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Malaria is caused by the Plasmodium sp. parasite that infects the red blood cells. Of the four types of malaria, the most serious type is transmitted by Plasmodium falciparum species. It can be life threatening. The other types of malaria (P. vivale, P. ovale and P. malariae) are generally less serious and are not life threatening. The existence of malaria as an enemy of humankind certainly predates written history. For thousands of years malaria has been a deadly scourge, and it remains one today. From American president John Adams who nearly succumbed to malaria in Amsterdam while on a diplomatic mission, back down to the timeline to the early Chinese, Indians, Greeks and Romans, malaria has not spared its victims, rich or poor. It wasn’t until the 19th Century that information about the true cause of malaria became known. Yet despite this knowledge, malaria still ravages Sub–Saharan Africa, South–East Asia and Latin America, taking as its victim’s mainly young children and pregnant women. However, without certain discoveries leading to a better understanding of malaria, new groundbreaking work wouldn’t be possible. Artemisinin and its derivatives are developing into a very important new class of antimalarial and their usage is becoming more common in the fight against malaria. The most commonly used and applied of these derivatives are artesunate, artemether, arteether and dihydroartemisinin. The discovery of artemisinin as the pharmacological active ingredient in an age old Chinese herb, Artemisia annua, was a major breakthrough in malaria chemotherapy. Discovery of qinghaosu in the 1970s sparked a new age for chemotherapy of malaria, and greatly inspired further research on organic peroxides. This generated widespread interest and led to the design and synthesis of organic peroxides into a highly active area of organic chemistry. The artemisinin derivatives act quickly and are eliminated quickly. Their rapid onset makes them especially effective against severe malaria. Their rapid disappearance may be a key reason why artemisinin resistance has been so slow to develop, and may be the reason why recrudences are so common when these drugs are used in monotherapy. Since their isolation, artemisinins have had a substantial impact on the treatment of malaria. Although very potent, the use of artemisinins as prophylactic antimalarials is not recommended. The aim of this study was to synthesise ester derivatives of artemisinin, determine certain physicochemical properties such as aqueous solubility and partition coefficient, and to evaluate their antimalarial activity in comparison to dihydroartemisinin and chloroquine. In this study eight esters of dihydroartemisinin (DHA) were synthesised by substitution at C– 10. The structures of the prepared derivatives were confirmed by nuclear magnetic resonance spectroscopy (NMR) and mass spectrometry (MS). The new artemisinin esters were tested in vitro against the chloroquine sensitive strain of Plasmodium falciparum (D10). All the compounds tested showed activity against the D10 strain. All of the esters showed potency significantly better than chloroquine, except the octyl and decyl esters which were less active. The reason for the low activity could be ascribed to the fact that these two esters are both water immiscible oils, leading to solubility problems. The ethyl, butyl, phenyl and p–nitrophenyl esters all had similar IC50 values making their activity similar. The lowest IC50 value was displayed by the butyl ester with a value of 3.2 x 10– 3 uM. The poorest activity was recorded by the two oils, the octyl and decyl esters, with IC50 values of 38 x 10–3 uM and 90.2 x 10–3 uM respectively. All other compounds showed less antimalarial potency against the D10 strain compared with the other reference drug dihydroartemisinin, except the butyl ester. The butyl ester 12 displayed activity comparable to that of DHA (IC50; 3.2 x 10–3 uM versus 3.8 x 10–3 uM), and is thus worthwhile being further investigated in terms of pharmacokinetics in order to determine its half–life. Statistically it is impossible to make structure–activity relationship (SAR) deductions from the data received as the number of compounds in the series is too small. The butyl (12) (IC50 = 3.2 uM), 4–nitrobenzyl (16) (IC50 =15 uM), 2–(acetyloxy) acetyl (17) (IC50 = 8.6 uM), and 2–phenylacetyl (18) (IC50 = 12.4 uM) esters showed on a 0.05 level statistically significantly better activity against the chloroquine sensitive D10 strain of Plasmodium falciparum than chloroquine itself while the decyl ester (14) (IC50 = 90.2 uM) was statistically significantly less potent. The activity of the octyl (13) (IC50 = 38.0 uM) and benzyl (15) (IC50 = 25.7 uM) esters did not differ from that of chloroquine. In comparison to dihydroartemisinin the propyl (11) (IC50 = 24.1 uM), octyl (13) (IC50 = 38.0 uM), decyl (14) (IC50 = 90.0 uM), and benzyl (15) (IC50 = 25.7 uM) esters proved to be statistically significantly less potent than DHA while the activity of the butyl (12) (IC50 = 3.2 uM), 4– nitrobenzyl (16) (IC50 =15.3 uM), 2–(acetyloxy) acetyl (17) (IC50 = 8.6 uM), and 2–phenylacetyl (18) (IC50 = 12.4 uM) esters did not differ from that of DHA. / Thesis (M.Sc. (Pharmaceutical Chemistry))--North-West University, Potchefstroom Campus, 2012.

Malaria is caused by the Plasmodium sp. parasite that infects the red blood cells. Of the four types of malaria, the most serious type is transmitted by Plasmodium falciparum species. It can be life threatening. The other types of malaria (P. vivale, P. ovale and P. malariae) are generally less serious and are not life threatening. The existence of malaria as an enemy of humankind certainly predates written history. For thousands of years malaria has been a deadly scourge, and it remains one today. From American president John Adams who nearly succumbed to malaria in Amsterdam while on a diplomatic mission, back down to the timeline to the early Chinese, Indians, Greeks and Romans, malaria has not spared its victims, rich or poor. It wasn’t until the 19th Century that information about the true cause of malaria became known. Yet despite this knowledge, malaria still ravages Sub–Saharan Africa, South–East Asia and Latin America, taking as its victim’s mainly young children and pregnant women. However, without certain discoveries leading to a better understanding of malaria, new groundbreaking work wouldn’t be possible. Artemisinin and its derivatives are developing into a very important new class of antimalarial and their usage is becoming more common in the fight against malaria. The most commonly used and applied of these derivatives are artesunate, artemether, arteether and dihydroartemisinin. The discovery of artemisinin as the pharmacological active ingredient in an age old Chinese herb, Artemisia annua, was a major breakthrough in malaria chemotherapy. Discovery of qinghaosu in the 1970s sparked a new age for chemotherapy of malaria, and greatly inspired further research on organic peroxides. This generated widespread interest and led to the design and synthesis of organic peroxides into a highly active area of organic chemistry. The artemisinin derivatives act quickly and are eliminated quickly. Their rapid onset makes them especially effective against severe malaria. Their rapid disappearance may be a key reason why artemisinin resistance has been so slow to develop, and may be the reason why recrudences are so common when these drugs are used in monotherapy. Since their isolation, artemisinins have had a substantial impact on the treatment of malaria. Although very potent, the use of artemisinins as prophylactic antimalarials is not recommended. The aim of this study was to synthesise ester derivatives of artemisinin, determine certain physicochemical properties such as aqueous solubility and partition coefficient, and to evaluate their antimalarial activity in comparison to dihydroartemisinin and chloroquine. In this study eight esters of dihydroartemisinin (DHA) were synthesised by substitution at C– 10. The structures of the prepared derivatives were confirmed by nuclear magnetic resonance spectroscopy (NMR) and mass spectrometry (MS). The new artemisinin esters were tested in vitro against the chloroquine sensitive strain of Plasmodium falciparum (D10). All the compounds tested showed activity against the D10 strain. All of the esters showed potency significantly better than chloroquine, except the octyl and decyl esters which were less active. The reason for the low activity could be ascribed to the fact that these two esters are both water immiscible oils, leading to solubility problems. The ethyl, butyl, phenyl and p–nitrophenyl esters all had similar IC50 values making their activity similar. The lowest IC50 value was displayed by the butyl ester with a value of 3.2 x 10– 3 uM. The poorest activity was recorded by the two oils, the octyl and decyl esters, with IC50 values of 38 x 10–3 uM and 90.2 x 10–3 uM respectively. All other compounds showed less antimalarial potency against the D10 strain compared with the other reference drug dihydroartemisinin, except the butyl ester. The butyl ester 12 displayed activity comparable to that of DHA (IC50; 3.2 x 10–3 uM versus 3.8 x 10–3 uM), and is thus worthwhile being further investigated in terms of pharmacokinetics in order to determine its half–life. Statistically it is impossible to make structure–activity relationship (SAR) deductions from the data received as the number of compounds in the series is too small. The butyl (12) (IC50 = 3.2 uM), 4–nitrobenzyl (16) (IC50 =15 uM), 2–(acetyloxy) acetyl (17) (IC50 = 8.6 uM), and 2–phenylacetyl (18) (IC50 = 12.4 uM) esters showed on a 0.05 level statistically significantly better activity against the chloroquine sensitive D10 strain of Plasmodium falciparum than chloroquine itself while the decyl ester (14) (IC50 = 90.2 uM) was statistically significantly less potent. The activity of the octyl (13) (IC50 = 38.0 uM) and benzyl (15) (IC50 = 25.7 uM) esters did not differ from that of chloroquine. In comparison to dihydroartemisinin the propyl (11) (IC50 = 24.1 uM), octyl (13) (IC50 = 38.0 uM), decyl (14) (IC50 = 90.0 uM), and benzyl (15) (IC50 = 25.7 uM) esters proved to be statistically significantly less potent than DHA while the activity of the butyl (12) (IC50 = 3.2 uM), 4– nitrobenzyl (16) (IC50 =15.3 uM), 2–(acetyloxy) acetyl (17) (IC50 = 8.6 uM), and 2–phenylacetyl (18) (IC50 = 12.4 uM) esters did not differ from that of DHA. / Thesis (M.Sc. (Pharmaceutical Chemistry))--North-West University, Potchefstroom Campus, 2012.

Artemisinin compounds are the most potent anti-malarial drugs available in the market. Today, malaria treatment is largely relies on the artemisinin-based combination therapies. Artesunate (AS) is the most widely used artemisinin derivative. In this thesis, we characterized the population pharmacokinetics of AS and its active metabolite dihydroartemisinin (DHA) following oral administration of AS in different populations. In Chapter II, we developed a population pharmacokinetic model of AS and DHA in healthy subjects. These subjects received either single- or multiple-dosing of oral AS, as a monotherapy regimen or in combination with pyronaridine, with or without food. In Chapter III, we developed a population pharmacokinetic model of AS and DHA in adult and pediatric patients with uncomplicated falciparum and vivax malaria who were administered oral pyronaridine/artesunate combination once daily for 3 days. We modeled the AS and DHA data simultaneously using a parent-metabolite model that assumed complete conversion of AS to DHA. Following oral administration, AS is rapidly absorbed with maximum concentrations reached at about 0.5 hours post-dose. AS is rapidly converted to DHA. DHA then undergoes rapid metabolism, with an elimination half-life of about 0.8 hours in malarial patients. Inter-individual variability for almost all pharmacokinetic parameters and residual variability for both compounds were estimated by the models. Substantial variability was seen in the pharmacokinetic parameters between the subjects. In healthy subjects, intake of food with the dose was found to delay the absorption of AS significantly, but not the extent of absorption. Weight was also included in this model as a determinant of DHA clearance. When modeling the data from patients, we included weight as part of the model a prioria priori using an established allometric function. No other covariates examined in the analysis were statistically significant. The performance of final models was evaluated using non-parametric bootstrap technique and visual predictive check. The models were found to adequately described the data at hand, and robust with sufficient predictive power. The results can be used as the base to develop a population pharmacokinetic-pharmacodynamic model and as prior information in guiding the selection of optimal sampling schedule for future pharmacokinetic studies of AS.

Artemisinin derivatives are key to the current global treatment approach for malaria. However, much remains unknown regarding the pharmacokinetics of these agents, particularly in children and pregnant women, two groups highly vulnerable to development of severe malaria infection. In this thesis, nonlinear mixed effects modeling is used to characterize the pharmacokinetics of the artemisinin derivative artesunate and its active metabolite, dihydoartemisinin (DHA), in children and in pregnant women. Chapter 1 of this thesis contains a general review of the clinical pharmacokinetic findings for artesunate and DHA following artesunate administration by the intravenous, intramuscular, oral and rectal routes. Chapter 2 presents a population pharmacokinetic model utilizing both pediatric and adult data from one Phase II and four Phase III clinical trials evaluating the combination agent pyronaridine tetraphosphate/artesunate. The focus of the modeling described in this chapter is the evaluation of the effects of body size and gender on the pharmacokinetics of artesunate and DHA in pediatric patients with uncomplicated malaria. Chapter 3 consists of a population pharmacokinetic model built utilizing plasma artesunate and DHA concentrations from 26 parasitemic second and third trimester pregnant women and 25 parasitemic non-pregnant female controls in the Democratic Republic of Congo who received 200 mg oral artesunate. The model described in Chapter 2 is a simultaneously implemented parent-metabolite model consisting of a one compartment model for artesunate, a one compartment model for DHA, and first-order artesunate absorption. Various approaches for incorporating body size on artesunate and DHA apparent clearance and volume of distribution parameters were evaluated, with a linear body surface area model and an allometric scaling model both proving satisfactory. The effect of gender was modeled on artesunate and DHA apparent clearance and volume terms. Only the effect of gender on DHA apparent clearance could be estimated with reasonable precision, with the 95% confidence interval for the effect being almost wholly contained within the predefined 0.75 to 1.25 no relevant clinical effect interval. The model described in Chapter 3 consists of a one compartment model for artesunate, a one compartment model for DHA, and mixed zero-order, lagged first order absorption of artesunate. In this model, pregnancy was found to have a marked effect on DHA apparent clearance, with a pregnancy-associated increase in DHA apparent clearance of 42.3%. The models described in this thesis indicate that, for a given mg/kg dose of artesunate, both young children and pregnant women would be expected, on average, to display lower DHA concentrations than would be observed following administration of the same mg/kg dose to non-pregnant adults. Suboptimal dosing has clinical implications for the individual as well as potential implications regarding parasite susceptibility. Given this, the findings of the research described in this thesis highlight the necessity of investigations designed to comprehensively characterize the pharmacokinetics of artesunate and DHA in these two highly susceptible populations.

Malaria is endemic in 106 countries worldwide. This disease is caused by a parasite from the genus Plasmodium. Of the five species that infect humans, Plasmodium falciparum is the most virulent, with over three billion people at risk and around 660 000 deaths reported in 2011. Of these deaths, 91% were in the African region, while 86% were children under the age of five. In light of the widespread development of resistance by malaria parasites against the classic antimalarial drugs, such as chloroquine (CQ) and now the established tolerance towards the widely used artemisinins, an immense need exists for identifying and developing new and effective antiplasmodial drugs. In search for such new antimalarial drugs, three chalcone based series of compounds were prepared and investigated during this study. The first series (Chapter 3) comprised 4-aminoquinolinyl-chalcone amides, which were synthesized through amidation of carboxylic acid-functionalised chalcone with aminoquinolines, using 1,1'-carbonyldiimidazole (CDI) as coupling agent. These compounds were screened alongside CQ against the CQ sensitive (3D7) and CQ resistant (W2) strains of P. falciparum. Cytotoxicity was assessed against the WI-38 cell line. The amide, featuring the 1,6-diaminohexane linker, was found the most active of all these new novel compounds tested. It was found to be as potent as CQ against 3D7, while displaying a two-fold higher activity than CQ against the W2 strain, coupled with good selective antimalarial activity (SI = 435) towards the parasitic cells. The second series (Chapter 4) consisted of aminoferrocenyl-chalcone amides, synthesized through condensation of a chalcone with an aminoferrocenyl. These compounds were screened against the 3D7, and antifolate- and CQ resistant (FCR3) strains of P. falciparum and cytotoxicity was determined against the WI-38 line. The most active compound of this series was the amide, containing the 1,2-diaminoethane linker, which showed 130- and 42 times less potency than CQ against the 3D7 and W2 strains, respectively. The third series of antimalarials (Chapter 5) involved dihydroartemisinyl-chalcone esters, synthesized through esterification of chalcones with DHA. These compounds were screened against 3D7 and W2 strains of P. falciparum, while the cytotoxicity was determined against the WI-38 line. Those esters featuring oxygenated aryl rings were three- to four-fold more potent than current clinically used artesunate against both P. falciparum strains. They were also screened in vitro against a panel of three cancer cell lines consisting of TK-10, UACC-62 and MCF-7. Thermogravimetric analysis revealed that the targeted hybrids were all thermally more stable than DHA as a result of the presence of the chalcone moiety in their structures. This could prove beneficial to the high temperature storage conditions that prevail in most malaria endemic countries. This study resulted in a number of compounds with varying antiplasmodial activity ranges. The compounds in series 3 were overall the most active, due to the incorporation of the highly active dihydroartemisinin pharmacophore. The chalcone moiety, especially, demonstrated a large scope for future development, owing to the ease of synthesis and the relatively low costs involved. The most active compounds of the three series could serve as potential lead compounds in the future development of more effective antimalarial drugs. / PhD (Pharmaceutical Chemistry), North-West University, Potchefstroom Campus, 2014

Malaria is endemic in 106 countries worldwide. This disease is caused by a parasite from the genus Plasmodium. Of the five species that infect humans, Plasmodium falciparum is the most virulent, with over three billion people at risk and around 660 000 deaths reported in 2011. Of these deaths, 91% were in the African region, while 86% were children under the age of five. In light of the widespread development of resistance by malaria parasites against the classic antimalarial drugs, such as chloroquine (CQ) and now the established tolerance towards the widely used artemisinins, an immense need exists for identifying and developing new and effective antiplasmodial drugs. In search for such new antimalarial drugs, three chalcone based series of compounds were prepared and investigated during this study. The first series (Chapter 3) comprised 4-aminoquinolinyl-chalcone amides, which were synthesized through amidation of carboxylic acid-functionalised chalcone with aminoquinolines, using 1,1'-carbonyldiimidazole (CDI) as coupling agent. These compounds were screened alongside CQ against the CQ sensitive (3D7) and CQ resistant (W2) strains of P. falciparum. Cytotoxicity was assessed against the WI-38 cell line. The amide, featuring the 1,6-diaminohexane linker, was found the most active of all these new novel compounds tested. It was found to be as potent as CQ against 3D7, while displaying a two-fold higher activity than CQ against the W2 strain, coupled with good selective antimalarial activity (SI = 435) towards the parasitic cells. The second series (Chapter 4) consisted of aminoferrocenyl-chalcone amides, synthesized through condensation of a chalcone with an aminoferrocenyl. These compounds were screened against the 3D7, and antifolate- and CQ resistant (FCR3) strains of P. falciparum and cytotoxicity was determined against the WI-38 line. The most active compound of this series was the amide, containing the 1,2-diaminoethane linker, which showed 130- and 42 times less potency than CQ against the 3D7 and W2 strains, respectively. The third series of antimalarials (Chapter 5) involved dihydroartemisinyl-chalcone esters, synthesized through esterification of chalcones with DHA. These compounds were screened against 3D7 and W2 strains of P. falciparum, while the cytotoxicity was determined against the WI-38 line. Those esters featuring oxygenated aryl rings were three- to four-fold more potent than current clinically used artesunate against both P. falciparum strains. They were also screened in vitro against a panel of three cancer cell lines consisting of TK-10, UACC-62 and MCF-7. Thermogravimetric analysis revealed that the targeted hybrids were all thermally more stable than DHA as a result of the presence of the chalcone moiety in their structures. This could prove beneficial to the high temperature storage conditions that prevail in most malaria endemic countries. This study resulted in a number of compounds with varying antiplasmodial activity ranges. The compounds in series 3 were overall the most active, due to the incorporation of the highly active dihydroartemisinin pharmacophore. The chalcone moiety, especially, demonstrated a large scope for future development, owing to the ease of synthesis and the relatively low costs involved. The most active compounds of the three series could serve as potential lead compounds in the future development of more effective antimalarial drugs. / PhD (Pharmaceutical Chemistry), North-West University, Potchefstroom Campus, 2014

Malaria is the most important parasitic disease in man and it kills approximately 2,000 people each day. Pregnant women are especially vulnerable to malaria with increased incidence and mortality rates. There are indications that pregnancy alters the pharmacokinetic properties of many antimalarial drugs. This is worrisome as lower drug exposures might result in lowered efficacy and lower drug exposures can also accelerate the development and spread of resistant parasites. The aim of this research was to study the pharmacokinetics and pharmacodynamics of the most commonly used drugs for the treatment of uncomplicated Plasmodium falciparum malaria during the second and third trimester of pregnancy using a pharmacometric approach. This thesis presents a number of important findings that increase the current knowledge of antimalarial drug pharmacology and that may have an impact in terms of drug efficacy and resistance. (1) Lower lumefantrine plasma concentrations at day 7 were evident in pregnant women compared to that in non-pregnant patients. Subsequent in-silico simulations with the final pharmacokinetic-pharmacodynamic lumefantrine/desbutyl-lumefantrine model showed a decreased treatment failure rate after a proposed extended artemether-lumefantrine treatment. (2) Dihydroartemisinin exposure (after intravenous and oral administration of artesunate) was lower during pregnancy compared to that in women 3 months post-partum (same women without malaria). Consecutive in-silico simulations with the final model showed that the underexposure of dihydroartemisinin during pregnancy could be compensated by a 25% dose increase. (3) Artemether/dihydroartemisinin exposure in pregnant women was also lower compared to literature values in non-pregnant patients. This further supports the urgent need for a study in pregnant women with a non-pregnant control group. (4) Quinine pharmacokinetics was not affected by pregnancy trimester within the study population and a study with a non-pregnant control group is needed to evaluate the absolute effects of pregnancy. (5) Finally, a data-dependent power calculation methodology using the log likelihood ratio test was successfully used for sample size calculations of mixed pharmacokinetic study designs (i.e. sparsely and densely sampled patients). Such sample size calculations can contribute to a better design of future pharmacokinetic studies. In conclusion, this thesis showed lower exposures for drugs used to treat uncomplicated Plasmodium falciparum malaria during the second and third trimester of pregnancy. More pharmacokinetic studies in pregnant women with a non-pregnant control group are urgently needed to confirm the current findings and to enable an evidence-based dose optimisation. The data-dependent power calculation methodology using the log likelihood ratio test can contribute to an effective design of these future pharmacokinetic studies.