Parasitol Res (2003) 90: 7179 DOI 10.

1007/s00436-002-0799-9

O R I GI N A L P A P E R

Andrew Jonathan Nok

Arsenicals (melarsoprol), pentamidine and suramin in the treatment of human African trypanosomiasis

Received: 23 October 2002 / Accepted: 31 October 2002 / Published online: 31 January 2003 Springer-Verlag 2003

Abstract Human African trypanosomiasis (HAT), otherwise known as sleeping sickness, has remained a disease with no eective treatment. Recent progress in HAT research suggests that a vaccine against the disease is far from being successful. Also the emergence of drugresistant trypanosomes makes further work in this area imperative. So far the treatment for the early stage of HAT involves the drugs pentamidine and suramin which have been very successful. In the second stage of the disease, during which the trypanosomes reside in the cerebrospinal uid (CSF), treatment is dependent exclusively on the arsenical compound melarsoprol. This is largely due to the inability to nd compounds that can cross the blood brain barrier and kill the CSF-residing trypanosomes. This review summarises our current understanding on the treatment of HAT.

ability to cross the BBB and kill the CSF residing Trypanosoma brucei gambiense and Trypanosoma brucei rhodesiense parasites. Melarsoprol was rst introduced in 1949 for the treatment of late stage trypanosomiasis, and has remained the main drug of choice to this day (Friedheim 1949). Structure and activity Melarsoprol As shown in the Fig. 1, melarsoprol contains the trivalent arsenic element with a markedly reactive arsenoxide group. The presence of the arsenoxide confers the physicochemical ability of lipid solubility that allows passage across the BBB (Peppin and Milord 1994). Apart from its transport function, the arsenoxide group mediates in the killing of the parasites in the CSF. Modication of the melarsoprol parent ring to generate other analogous compounds can have a signicant impact on its trypanocidal ecacy. Figure 2 summarises the trypanocidal properties of several derivatives of melarsoprol on Trypanosoma brucei (Keiser et al. 2000). The trivalent arsenicals; melarsoprol, melarsen oxide and phenylarsen are highly active with a minimum inhibitory concentration (MIC) of 16.5 ng/ml. The pentavalent form of melarsen and inorganic arsenic oxide are considerably less active with MIC 50 lg and 111 ng/ml, respectively. Melamie and dimercaprol are non-arsenical chemical constituents of melarsoprol and are completely inactive towards the parasites. It is clear from the table that it is the arsenical part of the compound that mediates the action of melarsoprol. Pharmacology and target of melarsoprol Although melarsoprol has been the standard drug of choice for the treatment of second stage human trypanosomiasis since 1949 (Friedheim 1949), it was introduced without undergoing basic pharmacokinetic investigation.

Treatment of human African trypanosomiasis

Arsenicals (melarsoprol) In the rst stage of human African trypanosomiasis, parasites are restricted to the haemolymphatic system and the treatment of the disease is dependent on drugs like suramin and pentamidine which remain within the lymphatic system. The major problem with the treatment of human African trypanosomiasis is the second stage of the disease, during which the trypanosome parasites reside in the cerebrospinal uid (CSF). At this stage of the disease, drugs for the treatment must have the ability to cross the blood brain barrier (BBB). Whereas a lot of work has been done in search of such drugs, to date it is only the arsenical compound melarsoprol that has shown the remarkable
A.J. Nok Department of Biochemistry, Ahmadu Bello University, Zaria, Nigeria E-mail: jandrew@skannet.com Tel.: +234-69-550510

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Fig. 3 Chemical formula of cymelarsen Fig. 1 Chemical formula of melarsoprol

Fig. 2 Structurally modied melarsoprol derivatives and their minimum inhibitory concentration (MIC) (Keiser et al. 2000)

By using the bioassay and enzyme linked immunouorescence sandwich assay (ELISA) techniques, melarsoprol has been reported to have an elimination half life of 35 h in the blood (Burri et al. 1993). When a single dose of 2.2 mg/kg is administered to T. b. rhodesiense patients, the maximum level of melarsen oxide concentration (Cmax range 461848 ng/ml) is detected 15 min after administration. After 8 h, the concentration diminishes to 711% of this level and thereafter has a halflife of 3.9 h with an average clearance rate of 21.5 ml/ min. Usually the transport of melarsoprol into the trypanosome parasite is accomplished by purine tranporters (Carter and Fairlamb 1993). Purine transport is highly developed in trypanosomes as they do not synthesize nucleic acids and must directly acquire them from their hosts. As such, melarsoprol acts as a potentially competing ligand for the purine site on the transport protein. The potential targets of the drug on the parasite are thiol containing enzymes like glycerol-3-phosphate dehydrogenase (G3PDH) which is reported to have very strong binding characteristics to cymelarsen (Fig. 3), an analogue of melarsoprol used in the treatment of animal trypanosomiasis. Also, trypanothione (N1-N8-bis glutathionylspermidine), an unusual, low molecular weight thiol compound, forms a very stable complex with arsenic (Harder et al. 2001). This report suggests that the interaction with the thiol group could be a feasible basis for chemotherapy. Metabolism of melarsoprol Although melarsoprol is known to be an active trypanocide, the active metabolite predominant in the body is melarsen oxide (Fig. 2), which has a half-life of less than 30 min (Keiser et al. 2000). Melarsen oxide is rapidly formed in the body and reaches a maximum level after 15 min before disappearing from the plasma with an apparent half-life of 3.8 h. However, in vitro studies

showed that the breakdown of melarsoprol, measured at room temperature, is a slow process with a half-life of 3 days (Ericsson et al. 1997), suggesting the possible contribution of an enzymatic process in vivo in the rapid conversion of melarsoprol to melarsen oxide. In addition, a product of melarsoprol metabolism is bound to plasma proteins with a size of more than 20 kDa. This is proposed as the mechanism that leads to the accelerated conversion of melarsoprol to melarsen in vivo (Keiser et al. 2000). The proteins in this category include albumin (66 kDa) and a1-glycoprotein (44 kDa). The protein-melarsen complex acts like a reservoir that is slowly depleted. Melarsoprol has a long half-life of 3.5 h in the serum and a very slow elimination time from the CSF with a half-life of 120 h. This kinetic behaviour has been used to explain the delayed increase of the drug in the CSF (Burri et al. 1993). Administration The treatment regimen for human African trypanosomiasis with melarsoprol is complicated . Patients are usually given three series of four intravenous injections with interval of 10 days between each series (Freidheim 1949). This schedule may not be the most eective and could possibly account, in part, for the frequently reported side eects, the most severe being reactive encephalopathy in up to 10% of the treated patients with a mortality rate of up to 5% (Pepin and Milord 1994). The drug is administered at a dose of 3.6 mg/kg according to the following treatment schedule: day 1, a third of the dose; day 2, two thirds of the dose; day 3, a full dose, and day 4 a full dose. Melarsoprol is only soluble in propylene glycol and is marketed as a 3.6% solution in propylene glycol. Once the vial is opened, it begins to deteriorate. Administering the drug is painful and thrombophlebitis at the site of injection is a common feature. Little attention has been given to the improvement of the method of administering the drug, though earlier small scale studies showed oral activity (Onyango et al. 1968). Melarsen oxide is more polar, a 36 mg/ml stock solution in propylene glycol could be diluted 1:2 in water compared with 1:500 for melarsorprol. Though melarsen is toxic to trypanosomes, safety considerations prevented its approval in clinical practice (Freidheim 1949). Interestingly, melarsoprol and melarsen oxide accumulate in the CSF to only 25% of the level found in the blood. This level is only sucient to kill trypanosomes in this compartment as the minimal inhibitory concentration against most

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wild-type trypanosomes is close to the levels obtained in the CSF. Based on pharmacokinetic analysis, computer simulation and an extensive literature search covering schedules previously used and tested, a new schedule for administering melarsoprol has recently been proposed and tested (Burri et al. 2000). The pharmacokinetic model was validated in uninfected vervet monkeys with no unexpected drug accumulation or systemic toxic effects. This schedule has been tested in a small group of T. b. gambiense patients in the Congo and in an open randomised clinical trial conducted on 500 patients in Angola with 100% parasitological cure. There has been a consistent search for a less strenuous and shorter drug regimen. Some workers have described a course of two injections each equivalent to 10 mg/kg separated by a 2-day rest. This regimen was successfully tested in T. b. gambiense infected mice (Mbati et al. 2000) and could prove valuable if it suces for human subjects. Adverse eects The treatment of late stage human African trypanosomiasis with melarsoprol is accompanied by some adverse eects such as cutaneous reactions, polyneuropathy, diarrhoea and fever (Kuzoe 1993). The worst of the eects associated with melarsoprol treatment is that up to 19% of the patients suer reactive encephalopathy (Barret 2000). The debate as to whether this reactive encephalopathy is drug induced or related to immunological reactions against residual or dying parasites has been resolved. Following observations that arsenic oxide was eective against selected leukaemias, the arsenic based trypanocide melarsoprol was used in a clinical trial against leukaemia (Soignet et al. 1999). These authors showed that a high proportion of the patients suered neurological seizures demonstrating that melarsoprol is itself responsible for the reactive encephalopathy. The development of adverse eects in melarsoprol treatment could arise because of the covalent binding of the drugs to known molecules, leading to immune reactions of types IIV, which may turn into immunogens (Keiser et al. 2000). The severity of such reactions is usually inuenced by genetic or host factors and the disposition of the conjugate (Park and Kitteringham 1996). Although the trigger for the encephalopathic syndromes is still unknown, the involvement of the immune system has been proposed. Nevertheless, the causes remain controversial. Hunter et al. (1992) suggested that insucient drug levels which lead to the complete elimination of trypanosomes from blood stream but not from the CSF might provoke a violent inammatory response of activated astrocytes after the restoration of immune competence. In addition, high initial doses can lead to a massive dying of trypanosomes, leading to the release of immunocytes that subsequently attract an array of antibodies (Pepin and Milford 1994).

Resistance The failure of melarsoprol to cure 10% of the late stage sleeping sickness patients possibly relates to the fact that these individuals accumulate sub-curative levels of the drug in the brain (Burri and Keiser 2001). However, one study has indicated that the levels of drug are similar in the CSF of relapsing and non-relapsing patients, so parasites resident at other extravascular sites may be a key to treatment failure (Burri et al. 2000). In some regions, treatment failures have reached an alarming level of 30% (Legros et al. 1999). Parasites retrieved from the patients with these treatment failures were less responsive to melarsoprol than parasites isolated from other foci (Matovu et al. 2001a). This clearly points to some form of mutation towards resistance. Indeed arsenic refractory parasites do possess an unusual amino purine transporter which accumulates melarsoprol and the loss of this transporter in the parasite leads to drug resistance (Carter and Fairlamb 1993, Barret and Fairlamb 1999). T. brucei contains several of the purine nucleoside transporter activities that includes P2, which carries adenosine and the nucleobase adenine, and the second P1, which appears to be a general purine nucleoside transporter (Carter and Fairlamb 1993). The P2 transporter also interacts with melaminophenyl arsenicals and diamidines and has been conrmed to be critical in the uptake of arsenicals (Carter and Fairlamb 1993). The incidence of drug resistance to melarsoprol is centred on the mutation of these transporters. Since nucleic acid transporters are involved in the uptake of melarsoprol, a mutation will have a direct consequence on the response of the parasite to the drug. Experiments on the genetic variants of the TbAT1 adenosine transporter have been conrmed in relapse infections following melarsoprol therapy (Matovu et al. 2001b). The analysis of 65 T. b. gambiense isolates with a high melarsoprol treatment failure from north-west Uganda, revealed that 38 had a mutated TbAT1. Furthermore, all of the individuals contained the same set of nine mutations in their TbAT1 gene. Of these, ve point mutations resulted from amino acid substitution, one resulted from the deletion of the amino acid of an entire codon and three were silent point mutations. Identical sets of mutations were also found in a drug resistant T. b. rhodesiense isolate from a relapsing patient from northern Angola. These observations demonstrate the unique conservation in the main set of mutations relevant to resistance to the drug, even from distant locations. Further to the role of the thiol group as a vital target for melarsoprol, the overexpression of the metal-thiol conjugate transporter TbMRPA has been reported to lead to melarsoprol resistance in T. b. brucei (Shahi et al. 2002). Pentamidine Pentamidine (Lomidine R) is an aromatic diamine [1,5-bis (4-amidi-phenoxypentane]) (Fig. 4). It has been

74 Fig. 4 Chemical formula of pentamidine

used for several decades in the chemotherapy of African trypanosomiasis, antimony-resistant leishmaniasis and Pneumocytis carini pneumonia (Sands et al. 1985). Pentamidine was developed after the observation that a related compound, synthalin, which induces hypoglycaemia in mammals, had profound antitrypanosomal activity (Sands et al. 1985). Pentamidine has remained one of the very popular drugs in the treatment of the early stage of gambiense sleeping sickness. The recommended pentamidine regimen for the treatment of early stage trypanosomiasis is seven to ten injections of 4 mg base/kg body weight given daily or on alternate days. The drug is ineective after the trypanosomes have entered the CNS. However, pharmacokinetic studies aimed at improving dose schedules for pentamidine measured in plasma, whole blood and CSF, showed small amounts of the drug in the CSF prompting suggestions that it may be useful during the early part of the late stage treatment of human African trypanosomiasis (Bronner et al. 1991, Doua et al. 1996). Mode of action and resistance Pentamidines (diamidines) are dicationic molecules that have very slow rates of diusion across biological membranes. Diamidines act directly against the parasites, independently of their physiological action against the host. Because the lethal intracellular concentrations of pentamidine are in excess of 1 mM (Damper and Patton 1976), the transport of diamines is a necessary rst step to drug action. The importance is underlined by the observation that in Leishmania spp. and T. b. brucei, pentamidine resistance is associated with a reduced uptake (Carter et al. 1995; Basselin et al. 1997), although at least one report shows a pentamidine resistant T. b. brucei strain which was not decient in drug accumulation (Berger et al. 1995). While studying the eect of pentamidine on the trypanosomatid Crithridia fasiculata, Gutteridge (1969) observed that it inhibited the accumulation of C14 lysine and speculated that pentamidine may selectively enter the parasite through a combined lysine/arginine transporter. In a related work, pentamidine was found to

inhibit the transport of several amino acids (Basselin et al. 1997) and polyamines (Basselin et al, 1997, 2000) in Leishmania spp. However, the inhibition was non-competitive, suggesting that the drug is a non-active site ligand or analog. The role of transporters is central in the mode of action of pentamidine in T. b. brucei and experiments have shown that pentamidine and berenil abrogated melarsoprol induced cell lysis in a similar manner to adenosine and adenine (Carter et al. 1995; Scott et al. 1996a). Likewise, adenine strongly inhibited (H3) pentamidine uptake (Carter et al. 1995), leading to the tentative conclusion that a P2 transporter is involved in the uptake of diamines and melaminophenyl arsenicals. The apparent paradox of a selective adenine/adenosine transporter with little anity for other purines/ pyrimidines while transporting such diverse structures as melarsoprol and pentamidine with Ki values of less than 1 mM (Carter et al. 1995; de Koning and Jarvis 1999) was resolved with the elucidation of its substrate recognition motif (Barret and Fairlamb 1999; de Koning 2001). Fig. 5 illustrates the substrate recognition motif for both the P1 and P2 nucleoside transporters of T. b. brucei. Both transporters bind to adenosine with similar anity, but interact with entirely dierent parts of the molecule, e.g. P1 appears to form 2-hydrogen bonds with hydroxyl groups of the ribose group leading to its inability to transport nucleobases, diamines or melanine compounds. On the other hand, P2 forms its main interactions with the nitrogen residue at position 1 and the amine group at position 6 of the purine ring, and does not bind to the ribose moiety (Carter and Fairlamb 1993; de Koning and Jarvis 1999). Some factors that contribute to the high anity binding of adenine are proposed to include Pi-Pi interactions between the purine ring and aromatic residues on the binding pocket and electrostatic interactions with N-9. The main H2N-C (R1)=NR2 motif can be integrated in a pyrimidine or purine ring and it is present in both diaminidines (amide group) and melaminophenyl arsenicals which are associated with aromatic systems and an electronegative group para to the amidine (de Koning and Jarvis 1999).

75 Fig. 5 Substrate recognition motifs of Trypanosoma brucei brucei P1 and P2 transporters. Parts of the adenosine molecule implicated in binding to the P1 or P2 transporter are indicated with arrows, and the probable mode of interaction is stated. Based on de Koning and Jarvis (1999)

This model favours P2 mediating the uptake of diaminidines in African trypanosomes. Experiments have shown that pentamidine inhibits P2 mediated adenosine uptake with Ki 0.5 lM, and even a 100-fold increase in adenosine has no eect on its uptake, lending support to the possible presence of multiple transporters on T. b. brucei. Indeed, the existence of such transporters has been reported in both procyclic and blood stream forms of the parasite. In the blood stream forms, pentamidine is transported by an adenosine sensitive pentamidine transporter (ASPT1), a high anity pentamidine transporter (HAPT1), and a low anity pentamidine transporter (LAPT1) with KM values of 0.26, 36 and 56 lM, respectively (de Koning 2001). The adenosine sensitive transporter is similar to P2, based on their identical Ki values for purine, melaminophenyl arsenicals and diamidines. Like P2, ASPT1 is expressed in the blood stream but not the procyclic forms of the parasite. The fractional contribution of each transporter is easily delineated from the KM and Vmax values. From the parameter values determined for each of the transporters, it was feasible to compute the contribution of each transporter to the total pentamidine inux at a given concentration. The plasma pentamidine concentration in patients treated with the standard 4 mg/kg dose has been determined to be within 0.52.5 lM. Over this range the contribution of LAPT1 increases from 13 to 35% of the total pentamidine uptake due to its very high capacity. It is only at concentrations less than 0.1 mM that HAPT1 contributes more than 10% of the total uptake (Waalkes and Devita 1970). Studies on this model of the three-pentamidine transporters have far reaching implications for the development of pentamidine resistance and for the potential cross-resistance with other trypanocides. As several transporters are present, there will be the tendency to switch over once there is a mutation on one transporter. It would thus seem that a transport-related pentamidine resistance might not develop readily as resistance to melarsoprol is dependent on uptake by P2 only. The role of these transporters and mutation related features on P2 are seen in melarsoprol resistance which is rapidly growing (Legros et al. 1999; Kaminsky and Maser 2000), whereas very few cases of pentamidine refractory T. b. gambiense have been reported (Kaminsky and Maser 2000). The use of pentamidine as a mass prophylactic against Gambian sleeping sickness by French, Belgian and Portuguese health authorities

(Berger et al. 1995), is a clear indication that even if there were mutations in the transporters, the eective uptake of the drug was still retained. Adverse eects Some treatment failures and adverse eects, such as nephrotoxicity and diabetes mellitus, have been reported. Pentamidine binds with great avidity to imidazole receptors (Wood. et al. 1998), which explains the hypotension and some other side eects characteristically induced by the drug. The P2 transporter already described is capable of transporting diamidine drugs, and the selection of parasites resistant to one of these classes of drugs can often underlie cross-resistance in others (Barret and Fairlamb 1999). Other targets and metabolism Pentamidine interacts with a number of cellular anions and binds tightly to the minor groove of DNA, inhibiting nucleic acid replication (Nun and Neide 1995). One possible target in trypanosomes is the highly intercalated network of circular DNA molecules that comprise the mitochondrial genome or kinetoplast (Shapiro 1993). Pentamidine inhibits the self splicing of the GpI intron CaLSM from transcripts of the 26rRNA gene of Candida albicans and prevents the formation of the catalytically active F-band conformation of the precursor RNS by altering the ribonuclease cleavage pattern of CaLSM RNA. The drug has also been shown to specically inhibit S-adenosyl methionine decarboxylase (Ado-MetDC), suggesting that direct inhibition of polyamine synthesis could be involved. Moreover, it has been shown to inhibit trypanothione metabolism (Bitonti et al. 1986). Trypanosomes do not metabolise pentamidine. In contrast, the rat liver microsomes readily metabolise the drug via the cytochrome P450 dependent mixed function oxidases to form a number of products including 2- and 3-hydroxy pentamidine, the amidoxims, N-hydroxypentamidines and N1N1-dihydroxypentamidines and the 1-hydroxylation products parahydroxybenzidine, 5-(4-amidine phenoxy) pentoic acid and, 5-(4-amidine phenoxy) 1-1-pentanol. Recent research has demonstrated that pentamidine resistance by parasites is not associated with the transformation of the drug (Berger et al. 1993).

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Suramin Suramin (Germin, Naganol, Morany, Bayer 205) is a colourless, polysulphonated, symmetrical naphthalene derivative (Fig. 6). This drug was initially developed by a German scientist and rst used against sleeping sickness in 1922 (Voogd et al. 1993). The development of suramin followed observations that a number of similar naphthalene dye substances including trypan red and trypan blue had a marked trypanocidal activity. Suramin has six negative charges at physiological pH, thereby precluding its diusion across biological membranes and possible use against late stage trypanosomiasis because it does not cross the BBB (Hawking 1940). Suramin is generally considered the drug of choice for the early stages of human African trypanosomiasis, especially T. b. rhodesiense infections. In humans it is also used against adult forms of the larial parasite
Fig. 6 Structural formula of suramin

Onchocerca volvulus. In domestic animals, suramin is used primarily as a curative drug against Trypanosoma evansi infection in camels and T. evansi and Trypanosoma equiperdum infections in equines (Zhang and Baltz 1991). Mode of action The combination of size and charge makes suramin thermodynamically unsuitable to have a specic transporter. It was therefore hypothesized that the drug is taken up by endocytosis in trypanosomes (Vansterkenurg et al. 1993). On account of its charge, suramin easily binds to many serum proteins. At the attainable levels of 75100 lM (Collins et al. 1986), more than 75% of suramin is bound to serum proteins (Vansterkenurg et al. 1993).

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Some serum proteins, including low-density lipoproteins (LDL) and transferin but not albumin, are taken up through a receptor-mediated endocytosis (Collins et al. 1986) and suramin enters the parasite specically bound to LDL (Vansterkenurg et al. 1993). Accumulation of the drug in trypanosomes is relatively slow. LDL is a molecule consisting of a non-polar core of approximately 1,500 cholesterol molecules esteried to a long chain fatty acid. The cholesterol core is surrounded by a polar core of phospholipid together with unesteried cholesterol and apoprotein (Goldstein et al. 1979). Because of the presence of the positively charged arginine and lysine residues of apoprotein B, it is able to interact with negatively charged amino acid residues located on the LDL receptor of mammalian cell lines (Mahley et al. 1977, Brown et al. 1980). In essence, suramin directly interferes with the complex formed between LDL and its receptors, thereby inhibiting the transport of LDL. Schneider et al. (1982) isolated the LDL receptor from bovine adrenal cortex membranes and showed that suramin, inhibits the binding of LDL to this receptor. It also displaces the ligand from the receptor. This nding implies that there could be a correlation between suramin toxicity for certain cell types and the abundance of LDL receptors on their membranes. The LDL receptor varies within tissues (Mahley et al. 1977); while there are hardly are any receptors on erythrocytes, the membrane of the adrenal cortex contains them in large numbers. This probably explains why suramin does not interfere with erythrocyte function (Hawking 1978). This is corroborated by the fact that there is no report of any change in the haematological parameters of patients treated with suramin, but adrenal failure is one of its most serious side eects, especially after prolong treatment (Feuillan et al. 1987). Several studies have shown that suramin may interfere with the metabolism of trypanosomes in at least three dierent ways: 1. It hampers the receptor mediated uptake of LDL, the carrier of cholesterol which is required for parasite growth, by the formation of a complex with LDL. 2. Suramin, which enters via receptor mediated endocytosis in association with LDL, is most likely to accumulate inside the lysosome. In this respect it is noteworthy that several of the enzymes that suramin encounters on its way from the extracellular uid to the secondary lysosomes (i.e. 3-nucleotidase and protein kinase, both bound to the plasma membrane of the trypanosome, acid phosphatase and acid pyrophosphatase in the agellar pocket and phospholipase A1) are all inhibited by suramin (Opperdoes et al. 1987). 3. Many glycolytic enzymes located inside the glycosome on the African trypamosome carry a high positive charge, and as a consequence are all inhibited by micromolar levels of suramin (Williamson et al. 1993). In comparison to less positively charged homologous enzymes from other organisms and

mammals, they are insensitive to suramin inhibition (Misset et al. 1986; Misset and Opperdoes 1987; Wierenga et al. 1987;Williamson et al. 1993). Resistance The metabolism of suramin does not seem to be a cause of resistance, given that it is extremely stable in vivo remaining in the blood stream with a half life of 4454 days after which it is nally excreted (Collins et al. 1986). In contrast to diaminidines and melarsoprol, it is unlikely that the mechanism leading to suramin resistance will be the result of a dramatic change in uptake. This is because LDL is essential for the proliferation of T. b. brucei, which is unable to synthesize fatty acid and cholesterol de novo (Dixon et al. 1971). Alternatively, suramin resistance has been postulated to develop as a result of changes in the drug target by expression of a drug extrusion mechanism (de Koning 2001). The induction of suramin resistance in vivo with a stable phenotype after transmission by the tsetse y has been reported (Scott et al. 1996b). Dosage For prophylactic purposes, doses of 12 g repeated at 10-day intervals are recommended. Complexes of suramin with cationic drugs, especially quinapyramine, are more eective in prophylaxis than suramin alone. But such complexes have not been widely accepted for eld use because of the high cost of the complex formulations and the severe tissue reactions that occur at the site of injection (Scott 1996b). It is unclear whether the improved prophylactic activity is due to pharmacodynamic or pharmacokinetic factors. However, it is clear that suramin is synergistic with a number of drugs including tryparsamide, puromycin and diaminazine (Scott 1996b). Action on other diseases At physiological pH, suramin bears six negative charges, thus giving it a good capacity to inhibit many enzymes. A key enzyme of the human immunodeciency virus (HIV), reverse transcriptase, is known to be strongly inhibited by suramin (De Clercq 1979). It also prevents HIV from penetrating CD4+ cells. This observation prompted the American AIDS lobby to push hard for a clinical trial of suramin against AIDS in the 1980s. The drug had no impact on the progression of AIDS, although minor eects on some incidences of the AIDSassociated cancer Kaposis sarcoma were observed (Collins et al. 1986). The drug was then tested against neoplastically transformed cell lines and went into trial against a variety of cancers where it was shown to be of value against hormone refractory prostrate cancer, a leading cause of cancer-related death in men. A clear mechanism of action against cancer remains elusive, although a variety of ideas have some support, including

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an anti-angiogenesis eect as well as the interaction of drugs with a growth factor receptor with a consequent eect on the cell signalling pathway. Suramin has recently been shown to induce the reversal of chronic cerebral vasospasm in experimental subaracnoid haemorrhage. The mode of action involves the inhibition of growth factor receptors (receptor tyrosine kinases) and G protein coupled P2Y receptors (Kimura et al. 2002).

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