A Novel Endogenous Antimalarial:  Fe(II)-Protoporphyrin IXα (Heme) Inhibits Hematin Polymerization to β-Hematin (Malaria Pigment) and Kills Malaria Parasites

Abstract

The polymerization of hemoglobin-derived ferric-protoporphyrin IX [Fe(III)PPIX] to inert hemozoin (malaria pigment) is a crucial and unique process for intraerythrocytic plasmodia to prevent heme toxicity and thus a good target for new antimalarials. Quinoline drugs, i.e., chloroquine, and non-iron porphyrins have been shown to block polymerization by forming electronic pi-pi interactions with heme monomers. Here, we report the identification of ferrous-protoporphyrin IX [Fe(II)PPIX] as a novel endogenous anti-malarial. Fe(II)PPIX molecules, released from the proteolysis of hemoglobin, are first oxidized and then polymerized to hemozoin. We obtained Fe(II)PPIX on preparative scale by electrochemical reduction of Fe(III)PPIX, and the reaction was monitored by cyclic voltammetry. Polymerization assays at acidic pH were conducted with the resulting Fe(II)PPIX using a spectrophotometric microassay of heme polymerization adapted to anaerobic conditions and the products characterized by infrared spectroscopy. Fe(II)PPIX (a) did not polymerize and (b) produced a dose-dependent inhibition of Fe(III)PPIX polymerization (IC(50) = 0.4 molar equiv). Moreover, Fe(II)PPIX produced by chemical reduction with thiol-containing compounds gave similar results: a dose-dependent inhibition of heme polymerization was observed using either L-cysteine, N-acetylcysteine, or DL-homocysteine, but not with L-cystine. Cyclic voltammetry confirmed that the inhibition of heme polymerization was due to the Fe(II)PPIX molecules generated by the thiol-mediated reduction of Fe(III)PPIX. These results point to Fe(II)PPIX as a potential endogenous antimalarial and to Fe(III)PPIX reduction as a potential new pharmacological target.

Reaction of artemisinin with haemoglobin: implications for antimalarial activity

“…In addition, HAA has a markedly lowered oxidative potential, which can tilt the haemin (Fe 3+ ) to haem (Fe 2+ ) ratio in favour of the latter. Since haem is a potent inhibitor of haemozoin biosynthesis [45], HAA-mediated inhibition of haemozoin synthesis (indicated by 'I') is likely to occur also by creation of a more reducing environment in the food vacuole. The broken triangle at the bottom suggests the inverse relationship between the concentration of haemarts and the yield of haemozoin biosynthesis.…”

“…A reducing environment created by HA/HAA formed because of the alkylation of haem by artemisinin could shift the haem-haemin equilibrium towards the former. Such a mileu, which causes enrichment of haem, would antagonize haemozoin formation, since haem is known to be a very potent inhibitor of haemozoin synthesis [45].…”

Effects of Amino Acids on Malarial Heme Crystallization

“…Consequently, this state of iron does not allow the formation of the carboxylate bond needed for BH formation. 37) In addition, heme degradation by reduced glutathione has been detected under physiological conditions. Therefore, the degradation of heme by cysteine was further examined in an Strong or weak effect is indicated by the number of arrows, based on the effect of amino acids at their nearly 50% saturated concentration in comparison with control sample.…”

Papel do IP3 na transdução de sinal e função da heme oxigenase em Plasmodium falciparum.

“…Metaloprotoporfirinas são potentes inibidores da polimerização in vitro da hemina (Basilico et al, 1997;Martiney et al, 1996;Monti et al, 1999). O íon metálico localizado no centro da protoporfirina IX tem importante função em inibir as interações π-π formadas entre a hemina e o íon metálico (Cole et al, 2000).…”

“…Após estudos que comprovaram que metaloprotoporfirinas são capazes de inibir a hemozoína (Basilico et al, 1997;Martiney et al, 1996;Monti et al, 1999), houve grande interesse em testar a viabilidade dos parasitas da malária na presença destes compostos (Begum et al, 2003). Interessante ressaltar que os polímeros e micelas podem ser usadas para aumentar a permeabilidade celular das metaloprotoporfirinas (Ding et al, 2011;Engelmann et al, 2007).…”