Current chemotherapy of human African trypanosomiasis

Abstract Human African trypanosomiasis is a fatal dis- ease caused by Trypanosoma brucei gambiense and Try- panosoma brucei rhodesiense that has re-emerged in recent years. However, very little progress has been made in the development of new drugs against this disease. Most drugs still in use were developed one or more decades ago, and are generally toxic and of limited effectiveness. The most recently introduced compound, eflornithine, is only useful against sleeping sickness caused by T. b. gambiense, and is prohibitively expensive for the African developing countries. We present here an overview of today’s ap- proved and clinically used drugs against this disease.

Drugs available

There are currently four drugs available for treating human African trypanosomiasis: suramin, pentamidine, melarsorprol and eflornithine (Fig. 1). Most of these are toxic, and encounter parasite resistance. The drugs used for the treatment of first-stage sleeping sickness (pen- tamidine and suramin) were both introduced more than half a century ago. Melarsoprol has been in use since 1949, while eflornithine was registered in 1990.

Suramin

Suramin (trade name Bayer 205) is a symmetrical, polysulfonated naphthylamine polyanionic compound first used against sleeping sickness in 1922 (Wang 1995). It is soluble in water, colorless, and has six positive charges at physiological pH. It was developed because two close analogues, trypan blue and trypan red, were shown to be effective antitrypanosomal drugs in the early twentieth century. Like suramin, these compounds cannot pass through intact membranes and are therefore used routinely as biological stains for indicating the vi- ability of cells. The ability of trypanosomes to take up these drugs may be the reason for their selective action (Wang 1995). Suramin, because of its charged nature, binds to many serum proteins and it was postulated that it would be taken up by receptor-mediated endo- cytosis while bound to low-density lipoprotein (LDL) (Vansterkenburg et al. 1993). Recent results, however, suggest that suramin entry into trypanosomes in not mediated via a LDL-specific receptor (Pal et al. 2002) but by a distinct entity not yet identified. The binding to many serum proteins is also responsible for its slow excretion. It can be found in the blood for up to 3 months after injection and has one of the longest half- lives ever documented for drugs given to humans (about 50 days) (Pe´pin and Milord 1994). Binding to numerous enzymes, such as dihydrofolate reductase, thymidine kinase and glycolytic enzymes, by electrostatic interac- tions, results in their inhibition, but it is still uncertain whether any of these inhibitions is responsible for its trypanocidal action. It has been postulated that suramin could be inhibiting the transport of glycolytic enzymes into the glycosomes, thus explaining its slow inhibitory activity against trypanosomes, killing them in a matter of days (Wang 1995). Inhibition of the human immun- odeficiency virus reverse transcriptase led to an unsuc- cessful clinical trial of suramin against AIDS in the 1980s. Since it had some effects on the AIDS-associated cancer, Kaposi’s sarcoma, it was tested against a variety of cancers and was found to have some value against hormone-refractory prostate cancer (Barrett and Barrett 2000).

The present recommended regimen for human tryp- anosomiasis is 5 mg/kg body weight (b.w.) at day 1,10 mg/kg b.w. at day 3, and 20 mg/kg b.w. at days 5, 11, 23, and 30, given by slow intravenous injection. This is mainly used for the treatment of early stage Trypanosoma brucei rhodesiense human sleeping sickness (Legros et al. 2002). Subcutaneous or intramuscular injections are not recommended because they cause local inflammation and necrosis (Pe´pin and Milord 1994). For many reasons (treatment failures, duration of treatment, cost and the need for intravenous injection) suramin monotherapy has rarely been used recently for Trypanosoma brucei gambiense early-stage disease (Pe´pin and Milord 1994). Because of its poor central nervous system penetration, suramin is not an efficient treatment for late-stage trypanosomiasis, although it has been used as a pre-treatment to reduce the toxicity of melarsoprol or to ‘‘sterilize’’ patients until they reach the hospitals where melarsoprol will be administered. Although it has shown some synergistic effects with other trypanocidal drugs, including eflornithine, nifur- timox and 5-nitroimidazoles, in animal studies, such synergism has not been demonstrated in humans (Pe´pin and Milord 1994).

Renal toxicity is the most common problem of suramin monotherapy, although it is usually mild. Poly- neuropathy and stomatitis have also been described. A test dose of 200 mg is sometimes recommended to pre- vent idiosyncratic reactions (Pe´pin and Milord 1994). Trypanosomal resistance to suramin has not been a se- rious problem after 80 years of treating trypanosomiasis with it.

Pentamidine is an aromatic diamidine introduced in 1937. It is soluble in water and is used as pentamidine isethionate (trade name Pentacarinate). Pentamidine is active against the early stage of T. b. gambiense human infection. The drug has also been used for treating an- timony-resistant leishmaniasis and Pneumocystis carinii pneumonia (Wang 1995). It was developed after the observation that a related compound that induces hyp- oglucemia in mammals, synthalin, had potent anti-ty- panosomal activity (Demise and Barrett 2001). Since it is a di-cationic molecule, it has very slow rates of diffusion across biological membranes. The drug is accumulated intracellularly in excess of 1 mM (Damper and Patton 1976) and has a slow trypanocidal action. Transport of pentamidine occurs through at least three transporters with different affinities for adenosine and other diami- dines. In the bloodstream forms of T. b. brucei, adeno- sine-sensitive (ASPT1), high affinity, and low affinity pentamidine transporters transport pentamidine. The ASPT1 is almost certainly identical to the P2 trans- porter, which is also involved in the transport of ade- nosine, adenine, melaminophenyl arsenicals and other diamidines (berenil, propamidine, and stilbamidine) (de Koning 2001). The presence of multiple transporters explains why in T. b. brucei the loss of P2 transporter activity results in high resistance to melaminophenyl arsenicals, stilbamidine and berenil, medium resistance to propamidine, and very low levels of resistance to pentamidine (Fairlamb et al. 1992).

The mode of action of pentamidine has not been es- tablished. As a di-cation, the molecule interacts elec- trostatically with cellular polyanions, in particular with nucleic acids and the network of circular DNA mole- cules present in the kinetoplast of the parasites, dis- rupting their structure (diskinetoplatidy). However, this may not account for its anti-trypanosomal action be- cause the generation of diskinetoplastic trypanosomes in infected mammalian hosts is not expected to cure the disease (Wang 1995). Given that the drug reaches mil- limolar concentrations within the cells (Damper and Patton 1976), it could be that its toxic effect arises from the inhibition of multiple cellular targets. Interestingly, pentamidine has been shown to decrease the mitoc- hondrial membrane potential of trypanosomatids (Vercesi and Docampo 1992) and act as an uncoupler of oxidative phosphorylation in mammalian mitochondria (Moreno 1996). It also selectively inhibits the plasma membrane Ca2+-ATPase of Trypanosoma brucei brucei (Benaim et al. 1993).

The typical protocol for the treatment of early stage human African trypanosomiasis is seven to ten doses of 4 mg/kg b.w. per day given intramuscularly once daily or every other day (Legros et al. 2002). Pentamidine injection sites are extremely tender and sterile gluteal abscesses are not uncommon. Pruritus, rash, tachycar- dia, nausea and vomiting are also seen. Hypoglucemia as a result of insulin release by the pancreas, hypocalcemia and renal failure have also been reported (Pe´pin and Milord 1994). Pentamidine is not as effective for the treatment of late stage human African trypanosomiasis. It is also not mutagenic or genotoxic and has no adverse effects on pregnancy (Pe´pin and Milord 1994). Resis- tance to pentamidine has been induced in laboratory models and also been found in the field. It is generally due to a diminished ability to import the drug (Wang 1995).

Melarsoprol

Melarsoprol, also known as Mel B, is a trivalent organic melaminophenyl arsenical introduced in Africa in 1949 (trade name Asobal). It is useful for the treatment of late stage T. b. gambiense and T. b. rhodesiense infections. It was originally synthesized by the addition of the heavy metal chelator BAL (British antilewisite) to the arsenic of melarsen oxide (Friedheim 1949). It is poorly soluble in water, alcohol or ether, and is administered intrave- nously, dissolved in propylene glycol. Because of its solvent, melarsoprol injections are very painful. Water- soluble melaminophenyl arsenicals, such as melarsen oxide, are transported into trypanosomes by the P2 adenosine/adenine transporter, which has recently been cloned, but it is not known whether the lipophilic Mel B is also transported by the same mechanism (de Koning 2001). Arsenicals lead to the rapid lysis of trypanosomes but their mechanism of action is not completely under- stood. These drugs may be nonspecific inhibitors of different enzymes forming adducts with a variety of dithiols, such as trypanothione and dihydrolipoate. This would explain their toxic effects and the necessity of using melarsen adducts as trypanocidal agents instead of melarsen oxide itself. The inhibition of glycolytic en- zymes, leading to a block of glycolysis and lysis, is the most likely mode of action (Wang 1995).

The typical treatment protocol consists of three to four series of three or four injections (one per day) separated by rest periods. A new schedule of ten daily doses of 2.2 mg/kg b.w. has been proposed, based in pharmocokinetic investigations and computer modeling, for the treatment of late stage T. b. gambiense. This schedule was demonstrated to be as effective as the previous protocol (Blum and Burri 2002). Although skin reactions were more common with this protocol, the treatment schedule was shorter, decreasing the amount of drug used and the hospital stay, which is of great practical advantage (Blum and Burri 2002). The major problem of therapy with melarsoprol is its toxicity and the possibility of relapse. The worst adverse event is reactive encephalopathy, which occurs in 5–10% of the patients treated, and results in death in 10–50% of those in whom it develops. It has been proposed that this be due to an auto-immune reaction rather than a direct toxic effect of the drug. The concomitant administration of prednisolone reduced the frequency of reactive encephalopathy in some patients. Other severe reactions reported are polyneuropathy (10% of patients) and exfoliative dermatitis (1% of patients). Febrile reactions, diarrhea, pruritus, and abdominal and chest pain also occur. Extravasation of the drug can cause a chemical cellulitis (Pe´pin and Milord 1994). The increasing Mel B resistance in the field, as well as resistance induced in the laboratory, appears to be the result of a loss of P2 transporter function (Kaminsky and Ma¨sser 2000).

Eflornithine

Eflornithine (DL-alpha-difluoremethylornithine) is an analogue of ornithine, first synthesized as an anti-cancer agent, although not yet registered for that use. It has been used against late stage human T. b. gambiense in- fection since the 1980s and was registered in 1990 (trade name Ornidyl). Uptake of eflornithine in T. brucei is possibly a combination of passive and mediated diffu- sion, depending on the extracellular concentration, and it can be accumulated intracellularly to millimolar levels (de Koning 2001). It has a slow action and is trypano- static rather than trypanocidal (Pe´pin and Milord 1994). As a fluorinated amino acid derivative, it is a zwitterion at physiological pH, and therefore poorly absorbable and rapidly excreted in the urine, which explains the brief duration of its action (Wang 1995). Its mode of action is through the suicide inhibition of ornithine decarboxylase, the key enzyme in the pathway leading to the biosynthesis of polyamines: putrescine, spermidine, and spermine, which are essential for cell proliferation. Its selectivity against trypanosomes is due to the shorter half-life of ornithine decarboxylase in these organisms, which leads to the complete inhibition of its activity and depletion of polyamines. Trypanosomes are then trans- formed into the non-dividing stumpy forms which are more vulnerable to the host immune reaction since they cannot change their variant surface glycoprotein (Wang 1995). The requirement of a normal immune response explains why the drug is almost useless in immunosup- pressed patients (Pe´pin and Milrod 1994). Eflornithine is difficult to administer, requiring 400 mg/kg b.w. per day in four daily intravenous infusions for 7–14 days. Re- lapses are more frequent in children, probably due to their more rapid elimination of the drug. Oral eflorni- thine is not as effective and is used only when intrave- nous administration is not possible, such as in congenital trypanosomiasis (Pe´pin and Milord 1994). The drug is relatively safe and the most frequent adverse effects are bone marrow suppression with anemia, convulsions and other neurological effects, as well as osmotic diarrhea (Pe´pin and Milord 1994). Although drug-resistant mu- tants due to the deficient uptake of eflornithine have been isolated in the laboratory, no drug resistance has yet been reported in the field.