Malaria continues to threaten human beings, causing a staggering number of more than 400,000 deaths each year. Although effective treatment and prevention methods are available, rapidly emerging resistance towards existing drugs is of great concern, and the need for novel antimalarial compounds are still urgent. The Malaria Box lead molecules MMV008138 and MMV665831 are promising in this regard, due to their apparently novel antimalarial mechanisms of action. The target of MMV008138 is the PfIspD enzyme in the MEP pathway, which is absent in humans. This difference makes the PfIspD a great target. However, while MMV008138 shows potency against Plasmodium falciparum-infected human erythrocytes in vitro, no efficacy was observed in a humanized mouse model or a P. berghei infected mouse in vivo. In order to block potential metabolic spots and to probe for steric demand, a series of analogous featuring C1-deuteration, methyl substitution on B- and C-ring, and an ethylene bridge were prepared. The effect of various substitution on the tetrahydro-β-carboline conformation and D-ring orientation was studied. In the course of preparing the C1-Me analog of MMV008138 featuring 2',4'- dichloro substitution, unexpected ring-expanded azepane products were isolated. Later it was found that the desired product could be isolated when the imine formed was treated with acid at lower temperature. Other intermediates possessing a 2ʹ- substituent were also isolated under the low temperature acid treatment protocol, which upon heating in acid gave the ring-expanded azepane we initially isolated. A mechanism was proposed to account for the formation of the azepane as well as other intermediates. The driving force of the expansion reaction was explored, and the hypothesis that the steric interaction between the 2ʹ-substituent and the C1-Me was supported via DFT calculation and conformational analysis. MMV665831 is another potent hit from the Malaria Box, and it appears to inhibit the hemoglobin endocytosis process of P. falciparum. The structure–activity relationship of MMV665831 was studied with analogues featuring modifications on C2-benzamide, C3-ester, C-7 phenol, as well as the phenolic Mannich base moiety. Modifications at phenolic Mannich base moiety leads to the discovery of an analogue that is twice as potent toward cultured P. falciparum compared to MMV665831. We were worried the phenolic Mannich base moiety might act as the precursor of toxic quinone methide intermediates, and designed two analogs to block this potential toxicophore. Although the modification resulted in reduced potency, this result proved that the potency of MMV665831 does not stem from the formation of quinone methides. Unfortunately, MMV665831 did not reduce parasitemia in P. berghei- infected mice. Fast hepatocyte metabolism was observed for MMV665831, and the loss of in vivo efficacy was discussed in comparison with other phenolic Mannich bases with similar hepatocyte stability. / Doctor of Philosophy / In the fight against malaria, one concerning issue is the rapidly emerging resistance towards existing drugs. The continuous development of antimalarials with novel mechanism of action is greatly needed. To accelerate the development of novel antimalarials, an open access ensemble of 400 compounds that are toxic to the malaria parasite known as the Malaria Box, has been made available. My work involves the optimization of two compounds from this ensemble, MMV008138 and MMV665831. MMV008138 kills the malaria parasite by inhibiting an enzyme named PfIspD, which is absent in human. In the parasite an enzyme called PfIspD is responsible for the biosynthesis of IPP and DMAPP, two chemical building blocks that are essential for all cells. It is unlikely that MMV008138 will interrupt with the biosynthesis of IPP and DMAPP in human, since we use another enzyme to synthesize them. Although MMV008138 shows great in vitro potency, but did not protect a live mouse from malaria infection. The lack of in vivo efficacy could stem from the rapid metabolism of MMV008138, and analogs aimed to prevent metabolism were designed and prepared. While preparing analogs featuring 2ʹ-substitution, the desired product was not found, but other unexpected by-products were isolated. The conditions that leads to both the desired products and the by-products were found, and the mechanistics detail of this unexpected reaction were studied. During the blood-stage, which causes malaria symptoms in human, the Plasmodium falciparum parasite invades and feeds on human red blood cells (erythrocytes). The parasite destroys human hemoglobin through a multistep process that begins by transporting the hemoglobin from the red blood cell into itself, a process called endocytosis. MMV665831 appears to interfere with this endocytosis process of P. falciparum, thus starving the parasite of its food. Analogs of MMV665831 were prepared to probe for the effect on potency, and one compound that is twice as potent in cultured parasites was found. The structure of MMV665831 contains a potentially unstable moiety, which might lead to toxicity in humans. Two analogs with the problematic moiety removed were designed and prepared, and one still shows antimalarial activity, showing that the reactivity of the potentially unstable moiety is not the reason for the antimalarial activity of MMV665831. However, MMV665831did not protect P. berghei-infected mice (murine malaria) in vivo, and the reason for the loss of efficacy was discussed.