Article

Potentiation of antimalarial drug action by chlorpheniramine against multidrug-resistant Plasmodium falciparum in vitro.

Department of Biochemistry, Faculty of Science, Mahidol University, Rama VI road, Bangkok 10400, Thailand.
Parasitology International (Impact Factor: 2.11). 10/2006; 55(3):195-9. DOI: 10.1016/j.parint.2005.11.058
Source: PubMed

ABSTRACT Chlorpheniramine, a histamine H1 receptor antagonist, was assayed for in vitro antimalarial activity against multidrug-resistant Plasmodium falciparum K1 strain and chloroquine-resistant P. falciparum T9/94 clone, by measuring the 3H-hypoxanthine incorporation. Chlorphenirame inhibited P. falciparum K1 and T9/94 growth with IC50 values of 136.0+/-40.2 microM and 102.0+/-22.6 microM respectively. A combination of antimalarial drug and chlorpheniramine was tested against resistant P. falciparum in vitro. Isobologram analysis showed that chlorpheniramine exerts marked synergistic action on chloroquine against P. falciparum K1 and T9/94. Chlorpheniramine also potentiated antimalarial action of mefloquine, quinine or pyronaridine against both of the resistant strains of P. falciparum. However, chlorpheniramine antagonism with artesunate was obtained in both P. falciparum K1 and T9/94. The results in this study indicate that antihistaminic drugs may be promising candidates for potentiating antimalarial drug action against drug resistant malarial parasites.

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    ABSTRACT: Die Protozoen des Genus Plasmodium verursachen weltweit rund 247 Millionen Malariafälle jedes Jahr. Da der Malariaparasit schnell und effektiv Resistenzen gegen neue Antimalaria-Medikamente entwickelt, ist es notwendig, stets innovative Wirkstoffe zu finden. Dabei spielen das Verständnis der grundlegenden Stoffwechselfunktionen und die Entdeckung neuer potenzieller drug targets wichtige Rollen in der präklinischen Forschung. Während ihrer Lebensphasen in menschlichen Erythrozyten und dem Mückenvektor sind die Malariaparasiten verschiedenen pro-oxidativen Umgebungen ausgesetzt. Daher haben die Parasiten ein komplexes Netzwerk aus antioxidativen schützenden Mechanismen entwickelt. Wie in vielen anderen Organismen repräsentieren redox-aktive Selenoproteine eine wichtige Komponente in diesem Netzwerk. In der vorliegenden Arbeit wurden die vier Selenoproteine aus Plasmodium falciparum biochemisch und funktionell charakterisiert. Mittels bioinformatischer Analysen, wie Alignments und Motiv-Scans, konnten Ähnlichkeiten zwischen PfSel1 und SelK, PfSel2 und SelT sowie PfSel4 und SelS herausgestellt werden. Hingegen erscheint PfSel3 einzigartig unter den bisher bekannten Selenoproteinen. Die Lokalisationsstudien mittels GFP-Fusions-Konstrukten unterstreichen die Ähnlichkeiten in den bioinformatischen Analysen. So befinden sich PfSel1, PfSel2 und PfSel4, wie SelK, SelT und SelS, im Endoplasmatischen Retikulum. Hingegen ist PfSel3 im Nukleus sowie im Apicoplasten aufzufinden und nimmt somit wieder eine Sonderstellung ein. Transkriptionsprofile der vier Selenoproteine und anderer plasmodialer redox-aktiver Proteine wurden nach dem Zusatz der Zellkultur mit Paraquat, Natriumnitroprussid, Methylenblau bzw. Natriumselenit erstellt. Die Ergebnisse zeigen, dass die Transkripte der Selenoproteine durch die Selenverfügbarkeit reguliert werden. Methylenblau hat keinen Einfluß auf die Transkription der untersuchten Gene. Vergleiche der Selenoproteine-Transkriptlevel mit den Transkriptleveln bekannter redox-aktiver Proteinen zeigten, dass PfSel1 und PfSel4 ähnlich wie PfTPx1, PfTPxGl und PfTrx1 durch oxidativen und nitrosativen Stress während des Wechels von Ring zu Trophozoiten stark reprimiert werden. Im Ringstadium werden hingegen die mRNA-Level von PfSel3, PfSPS und PfAOP extrem durch nitrosativen Stress reprimiert. Im Schizontenstadium reguliert Plasmodium ebenfalls durch nitrosativen Stress fast alle redox-aktiven Transkripte, namentlich Pf1cys, PfAOP, PfGrx, PfTPxGl, PfTrx1 und PfTrxR, hoch. Weiterhin wurde die heterologe Überexpression der plasmodialen Selenoproteine als Sec>Cys-Mutanten in E. coli optimiert. Mittels Gelfiltrationschromatographie und Quervernetzungs-experimenten konnten erste Eindrücke auf die Oligomerisierungszustände der rekombinanten Proteine gewonnen werden. Erste funktionelle Assays zeigen eine mögliche Redoxfunktion von PfSel1 und PfSel4. Die Überexpression von PfSel2 in Plasmodium falciparum führt zu einem Anstieg des freien cytosolischen Calciums. Daneben konnten einige Interaktionspartner mittels pull-down-Analysen für PfSel1 und PfSel4 gefangen werden. Im Rahmen dieser Arbeit konnten neue Informationen über die plasmodialen Selenoproteine gewonnen werden. Sie stellen potenzielle drug targets dar, die in weiteren Experimenten genauer analysiert werden müssen. The protozoa of the genus Plasmodium cause 250 million cases of malaria worldwide each year. Because the malaria parasite develops resistance to antimalarial drugs rapidly and effectively, there is an urgent need to discover innovative active agents. Therefore understanding basic metabolism and the detection of new potential drug targets play important roles in preclinical research. During their life cycle malaria parasites are exposed to various pro-oxidative environments in the human host and the mosquito vector. Consequently, the parasites have developed a complex network of antioxidative protective mechanisms. As in many other organisms, redox-active selenoproteins represent one important component of this network. In this present work, the four selenoproteins of Plasmodium falciparum have been characterised biochemically and functionally. Through bioinformatical analyses such as alignments and motif scans, similarities between PfSel1 and SelK, PfSel2 and SelT, and between PfSel4 and SelS were identified. In contrast, PfSel3 seems to be unique among all currently known selenoproteins. Localisation studies via GFP-fusion proteins underscore the similarities seen in the bioinformatical analyses. PfSel1, PfSel2, PfSel4, as well as SelK, SelT, and SelS are located in the endoplasmic reticulum. However, PfSel3 is located in the nucleus or apicoplast and therefore takes an exceptional position. Transcriptional profiles of the four selenoproteins and other plasmodial redox active proteins were prepared after supplementing the cell culture with sodium selenite, paraquat, sodium nitroprusside, and methylene blue, respectively. The results indicate that the transcripts of the selenoproteins are regulated via the availability of selenium. Methylene blue had no influence on the transcription of the examined genes. Comparisons of selenoprotein transcript levels with those of known redox-active proteins revealed a similar down-regulation of PfSel1 and PfSel4 to that of PfTPx1, PfTPxGl, and PfTrx1 under oxidative and nitrosative stress during the parasite’s transformation from ring to trophozoite stage. During the ring stage, the mRNA levels of PfSel3, PfSPS, and PfAOP were found to be extremely down-regulated after exposure to nitrosative stress. Almost all transcripts of redox active proteins, namely Pf1cys, PfAOP, PfGrx, PfTPxGl, PfTrx1, and PfTrxR, were also up-regulated under nitrosative stress in the schizont stage. Furthermore, the heterologous overexpression of the plasmodial selenoproteins as Sec>Cys mutants in E. coli was optimized. First impressions of the oligomerisation state of the recombinant proteins were gained via gel-filtration chromatography and cross-linking experiments. Initial functional assays showed a possible redox activity for PfSel1 and PfSel4. Overexpression of PfSel2 in Plasmodium falciparum leads to an increase of free cytosolic calcium. Moreover, some interacting proteins were captured via pull-down assays with PfSel1 and PfSel4. New informations about the plasmodial selenoproteins were obtained in this study. These proteins represent potential drug targets that have to be studied in further detail.
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    ABSTRACT: From the inoculation of Plasmodium sporozoites via Anopheles mosquito bites to the development of blood-stage parasites, a hallmark of the host response is an inflammatory reaction characterized by elevated histamine levels in the serum and tissues. Given the proinflammatory and immunosuppressive activities associated with histamine, we postulated that this vasoactive amine participates in malaria pathogenesis. Combined genetic and pharmacologic approaches demonstrated that histamine binding to H1R and H2R but not H3R and H4R increases the susceptibility of mice to infection with Plasmodium. To further understand the role of histamine in malaria pathogenesis, we used histidine decarboxylase-deficient (HDC(-/-)) mice, which are free of histamine. HDC(-/-) mice were highly resistant to severe malaria whether infected by mosquito bites or via injection of infected erythrocytes. HDC(-/-) mice displayed resistance to two lethal strains: Plasmodium berghei (Pb) ANKA, which triggers cerebral malaria (CM), and Pb NK65, which causes death without neurological symptoms. The resistance of HDC(-/-) mice to CM was associated with preserved blood-brain barrier integrity, the absence of infected erythrocyte aggregation in the brain vessels, and a lack of sequestration of CD4 and CD8 T cells. We demonstrate that histamine-mediated signaling contributes to malaria pathogenesis. Understanding the basis for these biological effects of histamine during infection may lead to novel therapeutic strategies to alleviate the severity of malaria.
    Journal of Experimental Medicine 03/2008; 205(2):395-408. DOI:10.1084/jem.20071548 · 13.91 Impact Factor

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