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La malaria est un des fléaux les plus dévastateurs dans les pays en voie de développement. L’absence d’un vaccin et la résistance aux agents antimalariaux disponibles démontrent le besoin urgent d’identifier de nouvelles cibles thérapeutiques. Les phosphoinositides (PIP) sont des composants essentiels des membranes cellulaires chez les eucaryotes jouant un rôle important dans la signalisation intracellulaire, la synthèse d’ADN et le trafic protéique, par exemple. Malgré leur importance chez les eucaryotes, on en connaît peu sur leurs fonctions chez le parasite de la malaria Plasmodium falciparum. Dans notre laboratoire, nous avons réalisé un criblage par inactivation génique de 36 effecteurs potentiels de la voie métabolique PIP pour identifier les gènes qui sont essentiels à la prolifération chez P. falciparum. Notre étude a montré que 72% des gènes potentiellement impliqués dans la voie métabolique des PIP ne pouvaient être inactivés et sont donc potentiellement essentiels pour la survie du parasite. L’analyse d’une souche knock-out pour la protéine PfPX, ayant un domaine de liaison aux PIP de type Phox, a démontré un ralentissement sévère de la croissance du parasite. La caractérisation de la protéine PfPX a révélé qu’elle se localisait à la membrane de la vacuole digestive, le site où le parasite digère l'hémoglobine (Hb) de l'hôte afin de subvenir à ses besoins en acides aminés. Nous avons montré que les parasites dépourvus de la protéine Phox accumulaient plus d'Hb et que celle-ci était piégée dans des vésicules à proximité de la vacuole digestive, suggérant un rôle pour cette protéine dans la fusion des vésicules d’Hb avec la membrane de la vacuole digestive. Globalement, nos résultats ont révélé que les PIP ont un rôle important dans le transport de l'Hb chez P. falciparum / Malaria is one of the most devastating curses in developing countries. The absence of a vaccine and resistance to available antimalarial agents demonstrate the urgent need to identify new therapeutic targets. Phosphoinositides (PIPs) are essential components of cell membranes in eukaryotes, playing an important role in intracellular signaling, DNA synthesis and protein trafficking, for example. Despite their importance in eukaryotes, little is known about their functions in the malaria parasite Plasmodium falciparum. In our laboratory, we screened 36 putative effectors of the PIP pathway by gene inactivation to identify the genes that are essential for proliferation in P. falciparum. Our studies showed that 72% of genes possibly involved in the PIP pathway could not be inactivated and are therefore potentially essential for parasite survival. Analysis of a knockout strain for PfPX protein, having a Phox-like PIP binding domain, demonstrated a severe slowdown in parasite growth. Characterization of the PfPX protein revealed that it was localized to the food vacuole membrane, the site where the parasite digests the hemoglobin (Hb) of the host in order to meet his needs in amino acids, and in vesicular type structures. We have shown that parasites lacking the Phox protein accumulate more Hb and that it is trapped in vesicles near the digestive vacuole, suggesting a role for this protein in the fusion of Hb vesicles with the membrane of the digestive vacuole. Overall, our results revealed that PIPs play an important role in the transport of P. falciparum Hb

En 2020, la malaria a provoqué 241 millions de cas d'infections et 627 000 morts. La faible efficacité du vaccin disponible et la résistance aux traitements rendent indispensable l'identification de cibles thérapeutiques. Plasmodium falciparum (Pf) envahit les érythrocytes pour s'y répliquer. Pour cela, de protéines contenues dans des organites d'invasion, tels que les micronèmes et les rhoptries rassemblés à un complexe apical, sont sécrétées. Le trafic des protéines entre l'appareil de Golgi et les organites d'invasion est médié par la PfSortiline. Dans les organismes modèles, la sortiline est recyclée entre les organites cibles et l'appareil de Golgi via le complexe protéique rétromère composé des protéines de tri vacuolaire (Vps) Vps26-29-35. Ce complexe est recruté via les complexes VpsC, composé de Vps11-16-18-33, CORVET composé de Vps3-8 et HOPS, composé de Vps39-41. Chez Pf, l'ensemble des composants des complexes VpsC et rétromère sont conservés. Seulement PfVps3 du complexe CORVET est retrouvée et le complexe HOPS est absent. L'hypothèse du projet est que ces protéines conservées sont impliquées dans la biogenèse des organites du complexe apical via le recyclage de la PfSortiline vers l'appareil de Golgi. Pour vérifier cela, des souches de parasites exprimant les protéines de fusion PfVps3-11-16-18-29 étiquetées à un domaine GFP ont été construites. Des techniques de Western Blot et de microscopie à fluorescence ont montré que ces protéines de fusion sont exprimées lors du cycle érythrocytaire. Il semble que PfVps29-GFP localise à des structures semblables aux endosomes et partiellement aux micronèmes. Les protéines PfVps16-18-GFP semblent localiser aux micronèmes et partiellement aux rhoptries et à l'appareil de Golgi. Finalement, des souches dans lesquelles les protéines PfVps3-16-29-GFP peuvent être délocalisées de façon conditionnelle ont été construites. Il a été montré que PfVps16-GFP semble essentielle à la survie de Pf. Ce projet participe à la caractérisation de nouvelles pistes thérapeutiques antipaludiques. / Malaria was responsible for 627,000 deaths and 241 million infections in 2020 alone. Drug resistance, and the poor efficacy of the only available vaccine, are strong arguments supporting the need to identify therapeutic targets. The malaria parasite Plasmodium falciparum (Pf) invades erythrocytes and multiplies inside them. To do so, it secretes invasion proteins located inside organelles, like micronemes and rhoptries, which are localised at an apical complex. Protein trafficking from the Golgi apparatus to these organelles is dependent on PfSortilin. In model organisms, this protein is recycled between the target organelles and the Golgi Apparatus by a protein complex called retromer. This complex is composed of Vacuolar Sorting Proteins (Vps)-26-29-35. The retromer complex is recruited by complexes, composed of Vps11-16-18-33, CORVET, composed of Vps3-8, and HOPS, composed of Vps39-41. In Pf, all components of the retromer and the VpsC complexes are conserved. However, only PfVps3 of the CORVET complex is conserved and the HOPS complex is absent. We hypothesized that conserved proteins play a key role in apical complex biogenesis by recycling PfSortilin to the Golgi apparatus. To verify the hypothesis, parasite strains coding the fusion proteins PfVps3-11-16-18-29 tagged with a GFP were generated. Western Blot and fluorescence microscopy showed that those proteins are expressed during the erythrocyte life cycle. PfVps29-GFP seemed to localize at endosome-like structures and partially at micronemes. PfVps16-18 seemed to localise at micronemes too and partially at rhoptries and at the Golgi apparatus. Finally, strains where PfVps3-16-29 could be functionally mislocalized have been generated. This technique showed that PfVps16-GFP were essential for Pf survival. Our work could lead to the characterization of new antimalarial drug targets.

Malaria is an infectious disease caused by Plasmodium parasites and transferred by Anopheles mosquitos. Due to Artemisinin resistance, new druggable targets identification and new drug development are urgently needed. Serine hydrolases (SHs) are one of the largest classes of enzymes having important roles in life processes. The deadliest malaria parasite, P. falciparum, encodes more than 50 SHs including proteases, lipases, esterase and others, while only several of them have been characterized. The study of uncharacterized SHs will shed light on future drug development to treat malaria. In this study, we applied chemical biology and genetic approaches to identify SHs important for the pathogenic asexual stage growth of P. falciparum parasites. We mainly focused on a depalmitoylase essential for merozoite invasion and lysophospholipases (LPLs) essential for acquiring fatty acids (FAs) from the host. Identifying essential metabolic enzymes will benefit the treatment to malaria. We focused on metabolic SHs and identified two SHs were refractory to knock out. We studied a likely essential SH named PfABHD17A, which is a human depalmitoylase homolog. PfABHD17A is localized on the rhoptry, an organelle essential for invasion. We expressed the recombinant PfABHD17A, conducted inhibitor screen and discovered that human depalmitoylase inhibitor ML211 inhibits PfABHD17A in vitro. ML211 inhibits merozoite invasion but not egress, which together with the localization of PfABHD17A on the rhoptries, suggested that PfABHD17A is essential in merozoite invasion. We also purified PfABHD17A and verified that PfABHD17A may exhibit depalmitoylase activity in vitro. LPLs are important for asexual stage parasites acquiring FAs from the host. The P. falciparum genome includes 17 putative LPLs while LPLs responsible for hydrolyzing FA from lysophosphatidylcholine (LPC) in the asexual stage are currently unknown. Using a chemical biology approach, we identified serine hydrolase inhibitor AKU-010 inhibits LPC hydrolysis effectively. Using activity-based protein profiling (ABPP) and genetic approaches, we identified that AKU-010 inhibits a series of SHs including Exported Lipases (XLs), Exported Lipases Homolog (XLH) and Plasmodium falciparum prodrug activation and resistance esterase (PfPARE). We generated a series of knockout parasite lines on the AKU-010 targets and identified that red blood cell (RBC)-localized XL2 and cytosolic XLH4 contribute to most LPC hydrolysis activity in the asexual stage. XLs and XLHs are important for parasites using LPC for growth and contribute to detoxification from accumulated LPC. XL2 and XL4 together are essential for parasite growth under high LPC concentration medium, such as human serum. XL/XLH-deficient parasites could still acquire FA from LPC, which is mainly contributed by parasite membrane- localized PfPARE. PfPARE has little impact on parasite growth and LPC metabolism with the existence of XLs and XLHs but is important after the loss of XLs and XLHs. Parasites deficient in PfPARE, XLs and XLHs have little ability to release FA from LPC and cannot use LPC as FAs source for growth. In summary, we identified metabolic SHs mediating protein depalmitoylation and lipid metabolism and in asexual stage Plasmodium falciparum, which may benefit future drug development to treat malaria. / Doctor of Philosophy / Malaria is an infectious disease caused by Plasmodium parasites and transferred by mosquitos. New druggable target identification and drug development are urgently needed to deal with the malaria issue. We focused on an understudied enzyme superfamily termed serine hydrolase (SHs), which includes more than 50 members in the deadliest malaria parasite, P. falciparum. We identified that several druggable enzymes, which can mediate protein depalmitoylation and lipid metabolism, are important for parasite growth in the pathogenic stage. Identifying essential metabolic enzymes will benefit the treatment to malaria. We screened eleven SHs and discovered that two of them are likely essential in the pathogenic stage. We focused on one human depalmitoylase homolog termed PfABHD17A. We screened the inhibitors on PfABHD17A and used the inhibitor to suggest that PfABHD17A is essential for the growth of pathogenic stage parasites. We also identified lipases important for acquiring fatty acids (FAs) from the host. Using chemical biology and genetic approaches, we discovered that three lipases are important for acquiring FAs form the host in the pathogenic stage. Inhibiting these enzymes may kill the parasite in the host.

La malaria, endémique dans 91 pays tropicaux et sub-tropicaux, est l’une des maladies infectieuses les plus mortelles chez l’humain. Le fardeau de cette maladie porte principalement sur l’Afrique, qui compte plus de 90% des cas d’infections ainsi que des morts enregistrés, la majorité étant des enfants en bas âge. Des cinq espèces de parasites du genre Plasmodium qui peuvent causer la maladie chez l’humain, Plasmodium falciparum est de loin la plus mortelle et la plus étudiée. La résistance aux médicaments actuels et l’absence d’un vaccin préventif procurant une immunité de longue durée démontrent l’urgent besoin de trouver de nouvelles cibles thérapeutiques. Chez les cellules eucaryotes, l’identité des organites cellulaires est définie par les phosphoinositides, des composants mineurs des membranes cellulaires, et maintenue grâce aux kinases et aux phosphatases impliquées dans leur métabolisme. Les rôles de certaines phospholipides-kinases dans plusieurs étapes critiques du cycle de vie de Plasmodium ont récemment été découverts, toutefois, rien n’est connu quant aux fonctions des phosphoinositides phosphatases de cet organisme. Les travaux décrits ci-dessous présentent une première caractérisation d’une protéine homologue à la famille des phosphoinositides phosphatases SAC1. Les résultats montrent que cette protéine est exprimée durant tout le cycle érythrocytaire asexué et qu’elle se localise au réticulum endoplasmique ainsi que potentiellement à l’appareil de Golgi. L’étude de lignées conditionnelles et knockout suggèrent qu’un niveau minimal de la protéine est nécessaire pour la survie du parasite durant le cycle érythrocytaire. En somme, la combinaison des résultats obtenus laisse penser que cette protéine pourrait avoir une fonction dans le système de sécrétion du parasite P. falciparum et qu’elle pourrait donc constituer une cible thérapeutique intéressante pour le développement de nouveaux antimalariaux. / Malaria is endemic in 91 tropical and sub-tropical countries and is one of the deadliest infectious human diseases. Africa has the highest burden with more than 90% of cases and malaria deaths registered yearly, mostly in children under 5 years-old. Despite the fact that infection in human can be caused by five Plamsodium species, infection by Plasmodium falciparum is the most severe and therefore the most studied. Resistance to antimalarials and the absence of a preventive vaccine show the urgent need of new therapeutic targets. In eukaryotic cells, organelles identity is defined by phosphoinositides, minor membranes components, and maintained by the kinases and phosphatases involved in their metabolism. The fact that certain kinases have roles in critical steps of Plasmodium life cycle has recently been acknowledged. However, the roles of the phosphatases are still unknown. My work presents a first characterization of a putative phosphoinositide phosphatase of the SAC1 family. Results provided show that the protein is expressed throughout the asexual blood stages and that it localizes to endoplasmic reticulum and potentially to the Golgi apparatus. Studies on knockdown and knockout strains suggest that a minimal amount of the protein is required during the asexual blood stages. In summary, the combination of the results presented suggests that the protein has an important function in the parasite P. falciparum secretion system and therefore, may represent an interesting potential target for drug development.

Malaria, a disease caused by members of the Plasmodium genus, remains a threat to global health. Despite the availability of therapeutics, Plasmodium's propensity for generating resistance-conferring mutations threatens the efficacy of these drugs. Therefore, it is essential to develop novel therapeutics, and one approach to discover such compounds is to repurpose current drugs as antimalarials. Human kinase inhibitors, most of which are developed as antineoplastics, are a valuable source of such novel compounds. Human kinase inhibitor research spans over twenty years, generating a wellspring of knowledge regarding compound design, mechanism, and tolerability that can be leveraged in the quest to develop new antiplasmodial drugs. Furthermore, the plasmodial kinome differs substantially from the human kinome, providing opportunities for selectivity and minimization of off-target effects in the host. To this end, we sought to identify and characterize compounds within human kinase inhibitor collections that have antiplasmodial effects. One library yielded a potent polo-like kinase 1 (PLK1) kinase inhibitor, BI-2536, which possessed potent antiplasmodial activity in both the asexual blood stage and liver stage and likely acts through involvement of amino acid starvation. Another library comprised exclusively of type II kinase inhibitors, designed to target kinases in the inactive conformation, produced several interesting lead compounds – TL5-135, YLIU-06-026-1, and the analog pair XMD13-99 and WZ9-034-2. These compounds were highly active against asexual blood stage parasites, killing rapidly while also possessing favorable selectivity and liver stage activity. In vivo, TL5-135 and YLIU-06-026-1 acted prophylactically by preventing infection, and therapeutically by resolving an established infection. Currently, investigations are underway to determine the mechanism of action of the lead compounds and to improve their druglike properties. In whole, this effort has not only yielded promising antiplasmodial compounds, but it also underscores the value of the repurposing approach in the quest for novel antimalarial drugs.

Plasmodium falciparum est un protozoaire parasite responsable du paludisme causant la mort d’environ un million de personnes par an. La résistance médicamenteuse du parasite augmente la pathogénicité de cette maladie. Il est question ici d’explorer les mécanismes moléculaires impliqués dans la mort cellulaire programmée (apoptose) du parasite en présence de chloroquine (CQ) et de tester l’hypothèse qu’une résistance à la CQ peut s’expliquer en partie par une défaillance de ce mécanisme de mort. Dans un premier temps l’étude des marqueurs de l’apoptose (TUNEL, JC1, formes pyknotiques) montre qu’une souche sensible de parasite (3D7) à la CQ peut subir une apoptose en présence de CQ alors qu’une souche résistante (7G8) présente un défaut d’apoptose. Dans un deuxième temps nous montrons que la protéine PfMCA1 (P. falciparum métacaspase 1) présente une structure et une maturation protéolytique proche de celui des caspases faisant de cette protéine un candidat potentiellement impliqué dans l’apoptose du parasite. Dans un troisième temps nous montrons que l’expression du domaine catalytique de PfMCA1 dans la levure induit une mort cellulaire et un retard de croissance de la levure. Nous montrons également que PfMCA1 présente une activité enzymatique de type arginase alors que les effets induit par sa surexpression peuvent être inhibés par l’ajout d’un inhibiteur de protéases spécifiques des aspartates. Ces résultats suggèrent que PfMCA1 pourrait agir comme une protéine initiatrice induisant l’action d’une protéase effectrice spécifique des aspartates conduisant à la mort cellulaire. Cette hypothèse testée chez la levure reste à confirmée chez P. falciparum / Plasmodium falciparum is a protozoan parasite responsible for malaria causing one million deaths per year. Drug resistance of the parasite increases the pathogenicity of this disease. In this thesis, it is question to explore the molecular pathway involved in programmed cell death (apoptosis) of the parasite in the presence of chloroquine (CQ) and to test the hypothesis that CQ resistance could be partly explained by a failure of such a mechanism. In a first step, we showed that a sensitive clone (3D7) exhibited the classical hallmarks of apoptosis (DNA fragmentation, mitochondrial depolarization) under a CQ pressure while a resistance clone failed to undergo apoptosis. In a second step we show that the protein PfMCA1 (P. falciparum metacaspase 1) has a structure and a processing similar to the well known caspases which are the key effectors of apoptosis for higher eukaryotic cells. In a third step we show that expression of the catalytic domain of PfMCA1 in yeast induces cell death and growth retardation of yeast. We show that PfMCA1 presented an arginine-specific protease activity while the effects induced by its overexpression were inhibited by an aspartate-specific protease inhibitor (z-VAD-fmk). These results suggest that PfMCA1 might act as an initiator protein inducing an aspartate-specific protease effector leading to cell death. This hypothesis tested in yeast remains to be confirmed in P. falciparum

Le parasite Plasmodium falciparum, responsable des formes graves du paludisme chez l'homme, est transmis par Anopheles gambiae, son principal vecteur en Afrique sub-saharienne. Les nouvelles stratégies de lutte contre la maladie visent à limiter ou à interrompre le développement du parasite chez le moustique vecteur, et il est donc nécessaire d'améliorer notre compréhension des interactions entre le vecteur, son environnement et le parasite. L'objectif de ce projet de thèse a été de caractériser la flore microbienne du vecteur An. gambiae en conditions naturelles de transmission, d'étudier le rôle des principales espèces bactériennes colonisant l'estomac du moustique sur le développement de P. falciparum et de mesurer l'influence des bactéries sur la réponse immunitaire des moustiques femelles et leur capacité à transmettre le parasite. Pour mener à bien ce projet, nous avons collecté des populations de moustiques sauvages au Cameroun pour la caractérisation de la flore microbienne, nous avons ensuite exposé des moustiques de la colonie de laboratoire Ngousso à des cultures bactériennes puis infecté ces moustiques avec des isolats naturels de P. falciparum. Notre étude a montré que les souches bactériennes naturelles de l'intestin du moustique Serratia, Pseudomonas et Escherichia réduisaient la prévalence et l'intensité de l'infection et que le degré d'inhibition variait selon les taxons bactériens et les porteurs de gamétocytes. L'analyse des flores bactériennes des différents épithéliums de l'insecte par pyroséquençage a révélé des similarités entre la flore intestinale et celles retrouvées dans les ovaires et les glandes salivaires pour un même moustique. Les analyses d'expression suggèrent que la régulation de l'expression des gènes l'immunité par les bactéries intestinales pourrait participer à la modulation de la réponse antiplasmodiale. Les mécanismes impliqués dans les interactions bactéries-Plasmodium-vecteur sont complexes et multifactorielle et la modélisation de l'ensemble des interactions qui permettent à P. falciparum d'accomplir son cycle chez le moustique vecteur sera nécessaire pour envisager de nouvelles méthodes de lutte efficaces et durables. / Plasmodium falciparum, the parasite responsible for the severe form of malaria, is transmitted by Anopheles gambiae, its major vector in sub-Saharan Africa. Novel strategies for malaria control envision interrupting the sporogonic development in An. gambiae, then it is important to better understand vector*environment*parasite interactions that underlie parasite transmission. The aim of this project was to characterize the microbial flora of An. gambiae in natural conditions, to study the role of the main bacterial strains on sporogonic development using natural isolates of parasites and to measure the influence of bacterial exposure on the mosquito immunity and its successive ability to transmit P. falciparum. To carry out this project, we used wild mosquito populations from Cameroon to characterize the mosquito microbial flora, next we challenged female mosquitoes of the Ngousso colony to bacterial strains and then infected the mosquitoes with natural isolates of P. falciparum. Our study showed that Serratia, Pseudomonas and Escherichia isolated from the mosquito midgut reduced infection prevalence and intensity and that the effect of the bacterial exposure on parasite infection levels varied between bacterial strains and gametocyte carriers. The analysis of the 454 sequencing of the different mosquito epithelia revealed intriguing similarities between bacterial communities in the midgut, ovaries and salivary glands of a single mosquito. Expression analyses suggested that immune gene regulation by midgut bacteria could help the mosquitoes to mount an effective antiplasmodial response. Mechanisms involved bacteria-Plasmodium-vector interactions are complex and rely on multiple factors. Deeper investigations on these interactions that allow P. falciparum to complete its cycle in the mosquito vector will be necessary for modeling parasite transmission in the field and for developing new methods for effective malaria control.

A malária, doença causada pelo protozoário do gênero Plasmodium, está entre as doenças que mais causam mortes os países subdesenvolvidosn. O hospedeiro é infectado por meio da picada do mosquito do gênero Anopheles, que introduz o parasita durante a hematofagia. As formas mais graves são causadas pelo Plasmodium vivax e o Plasmodium falciparum. As regiões mais afetadas por estas formas são África Subsaariana, Ásia, América Central e Sul. Desde o começo do século XXI, a Organização Mundial de Saúde (OMS) busca erradicar a doença, porém o P.falciparum se mostrou resistente aos fármacos antimaláricos existentes, dificultando a eficácia do tratamento. Isto, entre outros fatores, como mortalidade e alto índice de infecção, tornam necessárias novas pesquisas para a descoberta de novos fármacos mais seguros e eficazes contra a malária. Estudos têm mostrado como um alvo promissor para a criação de novos antimaláricos, a cisteína protease falcipaína, a qual se apresenta em três isoformas no parasita, sendo elas, falcipaína 1, 2 e 3. A falcipaína 2 está ligada com a hidrólise da hemoglobina, e seus inibidores vem sendo estudados como alternativas na busca de agentes antimaláricos. Derivados de semicarbazona, tais como o nitrofural e o hidroximetilnitrofural demonstraram atividade inibitória de cisteíno proteases parasitárias. Utilizando estratégias modernas de planejamento de fármacos e por meio da integração entre técnicas computacionais e experimentais, realizou-se o planejamento, síntese e avaliação biológica de compostos derivados dos ditiocarbazatos e tiossemicarbazonas, bioisosteros de semicarbazona, como inibidores da cisteíno protease falcipaína 2, no intuito de obter novos antimaláricos. Aplicaram-se técnicas de modelagem molecular em três séries de compostos (A, B e C), sendo a A e B derivados dos ditiocarbazatos e a C das tiossemicarbazonas. Estes estudos sugerem, três compostos da série A, quatro na série B e três na C com maior potencial para inibição da falcipaína 2. Isso devido aos resultados teóricos indicarem condições favoráveis ao ataque nucleofílico da cisteína 42 catalítica da falcipaína 2 às tiocarbonilass presentes nos compostos planejados. Estes derivados foram sintetizados, analisados por espectroscopia de ressonância magnética de 1H e 13C, espectroscopia de IV, ponto de fusão e pureza caracterizando sua formação. Após a obtenção, os compostos foram enviados para ensaios biológicos frente ao parasita P. falciparum. Os compostos testados não apresentaram inibição, porém é sabido que muitos inibidores enzimáticos não são ativos contra o parasita mesmo tendo alta potência contra a enzima, isto devido às barreiras a serem ultrapassadas até chegar ao alvo bioquímico, deste modo faz-se necessário ensaios contra a enzima para validar nossa hipótese. / Malaria, a disease caused by the protozoan of the genus Plasmodium, is among the most deadly diseases in poor countries. The host is infected through the bite of the mosquito of the genus ,i>Anopheles, which introduces the parasite during hematophagy. The most severe forms are caused by Plasmodium vivax and Plasmodium falciparum. The regions most affected by these forms are Sub-Saharan Africa, Asia, Central and South America. Since the beginning of the 21st century, the World Health Organization (WHO) has sought to eradicate the disease, but P. falciparum has been resistant to antimalarial drugs treatment. Among other factors, such as mortality and high infection rates, new research is needed to find new, safer and more effective drugs against malaria. Studies have shown as a promising target for the creation of new antimalarial drugs, the cysteine protease falcipain, which is present in three isoforms in the parasite: falcipain 1, 2 and 3. Falcipain 2 is linked to the hydrolysis of hemoglobin, and its inhibitors have been studied as alternatives in the search for antimalarial agents. Derivatives of semicarbazone such as nitrofural and hydroxymethylnitrofural demonstrated inhibitory activity of parasitic cysteine proteases. Using modern strategies for drug design and the integration of computational and experimental techniques, the design, synthesis and biological evaluation of compounds derived from dithiocarbazates and thiossemicarbazones, semicarbazone biosynthesis as inhibitors of cysteine protease falcipain 2 were carried out in order to new antimalarials. Molecular modeling studies were performed in three series of compounds (A, B and C), with A and B being derived from dithiocarbazates and C from thiossemicarbazones. These studies suggest three compounds in the A series, four in the B series, and three in the C group with the greatest potential for inhibition of falcipain 2. This is due to the theoretical results indicating favorable conditions for the nucleophilic attack of the catalytic cysteine of falcipain 2 on thionyls present in the compounds planned. These derivatives were synthesized, analyzed by 1H and 13C magnetic resonance spectroscopy, IR spectroscopy and melting point, characterizing their formation. After being obtained, the compounds were sent for biological assays against the P. falciparum parasite. The compounds tested did not show inhibition, but it is known that many enzyme inhibitors are not active against the parasite even though they have high potency against the enzyme, this is due to the barriers to be overcome until reaching the biochemical target, thus enzyme to validate our hypothesis.