Cysteamine, the natural metabolite of pantetheinase, shows specific activity against Plasmodium

Abstract

In mice, loss of pantetheinase activity causes susceptibility to infection with Plasmodium chabaudi AS. Treatment of mice with the pantetheinase metabolite cysteamine reduces blood-stage replication of P. chabaudi and significantly increases survival. Similarly, a short exposure of Plasmodium to cysteamine ex vivo is sufficient to suppress parasite infectivity in vivo. This effect of cysteamine is specific and not observed with a related thiol (dimercaptosuccinic acid) or with the pantethine precursor of cysteamine. Also, cysteamine does not protect against infection with the parasite Trypanosoma cruzi or the fungal pathogen Candida albicans, suggesting cysteamine acts directly against the parasite and does not modulate host inflammatory response. Cysteamine exposure also blocks replication of P. falciparum in vitro; moreover, these treated parasites show higher levels of intact hemoglobin. This study highlights the in vivo action of cysteamine against Plasmodium and provides further evidence for the involvement of pantetheinase in host response to this infection.

Fig. 1

The effect of cysteamine treatment on P. chabaudi infection in vivo. A/J mice were treated with CysH, infected with P. chabaudi (10⁵ pRBC i.v.) and monitored for parasitemia. (A) Parasite levels after prophylactic treatment with CysH (120 or 40 mg/kg) compared to PBS-injected controls, for female mice. Kaplan–Meier survival curves of treated and untreated mice are plotted in (B). Parasitemia levels from therapeutic cysteamine treatment starting at day +2 or day +4 are shown for males (C) and females (D), while survival is plotted in (E and F) for the same groups. Each dot represents a mouse. Statistically significant differences (unpaired t-test) are indicated by asterisks (*p < 0.05, **p < 0.01, ***p < 0.001).

Fig. 2

The effect of cysteamine treatment ex vivo on infectivity of P. chabaudi. P. chabaudi parasitized erythrocytes (pRBCs) were incubated with CysH for 1 h at 37 °C, ex vivo. Subsequently, mice were infected i.v. with each of the treated suspensions (1 × 10⁵ pRBC) and parasitemia was monitored (A); error bars represent SEM. The inhibitory effect of CysH treatment on development of parasitemia at day 8 is shown for individual animals (% parasitemia) and is calculated as a fraction of the untreated group (% inhibition) in (B).

Fig. 3

Cysteamine treatment in mouse models of T. cruzi and C. albicans infections. A/J mice were infected intra-peritoneally with Trypanosoma cruzi (10³ trypomastigotes; Y strain), treated with CysH (dashed line) or PBS alone (solid line) and the number parasites in the blood was monitored daily. CysH was given daily by intra-peritoneal injection (150 mg/kg) starting 2 days prior to infection and continuing up to day 14. The kinetics of the infection are shown in (A), expressed as parasites/mL of blood, and the survival of CysH or PBS-treated mice is shown in a Kaplan–Meier plot (B). A/J mice were infected intravenously with Candida albicans (3 × 10⁵ blastospores; SC5134) and 24 h later the animals were sacrificed and total organ fungal loads were determined in brain (C) and kidney (D). Animals were treated daily (solid bars) or not (empty bars; PBS treated) with CysH (120 mg/kg) starting 2 days prior to infection and ending at 24 h post-infection.

Fig. 4

The effect of DMSA and pantethine on P. chabaudi infection in vivo. The effect of thiol-based compounds DMSA (meso 2,3-dimercaptosuccinic acid) and pantethine was examined in susceptible A/J mice. The structure of each compound is shown in (A). Male and female mice were treated daily with 50 mg/kg of DMSA or 30 mg per mouse of pantethine, using a prophylactic regimen (staring 2 days prior to infection and continuing every day thereafter until day 12). Parasitemia, expressed as percent parasitized erythrocytes, is plotted at day 5 and day 7 post-infection for DMSA (B) and pantethine (D) with each dot representing a mouse. Filled circles are untreated (PBS) and empty circles are treated animals. Survival is shown as a Kaplan–Meier plot for DMSA (C) and pantethine (E) treated (dashed line) and control (solid line) groups.

Fig. 5

Growth inhibition of P. falciparum by cysteamine and cystamine in vitro. Growth of P. falciparum in human RBCs treated with CysH or Cys was monitored and is expressed as percentage of pRBC for a chloroquine-resistant isolate (ITG; A). The line graph represents the growth of treated and untreated cultures over three cycles of invasion and growth (indicated by grey and black bars, resp. along the bottom). Parasite growth was measured by SYBR-green analysis of Plasmodium DNA during treatment (B) and is expressed as a percentage of the untreated control at each time point. In (C), P. falciparum ITG cultures were treated for 3 days (d1–d3) with CysH (200 μM) or Cys (150 μM). CysH/Cys treatment was stopped at day 4 and the culture was continued for 4 days (d4–d7) with daily medium replacement without addition of compound. Line graphs represent parasitemia levels of treated and untreated cultures and the dashed line represents the removal of drug.

Fig. 6

Inhibition of hemoglobin degradation by cysteamine and cystamine. Purified P. falciparum lysates from cultures treated or not with CysH (150 μM), Cys (75 μM), chloroquine (20 nM) or in combination, were assayed for Hb levels by immunoblotting (A). Uninfected whole blood lysate is shown in lane 1. Untreated parasite lysate is shown in lane 2, while treated lysates are present in lane 3–7. Twenty micrograms of total protein was loaded in each lane and the molecular mass markers are indicated in kilodaltons. A histogram plot (B) represents the quantification of the immunoblot in (A) showing the mean intensity units for the band after background subtraction. Abbreviations: CQ, chloroquine; CTR, whole blood control lysate.