Randomized Controlled Trial of Levamisole Hydrochloride as Adjunctive Therapy in Severe Falciparum Malaria With High Parasitemia

Severe malaria in adults has a high mortality rate between 15% and 30%, and no adjunctive treatment has been able to reduce this unacceptably high figure [1]. Pivotal in the pathogenesis of cerebral and severe Plasmodium falciparum malaria is sequestration of erythrocytes containing mature stages of the parasite, which is augmented by endothelial cell activation and dysfunction, culminating in microvascular obstruction [2, 3]. A cheap and safe drug targeting cytoadherence and thus sequestration in vital organs would be a promising candidate for adjunctive treatment in severe falciparum malaria. Cytoadherence is mediated by the parasite protein P. falciparum erythrocyte membrane protein 1 (PfEMP1) [4], and endothelial receptor molecules, including CD36 [5, 6] intercellular adhesion molecule 1 (ICAM-1) [7], E-selectin, and vascular cell adhesion molecule 1 (VCAM-1) [8]. Studies under physiological flow conditions have shown that the different adhesion molecules interact synergistically. The infected red blood cells tether and roll on endothelial receptors with a low affinity (ICAM-1, VCAM-1, and P-selectin), facilitating the subsequent more-definite adhesion to CD36 [9–12]. Peptide mapping studies have revealed that the critical region on PfEMP1 involved in binding to CD36 is localized to a 179–amino acid sequence within the cysteine-rich interdomain region 1 (CIDR-1). A recombinant 179–amino acid peptide expressed in yeast (PpMC-179) represents this minimal CD36-binding domain and is able to reduce cytoadherence of P. falciparum–infected red blood cells (IRBCs) by >80% in a human/severe combined immunodeficiency (SCID) mouse chimeric model [13]. It was shown more recently that binding of PpMC-179 to CD36 on human dermal microvascular endothelial cells activates a physically closely associated Src-family kinase [14, 15]. This activation results in an increased affinity of CD36 for IRBCs through activation of an ecto-alkaline phosphatase (ecto-ALP), causing dephosphorylation of the extracellular domain of CD36 [16]. Selective inhibition of the Src-family kinase by the pyrazolopyrimidine PP1 inhibits adhesion of IRBC by >70% in a flow-chamber assay and in a human/SCID mouse chimeric model. Levamisole hydrochloride is an old and cheap drug, primarily used to treat intestinal helminths. It is a specific alkaline-phosphatase inhibitor, and by targeting of the endothelial ecto-ALP it decreases the adhesion of IRBCs to CD36. We performed a small randomized controlled trial in 21 patients with uncomplicated falciparum malaria on the antiadhesive effect of a single oral dose of 150 mg of levamisole as adjunctive treatment to quinine. It was shown that in patients treated with levamisole, significantly more late-stage parasites could be detected in the peripheral blood over time, which was attributed to the inhibition of sequestration with levamisole [17]. No adverse effects of levamisole were observed in this study. Vomiting is a common side effect of levamisole when used in repeated high doses as an adjunctive therapy with 5-fluorouracil in colon cancer chemotherapy, but is uncommon in low doses as used in antihelminth treatment and this study [18]. Because of these encouraging results, we performed an open-label randomized controlled trial of levamisole hydrochloride as adjuvant treatment in adult patients with severe falciparum malaria to study its effect on sequestration of infected red blood cells. At the time of the study, artesunate had replaced quinine as first-line treatment for severe malaria, so levamisole was studied as an adjunct to artesunate rather than quinine [19].