Reaction of artemisinin with haemoglobin: implications for antimalarial activity

Peroxide antimalarials, including artemisinin, are important for the treatment of multidrug-resistant malaria. These peroxides are known to react with iron or heme to produce reactive intermediates that are thought to be responsible for their antimalarial activities. This study investigated the potential interaction of selected peroxide antimalarials with oxyhemoglobin, the most abundant form of iron in the human body. The observed stability of artemisinin derivatives and 1,2,4-trioxolanes in the presence of oxyhemoglobin was in contrast to previous reports in the literature. Spectroscopic analysis of hemoglobin found it to be unstable under the conditions used for previous studies, and it appears likely that the artemisinin reactivity reported in these studies may be attributed to free heme released by protein denaturation. The stability of peroxide antimalarials with intact oxyhemoglobin, and reactivity with free heme, may explain the selective toxicity of these antimalarials toward infected, but not healthy, erythrocytes.

Section: Figsupporting

confidence: 54%

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confidence: 99%

Stability of Peroxide Antimalarials in the Presence of Human Hemoglobin

Creek

Ryan

Charman

et al. 2009

“…Kannan et al (12) have reported that heme alkylation by artemisinin only marginally decreased the molar extinction coefficient of heme (27% decrease when the solvent was methanol), and it is expected that alkylation by trioxolanes would have a similar effect on absorptivity. The visible absorbance of the alkylated heme product (472 nm) from each active trioxolane in this study was equivalent to 79% (Ϯ 5%) of the decrease in heme absorbance (418 nm), suggesting highly efficient formation of the alkylated adduct.…”

Section: Discussionmentioning

confidence: 99%

Relationship between Antimalarial Activity and Heme Alkylation for Spiro- and Dispiro-1,2,4-Trioxolane Antimalarials

Creek

Charman

Chiu

et al. 2008

102

111

The reaction of spiro-and dispiro-1,2,4-trioxolane antimalarials with heme has been investigated to provide further insight into the mechanism of action for this important class of antimalarials. A series of trioxolanes with various antimalarial potencies was found to be unreactive in the presence of Fe(III) hemin, but all were rapidly degraded by reduced Fe(II) heme. The major reaction product from the heme-mediated degradation of biologically active trioxolanes was an alkylated heme adduct resulting from addition of a radical intermediate. Under standardized reaction conditions, a correlation (R 2 ‫؍‬ 0.88) was found between the extent of heme alkylation and in vitro antimalarial activity, suggesting that heme alkylation may be related to the mechanism of action for these trioxolanes. Significantly less heme alkylation was observed for the clinically utilized artemisinin derivatives compared to the equipotent trioxolanes included in this study.

“…It has also been suggested (21) that artemisinin may undergo reductive activation in the mitochondria of P. falciparum and thereby inhibit the growth of the parasite. Indeed, in addition to Hb (19,26,34,43), artemisinin alkylates other hemoproteins, such as cytochrome c (43). However, if the mitochondria were the primary plasmodial target of artemisinin (21), it is hard to explain why all other protozoa are so much less sensitive to artemisinin.…”

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confidence: 99%

Peroxide Bond-Dependent Antiplasmodial Specificity of Artemisinin and OZ277 (RBx11160)

Kaiser

Wittlin

Nehrbass-Stuedli

et al. 2007

Using nonperoxidic analogs of artemisinin and OZ277 (RBx11160), the strong in vitro antiplasmodial activities of the latter two compounds were shown to be peroxide bond dependent. In contrast, the weak activities of artemisinin and OZ277 against six other protozoan parasites were peroxide bond independent. These data support the iron-dependent artemisinin alkylation hypothesis.The antimalarial artemisinin contains a pharmacophoric peroxide bond within its 1,2,4-trioxane heterocycle. A longstanding hypothesis (17,24,29) to account for the antimalarial specificity of artemisinin is that the peroxide bond undergoes reductive activation by heme released by parasite hemoglobin (Hb) digestion. Although most (95%) of the heme released from parasite digestion is incorporated into the relatively inert hemozoin (12), it is estimated (8) that there is some 100 M of "free" heme/hematin in malaria-infected red blood cells, a more than adequate quantity to react with artemisinin. The irreversible redox reaction between artemisinin and heme produces carbon-centered free radicals or carbocations that alkylate heme (25,32,33) and membrane-associated parasite proteins (2), one of which is the translationally controlled tumor protein (PfTCTP) (4) and another is likely to be PfATP6, a SERCA-type Ca 2ϩ -ATPase (11). Eckstein-Ludwig et al. (11) studied the latter protein in some detail and provided evidence that it may be an important target of the artemisinins.The structural diversity of semisynthetic artemisinins (41) and synthetic peroxides (37) is quite remarkable and, we suggest, has implications for the mechanism of action for this class of antimalarial drugs. In this respect, we thought it might be useful to compare the activities of artemisinin and synthetic peroxide (1,2,4-trioxolane) OZ277 (RBx11160) (40) and their nonperoxidic counterparts deoxyartemisinin (6) and carbaOZ277 (Fig. 1) against a range of protozoan parasites. CarbaOZ277 (1,3-dioxolane) was prepared as a mixture of cis-trans isomers by an acidcatalyzed reaction (9) between the requisite adamantane diol and amino amide cyclohexanone. In vitro antiprotozoal assays for Plasmodium falciparum, Trypanosoma brucei rhodesiense, Trypanosoma cruzi, Leishmania donovani, Giardia duodenalis, Babesia divergens, and Neospora caninum were performed as previously described (1,5,16,28,35,38,40) with standard drug controls ( Table 1).The data in Table 1 shows a clear demarcation in the antiplasmodial activities of artemisinin and OZ277 versus their nonperoxidic analogs deoxyartemisinin (6) and carbaOZ277, confirming the peroxide bond-dependent activity of the former. For artemisinin and OZ277, there were losses of activity greater than 3 orders of magnitude against all of the other protozoa compared to activities against P. falciparum. In contrast, for deoxyartemisinin and carbaOZ277, there were only small differences in activity between P. falciparum and the other protozoa. Only for T. b. rhodesiense was there a significant difference between artemisinin and deoxyartemisinin ve...