Suramin: Effectiveness of analogues reveals structural features that are important for the potent trypanocidal activity of the drug.

Suramin was the first effective drug for the treatment of human African sleeping sickness. Structural analogues of the trypanocide have previously been shown to be potent inhibitors of several enzymes. Therefore, four suramin analogues lacking the methyl group on the intermediate rings and with different regiochemistry of the naphthalenetrisulphonic acid groups and the phenyl rings were tested to establish whether they exhibited improved antiproliferative activity against bloodstream forms of Trypanosomes brucei compared to the parent compound. The four analogues exhibited low trypanocidal activity and weak inhibition of the antitrypanosomal activity of suramin in competition experiments. This indicates that the strong trypanocidal activity of suramin is most likely due to the presence of methyl groups on its intermediate rings and to the specific regiochemistry of naphthalenetrisulphonic acid groups. These two structural features are also likely to be important for the inhibition mechanism of suramin because DNA distribution and nucleus/kinetoplast configuration analyses suggest that the analogues inhibit mitosis while suramin inhibits cytokinesis.

Suramin analogues protect cartilage against osteoarthritic breakdown by increasing levels of tissue inhibitor of metalloproteinases 3 (TIMP-3) in the tissue.

Osteoarthritis is a chronic degenerative joint disease affecting millions of people worldwide, with no disease-modifying drugs currently available to treat the disease. Tissue inhibitor of metalloproteinases 3 (TIMP-3) is a potential therapeutic target in osteoarthritis because of its ability to inhibit the catabolic metalloproteinases that drive joint damage by degrading the cartilage extracellular matrix. We previously found that suramin inhibits cartilage degradation through its ability to block endocytosis and intracellular degradation of TIMP-3 by low-density lipoprotein receptor-related protein 1 (LRP1), and analysis of commercially available suramin analogues indicated the importance of the 1,3,5-trisulfonic acid substitutions on the terminal naphthalene rings for this activity. Here we describe synthesis and structure-activity relationship analysis of additional suramin analogues using ex vivo models of TIMP-3 trafficking and cartilage degradation. This showed that 1,3,6-trisulfonic acid substitution of the terminal naphthalene rings was also effective, and that the protective activity of suramin analogues depended on the presence of a rigid phenyl-containing central region, with para/para substitution of these phenyl rings being most favourable. Truncated analogues lost protective activity. The physicochemical characteristics of suramin and its analogues indicate that approaches such as intra-articular injection would be required to develop them for therapeutic use.

100 years since the publication of the suramin formula.

Suramin was the first drug developed using the approach of medicinal chemistry by the German Bayer company in the 1910s for the treatment of human African sleeping sickness caused by the two subspecies Trypanosoma brucei gambiense and Trypanosoma brucei rhodesienese. However, the drug was politically instrumentalized by the German government in the 1920s in an attempt to regain possession of its former African colonies lost after the First World War. For this reason, the formula of suramin was kept secret for more than 10 years. Eventually, the French pharmacist Ernest Fourneau uncovered the chemical structure of suramin by reverse engineering and published the formula of the drug in 1924. During the Nazi period, suramin became the subject of colonial revisionism, and the development of the drug was portrayed in books and films to promote national socialist propaganda. Ever since its discovery, suramin has also been tested for bioactivity against numerous other infections and diseases. However, sleeping sickness caused by Trypanosoma brucei rhodesiense is the only human disease for which treatment with suramin is currently approved.

Synthesis and evaluation of antibacterial and trypanocidal activity of derivatives of monensin A.

The synthesis and biological evaluation of eleven derivatives of the natural polyether ionophore monensin A (MON), modified at the C-26 position, is presented. Eight urethane and three ester derivatives were tested for their antimicrobial activity against different strains of Staphylococcus aureus, Staphylococcus epidermidis, Escherichia coli and Pseudomonas aeruginosa. In addition, their antiparasitic activity was also evaluated with bloodstream forms of Trypanosoma brucei. The majority of the modified ionophores were active against a variety of Gram-positive bacterial strains, including methicillin-resistant S. epidermidis, and showed better antibacterial activity than the unmodified MON. The phenyl urethane derivative of MON exhibited the most promising antibacterial activity of all tested compounds, with minimal inhibitory concentration values of 0.25-0.50 µg/ml. In contrast, none of the MON derivatives displayed higher antitrypanosomal activity than the unmodified ionophore.

Trypanocidal and cell swelling activity of 20-deoxysalinomycin.

The naturally occurring polyether ionophore salinomycin was previously found to display promising anti-proliferative activity against bloodstream forms of Trypanosoma brucei. Here, we report the evaluation of 20-deoxysalinomycin, a naturally occurring homolog to salinomycin, for trypanocidal and cell swelling activity. The concentration of 20-deoxysalinomycin required to reduce the growth rate of bloodstream-form trypanosomes by 50% was determined to be 0.12 µM and found to be 8 times more trypanocidal than that of salinomycin. Moreover, 20-deoxysalinomycin and salinomycin displayed similar cytotoxic activity against human HL-60 cells. Measured as the ratio of cytotoxic to trypanocidal activity, 20-deoxysalinomycin thus exhibits a four-fold higher selectivity compared to salinomycin. The stronger trypanocidal activity of 20-deoxysalinomycin is attributed to an enhanced ability to induce cell swelling in trypanosomes. The findings support 20-deoxysalinomycin as a useful lead in the rational development of new and improved anti-trypanosomal drugs.

Scuticociliatosis caused by Philasterides dicentrarchi.

The ciliate Philasterides dicentrarchi has been previously identified as a new agent of scuticociliatosis in marine fish. The parasite can cause high mortalities in fish reared on farms or kept in aquariums. P. dicentrarchi is usually a free-living protozoan but can become an opportunistic histophagous parasite causing rapid lethal systemic infections in cultured fish. This review provides information about the morphology and biology of the scuticociliate P. dicentrarchi, as well as information about the pathological and immunological reactions of the host in response to the infection with the parasite. The epidemiology and the control strategies of the disease are also reviewed.

Gallic Acid Alkyl Esters: Trypanocidal and Leishmanicidal Activity, and Target Identification via Modeling Studies.

Eight gallic acid alkyl esters (1−8) were synthesized via Fischer esterification and evaluated for their trypanocidal and leishmanicidal activity using bloodstream forms of Trypanosoma brucei and promastigotes of Leishmania major. The general cytotoxicity of the esters was evaluated with human HL-60 cells. The compounds displayed moderate to good trypanocidal but zero to low leishmanicidal activity. Gallic acid esters with alkyl chains of three or four carbon atoms in linear arrangement (propyl (4), butyl (5), and isopentyl (6)) were found to be the most trypanocidal compounds with 50% growth inhibition values of ~3 µM. On the other hand, HL-60 cells were less susceptible to the compounds, thus, resulting in moderate selectivity indices (ratio of cytotoxic to trypanocidal activity) of >20 for the esters 4−6. Modeling studies combining molecular docking and molecular dynamics simulations suggest that the trypanocidal mechanism of action of gallic acid alkyl esters could be related to the inhibition of the T. brucei alternative oxidase. This suggestion is supported by the observation that trypanosomes became immobile within minutes when incubated with the esters in the presence of glycerol as the sole substrate. These results indicate that gallic acid alkyl esters are interesting compounds to be considered for further antitrypanosomal drug development.

On the Reversible and Irreversible Inhibition of Rhodesain by Curcumin.

Previously, it was suggested that the natural compound curcumin is an irreversible inhibitor of rhodesain, the major lysosomal cysteine protease of the protozoan parasite Trypanosoma brucei. The suggestion was based on a time-dependent inhibition of the enzyme by curcumin and a lack of recovery of activity of the enzyme after pre-incubation with curcumin. This study provides clear evidence that curcumin is a reversible, non-competitive inhibitor of rhodesain. In addition, the study also shows that the apparent irreversible inhibition of curcumin is only observed when no thiol-reducing reagent is included in the measuring buffer and insufficient solubilising agent is added to fully dissolve curcumin in the aqueous solution. Thus, the previous observation that curcumin acts as an irreversible inhibitor for rhodesain was based on a misinterpretation of experimental findings.

Trypanocidal activity of tetradentated pyridine-based manganese complexes is not linked to inactivation of superoxide dismutase.

Two tetradentated pyridine-based manganese complexes (Cpd2 and Cpd3) were previously reported to inhibit efficiently the growth of Trypanosoma cruzi in vitro and in vivo. Cpd3 was also shown to be a potent inhibitor of trypanosomal iron superoxide dismutase (Fe-SOD) and its trypanocidal activity linked to the inhibition of this enzyme. Here we investigated the anti-trypanosomal activity of the two compounds against bloodstream forms of Trypanosoma brucei. Both compounds displayed potent trypanocidal activity against T. brucei bloodstream forms with minimum inhibitory concentrations (MICs) and 50% growth inhibition (GI50) values of 1 µM and 0.2-0.3 µM, respectively. Cpd2 and Cpd3 also showed cytotoxicity against HL-60 cells but based on GI50 values the human cells were 14 and 87 times less sensitive indicating moderate selectivity. In contrast to previous observation, Cpd3 did not inhibit Fe-SOD within trypanosomes and Cpd2 inhibited the enzyme only by 34%. As Fe-SOD together with ornithine decarboxylase play vital roles in the antioxidant defence in bloodstream forms of T. brucei, inhibition of both enzymes should be synergistically. Therefore, the interaction of Cpd2 and Cpd3 with the ornithine decarboxylase inhibitor eflornithine was determined. Both compounds were found in combination with eflornithine to produce only an additive effect. Thus, the observed lack of synergy between Cpd2/Cpd3 and eflornithine can be regarded as further indication that both compounds are not very strong inhibitors of trypanosomal Fe-SOD. Nevertheless, tetradentated pyridine-based manganese complexes are interesting compounds with promising anti-trypanosomal activity.

Front-line glioblastoma chemotherapeutic temozolomide is toxic to Trypanosoma brucei and potently enhances melarsoprol and eflornithine.

Sleeping sickness is an infectious disease that is caused by the protozoan parasite Trypanosoma brucei. The second stage of the disease is characterised by the parasites entering the brain. It is therefore important that sleeping sickness therapies are able to cross the blood-brain barrier. At present, only three medications for chemotherapy of the second stage of the disease are available. As these trypanocides have serious side effects and are difficult to administer, new and safe anti-trypanosomal brain-penetrating drugs are needed. For these reasons, the anti-glioblastoma drug temozolomide was tested in vitro for activity against bloodstream forms of T. brucei. The concentration of the drug required to reduce the growth rate of the parasites by 50% was 29.1 µM and to kill all trypanosomes was 125 µM. Importantly, temozolomide did not affect the growth of human HL-60 cells up to a concentration of 300 µM. Cell cycle analysis revealed that temozolomide induced DNA damage and subsequent cell cycle arrest in trypanosomes exposed to the compound. As drug combination regimes often achieve greater therapeutic efficacy than monotherapies, the interactions of temozolomide with the trypanocides eflornithine and melarsoprol, respectively, was determined. Both combinations were found to produce an additive effect. In conclusion, these results together with well-established pharmacokinetic data provide the basis for in vivo studies and potentially for clinical trials of temozolomide in the treatment of T. brucei infections and a rationale for its use in combination therapy, particularly with eflornithine or melarsoprol.

Trypanosoma brucei: trypanocidal and cell swelling activities of lasalocid acid.

Chemotherapeutic treatment of human and animal trypanosomiasis is unsatisfactory because only a few drugs are available. As these drugs have poor efficacy and cause adverse reactions, more effective and tolerable medications are needed. As the polyether ionophore antibiotic lasalocid acid is used as medicated feed additive in cattle, the compound was tested for its trypanocidal and cytotoxic activity against bloodstream forms of Trypanosoma brucei and human myeloid HL-60 cells. The concentrations required of lasalocid acid to reduce the growth rate of trypanosomes by 50% and to kill the parasites were 1.75 and 10 µM, respectively. The ionophore displayed also cytotoxic activity against HL-60 cells but the human cells were about 10 to 14 times less sensitive indicating moderate selectivity. As the trypanocidal mechanism of action of polyether ionophore antibiotics is due to a sodium influx-induced cell swelling, the effect of lasalocid acid on cell volume change in bloodstream-form trypanosomes was investigated. Interestingly, lasalocid acid induced a much faster cell swelling in trypanosomes than the more trypanocidal related ionophore salinomycin. These results support further investigations of lasalocid acid and derivatives thereof as potential agents against African trypanosomiasis.

Cymantrenyl-Nucleobases: Synthesis, Anticancer, Antitrypanosomal and Antimicrobial Activity Studies.

The synthesis of four cymantrene-5-fluorouracil derivatives (1-4) and two cymantrene-adenine derivatives (5 and 6) is reported. All of the compounds were characterized by spectroscopic methods and the crystal structure of two derivatives (1 and 6), together with the previously described cymantrene-adenine compound C was determined by X-ray crystallography. While the compounds 1 and 6 crystallized in the triclinic P-1 space group, compound C crystallized in the monoclinic P21/m space group. The newly synthesized compounds 1-6 were tested together with the two previously described cymantrene derivatives B and C for their in vitro antiproliferative activity against seven cancer cell lines (MCF-7, MCF-7/DX, MDA-MB-231, SKOV-3, A549, HepG2m and U-87-MG), five bacterial strains Staphylococcus aureus (methicillin-sensitive, methicillin-resistant and vancomycin-intermediate strains), Staphylococcus epidermidis, and Escherichia coli, including clinical isolates of S. aureus and S. epidermidis, as well as against the protozoan parasite Trypanosoma brucei. The most cytotoxic compounds were derivatives 2 and C for A549 and SKOV-3 cancer cell lines, respectively, with 50% growth inhibition (IC50) values of about 7 µM. The anticancer activity of the cymantrene compounds was determined to be due to their ability to induce oxidative stress and to trigger apoptosis and autophagy in cancer cells. Three derivatives (1, 4 and 5) displayed promising antitrypanosomal activity, with GI50 values in the low micromolar range (3-4 µM). The introduction of the 5-fluorouracil moiety in 1 enhanced the trypanocidal activity when compared to the activity previously reported for the corresponding uracil derivative. The antibacterial activity of cymantrene compounds 1 and C was within the range of 8-64 µg/mL and seemed to be the result of induced cell shrinking.

Evaluation of Antiparasitc Activity of Mentha crispa Essential Oil, Its Major Constituent Rotundifolone and Analogues against Trypanosoma brucei.

Considering the pressing need for new drugs to treat sleeping sickness and Nagana disease, Mentha crispa essential oil, its principal constituent rotundifolone, and four related p-menthane-type monoterpenes (two stereoisomers of limonene epoxide, perillyl alcohol, and perillyl aldehyde) were investigated for their activity against bloodstream forms of Trypanosoma brucei. The general cytotoxicity of the compounds was determined with human myeloid HL-60 cells. The effect of the M. crispa essential oil and the monoterpenes on the growth of parasite and human cells was evaluated in cell cultures with the resazurin viability assay. Of all of the compounds tested, M. crispa essential oil, rotundifolone, and perillyl aldehyde showed the highest trypanocidal activities with 50 % growth inhibition (GI50) and minimum inhibitory concentration values of 0.3 µg/mL and 1 µg/mL, respectively. In contrast, HL-60 cells were considerably less sensitive to the compounds with minimum inhibitory concentration values of 100 µg/mL and GI50 values ranging between 3.4 to 13.8 µg/mL. As a consequence of this, GI50 and minimum inhibitory concentration ratios of cytotoxic to trypanocidal activity (selectivity index) of these three compounds were promising with values of 11-45 and 100, respectively. These results indicate that the p-menthane-type monoterpenes rotundifolone and perillyl aldehyde are interesting lead candidates for further rational antitrypanosomal drug development.

Trypanocidal and cysteine protease inhibitory activity of isopentyl caffeate is not linked in Trypanosoma brucei.

Previously, it was reported that caffeic acid esters inhibit the growth of bloodstream forms of Trypanosoma brucei and the activity of its major lysosomal cathepsin L-like cysteine protease, TbCATL. However, whether this trypanocidal activity is due to inactivation of TbCATL has not so far been demonstrated. Caffeic acid isopentyl ester (isopentyl caffeate) displayed antitrypanosomal activity against T. brucei bloodstream forms with minimum inhibitory concentration (MIC) and 50 % growth inhibition (GI50) values of 1 and 0.31 µg/ml, respectively. The ester also inhibited the activity of purified TbCATL but with a 27-fold higher half maximal inhibitory concentration (IC50) value of 8.5 µg/ml compared to its GI50 value. In contrast to previous suggestion, isopentyl caffeate did not interact with the active site of TbCATL but inhibited the enzyme in a non-competitive way. In addition, the ester was ineffective in blocking the proteolysis in the lysosome of the parasite, which, however, is a hallmark for inhibitors whose trypanocidal action is through inactivation of TbCATL. These results suggest that the antitrypanosomal activity of isopentyl caffeate (and probably of other caffeic acid esters) cannot be attributed to inhibition of TbCATL. Nevertheless, caffeic acid esters are interesting compounds with promising antitrypanosomal activity. This is supported by a more than 100 times less sensitivity of human HL-60 cells to isopentyl caffeate indicating that the ester has a favourable selectivity profile.

Membrane glucocorticoid receptors are localised in the extracellular matrix and signal through the MAPK pathway in mammalian skeletal muscle fibres.

KEY POINTS: Many studies have previously suggested the existence of stress hormone receptors on the cell membrane of many cell types, including skeletal muscle fibres; however, the exact localisation of these receptors and how they signal to the rest of the cell is poorly understood. In this study, we investigated the localisation and the mechanism(s) underlying the physiological functions of these receptors in mouse skeletal muscle cells. We found that the receptors were present throughout muscle development and that, in adult muscle fibres, they were localised in the extracellular matrix, satellite cells (muscle stem cells) and close to mitochondria. We also found that they signalled to the rest of the cell by activating enzymes called mitogen-activated protein kinases. From these results we suggest that, at physiological concentrations, stress hormones may be important in skeletal muscle differentiation, repair and regeneration. ABSTRACT: A number of studies have previously proposed the existence of glucocorticoid receptors on the plasma membrane of many cell types, including skeletal muscle fibres. However, their exact localisation and the cellular signalling pathway(s) they utilise to communicate with the rest of the cell are still poorly understood. In this study, we investigated the localisation and the mechanism(s) underlying the non-genomic physiological functions of these receptors in mouse skeletal muscle cells. The results show that the receptors were localised in the cytoplasm in myoblasts, in the nucleus in myotubes, in the extracellular matrix, in satellite cells and in the proximity of mitochondria in adult muscle fibres. Also, they bound laminin in a glucocorticoid-dependent manner. Treating small skeletal muscle fibre bundles with the synthetic glucocorticoid beclomethasone dipropionate increased the phosphorylation (= activation) of extracellular signal-regulated kinases 1 and 2, c-Jun N-terminal kinase and p38 mitogen-activated protein kinase. This occurred within 5 min and depended on the fibre type and the duration of the treatment. It was also abolished by the glucocorticoid receptor inhibitor, mifepristone, and a monoclonal antibody against the receptor. From these results we conclude that the non-genomic/non-canonical physiological functions of glucocorticoids, in adult skeletal muscle fibres, are mediated by a glucocorticoid receptor localised in the extracellular matrix, in satellite cells and close to mitochondria, and involve activation of the mitogen-activated protein kinase pathway.

Evaluation of trypanocidal activity of combinations of anti-sleeping sickness drugs with cysteine protease inhibitors.

Chemotherapy of human African trypanosomiasis (HAT) is unsatisfactory because only a few drugs, with serious side effects and poor efficacy, are available. As drug combination regimes often achieve greater therapeutic efficacy than monotherapies, here the trypanocidal activity of the cysteine protease inhibitor K11777 in combination with current anti-HAT drugs using bloodstream forms of Trypanosoma brucei was investigated. Isobolographic analysis was used to determine the interaction between cysteine protease inhibitors (K11777, CA-074Me and CAA0225) and anti-HAT drugs (suramin, pentamidine, melarsoprol and eflornithine). Bloodstream forms of T. brucei were incubated in culture medium containing cysteine protease inhibitors or anti-HAT drugs alone or in combination at a 1:1 fixed-dose ratio. After 48 h incubation, live cells were counted, the 50% growth inhibition values determined and combination indices calculated. The general cytotoxicity of drug combinations was evaluated with human leukaemia HL-60 cells. Combinations of K11777 with suramin, pentamidine and melarsoprol showed antagonistic effects while with eflornithine a synergistic effect was observed. Whereas eflornithine antagonises with CA-074Me, an inhibitor inactivating the targeted TbCATL only under reducing conditions, it synergises with CAA0255, an inhibitor structurally related to CA-074Me which inactivates TbCATL independently of thiols. These findings indicate an essential role of thiols for the synergistic interaction between K11777 and eflornithine. Encouragingly, the K11777/eflornithine combination displayed higher trypanocidal than cytotoxic activity. The results of this study suggest that the combination of the cysteine protease inhibitor K11777 and eflornithine display promising synergistic trypanocidal activity that warrants further investigation of the drug combination as possible alternative treatment of HAT.

Trypanotoxic activity of thiosemicarbazone iron chelators.

Only a few drugs are available for treating sleeping sickness and nagana disease; parasitic infections caused by protozoans of the genus Trypanosoma in sub-Saharan Africa. There is an urgent need for the development of new medicines for chemotherapy of these devastating diseases. In this study, three newly designed thiosemicarbazone iron chelators, TSC24, Dp44mT and 3-AP, were tested for in vitro activity against bloodstream forms of Trypanosoma brucei and human leukaemia HL-60 cells. In addition to their iron chelating properties, TSC24 and Dp44mT inhibit topoisomerase IIα while 3-AP inactivates ribonucleotide reductase. All three compounds exhibited anti-trypanosomal activity, with minimum inhibitory concentration (MIC) values ranging between 1 and 100 µM and 50% growth inhibition (GI50) values of around 250 nM. Although the compounds did not kill HL-60 cells (MIC values >100 µM), TSC24 and Dp44mT displayed considerable cytotoxicity based on their GI50 values. Iron supplementation partly reversed the trypanotoxic and cytotoxic activity of TSC24 and Dp44mT but not of 3-AP. This finding suggests possible synergy between the iron chelating and topoisomerase IIα inhibiting activity of the compounds. However, further investigation using separate agents, the iron chelator deferoxamine and the topoisomerase II inhibitor epirubicin, did not support any synergy for the interaction of iron chelation and topoisomerase II inhibition. Furthermore, TSC24 was shown to induce DNA degradation in bloodstream forms of T. brucei indicating that the mechanism of trypanotoxic activity of the compound is topoisomerase II independent. In conclusion, the data support further investigation of thiosemicarbazone iron chelators with dual activity as lead compounds for anti-trypanosomal drug development.

A cultivation method for growing bloodstream forms of Trypanosoma brucei to higher cell density and for longer time.

A novel cultivation technique using transwells allowing the growth of bloodstream forms of Trypanosoma brucei for longer periods of time and to higher cell densities is described. Trypanosomes cultured in transwells placed in cups of tissue culture plates containing sixfold excess of medium grew within 4 days to maximum cell densities of 2 × 10(7) parasites per milliliter. Compared to control cultures, the exponential growth of trypanosomes was 1 day longer and the maximum cell concentration was increased fourfold. The new culture system may be useful in drug screening assays and analysing the T. brucei secretome.

In vitro and in vivo studies of trypanocidal activity of dietary isothiocyanates.

Six dietary isothiocyanates, allyl-isothiocyanate, benzyl-isothiocyanate, phenylethyl-isothiocyanate, sulforaphane, erucin, and iberin, were tested for their trypanocidal activities in vitro using culture-adapted bloodstream forms of Trypanosoma brucei. All isothiocyanates showed a dose-dependent effect on the growth of trypanosomes. Five compounds displayed MIC values of 10 µM and GI50 values of around 1.5 µM, while allyl-isothiocyanate exhibited values of 100 and 11 µM, respectively. The compounds showed similar cytotoxic activities against human HL-60 cells with GI50 values of 1-4 µM and MIC values of 10-100 µM. Short-term experiments revealed that, with the exception of allyl-isothiocyanate, isothiocyanates at a concentration of 10 µM kill trypanosomes within 1-4 h of incubation. In contrast, HL-60 cells were not affected by any of the compounds in short-term incubation experiments. Sulforaphane, the most intensively studied isothiocyanate, was also investigated for its in vivo trypanocidal activity. However, administration of 50 mg/kg sulforaphane orally or intraperitoneally for four days had no effect on the parasitaemia in mice infected with T. brucei compared to control animals treated with vehicle alone.

Conjugation to 4-aminoquinoline improves the anti-trypanosomal activity of Deferiprone-type iron chelators.

Iron is an essential growth component in all living organisms and plays a central role in numerous biochemical processes due to its redox potential and high affinity for oxygen. The use of iron chelators has been suggested as a novel therapeutic approach towards parasitic infections, such as malaria, sleeping sickness and leishmaniasis. Known iron chelating agents such as Deferoxamine and the 3-hydroxypyridin-4-one (HPO) Deferiprone possess anti-parasitic activity but suffer from mammalian toxicity, relatively modest potency, and/or poor oral availability. In this study, we have developed novel derivatives of Deferiprone with increased anti-parasitic activity and reduced cytotoxicity against human cell lines. Of particular interest are several new derivatives in which the HPO scaffold has been conjugated, via a linker, to the 4-aminoquinoline ring system present in the known anti-malaria drug Chloroquine. We report the inhibitory activity of these novel analogues against four parasitic protozoa, Trypanosoma brucei, Trypanosoma cruzi, Leishmania infantum and Plasmodium falciparum, and, for direct comparison, against human cells lines. We also present data, which support the hypothesis that iron starvation is the major cause of growth inhibition of these new Deferiprone-Chloroquine conjugates in T. brucei.