergman LW - Nat Commun (2014)

Bottom Line: Investigations involving pyrazoleamide-resistant parasites, whole-genome sequencing and gene transfers reveal that mutations in two proteins, a calcium-dependent protein kinase (PfCDPK5) and a P-type cation-ATPase (PfATP4), are necessary to impart full resistance to these compounds.A pyrazoleamide compound causes a rapid disruption of Na(+) regulation in blood-stage Plasmodium falciparum parasites.Similar effect on Na(+) homeostasis was recently reported for spiroindolones, which are antimalarials of a chemical class quite distinct from pyrazoleamides.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Center for Molecular Parasitology, Drexel University College of Medicine, 2900 Queen Lane, Philadelphia, Pennsylvania 190129, USA.

ABSTRACT
The quest for new antimalarial drugs, especially those with novel modes of action, is essential in the face of emerging drug-resistant parasites. Here we describe a new chemical class of molecules, pyrazoleamides, with potent activity against human malaria parasites and showing remarkably rapid parasite clearance in an in vivo model. Investigations involving pyrazoleamide-resistant parasites, whole-genome sequencing and gene transfers reveal that mutations in two proteins, a calcium-dependent protein kinase (PfCDPK5) and a P-type cation-ATPase (PfATP4), are necessary to impart full resistance to these compounds. A pyrazoleamide compound causes a rapid disruption of Na(+) regulation in blood-stage Plasmodium falciparum parasites. Similar effect on Na(+) homeostasis was recently reported for spiroindolones, which are antimalarials of a chemical class quite distinct from pyrazoleamides. Our results reveal that disruption of Na(+) homeostasis in malaria parasites is a promising mode of antimalarial action mediated by at least two distinct chemical classes.

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In vivo efficacy of PA21A092 against P. falciparum.Four indicated doses of PA21A092 were administered orally to groups of three NOD/scid/IL2Rγ mice each engrafted with human erythrocytes and infected with P. falciparum. The compound was administered starting on day 3 post infection for 4 consecutive days. Parasitemia was assessed each day from day 3 post infection up to 7 days (a). Concentrations of PA21A092 were measured by LC/MS in each mouse at 0.25, 0.5, 1, 2, 4, 7, 10 and 23 h after the first dose (b). The estimated drug exposure necessary to inhibit P. falciparum parasitemia on day 7 post infection by 90% (AUCED90) was 0.24 μg h ml−1 per day−1 (c). Comparison of morphology of parasitized human RBC (as observed in Giemsa-stained thin blood smears prepared on day 7 after infection) in vehicle-treated (d) and PA21A092-treated (e) mice revealed normal stages of parasites in control but erythrocytes with only highly pyknotic staining nuclei fragments in treated mice (scale bars, 10 μm). In a–c, each symbol represents individual mouse with the dose of compound indicated in the inset on the right. Open circles in a are mice treated with the vehicle only.
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f2: In vivo efficacy of PA21A092 against P. falciparum.Four indicated doses of PA21A092 were administered orally to groups of three NOD/scid/IL2Rγ mice each engrafted with human erythrocytes and infected with P. falciparum. The compound was administered starting on day 3 post infection for 4 consecutive days. Parasitemia was assessed each day from day 3 post infection up to 7 days (a). Concentrations of PA21A092 were measured by LC/MS in each mouse at 0.25, 0.5, 1, 2, 4, 7, 10 and 23 h after the first dose (b). The estimated drug exposure necessary to inhibit P. falciparum parasitemia on day 7 post infection by 90% (AUCED90) was 0.24 μg h ml−1 per day−1 (c). Comparison of morphology of parasitized human RBC (as observed in Giemsa-stained thin blood smears prepared on day 7 after infection) in vehicle-treated (d) and PA21A092-treated (e) mice revealed normal stages of parasites in control but erythrocytes with only highly pyknotic staining nuclei fragments in treated mice (scale bars, 10 μm). In a–c, each symbol represents individual mouse with the dose of compound indicated in the inset on the right. Open circles in a are mice treated with the vehicle only.

Mentions: We used nonobese/severe combined immunodeficiency/IL2Rγ (NOD/SCID/IL2Rγ) mice engrafted with human erythrocytes and infected with an adapted line of P. falciparum as a model to assess in vivo efficacy of the pyrazoleamide compounds1314. This model has been used previously for examining many different antimalarials for their in vivo activity against the human malaria parasite P. falciparum15. Groups of three mice each were treated once a day for 4 consecutive days, beginning on day 3 post infection, with four oral dose levels of PA21A092. As shown in Fig. 2a, PA21A092 was highly effective when administered orally with effective dose for 90% parasitemia reduction (ED90) of 2.5 mg kg−1. Compound PA21A050 had ED90 of 0.9 mg kg−1 (Supplementary Fig. 1), whereas PA21A102 had an ED90 of 4 mg kg−1 (Supplementary Fig. 2). The pharmacokinetics of PA21A092 in the mice employed in the therapeutic efficacy study was studied by taking 25 μl serial blood samples up to 23 h after the first dose. As shown in Fig. 2b and Supplementary Table 2, oral pharmacokinetics were approximately linear in the dose range used. Assuming no significant accumulation during treatment, the estimated drug exposure necessary to inhibit P. falciparum parasitemia on day 7 post infection by 90% with respect to control mice (area under the curve at 90% effective dose, AUCED90) was 0.24 μg h ml−1 per day (Fig. 2c). In this in vivo assay, the AUCED90 is the average daily exposure necessary to achieve no net parasite growth on day 7. The maximum parasite clearance was achieved at PA21A092 blood exposure of above 1 μg h ml−1 per day and was comparable, in this model, to that seen with artesunate, which is thus far the fastest-acting antimalarial in use. Erythrocytes with only remnants of parasite nuclei were seen following 2-day treatments with the pyrazole compounds (Fig. 2d,e).


Pyrazoleamide compounds are potent antimalarials that target Na+ homeostasis in intraerythrocytic Plasmodium falciparum.

Vaidya AB, Morrisey JM, Zhang Z, Das S, Daly TM, Otto TD, Spillman NJ, Wyvratt M, Siegl P, Marfurt J, Wirjanata G, Sebayang BF, Price RN, Chatterjee A, Nagle A, Stasiak M, Charman SA, Angulo-Barturen I, Ferrer S, Belén Jiménez-Díaz M, Martínez MS, Gamo FJ, Avery VM, Ruecker A, Delves M, Kirk K, Berriman M, Kortagere S, Burrows J, Fan E, Bergman LW - Nat Commun (2014)

In vivo efficacy of PA21A092 against P. falciparum.Four indicated doses of PA21A092 were administered orally to groups of three NOD/scid/IL2Rγ mice each engrafted with human erythrocytes and infected with P. falciparum. The compound was administered starting on day 3 post infection for 4 consecutive days. Parasitemia was assessed each day from day 3 post infection up to 7 days (a). Concentrations of PA21A092 were measured by LC/MS in each mouse at 0.25, 0.5, 1, 2, 4, 7, 10 and 23 h after the first dose (b). The estimated drug exposure necessary to inhibit P. falciparum parasitemia on day 7 post infection by 90% (AUCED90) was 0.24 μg h ml−1 per day−1 (c). Comparison of morphology of parasitized human RBC (as observed in Giemsa-stained thin blood smears prepared on day 7 after infection) in vehicle-treated (d) and PA21A092-treated (e) mice revealed normal stages of parasites in control but erythrocytes with only highly pyknotic staining nuclei fragments in treated mice (scale bars, 10 μm). In a–c, each symbol represents individual mouse with the dose of compound indicated in the inset on the right. Open circles in a are mice treated with the vehicle only.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4263321&req=5

f2: In vivo efficacy of PA21A092 against P. falciparum.Four indicated doses of PA21A092 were administered orally to groups of three NOD/scid/IL2Rγ mice each engrafted with human erythrocytes and infected with P. falciparum. The compound was administered starting on day 3 post infection for 4 consecutive days. Parasitemia was assessed each day from day 3 post infection up to 7 days (a). Concentrations of PA21A092 were measured by LC/MS in each mouse at 0.25, 0.5, 1, 2, 4, 7, 10 and 23 h after the first dose (b). The estimated drug exposure necessary to inhibit P. falciparum parasitemia on day 7 post infection by 90% (AUCED90) was 0.24 μg h ml−1 per day−1 (c). Comparison of morphology of parasitized human RBC (as observed in Giemsa-stained thin blood smears prepared on day 7 after infection) in vehicle-treated (d) and PA21A092-treated (e) mice revealed normal stages of parasites in control but erythrocytes with only highly pyknotic staining nuclei fragments in treated mice (scale bars, 10 μm). In a–c, each symbol represents individual mouse with the dose of compound indicated in the inset on the right. Open circles in a are mice treated with the vehicle only.
Mentions: We used nonobese/severe combined immunodeficiency/IL2Rγ (NOD/SCID/IL2Rγ) mice engrafted with human erythrocytes and infected with an adapted line of P. falciparum as a model to assess in vivo efficacy of the pyrazoleamide compounds1314. This model has been used previously for examining many different antimalarials for their in vivo activity against the human malaria parasite P. falciparum15. Groups of three mice each were treated once a day for 4 consecutive days, beginning on day 3 post infection, with four oral dose levels of PA21A092. As shown in Fig. 2a, PA21A092 was highly effective when administered orally with effective dose for 90% parasitemia reduction (ED90) of 2.5 mg kg−1. Compound PA21A050 had ED90 of 0.9 mg kg−1 (Supplementary Fig. 1), whereas PA21A102 had an ED90 of 4 mg kg−1 (Supplementary Fig. 2). The pharmacokinetics of PA21A092 in the mice employed in the therapeutic efficacy study was studied by taking 25 μl serial blood samples up to 23 h after the first dose. As shown in Fig. 2b and Supplementary Table 2, oral pharmacokinetics were approximately linear in the dose range used. Assuming no significant accumulation during treatment, the estimated drug exposure necessary to inhibit P. falciparum parasitemia on day 7 post infection by 90% with respect to control mice (area under the curve at 90% effective dose, AUCED90) was 0.24 μg h ml−1 per day (Fig. 2c). In this in vivo assay, the AUCED90 is the average daily exposure necessary to achieve no net parasite growth on day 7. The maximum parasite clearance was achieved at PA21A092 blood exposure of above 1 μg h ml−1 per day and was comparable, in this model, to that seen with artesunate, which is thus far the fastest-acting antimalarial in use. Erythrocytes with only remnants of parasite nuclei were seen following 2-day treatments with the pyrazole compounds (Fig. 2d,e).

Bottom Line: Investigations involving pyrazoleamide-resistant parasites, whole-genome sequencing and gene transfers reveal that mutations in two proteins, a calcium-dependent protein kinase (PfCDPK5) and a P-type cation-ATPase (PfATP4), are necessary to impart full resistance to these compounds.A pyrazoleamide compound causes a rapid disruption of Na(+) regulation in blood-stage Plasmodium falciparum parasites.Similar effect on Na(+) homeostasis was recently reported for spiroindolones, which are antimalarials of a chemical class quite distinct from pyrazoleamides.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Center for Molecular Parasitology, Drexel University College of Medicine, 2900 Queen Lane, Philadelphia, Pennsylvania 190129, USA.

ABSTRACT
The quest for new antimalarial drugs, especially those with novel modes of action, is essential in the face of emerging drug-resistant parasites. Here we describe a new chemical class of molecules, pyrazoleamides, with potent activity against human malaria parasites and showing remarkably rapid parasite clearance in an in vivo model. Investigations involving pyrazoleamide-resistant parasites, whole-genome sequencing and gene transfers reveal that mutations in two proteins, a calcium-dependent protein kinase (PfCDPK5) and a P-type cation-ATPase (PfATP4), are necessary to impart full resistance to these compounds. A pyrazoleamide compound causes a rapid disruption of Na(+) regulation in blood-stage Plasmodium falciparum parasites. Similar effect on Na(+) homeostasis was recently reported for spiroindolones, which are antimalarials of a chemical class quite distinct from pyrazoleamides. Our results reveal that disruption of Na(+) homeostasis in malaria parasites is a promising mode of antimalarial action mediated by at least two distinct chemical classes.

Show MeSH
Related in: MedlinePlus