A Brief Historical Overview of the Antimalarials Chloroquine And

Iowa State University Capstones, Theses and Creative Components Dissertations

Spring 2021

A brief historical overview of the antimalarials chloroquine and artemisinin: An investigation into their mechanisms of action and discussion on the predicament of antimalarial drug resistance

Ekaterina Ellyce San Pedro

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A Brief Historical Overview of the Antimalarials Chloroquine and Artemisinin: An

Investigation into their Mechanisms of Action and Discussion on The Predicament of

Antimalarial drug resistance

By Ellyce San Pedro

Abstract:

Malaria is a problem that has affected humanity for millenia. As a result, two important antimalarial drugs, chloroquine and artemisinin, have been developed to combat Malaria.

However, problems with antimalarial resistance have emerged. The following review discusses the history of Malaria and the synthesis of chloroquine and artemisinin. It discusses both drugs’ mechanisms of action and Plasmodium modes of resistance. It also further discusses the widespread predicament of antimalarial drug resistance, which is being combated by artemisinin-based combination therapy. Lastly, a case study is discussed concerning the reintroduction of chloroquine in areas where it has previously failed due to resistance.

Introduction:

Malaria or in Italian, “Mal ‘aria’” (Hempelmann & Krafts, 2013) has plagued humanity for millenia. From the ancient Egyptians in 1550 B.C. (Fagan, 2000) to the present day, historical records detail a deadly disease that results in symptoms such as shaking, fever, fatigue and death

(CDC 2021). In the present day, Malaria has persisted, resulting in around 409,000 deaths in

2019 alone (CDC 2021). Prior to the modern day, Malaria’s origins were considered mysterious and miasmic in nature. But with the help of modern technology, great clinical advancements San Pedro 2 have been made in decreasing its potency. In particular, the antimalarial drugs chloroquine and artemisinin are today’s front-line defense against this dangerous disease.

Parasite Life Cycle:

Malaria is caused by a protozoan parasite. Malaria can be commonly contracted from four specific sporozoan species: P. malariae, P. falciparum, P. vivax and P. ovale (Fagan, 2000).

The most common malarial infections are derived from the P. falciparum and P. vivax species

(Fagan, 2000).

The following parasite cycle can be used to generally describe the life cycles of the above listed parasites. The parasite life cycle requires a human host and a mosquito host (CDC 2021).

The cycle begins when an infected female mosquito partakes in a human blood meal, which allows the entry of sporozoites into the skin (CDC 2021). The sporozoite then moves into the circulation (CDC 2021). Sporozoites are ambulatory parasites derived from a female anopheline mosquito that infects humans and targets human hepatocytes (WHO 2015).

After sporozoite entry, a sporozoite infection of liver cells occurs. Sporozoites are obligate intracellular parasites that develop within the host cell during the vertebrate stage of the life cycle. The sporozoites then mature into schizonts (WHO 2015). Schizonts are mature malarial parasites that reside in the host’s liver cells where they undergo nuclear division (WHO

2015). Upon their maturation, the schizonts burst and release merozoites (CDC 2021).

Merozoites are parasites that are released into the bloodstream when a liver cell bursts (WHO

2015). Upon formation, the merozoites proceed to invade erythrocytes. It is also important to note, that as the parasite grows within the erythrocyte, it must digest the host cell’s hemoglobin to acquire amino acids to support its metabolism and also to create more space in the host cell to San Pedro 3 grow. The sporogonic cycle or the sexual reproduction portion of the parasite life cycle occurs when the parasites multiply within the mosquito (CDC 2021). This occurs when parasite gametes are ingested by a mosquito in another blood meal. During the sporogonic cycle, the male microgametes fuse with female macrogametes to produce a zygote (CDC 2021). These zygotes will eventually mature into oocytes that will become sporozoites (CDC 2021). These sporozoites will be released from the mosquito’s salivary glands (CDC 2021). This continues the parasite’s life cycle as the mosquito continues taking blood meals.

Past Explanations for Malaria:

In the past, many explanations have been attributed to the cause of Malaria. Malaria was first attributed to miasma. Miasma was thought to be a dangerous cloud of particles that caused diseases (Hempelmann & Krafts, 2013). This led to the coinage of the Italian phrase “mal’aria”, indicating bad air (Hempelmann & Krafts, 2013).

The idea of a malarial parasite was not introduced until microscopic staining techniques were able to detect the presence of the parasites in blood (Hempelmann & Krafts, 2013). Dr.

Ronald Ross, a British army surgeon, was the first individual to prove that mosquitoes transmitted Malaria (Hempelmann & Krafts, 2013). In 1897, Ross obtained mosquitoes and gave them blood that had crescent-shaped cells (Murray, 1923). After feeding, Ross saw that there were pigmented crystals in the stomach wall (Murray 1923). Ross realized that mosquitoes do not normally produce the pigment known as hemozoin and he deduced that this pigment could be related to Malaria (Murray, 1923). Prior to this in 1880, Dr. Alphonse Laveragen discovered that

Malaria was caused by a protozoan parasite (Nye, 2002). This discovery occurred when Dr. San Pedro 4

Laveragen, a military surgeon, visualized the malarial parasite through the use of blood smears

(Nye, 2002).

Past Treatments/Drugs Used to Treat Malaria:

Antimalarial drugs have had a colorful history. Alternatives to quinine (the derivative for chloroquine), have included methylene blue dye, Prontosil and Atabrine (Lowe, 2020).

Unfortunately, these drugs were not highly effective. They have also had the unfortunate side effect of turning patients into a whole spectrum of colors (Lowe, 2020). Methylene blue gave patients a blue tint. Prontosil turned patients into a permanent shade of red (Lowe, 2020).

Atabrine gave individuals a yellow hue, while also causing depression, psychosis, seizures, and other problems (Lowe, 2020). In terms of today’s antimalarial treatments, the CDC states that common drugs that are used to treat Malaria include: atovaquone/Proguanil, and chloroquine

(CDC 2021). However, chloroquine and artemisinin remain as the two frontline drugs that are most commonly used to combat the effects of Malaria.

Chloroquine - A History:

Chloroquine was discovered in 1934 by Johnann Hans Andersag (Lowe, 2020) from cinchona trees that are located in South America. Chloroquine is a weak base drug and it belongs to the family of 4-aminoquinolines (Coban, 2020). Quinolone was extracted in 1820 by researchers Pelletier and Caventou (Lowe, 2020). Quinine was synthesized in 1944 (Lowe,

2020). Unlike its predecessors, chloroquine possesses strong antimalarial activity and does not change the appearance of a patient's skin tones (Lowe, 2020). San Pedro 5

Chloroquine Mechanism of Action:

Chloroquine's mechanism of action is still not fully understood. However, there are several suppositions that may explain how chloroquine functions in infected erythrocytes. An early theory for chloroquine’s mechanism of action focused on chloroquine’s ability to bind

DNA and RNA (Coban, 2020). It was previously thought that chloroquine would bind to the host cell’s DNA and RNA through hydrogen bonds and electrostatic forces (Coban, 2020). However,

DNA-chloroquine interactions required an excessively high concentration of chloroquine to occur (Coban, 2020). This amount was much higher than needed to eliminate parasites, therefore making it unlikely that this supposition was true (Coban, 2020).

The second and more substantial mechanism of action relies on chloroquine’s interactions with free-heme in the infected erythrocyte. Chloroquine weakens the malarial parasite by reducing its ability to synthesize hemozoin, a substance that is significant to Plasmodium survival (Pilat et al, 2020).

In the absence of antimalarial drugs, the malarial parasite performs hemoglobin degradation using proteases such as plasmepsins and falcipains (Coban, 2020). Plasmepsins and falcipains are a part of Plasmodium-specific families of aspartic proteases and cysteine proteases.

These proteases catabolize hemoglobin to release peptides and nutrients that the parasite requires to survive (Wicht et al, 2020). However, it has also been shown that many of these amino acids are returned to the serum. Thus, hemoglobin degradation is also surmised to create space in the infected erythrocyte for parasite growth (Krugliak et al, 2002). San Pedro 6

During the process of hemoglobin degradation, heme is released and converted into alpha-hematin (ferriprotoporphyrin IX) which is toxic to the parasite (Coronado et al, 2015). The parasite normally evades this problem by transforming alpha-hematin into hemozoin (or beta-hematin), which is non-toxic to the parasite (Coronado et al, 2015).

Chloroquine inhibits hemozoin synthesis in the following manner. Chloroquine moves from the cell cytosol into the digestive vacuole within the infected cell (Wicht et al, 2020). Upon entrance into the digestive vacuole, the weak base chloroquine is protonated and becomes unable to exit the digestive vacuole membrane (Wicht et al, 2020). The protonated chloroquine then binds to the alpha-hematin and to the surfaces of already existing hemozoin crystals (Wicht et al,

2020). Once this binding occurs, hemozoin formation is inhibited and leads to swelling of the digestive vacuole and the accumulation of toxic free alpha-hematin in the vacuole (Wicht et al,

2020). Additionally, chloroquine damages the digestive vacuole’s membrane and allows for the release of proteases into the cell (Coban 2020).

Methods used to Determine Anti-Malarial Drug Resistance:

Anti-malarial drug resistance is the parasite’s ability to survive despite antimalarial drugs

(Basco, & Ringwald, 2000). Anti-malarial drug resistance is a problem that has appeared in many areas of the world. Examples include P. falciparum resistance to chloroquine in Southeast

Asia, Colombia, and Papua New Guinea (Mita et al, 2009).

As such, several tools have been developed to survey antimalarial resistance. The three most commonly used approaches are: in vivo studies, in vitro/ex vivo studies and molecular assays. In vivo studies are used for determining how effective the drug is in patients. In vitro/ex San Pedro 7 vivo studies determine how susceptible a parasite is to the drug in question. Molecular assays are used to find gene mutations that may be related to drug resistance (Nsanzabana et al, 2018).

In vivo studies are conducted by prescribing a specific dosage of the drug to patients who are experiencing uncomplicated Plasmodium infections (Nsanzabana et al, 2018). After treatment, a clinical follow up is conducted for a specific time period to assess if the treatment was successful or not (Nsanzabana et al, 2018). In vitro/ex vivo studies use parasite samples that have been acquired from patients or from cultures (Trager & Jensen, 1976). The parasites are then cultured in either different concentrations of the anti-malarial drug or through exposure to high concentrations of the antimalarial for a short interval of time to observe its effects

(Witkowski et al, 2013). Different methods that are used to measure parasite growth include: microscopy, isotopic tests, ELISA, fluorescent markers, flow cytometry and RSA or ring-stage survival assays (Nsanzabana et al, 2018). Molecular methods are also used to determine polymorphisms that are related to anti-malarial drug resistance (Nsanzabana et al, 2018). This technique involves the detection of (SNPs) single nucleotide polymorphisms (Nsanzabana et al,

2018), which can be determined in a variety of different ways.

Anti-Chloroquine Resistance

Selective pressure is defined as a factor involving natural selection that selects for one group over another (Hilaris 2021). As previously mentioned, chloroquine and artemisinin have been used as the drugs of choice to eliminate malarial parasites. However, this has also resulted in the emergence of chloroquine and artemisinin resistance. This section will discuss chloroquine resistance. San Pedro 8

Chloroquine has been used as the antimalarial drug of choice for decades since its inception. Over time, it has been deemed as a safe and affordable antimalarial. This was the case until resistance against chloroquine was seen in Southeast Asia and also in South America (Kim et al, 2019).

There are also several theories for anti-chloroquine resistance. The most supported mechanism is that chloroquine resistance is associated with a series of point mutations in the

PfCRT or the P. falciparum CQ-resistant transporter (Kim et al, 2019). The PfCRT is a transporter whose structure includes 424 amino acids as well as 10 transmembrane helix domains

(Wicht et al, 2020). These components create a negatively charged chamber that faces the digestive vacuole (Wicht et al, 2020). The cavity is significant because it is believed to attract positively charged drugs for transport out of the cytosol. In this case, it is believed to transport the positively-charged chloroquine out of the cytosol (Wicht et al, 2020). Once chloroquine is removed from the digestive vacuole, the parasite can continue to detox by synthesizing the nontoxic hemozoin.

The important mutation involved changes a lysine to a threonine at the 76th position

(Wicht et al, 2020). The K76T mutation is important because this mutation arises with the presence of other mutations that are highly specific to region or location (Wicht et al, 2020). The structure of the PfCRT showed that the mutations C72S, N326D , K76T, A220S, I356L and

A220S are inside the drug-binding chamber (Wicht et al, 2020). This is important because four of these mutations are necessary for chloroquine resistance (Wicht et al, 2020).

Artemisinin - A History: San Pedro 9

Artemisinin was created in 1972 amidst controversy in the east (History of antimalarials

2021). In 1955, The United States and Vietnam were at war (History Channel Editors, 2017). But

Vietnam was also fighting a war on a second front. Vietnam was struggling against Malaria. As such, Vietnam turned to its ally, China for aid (Faurant, 2011). In response, on May 23, 1967,

Mao Tse-tung began the 523 research program, which led to the discovery and development of artemisinin (Faurant, 2011). Youyou Tu from the Chinese Academy of Medical Sciences led the project (Krungkai & Krungkai, 2016). During the project, the team investigated and researched

2,000 traditional Chinese recipes and 380 herbal extracts in a bid to find a compound that possessed strong antimalarial properties (Krungkai & Krungkai, 2016).

Artemisinin was founded from ancient Chinese medicine (Krungkai & Krungkai, 2016).

An ancient text described sweet wormwood as a treatment for Malaria and it was identified as an effective anti-malarial (Faurant, 2011). However, at first there was confusion between the forms of quinghao or huanghuahao, wherein quinghao or artemisinin emerged as the effective version

(White et al, 2015).

Artemisinin was also developed against the backdrop of the Chinese Cultural revolution in 1967 (Faurant, 2011). After artemisinin was developed, the relationship between China and

Vietnam soured. For a time, artemisinin’s significance was lost.

Artemisinin Mechanism of Action:

Artemisinin is believed to function by utilizing free heme-bound iron to catalyze parasite-toxic free radicals. (Krungkai & Krungkai, 2016). Upon entry into the digestive vacuole, artemisinin is activated when its endoperoxide bridge is cleaved by iron protoporphyrin IX or alpha-hematin (Wicht et al, 2020). As discussed above, iron protoporphyrin IX or Fe2+ heme is a San Pedro 10 byproduct of the parasite-digested hemoglobin (Wicht et al, 2020). Artemisinin’s endo-peroxide bridge is then reduced from Fe2+ to the ferrous iron Fe3+ (Krungkai & Krungkai, 2016). The

Fe2+-heme-artemisinin carbon-centered radicals then cause damage to the parasite by damaging its proteins, lipids and other components (Wicht et al, 2020).

There are two theories for the activation of artemisinin, the mitochondrial and the heme-degradation pathway (Muangphrom et al, 2016). As stated above, artemisinin activation occurs when the endoperoxide bridge has been cleaved. In the mitochondrial pathway, artemisinin is activated by the mitochondria and is associated with lipid peroxidation due to the synthesis of reactive oxygen species. It is also thought to cause the depolarization of plasma membranes and mitochondria (Muangphrom et al, 2016). The heme-mediated pathway is a bit more complex because it involves two proposed activation models, the reductive scission model and the open peroxide model (Muangphrom et al, 2016).

In the Reductive Scission Model it has been proposed that oxygen centered radicals change conformation to create carbon centered radicals (O’Neil et al, 2010). Ferrous heme was also seen to bind to artemisinin and to eventually cause a cleavage of the peroxide bridge that created oxygen centered radicals that would change shape to create carbon centered radicals

(O’Neil et al, 2010).

In the Open Peroxide model, it is suggested that protonation drives ring opening (O’Neil et al, 2010). It is suggested that iron works as a Lewis acid by promotion of the ionic bioactivation of artemisinin (O’Neil et al, 2010). It has also been suggested that non-peroxide oxygen aids with ring opening to create hydroperoxide (O’Neil et al, 2010). The oxygen atom stabilizes positive charges and reduces the energy that is needed for ring opening. Cleavage of the endoperoxide bridge and inclusion of water creates an unsaturated hydroperoxide that San Pedro 11 modifies proteins using direct oxidation (O’Neil et al, 2010). This degradation of hydroperoxide creates a hydroxyl radical that oxidizes certain amino acids (O’Neil et al, 2010 ). The open peroxide model could possibly create several ROS or reactive oxygen species that affect antimalarial activity (O’Neil et al, 2010).

Other targets of artemisinin include the creation of reactive oxygen species through activation of the mitochondrial electron transport chain (Krungkai & Krungkai, 2016).

Artemisinin Resistance and K-13:

Artemisinin has been highly effective in treating Malaria. Artemisinin resistance was first reported in 2009 in falciparum malaria patients located on the Thai-Cambodian border (Krungkai

& Krungkai, 2016). More cases of artemisinin resistance were seen in 2014, where it was reported that resistance was spreading from Vietnam to Myanmar (Krungkai & Krungkai, 2016).

One of the main players in artemisinin resistance is the Kelch-13 protein. The Kelch-13 protein or K-13 protein is important for the development of the parasite in the erythrocytic cycle

(Wicht et al, 2020). The K-13 protein is important because it controls the endocytosis of hemoglobin into the digestive vacuole (Birnbaum et al, 2020). Digested hemoglobin is partially used for the synthesis of amino acids that are used to create the parasite’s proteins, and its digestion results in the creation of toxic, alpha-hematin (Krugliak et al, 2001). Alpha-hematin is important because it activates artemisinin through the cleavage of its endoperoxide bridge.

Mutations that resulted in decreased levels of K-13 were seen to reduce the endocytosis of hemoglobin, which resulted in reduced levels of alpha-hematin or Fe(II)PPIX for artemisinin activation, leading to artemisinin resistance (Birnbaum et al, 2020). More research also showed San Pedro 12 that disruption of the pathway through the interruption of falcipain 2a or the addition of E64 (a protease inhibitor) can increase the parasite’s resistance to artemisinin (Wicht et al, 2020).

K-13 has also been seen to change the cell cycle in K-13 C580T clinical isolates during the ring-to-trophozoite stage, leading to additional reduction of alpha-hematin for artemisinin activation (Wicht et al, 2020). It has also been theorized to disrupt proteins and cause dysregulation of PfPK4 phosphorylation (Wicht et al, 2020). It has also been suggested that

K-13 uses the UPR or unfolded protein response to dispose of damaged proteins as a way to respond to cell stress (Wicht et al, 2020). However, there is also evidence that K-13 mutations do not cause resistance through the modulation of proteasomal activity (Wicht et al, 2020). K-13 mutations have also been linked to the mitochondria of the parasite. (Wicht et al, 2020).

ART Resistance Non-Related to K-13:

K-13 is not the only gene that has been associated with artemisinin resistance. Other antimalarial suspects include: UBP-1, V3275F mutation in pfubp1 and the T381 mutation (Wicht et al, 2020). The UBP-1 is thought to be significant in artemisinin resistance because of a linkage group selection analysis of a genetic cross that involved Plasmodium chabaudi between a selected artemisinin-resistant parent and an artemisinin-sensitive parent (Wicht et al, 2020).

According to Wicht et al, the V3275F mutation for pfubp1 was correlated with resistance in vitro, but not in in vivo experiments. It is also important to note that the UBP-1 and the V3275F mutation have not been found to be correlated with reduced clearance of P. falciparum infections after artemisinin treatment (Wicht et al, 2020). The T381 mutation is considered to be more compelling because of a GWAS related to the China-Myanmar border that entailed the P. falciparum’s weakness to artemisinin (Wicht et al, 2020). San Pedro 13

Combination Therapy as a Solution to Anti-Malarial Drug Resistance

As discussed above, new solutions are necessary to overcome anti-malarial drug resistance in Asia, Africa and other areas of the world (Nyunt & Plowe, 2007). Combination drugs therapies are therefore used to block and deal with drug resistance (Nyunt & Plowe, 2007).

Combination therapy was suggested in 1955 (ter Kile et al, 2007) and has been supported by multiple studies, including those by Peters (Covell et al, 1955), White and Olliaro (Nyunt &

Plowe, 2007) and White et al (Nyunt & Plowe, 2007).

An early test of this theory was the combination of chloroquine and pyrimethamine in the

1950s and 1960s (Nyunt & Plowe, 2007). This test failed when resistance evolved in response to chloroquine and pyrimethamine usage (Nyunt & Plowe, 2007). This combination was also used in Asia and in Africa, but only led to anti-malarial resistance (Nyunt & Plowe, 2007).

Today’s new approach is to use drugs that have different mechanisms of action (Nyunt &

Plowe, 2007). The mainstay of this theory is artemisinin in ACTs or artemisinin-based combination therapies (Nyunt & Plowe, 2007). Artemisinins have long been considered to be the most potent antimalarial drug. Combining different drugs with artemisinin has led to high efficacy rates, even with antimalarials that were not considered as potent as artemisinin (Van der

Pljim et al, 2021). In this treatment, a large percentage of parasites are eliminated by artemisinin and the remaining parasites are dealt with by the partner drug (Van der Pljim et al, 2021). As such, ACTs are considered a significant treatment against Malaria in areas where other drugs have been compromised by drug resistance (Nyunt & Plowe, 2007). San Pedro 14

Other combination therapies recommended by WHO include artesunate-mefloquine and artesunate-amodiaquine, among others (Nyunt & Plowe, 2007). However, in these combinations, sometimes the non-artemisinin based drug has a longer half life and is somewhat compromised by resistance (Nyunt & Plowe, 2007). This implies that the non-ART partner drug is at risk of resistance during the time period after treatment, when new parasites can emerge from the liver (Nyunt & Plowe, 2007). Unfortunately, this means that resistance may still occur and may be accelerated in regions where anti-malarial drug treatment is common (Nyunt &

Plowe, 2007). To combat this problem, it is highly important to choose ideal partner drugs and to use them in the correct time frame (Nyunt & Plowe, 2007) .

Phenomenon of older anti-malarial drugs being brought back to fight against Malaria.

The previous discussion has involved different types of chloroquine and artemisinin resistance, which has reduced the efficacy of these antimalarial drugs in certain regions of the world. In 1993, physicians in Mawai had to replace chloroquine with pryimethamine and sulfadoxine due to chloroquine resistance (Laufer et al, 2006). At the time, chloroquine efficacy had been reduced to less than 50% (Laufer et al, 2006). However, by 2001, chloroquine resistant falciparum Malaria became undetectable (Laufer et al, 2006). It was hypothesized that chloroquine could return as an effective anti-malarial drug in Malawi (Laufer et al, 2006).

In the study, a randomized clinical trial was held with 210 individuals that had uncomplicated Plasmodium falciparum Malaria (Laufer et al, 2006). The individuals were treated with either chloroquine or sulfadoxine-pyrimethamine to test for antimalarial efficacy of the drugs (Laufer et al, 2006). The results showed that treatment failure occurred in 1/80 cases that were assigned to chloroquine (Laufer et al, 2006). In other words, the cumulative efficacy of San Pedro 15 chloroquine had risen to 99% compared with the less than 50% efficacy rate of chloroquine in

1993 (Laufer et al, 2006). In comparison, sulfadoxine-pyrimethamine treatment had failed in

71/87 cases (Laufer et al, 2006). The cumulative efficacy of sulfadoxine-pyrimethamine was

21% (Laufer et al, 2006).

It was previously suggested that reduction of chloroquine useage could result in the return of a chloroquine-sensitive P. falciparum parasite. This was hypothesized to be due to a mutation in the PfCRT gene that confers chloroquine resistance but also comes at the expense of a fitness cost to the parasite. Specifically, chloroquine resistance could be linked to defective hemoglobin catabolism, which could eventually eliminate or reduce the presence of the chloroquine-resistant parasite in the absence of drug pressure (Lewis et al, 2014).

The point mutation in the PfCRT gene is associated with chloroquine-resistant falciparum

Malaria (Laufer et al, 2006). This marker was measured before and after reducing the use of chloroquine. Results showed that from 1992-2000 the marker for PfCRT decreased as the use of chloroquine was reduced (Laufer et al, 2006). In 2001, the marker was not seen. This is in contrast to countries that still use chloroquine and have infections where more than 90% of the P. falciparum infections are due to chloroquine-resistant parasites (Laufer et al, 2006). This strongly suggests that chloroquine can and should be used again as an effective anti-malarial in the Malawi region and this method can possibly be used in other locations to help reduce drug-resistant malaria.

Conclusion:

Over the centuries, Malaria and humanity have had an interesting and colorful relationship. Artemisinin and chloroquine are antimalarial drugs that have been developed in San Pedro 16 response to malarial pressure. Antimalarial resistance has developed in both drugs, which poses problems for preventing and treating antimalarial resistance. As such, ACT’s or artemisinin based combination therapies have been used to offset antimalarial resistance. Additionally, it may be possible to reuse chloroquine or other antimalarials in areas where there was previous antimalarial resistance to the drug that has been eliminated.

However, new therapies will still be needed to treat Malaria in the future. The new frontier of antimalarial treatment may be rooted in gene therapy through the concept of drug targeting of single lethal pairs for drug resistant malaria (Lee et al, 2013). If tangible, it may be possible to eliminate Plasmodium cells while also protecting human cells against elimination

(Lee et al, 2013), In the future, this type of treatment may lead to the reduction of unpleasant symptoms for patients as well as the possible future elimination of Malaria. San Pedro 17

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