Chloroquine and Post Malaria Anaemia Study
Condition: Malaria Anaemia
MeSH terms: Malaria; Anemia
Study type: Interventional

Summary: 
      The pathogenesis of post-malaria anaemia is multifactorial. Iron supplementation remains the
      mainstay of management of moderate and severe anaemia; however the management of mild anaemia
      (Hb 80-110g/l) is problematic as population supplementation studies of children in malaria
      endemic areas demonstrate adverse effects in children with mild anaemia. We hypothesize that
      the anti-inflammatory, anti-malarial and anti-macrophageal iron loading effects of
      chloroquine could make it a useful drug in the management of mild post malaria anaemia. To
      test this hypothesis, we plan to randomize children (aged 12 months to 6 years) with post
      malaria anaemia (Hb 70-110g/l) to receive a standard anti-malarial treatment, co-artemether .
      All children with parasitologic cure after three days on treatment will be randomised to
      receive either weekly chloroquine or weekly placebo starting from day 10 till day 90. By
      comparing the curve of haemoglobin change between day 3 and day 30 in the placebo arms of the
      two groups, we will test the effect of chloroquine vs. ACT treatment on macrophageal iron
      loading and release in acute clinical malaria. By comparing the haemoglobin change between
      day 3 and day 90 between the weekly chloroquine arms and the weekly placebo arms we will test
      the longer-term anti-inflammatory and anti- malarial effects of weekly chloroquine
      prophylaxis. In addition to the primary endpoint, we plan to assess potential mechanisms of
      action by determining parasite clearance, peripheral cytokine production and iron flux
    

Detailed description: 
      Pathogenesis of Malaria Anaemia

      Although the pathogenesis of malaria anaemia is not completely understood (1), mechanisms
      that have been proposed include immune and non-immune-mediated hemolysis of parasitized and
      non-parasitized red blood cells, bone marrow dysfunction and iron delocalisation (2-4).
      Plasmodium falciparum infection is associated with changes in the cell membranes of infected
      and uninfected erythrocytes causing alterations in membrane permeability and susceptibility
      to early hemolysis. It is also associated with bone marrow dysfunction probably due to
      malaria-induced abnormalities in erythroid progenitors, CFU-e and BFU-e (5,6). Jason et a
      (l7) have shown that malaria exerts a pro-inflammatory immune response in children and is
      associated with release of cytokines which act at various levels of the erythropoietic
      pathway to cause anaemia. These cytokines include human tumour necrosis factor (TNF)(8),
      interleukin (IL)-12, IL-10, IL-6 and IL-4 (7,9), and interferon-γ (IFN-γ) (9). In addition to
      the absolute concentrations of the cytokines, the balance between opposing anti- and
      pro-inflammatory responses may determine the clinical characteristics of malaria, including
      the level of anaemia (7) e.g. high TNF/IL-10 ratio may contribute to the reversible bone
      marrow suppression seen in malaria patients (10,11). Several investigators have reported the
      persistence of a fall in haemoglobin after successful treatment of malaria1 (2). Although the
      precise mechanism for this persistent anaemia is unclear, it is likely to be multifactorial.
      Studies conducted among Gambian children1 (3), French adults (14), and Thai adults (15) show
      that cytokine levels remain elevated for one to four weeks after a successful treatment for
      malaria; and Camacho et al (12) reported more than 80% of Thai adults with malaria had
      persisting anaemia on days 7, 14, and 21 after successful treatment, while 55% were still
      anaemic on day 28. On day 28, 46% of these subjects had hypoproliferative erythropoiesis
      while 7% had ineffective erythropoiesis (12). Although the study did not explore the
      mechanisms behind these findings, it is likely that persisting elevation of inflammatory
      cytokines reported by earlier workers (13-15) could be contributory. Greenberg and colleagues
      (16) have suggested that unusually strong and prolonged Th-1 response in conjunction with an
      inadequately developed Th-2 response may contribute to persistent anaemia after clearance of
      parasitaemia. Other workers have reported that persistent microscopically undetectable
      parasitaemia following successful treatment with an antimalarial was associated with
      protracted bone marrow suppression (17).

      Macrophages and Malaria Anaemia Macrophages are responsible for the removal of senescent red
      blood cells from the body. They process these erythrocytes to release the heme iron which is
      then transported to the peripheral tissues including the bone marrow. The mononuclear
      phagocyte system plays two major roles in iron metabolism (18) - iron recycling from
      senescent erythrocytes and serving as a large storage depot for excess iron. This
      macrophageal recycling accounts for most of the 20-24mg of iron required daily for
      haemoglobin production. The cytokines released as part of the body's response to infection
      with Plasmodium cause the macrophages to release oxygen and nitrogen radicals leading to
      oxidative damage to parasitized and non-parasitized red blood cells, enhancing their removal
      from the circulation and contributing to anaemia (19). In addition, malaria-induced
      inflammation impedes release of iron from the macrophage-monocyte system hampering the supply
      of iron to the erythropoietic tissues. This causes considerable delocalisation of iron within
      the macrophage/monocyte system which makes iron unavailable to the marrow cells for
      erythropoiesis (18). The clinical practice of administration of iron for malaria anaemia is a
      result of the observation that serum iron is often low in such patients, as demonstrated in
      several animal and human studies (20,21). That the hypoferraemia observed in malaria could be
      largely due to iron sequestration in the macrophages is supported by reports of decreased
      peripheral iron levels despite normal or increased bone marrow iron (22,23).

      Iron supplementation versus macrophageal iron mobilization The recent controversy regarding
      routine iron supplementation of children in malaria endemic areas with high prevalence of
      anaemia has further highlighted the need for alternative therapeutic regimens in children
      with malaria-associated anaemia. The genesis of the 'iron supplementation controversy' arose
      from several reports of increased morbidity and mortality from malaria and other infections
      in children from malaria endemic regions supplemented with iron. Recent evidence (reviewed by
      Prentice et al, Position paper for WHO Expert Consultation, Lyon, June 2006) suggests that
      the risk of adverse outcomes is less in more anaemic children. However, a growing body of
      evidence points to the poor rationale for giving iron supplements to children with mild
      malaria anaemia. First, during acute malaria, there is reduced iron absorption (Doherty et
      al. submitted, AJCN, 2006) and at least initially, erythropoietic iron supply is likely to
      come from reticuloendothelial macrophages rather than iron supplements. Secondly, the
      hypoferreamia associated with malaria has been shown to be due to iron delocalisation rather
      than absolute iron lack, reviewed in 18. Finally, a number of studies have found little or no
      benefit of giving iron supplements to children with malaria anaemia compared to other
      alternative regimens (24-27). While the management of moderate to severe malaria anaemia (Hb
      <80g/L) is not in contention, the management of mild anaemia remains an enigma because iron
      supplementation in these children, apart from providing questionable benefit, might in fact
      be harmful. Results from a large clinical trial of routine iron supplementation of children
      in an area of both iron deficiency and malaria transmission in Tanzania showed that among
      24,076 children recruited those who received iron and folic acid, with or without zinc, were
      significantly more likely to die or experience adverse events than children who did not
      receive iron and folic acid (28).

      Presently, there are no clear guidelines on the management of children with mild malaria
      anaemia. Mild anaemia in malaria endemic areas is likely due to either or both malaria and
      iron deficiency but distinguishing the iron delocalization of malaria from iron deficiency is
      difficult. It is important to optimize a child's iron nutrition to promote optimal cognitive
      development (29); however iron supplementation of this group is potentially dangerous hence
      the urgent need for alternative management strategies for a very common clinical scenario in
      Africa. Such interventions should take into consideration the complex pathogenesis of malaria
      anaemia including the mechanisms of iron flux and macrophageal iron delocalization during
      Plasmodium falciparum infection. It is likely that reduction of malaria-induced macrophageal
      iron sequestration and inflammation will enhance erythropoietic recovery post-malaria. We
      hypothesize that the anti-inflammatory, anti-macrophageal iron loading and anti-malarial
      effects of chloroquine could make it a useful drug in the management of mild post malaria
      anaemia by reducing macrophageal iron sequestration and interrupting the malaria-induced
      inflammatory process.

      Chloroquine as a macrophageal iron mobilization agent Although resistance has reduced its
      effectiveness in the prevention and treatment of malaria, the non-antimalarial
      pharmacological properties of chloroquine make it a potentially useful therapeutic agent for
      other conditions. Chloroquine has antipyretic and anti-inflammatory properties (30-32).
      Chloroquine exerts a steroid-sparing effect (33), and inhibits the replication of a number of
      viruses such as HSV-1 virus (34), HIV-1 and several AIDS related opportunistic microorganisms
      (35,36). By inhibiting phospholipase A2 and tumour necrosis factor, chloroquine acts as an
      immunomodulator (37,38); and also acts as a lysosome-stabilizing agent. Clinically,
      chloroquine is used as a second line anti-inflammatory drug in chronic conditions like
      rheumatoid arthritis (39).

      Chloroquine and iron metabolism Although the role of chloroquine in iron metabolism is still
      poorly understood, it is likely that many of the effects of chloroquine result from the
      interference with intracellular free iron. Chloroquine, a weak base, accumulates in acid
      intracellular compartments increasing the intracellular pH. Legssyer and co-workers have
      shown that chloroquine significantly reduces incorporation of iron into the liver (20%
      reduction), spleen (20%) and alveolar (400%) macrophages of rats loaded in vivo with iron
      dextran40.

      Chloroquine and post malaria anaemia Chloroquine (CQ) may likely have three effects at
      different time points during the erythropoietic response to malaria. Firstly it may block the
      acute incorporation of iron into reticuloendothelial macrophages during clinical malaria
      associated with haemolysis and iron delocalization. Secondly it may have an anti-inflammatory
      effect. Increased serum levels of TNF-α, IFN-γ and nitric oxide depress erythropoiesis via
      bone marrow depression, dyserythropoiesis and erythrophagocytosis. Continuing inflammation
      after a malarial event may contribute to the slow resolution of anaemia (13-15) and
      chloroquine's anti-inflammatory effect might be a useful adjunctive therapy to continue to
      utilize after its initial antimalarial effect. Lastly chloroquine will have a continuing
      direct anti-malarial effect to both clear microscopically undetectable persistent infection
      and prevent further episodes until haematological recovery is optimized.

      The anti-anaemic effects of chloroquine have been reported by a number of clinical studies.
      Salihu and colleagues41 reported a significant anti-anaemia effect of chloroquine given
      weekly to pregnant women in Cameroon compared to women not given any prophylaxis, even after
      controlling for possible confounders. Other studies among pregnant women in Cameroon (42),
      Burkina Faso (42), Uganda (43) and Thailand (44) all showed significant benefit of weekly
      chloroquine on maternal haemoglobin levels compared to controls. Although these studies were
      carried out on pregnant women, it is likely that similar benefits will occur in children.

      Aim of Study and Hypothesis to be tested In exploring the effect of chloroquine on post
      malaria anaemia, we hypothesize that post-malarial CQ improves erythropoietic recovery after
      standard malarial treatment. We further hypothesize that post-malarial CQ improves
      erythropoietic recovery after co-artemether treatment, by a mechanism other than its
      anti-malarial effect in controlling residual parasitaemia. To test these hypotheses, we plan
      to randomize children (aged 12 months to 6 years) with acute malaria to receive either
      standard anti-malarial treatment (chloroquine plus sulphadoxine/pyrimethamine) or
      artemisinine combination therapy. Three days after commencement of antimalarial treatment,
      the children in each of the two arms, whose parasites have been cleared, will be randomised
      to receive either weekly chloroquine or weekly placebo. By comparing the curve of haemoglobin
      change between day 3 and day 30 in the placebo arms of the two groups, we will test the
      effect of chloroquine vs. ACT treatment on macrophageal iron loading and release in acute
      clinical malaria. By comparing the haemoglobin change between day 3 and day 90 between the
      weekly chloroquine arms and the weekly placebo arms we will test the longer-term
      anti-inflammatory and anti- malarial effects of weekly chloroquine prophylaxis. In addition
      to the primary endpoint (haemoglobin change), we plan to assess potential mechanisms of
      action by determining parasite clearance by PCR detection, peripheral cytokine production (&
      markers of inflammation), and indicators of monocyte iron loading and iron flux.

      **Additional information: During the course of the study, the Gambian Government changed the
      antimalarial drug policy making artemisinine the first line antimalarial and discontinuing
      the use of chloroquine. We were therefore forced to alter the protocol to remove the initial
      chloroquine treatment.
    

Intervention name(s): Chloroquine; Placebo
Intervention type(s): Drug; Drug
Intervention description(s): This is an orange syrup in a 60ml amber coloured glass bottle containing 50mg of chloroquine base per 5mls as the chloroquine phosphate. The syrup was manufactured by Medreich Sterilab Ltd, Avalahalli, Bangalore, India. Chloroquine: weekly treatment of 7.5mg/kg for 90 days; The placebo is an orange syrup in a 60ml amber coloured glass bottle containing sucrose syrup base. The syrup was prepared by the Pharmacy department of the Royal Victorial Teaching Hospital and Atlantic Pharmaceuticals Limited, Banjul
Intervention MeSH terms: Chloroquine
Allocation: Randomized
Intervention model: Parallel Assignment
Masking: Quadruple (Participant, Care Provider, Investigator, Outcomes Assessor)
Primary purpose: Treatment
Keywords: malaria; anaemia; chloroquine; iron delocalisation; macrophages