key: cord-0047167-ddv53bip
authors: Adebayo, J. O.; Tijjani, H.; Adegunloye, A. P.; Ishola, A. A.; Balogun, E. A.; Malomo, S. O.
title: Enhancing the antimalarial activity of artesunate
date: 2020-07-07
journal: Parasitol Res
DOI: 10.1007/s00436-020-06786-1
sha: fb32f147a860ab957f46d1198f6b1c13f84d0c04
doc_id: 47167
cord_uid: ddv53bip

The global challenge to the treatment of malaria is mainly the occurrence of resistance of malaria parasites to conventionally used antimalarials. Artesunate, a semisynthetic artemisinin compound, and other artemisinin derivatives are currently used in combination with selected active antimalarial drugs in order to prevent or delay the emergence of resistance to artemisinin derivatives. Several methods, such as preparation of hybrid compounds, combination therapy, chemical modification and the use of synthetic materials to enhance solubility and delivery of artesunate, have been employed over the years to improve the antimalarial activity of artesunate. Each of these methods has advantages it bestows on the efficacy of artesunate. This review discussed the various methods employed in enhancing the antimalarial activity of artesunate and delaying the emergence of resistance of parasite to it.

Malaria is an infectious disease caused by the Plasmodium species, of which P. falciparum is the most virulent and is responsible for majority of the malaria morbidity and mortality recorded worldwide (Peter and Anatoli 1998) . Other species of the genus Plasmodium that infect humans are P. vivax, P. knowlesi, P. ovale, and P. malariae. About 228 million cases of malaria were recorded globally in 2018 (World Health Organization 2019) . This resulted in about 405,000 deaths, 94% of which occurred in the African region where the burden is highest (World Health Organization 2019). In addition to being a threat to human health, malaria is also a threat to the economy and development of sub-Saharan Africa (Sachs and Malaney 2002) . The funding for malaria in 2018 was estimated at US$2.7 billion (World Health Organization 2019), representing less than 41% of the total malaria funding of US$6.5 billion required annually to meet the 2030 global malaria target (World Health Organization 2017). To eliminate malaria and its associated burden, some key approaches are employed: (i) the use of vector control and (ii) the use of antimalarial drugs for chemoprophylaxis or chemotherapy (Rappuoli and Aderem 2011) . Hence, the target research areas include vector control, chemotherapy and vaccine development for the disease. However, resistance to insecticides and antimalarial drugs has hindered research breakthrough in vector control and chemotherapy respectively, while the most advanced malaria vaccine developed (RTS,S/AS01), for Section Editor: Berit Bangoura which the 3rd phase of clinical trial was completed in 2014 (Agnandji et al. 2015) , is yet to guarantee up to 50% protection and is best advised as a complementary malaria control tool rather than as a replacement package (World Health Organization 2020). The resistance developed to conventional antimalarial drugs, including the potent artemisinin derivatives, has led to the discouragement of monotherapy of such drugs. Thus, various efforts are being put in place globally to enhance the efficacy of artemisinin derivatives and delay the emergence of resistance to these drugs.

Artemisinin (1) is a sesquiterpene lactone peroxide isolated from the leaves of the shrub Artemesia annua, better known indigenously as Qinghao in China (Mohammadi et al. 2020) . Artemisinin is a 15-carbon compound with an endoperoxide bridge, which is crucial to its activity against Plasmodium species (Butler and Wu 1992) . Artemisinin is effective against blood and early gametocyte stages of P. falciparum (Piyaphanee et al. 2006; White 2008) . However, this effectiveness is endangered by the emergence of resistance of P. falciparum to artemisinin as reported in six countries/regions: Cambodia, Lao People's Democratic Republic, Myanmar, Thailand, Viet Nam, and the Myanmar-China-India border area (Noedl et al. 2008; Thu et al. 2017; Ouji et al. 2018) . Likewise, parasite isolates from Nigeria and Madagascar have been reported to exhibit reduced sensitivity to artemisinin (Oduola et al. 1992; Randrianarivelojosia et al. 2001) . Regions, such as western Cambodia, with the common use of artemisinin as monotherapy have shown resistance to artemisinin in clinical studies (White 2008) . Decreased responses to treatments, specifically to artesunate and artemether, have also been reported in western Thailand (Luxemburger et al. 1998) , India (Gogtay et al. 2000) , and Sierra Leone (Sahr et al. 2001) . Preserving the efficacies of artemisinin derivatives for the treatment of malaria is now the challenge.

Due to some challenges with the solubility and bioavailability of artemisinin, derivatives with different solubilities in oil and water have been synthesized. Some of these artemisinin derivatives are dihydroartemisinin (2), artemether (3), arteether (4), and artesunate (5) (Fig. 1) . Artemether is oil-soluble and it i s u su a l ly a d m in i st e re d in t r a m us c u l a r l y , w h i l e dihydroartemisinin and artesunate are water-soluble and are usually administered orally, except for severe malaria, which requires artesunate being administered via intravenous route. After administration, the artemisinins are rapidly absorbed, distributed and metabolized. Due to the emergence of resistance to oral artemisinin monotherapy (oAMT), it was replaced with artemisinin-based combination therapies (ACTs) (World Health Organization 2006 , in which a fastacting artemisinin derivative is administered together with a slow-acting antimalarial drug of another class of antimalarials (Nigam et al. 2019; Mohammadi et al. 2020) . ACTs are highly effective and efficacious globally and currently remain the recommended first-line treatment for uncomplicated malaria (World Health Organization 2010 . The World Health Organization recommended the use of five different ACTs globally: artemether/ lumefantrine, artesunate/amodiaquine, artesunate/ sulfadoxine-pyrimethamine, artesunate/mefloquine, and dihydroartemisinin/piperaquine (World Health Organization 2017, 2019), while the sixth recommended ACT, artesunate/ pyronaridine (World Health Organization 2019), is used based on expression of resistance in regions. Close monitoring of resistance and necessary change of drug combinations due to change in response of infection to chemotherapy over time were also recommended (World Health Organization 2015). Also, in order to improve artemisinin activity, artemisinin dimers, trimers, and tetramers have been synthesized with each monomer retaining the peroxide bridge (Fröhlich et al. 2018) . Artesunate, a water-soluble derivative, is administered as an oral drug, a rectal suppository and intravenous and intramuscular injections. Thus, it is more widely used compared to other derivatives. Improving its activity can enhance its effectiveness and delay the emergence of resistance to it. Therefore, this review deals with the different means and methods by which the therapeutic efficacy of artesunate has been improved through the years.

Artemisinins are converted to active metabolite dihydroartemisinin, with the half-life of approximately 45 min. By this, they are by far the most rapidly eliminated antimalarial drugs, with broad-stage specificity in action. Several mechanisms have been proposed for the antimalarial activities of artemisinins, including the generation of a carbon-centered radical (though now untenable), inhibition of polymerization of heme, production of free radicals and alteration of membrane transport properties of the malaria parasite, which inhibits the nutrient flow in the parasite (Eckstein-Ludwig et al. 2003; Krishna et al. 2004; Posner and O'Neill 2004; Drew et al. 2006 Drew et al. , 2007 German and Aweeka 2008; Haynes et al. 2013 ).

Artemisinins bind to heme through alkylation to form hemeartemisinin adduct (Krishna et al. 2004 ). This adduct binds to HRP II, a protein involved in the polymerization of heme (Lee et al. 2019) , thereby preventing hemozoin formation (Kannan et al. 2002) . This leads to the accumulation of heme which is toxic to the parasite.

Artemisinin has been reported to be involved in covalent interaction with 25 kDa translationally controlled tumor protein (TCTP) homolog (Bhisutthibhan and Meshnick 2001) . This protein has also been reported to perform a role in hemozoin formation. Artemisinin has also been reported to alkylate cysteine proteases, such as falcipain in P. falciparum (Pandey et al. 1999) , thereby impairing its role in hemoglobin digestion. The alkylation of proteins by artemisinin is heme dependent.

Artemisinin has been reported to inhibit PfATP6, also known as sarco/endoplasmic reticulum Ca ATPase (SERCA) (Eckstein-Ludwig et al. 2003) , thereby disturbing intracellular calcium homeostasis. It is found in the endoplasmic reticulum membrane where it functions in transporting Ca 2+ ion from the cytoplasm into the endoplasmic reticulum, where it is stored.

The mechanism of action of artemisinin has been linked with the generation of reactive oxygen species (ROS) such as hydroxyl, superoxide, alkoxyl, and peroxyl radicals. These are formed from Fenton reaction in a Fe 2+ -dependent reaction. An increase in ROS leads to the depletion of antioxidant species in the parasite (Hunt and Stocker 1990) . Initially, it was believed that the conversion of artemisinin to carbon-centered radical was crucial to the generation of ROS, but recently, that hypothesis has been reported to be untenable (Haynes et al. 2013 ).

Artemisinin has been reported to accumulate in neutral lipids, inducing oxidative damage. The inhibition of heme polymerization leads to the accumulation of heme which also induces ROS generation. The generation of ROS induces oxidative damage of the membrane (Haynes et al. 2013 ).

Artesunate (5), also known as dihydroartemisinin-12-α-succinate, is a potent, semisynthetic antimalarial compound derived from its parent compound, artemisinin, in a two-step reaction i n v o l v i n g r e d u c t i o n a n d e s t e r i f i c a t i o n u s i n g diisobutylaluminum hydride (DIBAL) and succinic anhydride respectively (Chekem and Wierucki 2006) . Artesunate has improved solubility, absorption and pharmacokinetics (Li et al. 1998 ). This allows it to be recommended as an oral, Fig. 1 Artemisinin and its derivatives rectal, intramuscular and intravenous medication for uncomplicated and severe malaria (Barnes et al. 2004; Harin et al. 2006 ).

Artesunate has a short half-life of between 20 and 45 min by oral route (OS) (Morris et al. 2011 ) and is metabolized through esterase-catalyzed hydrolysis within this short time to dihydroartemisinin, which is the active metabolite responsible for its antimalarial activity (Teja-Isavadharm et al. 2001; Gautam et al. 2009 ). Dihydroartemisinin is converted to its inactive form through glucuronidation via UDPglucuronosyl transferase (Ilett et al. 2002) . A little amount of the DHA is also eliminated via the bile as glucuronides (Teja-Isavadharm et al. 2001 ).

Since the discovery of artesunate, several modifications have been done to this compound, in an attempt to improve its effectiveness and delay the emergence of resistance to it. The most widely accepted approach, as approved by the WHO, is the combination of the fast-acting artesunate with another slow-acting drug, usually from another class of antimalarials. Other approaches to improve its activity are also well documented. These include the synthesis of hybrid compounds, the use of drug delivery systems, chemical modification, etc. (Fig. 2 ).

The approaches discussed in this section include those that have yielded drugs that are currently undergoing clinical trials and those already in clinical use.

Sodium artesunate Sodium artesunate (5a) is a basic salt as opposed to the acidic form and the recommended form for the treatment of uncomplicated and severe malaria due to its solubility (Fig. 3 ). It is a water-soluble compound which can be administered either as an intravenous or intramuscular drug (Batty et al. 1996) . Sodium artesunate is more active in parasite clearance compared to quinine and some other antimalarials because of its fast action and better tolerance; however, it is less potent compared to its acidic form. Its activities against resistant malaria parasite strains are also well documented (Batty et al. 1996; White 1999a, b) . The administration of sodium artesunate was reported to increase urinary excretion of sodium, chloride and potassium ions in Wistar rats (Campos et al. 2001 ). In addition, Campos et al. (2001) reported that administration of sodium artesunate led to an increase in the concentrations of metabolites of nitric oxide (nitrite and nitrates), which have been linked with protection against malaria (Taylor-Robinson and Looker 1998). 

The resistance of malaria parasite to chloroquine in regions of Southeast Asia, South America and along the east coast of Africa led to the withdrawal of chloroquine as first-line treatment for uncomplicated P. falciparum malaria (World Health Organization 2001). Similarly, the use of artesunate monotherapy, as chloroquine replacement, could not be continued due to the emergence of parasite resistance to it (Sahr et al. 2001 ). This, therefore, led to the introduction of ACT (World Health Organization 2007). Different ACTs containing artesunate have been used since then with continued monitoring. ACTs are available as fixed dose combinations; the drug combinations are prepared in "ready to take formulation," also known as "co-blister." Unlike taking a series of drug regimens in multiple tablets, this ACT formulation approach improves patient's compliance during treatments (Lacaze et al. 2011) . The choice of partner drugs for combination with artesunate is guided by the following principles: (i) the drug should act synergistically with artesunate (Majori 2004) , (ii) it should be slow-acting, and (iii) it should exhibit a different mechanism of action in order to delay the emergence of drug resistance (White 1998) . Studies have indicated that the inclusion of artesunate does not influence the pharmacokinetics of other components (van Vugt et al. 1999) . When the sensitivity of a partner drug in the ACT decreases, such drug is replaced with another which possesses a higher sensitivity. Artesunatebased combination therapies containing primaquine, mefloquine, pyronaridine, amodiaquine, or piperaquine as partner drug have been formulated as single doses (Fig. 4) . Also, artesunate is combined with chlorproguanil-dapsone, sulfadoxine-pyrimethamine, atovaquone-proguanil, and some antibiotics like clindamycin, tetracycline, and doxycycline (Fig. 5) .

Primaquine-artesunate combination Primaquine (6) is an 8aminoquinoline, which has been in use since the 1950s. Primaquine is a hypnozoitocidal drug, which when given at the right dose, is also effective against asexual stages of P. vivax (Pukrittayakamee et al. 2000) . Primaquine is still the only drug currently used worldwide for the treatment of relapses from P. vivax malaria due to dormant hypnozoites (Waters and Edstein 2012) and for the inhibition of formation of gametocytes of the parasite (Waters and Edstein 2012) . Primaquine is generally absorbed but with short half-life of between 3.7 and 9.6 h, though longer than that of artesunate (Edwards et al. 1993; Cuong et al. 2006 ). Its combination with artesunate is highly effective against the gametocyte of P. falciparum and equally effective as a hypnozoitocidal drug for the treatment of P. vivax and P. ovale infections (Baird and Hoffman 2004; Graves et al. 2012; World Health Organization 2012; Galappaththy et al. 2013 ).

Amodiaquine-artesunate combination Amodiaquine (7) belongs to the class 4-aminoquinoline (Teixeira et al. 2014 ) and is a derivative of quinoline, with modifications in its side chain. It has both schizonticidal and gametocytocidal activities against Plasmodium species. After an oral administration of amodiaquine, it is rapidly absorbed and metabolized to its active metabolite called desethylamodiaquine. Amodiaquine and desethylamodiaquine have longer half-lives (5 h and above 6 days, respectively) (Krishna and White 1996) compared to artesunate. The combination of the two drugs is recommended for the treatment of uncomplicated falciparum malaria. This combination has been demonstrated to be more effective than amodiaquine monotherapy in African children by improving the cure rate and reducing gametocytemia (Adjuik et al. 2002) .The mechanism of action of amodiaquine is still not completely understood. However, earlier reports have shown that amodiaquine inhibits the polymerization of heme to the insoluble product, hemozoin, in the parasite (Ginsburg et al. 1998 ). (8) is another drug used in combination with artesunate for the treatment of uncomplicated P. falciparum and P. vivax malaria (Poravuth et al. 2011; Rueangweerayut et al. 2012) . Pyronaridine was synthesized in the 1970s, after the emergence of parasite resistance to chloroquine. It is a derivative of 10-phenyl aminobenzo[b] [1, 5] naphthyridine. The ratio of artesunate and pyronaridine in the combination is 1:3 respectively. Pyronaridine was reported to be active against erythrocytic stages of Plasmodium species in mouse models (Looareesuwan et al. 1996; Ye and Shao 1990) . Also documented are the in vitro activities of pyronaridine against the (5a) Fig. 3 Chemical structure of sodium artesunate multidrug-resistant P. falciparum isolates (Institute of Parasitic Diseases Chinese Academy of Medical Sciences 1980). Resistance to pyronaridine has been observed in some Plasmodium strains (Wu 1988; Li et al. 1995) . The mechanism of action of pyronaridine involves the inhibition of βhematin formation, resulting in the death of the parasite. It also involves the formation of a drug-hematin complex. This allows pyronaridine to inhibit the glutathione-dependent degradation of hematin and enhance hematin-induced lysis of red blood cell. Pyronaridine is active against all the asexual stages of P. falciparum, with higher potency than chloroquine (Croft et al. 2012 ).

Mefloquine-artesunate combination Mefloquine (9) was developed in the USA by the United States Army. Mefloquine is recommended for chemoprophylactic treatment of the infections of all Plasmodium species. Mefloquine was initially used in combination with sulfadoxine/pyrimethamine, to successfully treat resistant P. falciparum malaria (Nosten et al. 1987) , until resistance emerged some years later (Nosten et al. 1991) . The new formulation of artesunate-mefloquine was suggested by the WHO technical consultation team on antimalarial drug combination therapy in 2001 (World Health Organization 2001). The fixed dose of artesunate-mefloquine was recommended for daily use over a period of 3 days (ter Kuile et al. 1995; World Health Organization 2001) . Mefloquine has a half-life of about 2 weeks and rapid asexual blood-stage and gametocyte clearance rate (White 1998) . Mefloquine acts as an active schizonticidal drug but not as an effective gametocytocidal drug. Though its exact mechanism of action is not clear, it has high affinity for erythrocyte membrane and seems to act by inhibiting heme polymerization in malaria parasite (Palmer et al. 1993; Nosten et al. 2012; World Health Organization 2015) .

Sulfadoxine-pyrimethamine-artesunate combination Sulfadoxine (10) has a structure which is similar to paminobenzoic acid (PABA), which serves as the basis for its use as an antifolate. Sulfadoxine blocks the production of dihydrofolic acid, which is essential for the biosynthesis of nucleic acid, by inhibiting the key enzyme dihydropteroate synthase (Nzila 2012) . The combination of artesunate with sulfadoxine-pyrimethamine (SP) is among the antimalarial drugs strongly recommended by the WHO for the treatment of uncomplicated malaria in children and adult, except in the first trimester of pregnancy (World Health Organization 2015), which has been adopted in different countries with no adverse drug reactions (Obonyo et al. 2003; Marquiño et al. 2005) . Sulfadoxine/pyrimethamine (11) combination with mefloquine was previously used for the treatment of resistant P. falciparum malaria before the development of resistance to this combination (Nosten et al. 1987; Nosten et al. 1991) . Sulfadoxine-pyrimethamine-artesunate combination ensured a rapid decrease in fever and asexual parasite density; gametocytemia was also significantly lowered by the combination therapy compared to sulfadoxine-pyrimethamine treatment alone (Obonyo et al. 2003; Marquiño et al. 2005) .

Proguanil (12) is a prodrug, which is metabolized to cycloguanil (13). Proguanil is a strong schizonticidal drug and acts on asexual forms of the parasite. Proguanil inhibits dihydrofolate reductase (DHFR) activity of the parasite, thereby suppressing the reduction of dihydrofolate to tetrahydrofolate, which is necessary for the biosynthesis of amino acids and nucleic acids (Nzila 2012) . Atovaquone (14), on the other hand, interferes with the integrity of parasite mitochondria and depolarizes its electron transport, thereby blocking parasite cellular respiration. In various clinical trials conducted, artesunate-atovaquone-proguanil combination was highly effective and well tolerated in the treatment of multidrugresistant P. falciparum malaria (van Vugt et al. 1999 (van Vugt et al. , 2002 Tahar et al. 2014; Wojnarski et al. 2019 ; Clinicaltrials.gov/ct2/ show/NCT02297477). In the right combination as a single formulated drug, it provides alternative treatment for pregnant women infected with multidrug-resistant P. falciparum (McGready et al. 2003) . Although it may be expensive in its formulation, it provides an effective 3-day treatment for pregnant women (McGready et al. 2003) . In a preclinical study, the use of nanosuspension of atovaquone in triple combination, in order to reduce the high cost and dose of atovaquone, has been reported to be effective, increasing its bioavailability two-fold (Harsha et al. 2020 ).

Chlorproguanil-dapsone-artesunate combination Artesunate, dapsone (15), and chlorproguanil (16) were developed as an affordable, fixed dose ACT for use in tackling the emergence of resistance of parasites to chlorproguanildapsone combination in Africa. They are metabolized after or al a dmin i stra tion i nto t he ac tive m eta bol ites d i h y d r o a r t e m i s i n i n , m o n o a c e t y l d a p s o n e a n d chlorcycloguanil, respectively. The latter provides a longer half-life to the ACT (Zuidema et al. 1986; Edstein and Veenendaal 1987) . This antifolate combination is similar to SP, but with a rapid elimination from the body (Winstanley et al. 1997 ). This triple combination therapy has undergone phase III clinical trial (ClinicalTrials.gov Identifier: NCT00344006).

Artesunate-lumefantrine-amodiaquine is another triple combination that is currently undergoing phase III clinical trial (Thu et al. 2017) . Actually, the rationale for this triple combination therapy is to circumvent the limitations of the double combination therapy such as the emergence of resistance and high doses of partner drugs. This drug was effective when administered as a 3-day regimen (ClinicalTrials.gov Identifier: NCT02453308).

Combination with antibiotics Antibiotics are used for the prevention and treatment of diseases arising from microbial infection. Just like the malaria parasite, bacterial infection does not only pose health challenges but also economic burden due to the development of resistance by bacteria to conventional drugs (Gandra et al. 2014) . The combination of an antibiotic with an antimalarial, if effective and safe, has been recommended for malaria endemic regions (White and Olliaro 1996; World Health Organization 2015) . Clindamycin (18), tetracycline (19), and doxycycline (20) have been used specifically in combination with artesunate for the treatment of malaria (Fig. 4) .

Clindamycin is an effective antibacterial compound used for the treatment of pneumonia, inflammatory diseases and endocarditis. It is also active against methicillin-resistant Staphylococcus aureus (MRSA) (Daum 2007) . Clindamycin is well tolerated when used for the treatment of P. falciparum malaria. Quinine/clindamycin combination is particularly recommended for children and is the drug of choice for pregnant women Griffith et al. 2007 ). Clindamycin is not recommended for use as an antimalarial monotherapy because isolates of P. falciparum have exhibited resistance to it (Dharia et al. 2010) . However, it is used effectively in combination therapy because of its slow action when administered with a fast-acting drug, such as artesunate Griffith et al. 2007) . Similarly, doxycycline is used to treat infection caused by protozoans or bacteria. It is recommended as a prophylactic drug, co-administered with quinine or artesunate as follow-up treatment for severe malaria or in combination with other antimalarials for uncomplicated malaria (World Health Organization 2015) . Doxycycline is a preferred partner drug due to its longer half-life, reliable absorption and its renal safety profile (World Health Organization 2015). It acts by i m p a i r i n g P l a s m o d i u m a s e x u a l d e v e l o p m e n t (Pukrittayakamee et al. 2008) . Also, its use is associated with increase in gametocytes and clearance time as recorded for sulfadoxine-pyrimethamine combination (Pukrittayakamee et al. 2008) . Doxycycline is used to prevent malaria but it is not recommended for the initial treatment of malaria, even when the parasite is sensitive to it, since its antimalarial action is delayed (Dahl et al. 2006) .

Combination of clindamycin with artesunate or quinine has been recommended for the treatment of severe or uncomplicated malaria in patients, including women in the second trimester of pregnancy; its administration is for 7 days (World Health Organization 2010). The mechanism of action involves the inhibition of microbial protein synthesis by binding to the 50S ribosomal subunit and interfering with peptide chain initiation (Tenson et al. 2003; World Health Organization 2010) .

Ferroquine-artesunate combination Ferroquine (17) is a 4aminoquinoline derivative of chloroquine with ferrocenic attached to its side chain. Ferroquines are the most potent of the chloroquine derivatives with antimalarial properties. Ferroquine and its derivatives, such as hydroxyferroquines, trioxaferroquines and chloroquine-bridged ferrocenophanes, have been extensively studied due to their promising antimalarial properties (Wani et al. 2015) . They are used in combination with artesunate to ensure better activities and delay in the emergence of resistance to both compounds. Ferroquine alone is effective against the chloroquineresistant strain of P. falciparum (Biot 2004) . It is believed to act by inhibiting hemozoin formation in the parasite (Biot et al. 2005) .

Hybrids are chemical compounds with two or more different structures and biological functions (Meunier 2007) . Hybrid molecules are synthesized in order to bring a new pharmacophore, which is expected to bring a new property to the hybrid. Hybrid molecules with higher antimalarial activity or with a synergistic effect are reported to be good candidates for new antimalarial development.

Mefloquine-artesunate hybrid Mefloquine and artesunate have been used in combination therapy since 2001 (World Health Organization 2001), as well as with sulfadoxine/ pyrimethamine for the treatment of P. falciparum malaria (Nosten et al. 1987) . The choice of artesunate and mefloquine as ACT partners is ideal because fast-acting and slow-acting drugs are combined (Wiseman et al. 2006) . The quinolinic ring of mefloquine and the endoperoxide ring of artesunate were combined in a new hybrid molecule MEFAS (Varotti et al. 2008) . MEFAS (21) had new features different from the parent compounds and was more potent than the parent compounds when administered alone and in different mass proportions (Fig. 6) . The hybrid compound was effective against both chloroquine-sensitive (3D7, IC 50 1.1 ng/ml) and chloroquine-resistant (W2, IC 50 1.0 ng/ml) strains of P. falciparum parasites (Table 1) . MEFAS was active in vivo against P. berghei, with no recrudescence during the 30 days of parasite monitoring (Varotti et al. 2008 ). MEFAS did not have SERCA as its target but altered the pH gradient across the parasite's digestive vacuole (Varotti et al. 2008) .

Primaquine-artesunate hybrid Primaquine and artesunate (PRIMAS) is a hybrid molecule prepared from primaquine and artesunate in a similar manner to MEFAS. It is also a hybrid salt between the two parent compounds. The hybrid compound was designed to minimize the toxicity associated with primaquine (Boechat et al. 2014) . Studies on PRIMAS (22) showed that it was a potent drug with less toxicity compared to the parent compound, primaquine. Also, results of in vitro and in vivo experiments revealed it to be more active against drug-resistant isolates compared to the parent compounds (Boechat et al. 2014 ).

Indoloquinoline-artesunate hybrid Indoloquinolines are isolates from the root of the medicinal plant Cryptolepis sanguinolenta, which is a climbing shrub. The isolated phytochemicals, cryptolepine, isocryptolepine, and neocryptolepine, are promising antimalarial compounds (Kirby et al. 1995; Cimanga et al. 1997; Paulo et al. 2000) . Wang et al. (2014) synthesized a series of indoloquinoline hybrids (artesunate-indolo[2,3-b]quinoline, artesunateindolo[3,2-c]quinoline and artesunate-indolo[3,2-b-]quinolone) and evaluated their antimalarial properties. Indoloquinoline was linked to artesunate through the substitution of 3-aminopropylamino group of indoloquinolines with artesunate moiety to form an amide bond. The hybrid (23) expressed low cytotoxicity and increased antimalarial activity. They were active against chloroquine-sensitive (NF54) and chloroquine-resistant (K1) strains of P. falciparum with IC 50 values of 0.45 nM and 0.42 nM respectively. The in vivo studies revealed that the hybrids had good antiplasmodial activities, reducing parasitemia by 89.6% on the 4th day and expressing a mean survival time of 7.7 days (Wang et al. 2014 ). This hybrid molecule was a more potent inhibitor of β-hematin formation compared to its parent compounds, indicating it as one of its mechanisms of action.

Polyphenol-artesunate hybrids Recently, we synthesized antimalarial hybrid compounds from artesunate and some polyphenols present in Cocos nucifera husk, the decoction of which is used for the treatment of malaria in indigenous medicine in Nigeria (Adebayo et al. 2012 (Adebayo et al. , 2013 . The compounds synthesized were artesunate-ellagic acid and artesunateprocyanidin hybrid compounds with a ratio of 1:1 of the components in each hybrid compound (Adebayo et al., unpublished; Balogun et al., unpublished) . The artesunate-ellagic acid hybrid compound was found to be more active than artesunate against P. falciparum W2 in vitro, while artesunate-procyanidin hybrid compound was more active against P. berghei NK65 compared to artesunate in vivo (Adebayo et al., unpublished; Balogun et al., unpublished) . Use of drug delivery systems Nanohybrids have been used as drug delivery systems to release drugs into a particular site in the body at a specific rate. Different drug delivery systems have been developed, including liposomes, lipid nanoemulsions and polylactide-coglycolide (PLGA) and the use of natural polymers such as collagen and chitosan (Al-Qaraghuli et al. 2017; Obeid et al. 2017a, b) . Various strategies utilize liposomes and pheroids to improve the solubility and transport of antimalarial drugs to their target sites. Pheroids are biocompatible, biodegradable, and safe compounds used in medical applications. The poor solubility and erratic absorption of artemisone (25) following oral administration have been improved by using pheroids (Steyn et al. 2011) . The formulation consisted of ethyl ester, cremaphor, α-tocopherol, and butylated hydroxyanisole as oil phase, with nitrous oxide water as the water phase; the final pheroids obtained were 3.81 μm in size (Steyn et al. 2011) .

Liposomes were among the first drug delivery systems applied in disease therapy (Gregoriadis 1976) . They are made up of phospholipid bilayers surrounded by central aqueous phase and are self-assembling spherical colloidal entities. They are amphiphilic, shielding hydrophobic groups from aqueous environment while maintaining contact with aqueous phase through their hydrophilic head groups (Makino and Shibata 2006) . Liposomes have been used to encapsulate several antimalarial compounds, including artesunate, in order to improve their delivery to target organs. Gabriёls and Plaizier-Vercammen (2003) developed a liposome formulation to improve patient's compliance to the use of artesunate. To achieve this, liposome consisting of 4:3 molar ratio of phosphatidylcholine/cholesterol was used to develop a drug delivery system that ensured the slow release of artesunate. Kim et al. (2015) prepared artesunate nanohybrids by intercalating it in zinc salt in an attempt to increase its solubility and oral bioavailability and prevent its degradation in acidic medium. Kim et al. (2015) also encapsulated the artesunatezinc basic salt (ZBS) in an enteric coating agent to further decrease the release of artesunate and prevent its decomposition at intestinal pH. The synthesized nanohybrid significantly improved the aqueous solubility and chemical stability of artesunate in acidic conditions.

Recently, we also reported the use of mesoporous silica nanocarriers as effective drug delivery systems for artesunate and quinine (Amolegbe et al. 2018) . Mobil Composition of Matter (MCM)-41 was found to be the most effective in delivering artesunate and quinine to the target site, the red blood cells, compared to other delivery systems. However, it was more effective in delivering quinine than artesunate. The MCM-41 nanoparticle delivery system for quinine enhanced the antimalarial activity of quinine by about 240-fold, with 0.0625 mg/kg body weight of quinine encapsulated in MCM-41 nanoparticle exhibiting higher inhibition of parasite growth compared to 15 mg/kg body weight of quinine (Amolegbe et al. 2018) .

Drug solubility is very important for delivery to target sites. Drug candidates from biological base screening or combination chemistry are mostly lipophilic and are expected to exert their therapeutic action across biological membranes or membrane-associated proteins (Fahr and Liu 2007) .

Artesunate is a potent blood schizonticidal drug, the solubility of which has been described as poor. Its bioavailability is also low when administered through the oral route (Li et al. 1998) . These properties limit the therapeutic applications of artesunate (Zhang et al. 2009 ). Thus, improving its solubility could increase its delivery and therapeutic effects. Carbohydrate and noncarbohydrate matrices are employed to increase the solubility of drugs and enhance their delivery to target sites (Fahr and Liu 2007; Douroumis and Fahr 2013) . This process transforms hydrophobic drugs to more watersoluble forms, thereby improving their delivery. Chadha et al. (2011) investigated the inclusion of artesunate in a carbohydrate matrix. β-Cyclodextrin (β-CD) along with other matrix was prepared and evaluated for their physicochemical properties and therapeutic indices. Similarly, Muder and Sunderland (2014) investigated the inclusion of artesunate in hydroxypropylβcyclodextrin (HPβCD), which is a chemically modified β-CD with some hydroxyl groups substituted with hydroxypropyl groups. The artesunate-cyclodextrin molecules increased the solubilization strength of artesunate, which was highest in methyl-β-cyclodextrin (Me-β-CD) (Chadha et al. 2011 ). More so, the binary Me-β-CD-lyophilized suspensions were effective against P. berghei infection in mice with no mortality recorded throughout the duration of the study (Chadha et al. 2011) .

Synthesis of co-crystals of artesunate and nicotinamide using two separate methods of solvent evaporation and slurry increased the solubility and dissolution rate of artesunate but caused no significant increase in the in vivo antimalarial activity of artesunate alone (Setyawan et al. 2015) . Co-crystal formation has been used to design crystals with improved properties such as solubility, bioavailability and stability, amidst other properties without altering the effectiveness of the parent compound (Chadha et al. 2012) . The effect of nicotinamide in the crystal formed with artesunate seems to improve the dissolution of artesunate in water, but not its antiparasitic activities in vivo (Setyawan et al. 2015) .

Artemisone Artemisone (25), a polar heterocyclic compound, was synthesized from artesunate as a second-generation drug (Fig. 7) . The modification conferred on artemisone improved pharmacokinetics and prolonged half-life and also alleviated the neurotoxicity associated with artesunate (Haynes 2006) . Also, artemisone has been reported to be active against earlystage gametocytes and asexual stages (Coertzen et al. 2018 ). Its mechanism of action has been shown to include the inhibition of PfATPase6. It was also more efficient in killing malaria parasite than artesunate in animal studies. Further studies are still being carried out on the development of artemisone (Anthony et al. 2012; Schrader et al. 2012; Barnett and Guy 2014) . Artemisone has undergone phase II clinical trial (Held et al. 2015) .

Artesunate-polyamines The synthesis of polyamines of artesunate (26) and the evaluation of their activities against NF54 and K1 strains of P. falciparum were done by Pearce et al. (2017) . Among the active artesunate-polyamine compounds are (Bis)-Boc-(bis)-artesunate-polyamine and (tetra)-artesunate-polyamine conjugates which were active in vitro against the two strains with IC 50 values ranging from 0.3 to 1.1 nM. These polyamine conjugates also caused 95.5-99.8% reduction in parasitemia with no death recorded for two of them in experimental animal studies after a 30-day monitoring period (Pearce et al. 2017 ).

The peroxide bridge of artesunate is critical to its activity. Thus, hybrid compounds were synthesized from two or more compounds such that each monomer had an endoperoxide bridge and they were evaluated for their activities, relative to the monomers. Chaturvedi (2011) synthesized trimer and tetramer derivatives of artesunate (27), with some of the oligomers exhibiting good in vitro activities against chloroquinesensitive parasites. Some dimers were also synthesized and one of them had similar in vitro activity (IC 5 0 = 0.00077 μM) against chloroquine-sensitive Plasmodium species compared to artesunate (Paik et al. 2006) . Artesunate trioxane dimer (28) prepared by Conyers et al. (2015) gave promising results in an animal model of malaria (Fig. 7) . The Fig. 7 Chemical structures of modified artesunate compounds trioxane dimer was the most effective among the drugs tested, exhibiting complete parasite clearance and also extending the mean survival time of tested mice beyond 30 days. Reiter et al. (2015) prepared several artesunate dimers and trimers and evaluated their activities against P. falciparum 3d7. The dimeric derivatives of artesunate (29 and 30) were more active compared to the trimers, with IC 50 of 2.6 nM for both of them (Reiter et al. 2015) .

Artesunate triazine hybrids (31 and 32) synthesized by Cloete et al. (2014) were active antimalarial compounds against both chloroquine-resistant (Dd2) and chloroquinesensitive (NK65) strains of P. falciparum. The synthesized dimer derivatives of artesunate were more active than their monomeric counterparts (Cloete et al. 2014 ). Among the promising compounds, one of them (32) exhibited IC 50 of 0.0079 and 0.0102 mM against NF54 and Dd2 strains of P. falciparum, respectively. The compound exhibited higher activity against P. falciparum NF54 but was less active against P. falciparum Dd2 compared to artesunate and dihydroartemisinin (Cloete et al. 2014) .

Malaria still poses a high health risk to the populace, especially where the occurrence of the resistant strains of the parasite is widespread in areas with poor vector control. Several treatment approaches have been mired by the complex nature of the life cycle of malaria parasite and the emergence of drugresistant species. Researchers are keen on finding a compound that will be effective and possess a novel mechanism of action different from those of existing drugs, in order to avoid resistance by the parasite. Over the years, several methods have been used to improve existing antimalarial compounds. This review has highlighted some approaches that resulted in improved antimalarial activity of artesunate, such as combination therapy, the use of synthetic matrix, synthesis of hybrid compounds, and chemical modifications of the structure of artesunate. Reported cases of resistance or reduced sensitivity have made some of these combination therapies to be withdrawn. It is believed that modifications of the chemical structure of artesunate and the synthesis of its hybrid compounds may avail us the opportunity of getting an array of new compounds which are more effective than artesunate and also possess novel mechanisms of action which can be deployed sequentially in combating the emergence of artesunateresistant strains over a long period of time. This is expected to reduce the scourge of the disease, especially in Africa where majority of malaria deaths occur.

Conflict of interest The authors declare that they have no conflicts of interest.

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