Nature-derived Peptides as Promising Antiparasitic Agents against Neglected Tropical Diseases

Abstract

Neglected tropical diseases (NTDs) are a group of at least 20 infectious diseases that primarily affect tropical regions. Despite impacting 1.6 billion people worldwide, these diseases have not received adequate global priority and attention. Currently, NTDs caused by protozoa have limited therapeutic options, and the emergence of drug-resistant strains further exacerbates the situation. In recent years, several antimicrobial peptides (AMPs) have emerged as potential therapeutic candidates against NTDs. This review analyzes the contemporary trends of AMPs, explores their antiparasitic properties, and mechanisms of action against three parasitic protozoan NTDs: Chagas disease, human African trypanosomiasis, and leishmaniasis and one parasitic helminth NTD: lymphatic filariasis. Furthermore, notable drawbacks associated with AMPs are highlighted, and future research directions are proposed. Overall, this review points out the potential of AMPs as therapeutic agents for these three protozoan neglected tropical diseases and one parasitic helminth NTDs as well as emphasizes the imperative need for continued research in this field.

Introduction

Neglected tropical diseases (NTDs) are tropical diseases caused by protozoa, helminths, bacteria, and viruses that primarily affect marginalized populations in Africa, certain parts of Asia and South America (Joshi et al. 2021). The term “neglected” highlights the fact that these diseases are often overlooked by global health initiatives and are not prioritized in the Global Health Security Agenda (GHSA). This lack of priority is particularly concerning because NTDs disproportionately affect individuals living in rudimentary, impoverished areas and isolated rural regions, where access to necessities such as clean environments, personal hygiene, and quality healthcare is severely limited (Joshi et al. 2021). Additionally, due to their low priority in the GHSA, NTDs struggle to obtain funding, and are often excluded from global funding agencies, relegating them to the periphery of global health policy discussions (Engels and Zhou 2020; de Souza and Dorlo 2018). As a result, the incidence of these diseases continues to rise, with over 1 billion people recently affect worldwide (Mitra and Mawson 2017).

According to World Health Organization (WHO 2020), there are 20 infectious diseases have been classified as Neglected Tropical Diseases (NTDs) (accessed 2 November 2023). These diseases include dracunculiasis, echinococcosis, foodborne Trematodiases, onchocerciasis, schistosomiasis, soil-transmitted helminthiases, cysticercosis, buruli ulcer, leprosy, trachoma, yaws, mycetoma, dengue, scabies, snakebite envenoming, Chagas disease, human African trypanosomiasis, leishmaniasis, and lymphatic filariasis with the last 4 being the focus of this review (Rosenberg et al. 2016).

Currently, the treatment options for protozoal diseases, particularly in the case of Chagas disease, are limited. There are only two drugs, nifurtimox and benznidazole, have shown efficacy in combating the parasite Trypanosoma cruzi (T. cruzi). However, it is concerning to note that T. cruzi has developed resistance to benznidazole (Campos et al. 2014). Meanwhile, the current principal therapeutic strategies for leishmaniasis still harbor persistent issues, such as teratogenic effects caused by the drug miltefosine, high costs, and protracted treatment regimens (Parthasarathy and Kalesh 2020). Because of the limited availability of effective therapeutic agents, the emergence of drug-resistant parasites and the serious side effects of existing drugs, it is crucial to discover potent novel compounds that can act as antiparasitic agents against neglected tropical diseases (NTDs).

In recent decades, natural products have become a valuable source of alternative therapies for various NTDs. For instance, a neolignan known as eupomatenoid-5, isolated from Piper regnellii var. pallescens, has been shown to be effective against Trypanosoma cruzi (T. cruzi) and the promastigote, axenic, and intracellular amastigote forms of Leishmania amazonensis by depolarizing the mitochondrial membrane and disrupting the cell membrane of parasites (Vendrametto et al. 2010; Garcia et al. 2013; Lazarin-Bidóia et al. 2013). Other phytochemicals such as ferulic acid, a phenolic acid and rosmarinic acid, a polyphenol isolated from Pluchea carolinensis, have also demonstrated efficacy against intracellular amastigotes and promastigotes of Leishmania amazonensis (Montrieux et al. 2014). Additionally, demethylpraecansone B and praecansone B have been found to possess antiparasitic activities against Trypanosoma brucei rhodesiense (T. b. rhodesiense), the parasite responsible for human African trypanosomiasis (Tarus et al. 2002). Phytochemicals are well-known for their efficacy against bacteria, fungi, and parasites, including NTDs with substantial research has supported their therapeutic potential against these diseases. However, there is a growing interest in antimicrobial peptides which have shown broad-spectrum antimicrobial activities against bacteria, fungi and parasites (Chen et al. 2022). This emerging area of research holds promise for the development of novel therapeutic agents against NTDs that may surpass traditional phytochemicals in terms of efficacy and safety profile.

Numerous studies have reported on the significant antimicrobial properties of nature-derived peptides (Wang and Wang 2016). As the world is grappling with the onset of antimicrobial resistance (AMR), there is a growing pharmacological interest in antimicrobial peptides (AMPs) due to their effectiveness against various microbial infections and their low susceptibility to bacterial resistance evolution by targeting multiple targets in pathogens. In recent years, there has been a significant amount of research focused on using AMPs isolated from natural sources to combat NTDs (El Shazely et al. 2020). This review aims to analyze the current trend of AMPs, compile and evaluate the effectiveness of nature-derived antiparasitic peptides (APPs) stored in APD3 Antimicrobial Peptide Database against Chagas disease, human African trypanosomiasis and leishmaniasis. Additionally, we will explore the efficacy of nature-derived AMPs not mentioned in the database. These promising nature-derived peptides serve as a solid foundation for the development of innovative antiparasitic therapeutics against these diseases.

Trends of AMPs in Recent Times

Antimicrobial peptides (AMPs) are natural antibiotics, consisting of a class of bioactive small proteins that are naturally isolated from various living organisms, such as plants, fungi, bacteria, and a variety of animals, including amphibians, fish, insects, reptiles, and mammals (Santos et al. 2022). Occasionally, AMPs may transcend their initial use and find themselves appealing for other biological applications such as anticancer agents (Tornesello et al. 2020). The APD3 Antimicrobial Peptide Database currently contains 3940 identified AMPs, with 145 of these specifically recognized for their antiparasitic properties, classifying them as antiparasitic peptides (APPs) (accessed on 10 January 2024). The number of identified APPs has exhibited a substantial growth since 1965, as depicted in Fig. 1. This growth can be attributed to the sustained interest in APPs since the mid-1980s when the fight to eliminate NTDs gained momentum. This coincides with the launch of a network of research laboratories devoted to the study of parasitic diseases in the late 1970s by the Rockefeller Foundation (Molyneux et al. 2021). Therefore, the increasing identification of AMPs highlights their potential therapeutic applications.

Fig. 1

Discovery trend of APPs from 1965 to 2023 based on the APD3 Antimicrobial Peptide Database (accessed on 10 January 2024)

Classification of APPs Based on Sources

Among these 145 reported APPs, the majority peptides are derived from natural sources, with amphibians being the most significant peptide source contributing to 39% of the total, followed by insects (12%), mammals (12%), arachnids (10%), plants (8%), fish (6%), bacteria (2%), reptiles (1%), fungi (1%) and birds (1%). Moreover, approximately 8% of the identified APPs are synthetic peptides, as shown in Fig. 2.

Based on the APD3 Antimicrobial Peptide Database, amphibian sources make a critical contribution to the pool of APPs, with almost 40% of APPs, as shown in Fig. 2. Notable peptides derived from amphibians such as gaegurin, brevinine, temporin, magainin, ranalexin, caerulein precursor factor (CPF), xenopsin precursor factors (XPF), phylloseptin, and dermaseptin peptides. These peptides are primarily extracted from the skin secretions of various frog species, including the genera of Rana, Lithobates, Pelophylax, Xenopus, Phyllomedusa, Litoria, Cruziohyla, and Boana, which are found within the families of Ranidae, Pipidae, and Hylidae (Conlon 2011; König et al. 2015; Tian et al. 2021).

Besides, insects are also a significant source of APPs, accounting for approximately 12% of all APPs (Fig. 2). Notable examples of insect-derived peptides include cecropin and defensins, which have been isolated from the fly of Drosophila genus in the Drosophilidae family (Hanson and Lemaitre 2020). Besides, cecropin peptide have also been identified in moth species such as the Hyalophora genus in the Saturniidae family and the Galleria genus in the Pyralidae family (Steiner et al. 1981; Tonk et al. 2019). Additionally, mammal sources contribute to about 12% of APPs, with examples such as cathelicidin, intercrine, alpha- and beta-defensins peptides isolated from the human Homo genus in the Hominidae family, cathelicidin peptides from cattle Bos genus in the Bovidae family, and horse Equus genus in the Equidae family (Kościuczuk et al. 2012; Murakami et al. 2004; Selsted et al. 1985; Skerlavaj et al. 1996).

Fig. 2

APPs derived from different sources from year 1965 to 2023 based on the APD3 Antimicrobial Peptide Database (accessed on 10 January 2024)

The selectivity and potency of APPs against parasites are influenced by several factors, including peptide length, net charge, hydrophobicity, amino acid composition, and structural class. In terms of peptide length, Wang (2022) mentioned that APPs can be grouped into four groups based on their amino acid residues count: short (< 24 amino acids), medium (25 to 50 amino acids), long (50 to 100 amino acids) and protein (> 100 amino acids). Figure 3 shows that nearly half of the identified APPs (70 peptides) fall into the medium peptide category, with a length of 25 to 50 amino acids. Medium-length peptides have been found to have enhanced antiparasitic activity compared to short peptides, as increased peptide length has showed that they are more destructive to parasitic membranes (Sun et al. 2014). Additionally, medium-length peptides exhibited lower toxicity and greater stability against degradation than long peptides (Liu et al. 2007; Kim et al. 2014). Importantly, APPs require a minimum length of 7 to 8 amino acids to form an amphipathic structure (Bahar and Ren 2013). This structure is crucial for their ability to interact with parasitic membranes, and exhibit antiparasitic effects against parasites. Therefore, APPs with superior antiparasitic activities and low toxicity against human cells, they typically have lengths between 10 and 50 amino acids (Munk et al. 2014; Yan et al. 2020). This situation is evident in Fig. 3, where approximately 84% of APPs with antiparasitic properties have lengths between 10 and 50 amino acids.

Fig. 3

Classification of APPs based on peptide length. Abbreviation: aa, amino acid based on the APD3 Antimicrobial Peptide Database (accessed on 10 January 2024)

Besides peptide length, the net charge of a peptide is an important factor in determining its potency as an antiparasitic agent. Here, APPs can be categorized into three groups according to their net charge: cationic (net charge > 0), neutral (net charge = 0) and anionic (net charge < 0). Based on the APD3 Antimicrobial Peptide Database, the average net charge of the 145 APPs is 4.02, indicating a predominantly cationic nature with a net charge > 0 (accessed on 10 January 2024). Figure 4 illustrates this classification, with approximately 95% of APPs being cationic, and the highest number of entries (27 entries) having a + 4 charge. In contrast, only 3% of the APPs are anionic, and a mere 1% of the APPs are neutral. This predominance of cationic APPs with an overall positive charge is due to the presence of arginine, lysine, and histidine (Ulmschneider 2017). This characteristic enhances their binding affinity to negatively charged microbial membranes, providing them an advantage in exerting antiparasitic activity through membrane disruption (Benfield and Henriques 2020).

Fig. 4

Distribution of APPs based on net charge based on the APD3 Antimicrobial Peptide Database (accessed on 10 January 2024)

Beside net charge, the hydrophobic content of APPs is also an important factor determining their effectiveness against parasites. Li et al. (2021) mentioned the importance of achieving a balance between cationic charge and hydrophobicity, known as the amphiphilic balance, as it can greatly impact a peptide’s selectivity and its activity. Hydrophobic percentage of a peptide is calculated by determining the total number of hydrophobic residues (leucine, isoleucine, valine, methionine, phenylalanine, tryptophan, alanine, and cysteine), and dividing it by the total number of amino acids in the peptide (Bobde et al. 2021). Based on Fig. 5, most APPs (89%) have a hydrophobic content within the range of 31–70%, with the highest number of entries (44 entries) falling within the 41–50% range. This finding suggests that most APPs are amphipathic, containing both cationic and hydrophobic components, which is a critical feature contributing to their antiparasitic activities.

On the other hand, determining the standard amino acid composition in a peptide is important for understanding its biological activity (Mishra et al. 2012). Based on amino acids reference chart given by Merck, the 20 common amino acids can be categorized into four groups: hydrophobic side chain, unique amino acids, polar neutral side chains and electrically charged side chains (accessed on 12 January 2024). For instance, the amino acids with hydrophobic side chains include leucine (L), isoleucine (I), valine (V), methionine (M), phenylalanine (F), tryptophan (W), alanine (A), and cysteine (C). The unique amino acids consist of two amino acids, such as proline (P) and glycine (G). The amino acids with polar neutral side chains include serine (S), threonine (T), asparagine (N), glutamine (Q), and tyrosine (Y), while the electrically charged side chains include aspartic acid (D), glutamic acid (E), lysine (K), arginine (R), and histidine (H) (Merck; Chou and Fasman 1973). By analyzing the average percentage of the amino acid compositions in the 145 antiparasitic peptides from the APD3 Antimicrobial Peptide Database, it was found that leucine (L), glycine (G), serine (S), and lysine (K) are the most frequently occurring residues, as shown in Fig. 6. Remarkably, these amino acids, are also commonly present in antibacterial, antiviral, anticancer, and antifungal peptides, suggesting their important role in influencing peptide biological activity (Wang and Wang 2009). This information can be valuable for designing a new antiparasitic peptide with enhanced antiparasitic activity by using these four amino acids as a backbone for the peptide.

Fig. 5

Distribution of APPs based on their percentage of hydrophobicity based on the APD3 Antimicrobial Peptide Database (accessed on 10 January 2024)

Fig. 6

The average percentage of the 20 standard amino acids occurred in 145 of APPs based on the APD3 Antimicrobial Peptide Database (accessed on 10 January 2024). Red color bars represent hydrophobic side chain group; blue color bars represent unique amino acids group; yellow color bars represent polar neutral side chain group; green color bars represent electrically charged side chain group. Abbreviation: I, isoleucine; V, valine; L, leucine; F, phenylalanine; W, tryptophan; A, alanine; C, cysteine; P, proline; G, glycine; S, serine; T, threonine; N, asparagine; Q, glutamine; Y, tyrosine; D, aspartic acid; E, glutamic acid; K, lysine; R, arginine; H, histidine

Initially, APPs were classified into three groups based on their secondary structures: alpha (α)-helical, disulfide bonded beta (β)-sheet and those with high levels of specific amino acids such as proline, tryptophan, and histidine compositions (Boman 2003). However, a more detailed classification was later introduced by Wang (2010) into the APD3 Antimicrobial Peptide Database, which categorized the three-dimensional (3D) structures of APPs into four families based on their secondary structures (α-helix and β-sheet): alpha (α), beta (β), alpha beta (αβ) and non-alpha beta (non-αβ) families. The α family consists of peptides with α-helical structures, whereas the β family consists of peptides with β-sheet structures. The αβ family consists of peptides with both α-helical and β-sheet structures, and the non-αβ family does not have either α-helical or β-sheet structures. It is crucial to determine the 3D structure of peptide because it plays a significant role in understanding mechanism of actions of peptides (Wang 2023).

In the APD3 Antimicrobial Peptide Database, there are certain peptides that do not have 3D structure information, making it difficult to categorize them into the four mentioned families. To solve this issue, additional annotations such as the presence of disulfide bridges, unknown 3D structure, and richness in certain amino acids are used for grouping (Wang 2015). For example, certain peptides with cysteine knots, which are characterized by the presence of three disulfide bridges between six cysteine residues in their structures, providing stability to the protein structure (Daly et al. 2003). The peptides with these structural motifs are categorized as disulfide bridge group. Similarly, some of the peptides with high levels of specific amino acids, such as proline-rich peptides, are grouped as rich in certain amino acids. Those peptides without any of these characteristics mentioned are grouped as unknown in the database. Hence, the APD3 Antimicrobial Peptide Database has a total of seven groups to classify the 145 APPs: α-helix group, β-sheet group, αβ group, non-αβ group, disulfide bridge group, rich in certain amino acids group, and unknown structures group. Based on Fig. 7, most identified peptides have α-helical structures (45 entries), followed by peptides with disulfide bridges (20 entries), peptides with combined αβ structures (13 entries), peptides with β-sheet structures (5 entries), peptides with non-αβ structures (3 entries), and peptide rich in certain amino acids (2 entries). Meanwhile, there are 57 entries with unknown structures, reflecting the lack of mechanistic information for these peptides (Wang 2022). Most APPs exhibited an α-helical structure, which is the most common structural class that usually found in the extracellular fluids of living organisms, including amphibians, arthropods, and reptiles. These α-helical peptides typically have a linear, unstructured form and lack of cysteine residues (Ohtsuka and Inagaki 2020). When they interact with biological membranes, they easily transform into helical conformations (Sani and Separovic 2016). Additionally, these α-helical peptides have a high proportion of hydrophobic residues, making up approximately 50% of their composition. This characteristic enables them to transform into amphiphilic structures when in contact with biological membranes, thus giving them the ability to permeabilize parasitic membranes (Lewies et al. 2015).

Fig. 7

Classification of APPs based on structural class (accessed on 10 January 2024)

In 2015, Wang introduced a novel classification method, known as the universal classification (UC) system due to the limited known 3D structures of peptides (Wang et al. 2015). This system categorizes peptides based on their bonding patterns, which is an important factor as every chemical bond has their specific arrangements that significantly constrain the folding space of a protein chain (Wang 2022). These variant bonding patterns can give rise to a distinct protein conformation (Blackstock 1989), which are crucial in determining peptide structure and activity. The UC method consists of four classes: Class I (UCLL), Class II (UCSS), Class III (UCSB), and Class IV (UCBB). Class I (UCLL) includes the linear peptides that do not involve in chain connections with different amino acids. Class II (UCSS) includes the peptides with covalent bonds between different peptide side chains, which also known as sidechain-sidechain connections. Class III (UCSB) covers the peptides consisting of chemical bonds connected from peptide side chains to the backbone (N or C- terminus) between different amino acids, whereas Class IV (UCBB) includes the circular peptides with peptide bonds connected between the N and C-terminus (Wang 2015, 2022). In the APD3 Antimicrobial Peptide Database, there are a total of five classes, including UCLL, UCSS, UCBB, UCSB, and “unclassified”, used to categorize the APPs (accessed on 10 January 2024). Figure 8 shows a significant number of peptides (75 entries) are grouped in the UCLL class, followed by the UCSS class (37 entries), UCBB class (12 entries), and UCSB class (1 entry). Meanwhile, there are 20 entries of peptides grouped in the “unclassified” class. Hence, further research on those peptides is necessary to determine their bonding patterns which is critical to understanding their activities. Most peptides belong to the UCLL class, suggesting that most APPs with antiparasitic activities have linear structures.

Fig. 8

Classification of APPs based on their bonding patterns based on APD3 Antimicrobial Peptide Database (accessed on 10 January 2024)

Some of the nature-derived APPs mentioned in the APD3 Antimicrobial Peptide Database have shown good antiparasitic effects against several parasites, including those accountable for Chagas disease, human African Trypanosomiasis, and Leishmaniasis. This review explores the potential efficacy of these APPs in combating these diseases and aims to provide valuable insights that could aid the development of innovative antiparasitic therapeutics.

Chagas Disease

Chagas disease, also referred to as American trypanosomiasis, is a chronic parasitic disease caused by the protozoan parasite T. cruzi, which is endemic in Latin America (Echeverría et al. 2020). This pathogen is transmitted to humans through vector transmission by coming into contact the feces of infected triatomine bugs, a member of the Triatominae subfamily under the Reduviidae (Liu and Zhou 2015). These triatomine bugs are commonly found in the cracks and crevices in poorly constructed housing in rural areas, such as those made of palm thatch, mud, straw, and adobe (Gurevitz et al. 2011). The transmission of the disease normally occurs when an infected triatomine bug bites an individual and expels the contaminated feces, which contains flagellated trypomastigotes of the T. cruzi parasite, near the bite wound or mucous membranes, such as the eyes and mouth. T. cruzi enters the body when the individual unintentionally rubs the contaminated feces into an open wound or ingest it through the mouth or eyes (Reisenman et al. 2011). Besides, infections can occur through other modes of transmission, such as congenital transmission from an infected mother to baby, blood and blood products transfusion, organ transplantation from an infected donor, and oral transmission through the ingestion of food or drink contaminated with the feces of infected triatomine bugs or the consumption of raw meat from infected animals (Fearon et al. 2013; Coura 2015).

According to the Pan American Health Organization (PAHO), there are an estimated 6 million people are affected by Chagas disease, with 28 000 new cases and 12 000 death cases occurring every year in Latin America (accessed on 1 December 2023). The Global Burden of Disease (GBD) study has shown a significant decreased in the prevalence of Chagas disease globally from 1990 to 2019 (accessed 30 November 2023). For example, GBD study has shown the prevalence in Bolivia (South America) decreased from 12 393 cases per 100 000 in 1990 to 4630 cases per 100 000 in 2019. This decrease can be attributed to the implementation of public health programs, such as the Southern Cone Initiative, the Central America Initiative, and the Andean Pact Initiative, which aimed to eliminate the insect vector population that involved in the transmission of Chagas disease (Guzmán-Bracho 2001; Bonney 2014). Besides, efforts such as serological screening of donated blood have been critical in reducing the transmission of Chagas disease (Pati et al. 2022). While endemic countries, such as those in Latin America, have seen a reduction in cases, non-endemic countries, including the United States, Australia, Canada, Japan, and Europe, have witnessed an increase. For example, Spain (Europe) experienced an increased from 14.91 prevalent cases per 100 000 in 1990 year to 67.06 prevalent cases per 100 000 in 2019 year. Notably, the number of cases has reached its highest peak at 96.49 cases per 100 000 in 2010, as shown in GBD study. This increase in cases in non-endemic countries is attributed to the widespread migration of human populations from Latin America to non-endemic countries, expanding the geographical boundaries of this disease since the 1990s (Klein et al. 2012). Other factors contributing to the rise in the number of cases of Chagas disease in non-endemic regions include limited serological screening systems and insufficient training and experiences for physicians (Bonney 2014; Lidani et al. 2019). This situation has turned Chagas disease into a significant global health concern.

According to Salassa and Romano (2019), the life cycle of T. cruzi involves three distinct morphological forms: epimastigotes, amastigotes, and trypomastigotes. Epimastigotes and amastigotes are non-infective, meaning that they are only involved in the replicative stage present in the insect vector and mammalian cell, while the trypomastigote form is solely involved in the infective stage. This form consists of two different forms: metacyclic trypomastigotes, which are available in insect hindgut, and bloodstream trypomastigotes, which are available in the mammalian host blood (Pech-Canul et al. 2017). The first stage begins when triatomine bugs become infected with the epimastigote form of T. cruzi while feeding on an infected host, such as humans or animals. These infected triatomine bugs play a crucial role in transmitting T. cruzi to a new host. This transmission takes place when the infected triatomine bugs excrete their feces, which contains trypomastigotes. These trypomastigotes can enter the new host through various entry points, including the mouth, the conjunctiva of the eyes, and bite wounds. Once the host becomes infected, the trypomastigotes directly penetrate the host cells at the site of infection. They then transform into amastigotes and reproduce asexually through binary fission. These amastigotes subsequently transform into bloodstream trypomastigotes and enter the bloodstream to search for new host cells to invade at other infection sites. Once the trypomastigotes have entered other infection sites, they transform into amastigotes, thus continuing the replication cycle, as shown in Fig. 9 (Martín-Escolano et al. 2022).

Fig. 9

The lifecycle T. cruzi parasite in human stage and sandfly stage. Created with BioRender.com

Chagas disease has two phases: the acute phase, which lasts 2–8 weeks, and the chronic phase, which can extend over 10–30 years. During the acute phase, patients may either be asymptomatic or experience symptoms, such as headaches, tachycardia, fever, and lymphadenitis. In contrast, for chronically infected patients, most patients typically remain asymptomatic, but approximately 30% of patients developing to clinical manifestation such as gastrointestinal and heart defects, which known as Chagas cardiomyopathy (Bianchi et al. 2015). Currently, the main drugs used to treat the acute phase of Chagas disease are benznidazole and nifurtimox, but their efficacy against intracellular parasites during the chronic phase is limited (Bermudez et al. 2016). Additionally, concerns have arisen regarding adverse reactions related to benznidazole and nifurtimox such as pruritus, skin rash, anorexia, nausea, abdominal pain, and insomnia (Crespillo-Andújar et al. 2018; Aldasoro et al. 2018), the emergence of resistant strains of T. cruzi against both drugs (Campos et al. 2017; Mejia et al. 2012), and the prolonged treatment duration required for Chagas disease (Lascano et al. 2022; Viotti et al. 1994). The existence of a small subpopulation of T. cruzi (dormant parasite persisters in amastigote forms) able to survive when exposed to benznidazole is one of the reasons why benznidazole consistently fails to effectively kill all the T. cruzi parasites. This poses a significant challenge to effectively eradicate the disease using current treatment (Sánchez-Valdéz et al. 2018; De Rycker et al. 2023). As a result, it is an urgent need to develop novel and potent compounds to address these concerns related to current treatment.

In recent years, numerous studies have investigated the potential of APPs derived from animal sources, such as amphibians, insects, reptiles, and mammals, to combat T. cruzi parasites. The APD3 Antimicrobial Peptide Database reports approximately eleven APPs with antiparasitic effects against T. cruzi parasites (accessed on 10 January 2024). For example, three APPs: phylloseptin-O1 (APP ID: AP00759), phylloseptin-O2 (AP00760), and dermaseptin 01 (AP01389), isolated from the tree frog Phyllomedusa oreades, have shown significant antiparasitic activity against trypomastigotes of T. cruzi. Their respective half-maximal inhibitory concentration (IC₅₀) values were 5.1 µM (phylloseptin-O1) and 4.9 µM (phylloseptin-O2), as shown in Table 1 (Leite et al. 2005). Dermaseptin 01 was able to reduce trypomastigotes to non-detectable levels at a concentration of 16 µg/mL (Brand et al. 2002). Phylloseptin-O1 and phyllopseptin-O2 are short peptides with a length of around 20 amino acids, whereas dermaseptin 01 is a medium peptide with the length of 29 amino acids (Table 2). They belong to the UCLL class, suggesting a linear structure (Table 2). Besides, they are cationic with a net charge of + 4 (Table 2). However, their 3D structures remain unknown. Despite the lack of 3D structural information, their cationic linear structure allows them to directly kill the trypomastigotes by inducing membrane disruption without affecting host cells (Brand et al. 2002; Leite et al. 2005). Brand et al. (2002) also reported that two dermaseptin peptides: Dermaseptin-DI1 (AP00966) and Dermaseptin-DI2 (AP00958), isolated from Phyllomedusa distincta, were effective against trypomastigotes by reducing the number of trypomastigotes to non-detectable levels at concentration of 16 µg/mL after a 2-hour incubation, similar to the results shown by dermaseptin 01, as shown in Table 1. It is worth noting that the toxicity effects of both peptides were not evaluated in this study. Both peptides are medium peptides with lengths of 28 amino acids (Dermaseptin-DI2) and 33 amino acids (Dermaseptin-DI1). They are also cationic peptides with net charges of + 3 (Dermaseptin-DI2) and + 4 (Dermaseptin-DI1), and belong to the UCLL class due to their linear structure (Table 2). Both peptides were suggested to disrupt the negatively charged membrane of trypomastigotes in order to kill them, as the negatively charged membrane of trypomastigotes is a result of the presence of mucin and sialic acid (Leite et al. 2005). Mucins are known as surface glycoproteins that abundantly covers trypomastigotes and amastigotes, with O-linked oligosaccharides crucial in accepting sialic acid from the host (Giorgi and de Lederkremer 2011). T. cruzi parasite lack the ability to produce sialic acid themselves, so their trans-sialidase (TcTS) gains the sialic acid from host glycoconjugates, and transfers it to the parasites’ surface (dC-Rubin and Schenkman 2012; Fonseca et al. 2019). This process leads to the formation of negatively charged glycans, forming a protective coat for T. cruzi against proteases and human anti-α-galactosyl antibodies (Pereira-Chioccola et al. 2000). However, the positively charged APPs can effectively interact with the negatively charged membrane, and disrupt it, and ultimately kill T. cruzi trypomastigotes.

Table 1 Summary of the characteristics of APPs mentioned in APD3 database

Table 2 Summary of APPs mentioned in APD3 database, exhibiting antitrypanosomal activity against T. Cruzi

Numerous peptides have been isolated from amphibian sources, including temporin-SHd (from Sahara frog Pelophylax saharica; AP02118), figainin 1 (from the tree frog Boana raniceps; AP03248), and figainin 2 (also from the tree frog Boana raniceps; AP03190). These peptides have shown efficacy against T. cruzi epimastigotes, with IC₅₀ values of 16.8 µM (temporin-SHd), 15.9 µM (figainin 1) and 6.32 µM (figainin 2), as depicted in Table 1 (Abbassi et al. 2013; Santana et al. 2020a, b). They are classified as UCLL class, indicating their linear structure. They are also known as α-helix cationic peptides with net charges ranging from + 2 to + 5 (Table 2). Temporin-SHd possibly to interact directly with the parasite membrane, leading to membrane disruption by forming pores (Abbassi et al. 2013). However, the mechanisms of action of figainin 1 and figainin 2 remain unclear. Temporin-SHd and figainin 2 showed a low haemolytic effect against human erythrocytes with 50% haemolytic concentration, HC₅₀ value of 44 µM and 48.9 µM whereas figainin 1 showed high haemolytic effect against human erythrocytes with 50% lethal concentration, LC₅₀ = 10 µM, as shown in Table 1 (Abbassi et al. 2013; Santana et al. 2020a, b). This high haemolytic effect of figainin 1 may be attributed to its elevated hydrophobicity content and C-terminal amidation (Chen et al. 2007; Santana et al. 2020b). C-terminal amidation is known as a common post-translational modification exerted in AMPs (Xu and Lai 2015). Shyla et al. (2019) mentioned that peptides amidation not only increase antimicrobial efficacy but also causes an increase in haemolytic effects against human erythrocytes. In summary, temporin-SHd and figainin 2 show potential as promising candidates for the development of antiparasitic therapeutics. However, the significant haemolytic effect observed in figainin 1 poses a limitation for its therapeutic applications.

Apart from the peptides isolated from amphibians, the database also includes some of the peptides isolated from arachnids, reptiles, and insects, such as cupiennin 1a (AP00485) was isolated from arachnids Cupiennius salei, batroxicidin was isolated from snake Bothrops atrox and, melittin was isolated from honeybee Apis mellifera. Cupiennin 1a showed efficacy against T. cruzi amastigotes with an IC₅₀ value of 0.92 µM while batroxicidin showed an IC₅₀ value of 0.44 µM against trypomastigotes and an IC₅₀ value of 11.3 µM against epimastigotes of T. cruzi with a high selectivity (SI) value of 315, as shown in Table 1 (Kuhn-Nentwig et al. 2011; Mello et al. 2017). Notably, Mello et al. (2017) emphasized that batroxicidin (AP02423) significantly reduced the number of amastigotes around 400/100 LLC-MK2 cells when treated a concentration of 0.44 µM after 24 h, as shown in Table 1. The mechanisms of action of batroxicidin involves inducing necrosis to kill T. cruzi (Kuhn-Nentwig et al. 2011; Mello et al. 2017). The mechanism of action of cupiennin 1a against amastigotes of T. cruzi remains unclear. However, cupiennin 1a not only exhibited cytolytic effects against T. cruzi, but also on human erythrocytes, with half-maximal effective concentration, EC₅₀ value of 23 µM (Table 1). This cytolytic effect on human erythrocytes is due to the presence of sialic acid on the erythrocytes, resulting in the formation of negatively charged of human erythrocytes which are more susceptible to cupiennin 1a (Kuhn-Nentwig et al. 2011). Based on the database, Cupiennin 1a is classified as the UCLL class and consists of an α-helix structure with a net charge of + 8, suggesting their α-helical linear cationic features (Table 2). However, the 3D structure and bonding patterns of batroxicidin remain unknown, with the available information suggesting a net charge of + 15, indicating that it is a cationic peptide (Table 2).

In the study conducted by Adade et al. (2013), they investigated the effectiveness of melittin (AP00146), a linear cationic amphipathic peptide with a net charge of + 6 and consisting of 26 amino acids, that isolated from the honeybee Apis mellifera against T. cruzi, as shown in Table 1. Melittin showed a significant impact on the viability of all three forms of T. cruzi, with an IC₅₀ value of 2.44 µg/mL against epimastigote forms, 0.14 µg/mL against trypomastigote forms and a range of 0.15–0.22 µg/mL against amastigote forms (Table 2). The researchers also found that melittin multifaceted mechanisms of action, including inducing autophagic cell death in epimastigotes and amastigotes, and apoptotic action in trypomastigotes(Adade et al. 2013). However, it is important to note that melittin was found to be toxic to mammalian host cells at a concentration of 5 µg/mL. This concentration caused approximately 49% of LLC-MK2 mammalian cells to become non-viable after 24 h of treatment (Table 2).

Except for the APPs mentioned in the APD3 Antimicrobial Peptide Database for their efficacy against T. cruzi, there are also other AMPs not documented in the database have been reported to show antiparasitic effects against T. cruzi. For example, Pinto et al. (2013) highlighted the antiparasitic effects of AMPs found in the secretion of a tree frog, Phyllomedusa nordestina, which has shown in Table 3. These AMPs, specifically dermaseptin-1, dermaseptin-4, phylloseptins-7, and phylloseptins-8, demonstrated significant efficacy against T. cruzi. Among them, dermaseptin-4 is the most potent against trypomastigotes of T. cruzi (IC₅₀ = 0.25 µM), followed by Phylloseptin-7 (IC₅₀ = 0.34 µM), Phylloseptin-8 (IC₅₀ = 0.46 µM), and dermaseptin-1 (IC₅₀ = 0.68 µM), as summarized in Table 4. However, Phylloseptin-7 had the highest SI value of 101, while the other three peptides showed a lower SI value ranging from 20 to 40, indicating that phylloseptin-7 has more potential as a selective and promising antiparasitic candidate against trypomastigotes of T. cruzi. In comparison to the standard drug, benznidazole, these four peptides showed better activity and selectivity against T. cruzi (IC₅₀ value of benznidazole = 440.73 µM; CC₅₀ value = 469.93 µM; SI value = 1.07), suggesting their superior effectiveness in eliminating trypomastigotes of T. cruzi. This enhanced activity is attributed to their cationic and amphipathic properties. The cationic amphiphilic properties allowed them to bind to anionic phospholipids on the parasite membrane via electrostatic and hydrophobic interactions, which then allows them to permeabilize the parasitic membrane by forming pores (Lyu et al. 2023). This is a common advantage associated with AMPs. However, it is noticeable that this study only focused on the peptides’ effects against trypomastigotes of T. cruzi and did not evaluate effectiveness against amastigote and epimastigote forms. Therefore, future research should consider assessing these peptides against all forms of T. cruzi to provide more comprehensive understanding of their potential therapeutic applications.

Table 3 Summary of the characteristics of AMPs not mentioned in APD3 database

Table 4 Summary of AMPs not mentioned in APD3 database, exhibiting antitrypanosomal activity against T. Cruzi

In a separate study, Polybia-CP peptide and mastoparan peptide, both derived from the eusocial wasp Polybia paulista, were investigated their antiparasitic activities against T. cruzi. Polybia-CP peptide is a 12-residue α-helical, amphiphilic peptide that has been found to be highly effective against T. cruzi (Table 3). It showed an EC₅₀ of 9.38 µmol/L and 0.94 µmol/L against epimastigotes and trypomastigotes, respectively, and a high SI value of 106 against the trypomastigote form, as shown in Table 4 (Freire et al. 2020). Furthermore, at a concentration of 0.94 µmol/L, it was able to reduce amastigotes by 38% per 100 cells, which is considered more effective compared to the control drug, benznidazole (EC₅₀ of 218 µmol/L against epimastigotes and EC₅₀ of 282 µmol/L against trypomastigotes) (Freire et al. 2020; Wang et al. 2012). These findings suggest that Polybia-CP has the potential to be a selective and promising therapeutic agent against T. cruzi due to its low EC₅₀ and high SI value.

The mechanism of action of Polybia-CP differs from that of melittin, as it does not involve pore formation in the cell membrane. Instead, it induces apoptosis, triggering cell death in T. cruzi. Another peptide, mastoparan, which is isolated from Polybia paulista, has also shown antiparasitic effects against all three forms of T. cruzi. Its IC₅₀ values are 61.4 µM against epimastigote forms, 5.31 µM against trypomastigote forms, and a reduction of approximately 50% in amastigotes at a concentration of 10.62 µM, as summarized in Table 4 (Vinhote et al. 2017). Unlike melittin and Polybia-CP, the mechanism of action of mastoparan involves the inhibition of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in the T. cruzi parasite (Vinhote et al. 2017). This action is supported by the observed increase in levels of reactive oxygen species (ROS) and mitochondrial disruption (Vinhote et al. 2017).

Furthermore, researchers have evaluated the effectiveness of several AMPs derived from arachnids and insects. Two specific AMPs, VmCT1 isolated from the scorpion Vaejovis mexicanus and M-PONTX-Dq3a from the giant ant Dinoponera quadricep, have been studied for their activity against T. cruzi, as summarized in Table 3 (Lima et al. 2018; Pedron et al. 2020). Notably, both peptides have demonstrated efficacy against all three forms of T. cruzi by inducing necrosis as their mechanism of action (Table 4). These two peptides have a significantly higher SI value, ranging from 50 to 80, compared to benznidazole, which has a SI value of approximately 2. This significant increase in selectivity suggests that VmCT1 and M-PONTX-Dq3a have the potential to specifically target and eliminate the parasites, without harming the mammalian host cells.

Additionally, a cathelicidin-related peptide called crotalicidin (Ctn) has been isolated from the rattlesnake Crotalus durissus terrificus (Bandeira et al. 2018). Ctn is a 34-residue cationic amphipathic peptide with an α-helical structure. Notably, this peptide has shown strong antiparasitic effects against all three forms of T. cruzi, including a benznidazole-resistant strain. Ctn has a low EC₅₀ of 4.47 µM against epimastigotes, 0.22 µM against trypomastigotes, and reduce approximately 200 amastigotes per 100 cells after 24 h of treatment and approximately 900 amastigotes per 100 cells after 48 h of treatment at 0.22 µM, as shown in Table 4 (Bandeira et al. 2018). It is noteworthy that the antiparasitic effects of Ctn are more pronounced against trypomastigotes and amastigotes than epimastigotes. This variation in efficacy is due to the presence of negatively charged molecules, such as sialic acid and different type of mucin-like glycoproteins, extensively coat the membrane of trypomastigotes and amastigotes (Buscaglia et al. 2006). The O-linked glycans within mucin facilitates the acquisition of sialic acid from the host, especially considering that T. cruzi parasite lack the ability to produce sialic acid independently. Hence, the parasite’s TcTS extracts sialic acid from host glycoconjugates through a trans-glycosylation reaction, thereby transferring it into the glycans on the parasite surface. This process results in the formation of negatively charged glycans, providing a protection to T. cruzi from proteases and human anti-α-galactosyl antibodies (Freire-de-Lima et al. 2015; Giorgi and de Lederkremer 2011). Therefore, these negatively charged glycans are present on the surface of both forms, forming a negatively charged membrane that allows crotalicidin to selectively target and compromise their membrane integrity (Pech-Canul et al. 2017; Sabiá Júnior et al. 2019). Additionally, Ctn has been found to have a significantly higher SI value with a value of 203.2, surpassing that of benznidazole (SI of 2.18). This impressive SI value not only demonstrates the effectiveness of Ctn, but also its safety as a treatment against trypomastigotes when compared to benznidazole. In fact, study has revealed that Ctn eliminates the epimastigote form of T. cruzi through necrosis, as confirmed by flow cytometry experiments (Bandeira et al. 2018). These findings highlight the potential of Ctn as a promising therapeutic agent against T. cruzi, due to its effective antiparasitic effect and superior selectivity compared to benznidazole.

In addition, AMPs isolated from marine invertebrates have been shown to effectively kill T. cruzi parasites. One notable example is the peptide fragment Hmc364-382, which is derived from the hemocyanin of the shrimp species Penaeus monodon (Table 3). This 19-amino acid residue peptide has an α-helical structure and a net charge of + 1. It has been reported that this peptide fragment exhibits significant antiparasitic effects against T. cruzi due to its structural characteristics and its cationic properties (Monteiro et al. 2020). These properties allow it to disrupt membrane integrity and form pores, ultimately leading to cell death through necrosis. In their comprehensive study, Monteiro and colleagues tested the peptide fragment was tested on all three forms of T. cruzi, demonstrating significant antiparasitic effects with an IC₅₀ of 3.62 µM against trypomastigote forms and 4.79 µM against epimastigote forms. Additionally, at a concentration of 3.62 µM, it caused a reduction of approximately 61% of amastigotes per 100 host cells (LLC-MK2 cells) (Table 4). The peptide also has a high SI value (50), indicating its potential for further development and investigations.

In summary, the search for natural APPs against T. cruzi has yielded several promising candidates. The majority of identified APPs, derived from amphibian sources, have demonstrated antiparasitic activities against T. cruzi trypomastigotes, which is the infective form of the parasite. One notable peptide, temporin-SHd, isolated from an amphibian source, has shown efficacy against T. cruzi epimastigotes, which is the replicative form of the parasite. Besides, melittin, a peptide isolated from an insect source (Apis mellifera), has shown efficacy against all three forms of T. cruzi, indicating its potential for development as an antiparasitic therapeutic. However, melittin has been found to have toxic effects on LLC-MK2 mammalian cells, hindering its potential as an antiparasitic agent. In addition, six identified AMPs, not mentioned in the APD3 database, such as Polybia-CP, mastoparan, VmCT1, M-PONTX-Dq3a, crotalicidin, and Hmc364-382, have shown potential in reducing intracellular amastigotes and effectively combating all three developmental forms of T. cruzi. When compared to benznidazole, peptides such as dermaseptin-1, dermaseptin-4, phylloseptins-7, phylloseptins-8, and Polybia-CP have shown greater efficacy against T. cruzi with significantly lower IC₅₀ values. Notably, the Polybia-CP peptide was particularly effective against amastigotes, which are known to have low susceptibility to benznidazole. Additionally, the peptides VmCT1 and M-PONTX-Dq3a peptides showed higher SI values compared to benznidazole, indicating that a more targeted action against T. cruzi while minimizing toxicity against the mammalian host cells. Another promising peptide, crotalicidin showed antiparasitic activity against all three forms of T. cruzi, including a benznidazole-resistance strain. Crotalicidin peptide kills T. cruzi through necrosis, a mechanism different from the mechanism of action of benznidazole, which causes DNA damage through metabolites produced by nitroreductase (NTR). Therefore, these findings highlight that these peptides not only exhibit better efficacy against T. cruzi, but also have more favourable safety profiles compared to current treatments, suggesting their potential for advancing new treatment strategies for Chagas disease.

Human African Trypanosomiasis (HAT)

Human African trypanosomiasis (HAT), also known as sleeping sickness, is a parasitic infection caused by the T. brucei parasite and transmitted by tsetse flies. The infection is mainly localised in rural areas of Africa (Kennedy and Rodgers 2019). The T. brucei parasite has two subspecies, each responsible for distinct diseases: T. b. gambiense, which is typically endemic in western and central Africa and causes the West African trypanosomiasis, and T. b. rhodesiense primarily exists in eastern and southern Africa, leading to the East African trypanosomiasis (Table 5; Lun et al. 2010). At least 85% of HAT cases is attributed to T. b. gambiense and the remaining to T. b. rhodesiense (Venturelli 2022). Both types of the disease are primarily transmitted by tsetse flies of the Glossina genus, which carry Trypanosoma parasites and transmit them to reservoirs (Büscher et al. 2017). T. b. gambiense HAT is considered an anthroponosis, with humans being the main reservoir in the transmission cycle, as shown in Table 5 (Papagni et al. 2023). This means that the infected humans or mammalians with T. b. gambiense can transmit the disease to other humans through the bite of tsetse flies. On the other hand, T. b. rhodesiense HAT is a zoonosis, with animals such as livestock (cattle) and wild animals being the main reservoirs. Therefore, the transmission routes of T. b. rhodesiense HAT are mostly from infected animals to other animals through tsetse flies, from infected animals transmitted to humans through tsetse flies and from infected human transmitted to other human through tsetse flies (Table 5; Papagni et al. 2023). People who work with livestock, such as cattle, goats, and sheep, are at higher risk of contracting the disease through contact with infected animals (Ruiz et al. 2015). Additionally, HAT can also be transmitted through other routes, such as congenital transmission from an infected mother to baby, blood transfusion, accidental infection in laboratories, and organ transplantation from an infected donor (Sina et al. 1979; Herwaldt et al. 2001; Martín-Dávila et al. 2008; Gaillot et al. 2017). Besides, there have also been cases of T. b. gambiense being transmitted through sexual contact (Rocha et al. 2004).

Table 5 Comparison of T. b. gambiense and T. b. rhodesiense

According to the WHO, T. b. gambiense HAT is typically affected in 24 countries located at west and central Africa, which the reported incidence cases initially increased by 66%, rising from 12 756 to 37 385 between 1990 and 1998 (accessed 25 November 2023). Among these 24 countries, Congo is the most affected country, which attributed to 60% of total reported cases. While T. b. rhodesiense HAT is typically affected 13 countries in eastern and southern Africa, which approximately 1933 cases reported in 1990, as reported by WHO. However, based on WHO, there was a significantly decreased of 98% subsequently, dropping from 37 385 to less than 700 between 1998 and 2022. At the same time, the incidence cases of T. b. rhodesiense HAT in Africa drastically decreased from 1933 to 38 between 1990 and 2022.

The life cycle of HAT, as shown in Fig. 10, begins when an infected tsetse fly bites a mammalian host, injecting metacyclic trypomastigotes into the skin. These metacyclic trypomastigotes then transform into bloodstream trypomastigotes, which travel throughout the body. In the spinal fluid and lymph, they reproduce asexually through binary fission. Simultaneously, the tsetse fly becomes infected when it feeds on the blood of an infected host, and the bloodstream trypomastigotes transform into procyclic trypomastigotes in the fly’s midgut. After reproducing through binary fission, the procyclic trypomastigotes then transform into epimastigotes and migrate to the salivary glands to undergo reproduction. Finally, they transform into metacyclic trypomastigotes before being transmitted to another mammalian host (Quintana et al. 2021).

Fig. 10

The lifecycle T. brucei gambiense (T. b. gambiense) & T. brucei rhodesiense (T. b. rhodesiense) parasite in human stage and sandfly stage. Created with BioRender.com

The HAT infection progresses in two stages for both types of the disease: the initial haemolymphatic stage (first stage) and the meningoencephalitis stage (second stage) (Deeks 2019). In the first stage, symptoms include headache, arthralgia, fatigue, and pruritus are observed for both types of parasitic diseases. However, the second stage only occurs when the trypanosomes invade the central nervous system (CNS), causing neuropsychiatric manifestations (Bottieau and Clerinx 2019). Despite sharing a common mode of transmission and involving two stages of infection, HAT caused by T. b. rhodesiense and T. b. gambiense exhibit different pathological conditions. For example, T. b. rhodesiense HAT is an acute disease, characterized by the first stage symptoms, which typically developing 1–3 weeks after infection and progressing to the second stage within a few weeks, as shown in Table 5 (Kuepfer et al. 2011). In contrast, T. b. gambiense HAT presents as a chronic infection, with either asymptomatic or intermittent symptoms in the first month after infection. Second-stage symptoms emerge after several months or even a year, indicating CNS involvement and progression to meningoencephalitis (second stage), as shown in Table 5 (Pays and Nolan 2021).

The treatment options for HAT are tailored based on the type and stage of the disease. There are several treatment options available, including pentamidine, suramin, melarsoprol, eflornithine, and fexinidazole (Kennedy 2019). Pentamidine is typically used for treating the first stage of T. b. gambiense HAT and is generally well-tolerated. However, it may cause certain reported side effects such as heart problems, hypotension, nephrotoxicity, hypoglycemia, pancreatitis and hepatic dysfunction (Babokhov et al. 2013). On the other hand, suramin is prescribed for the first stage of T. b. rhodesiense HAT and may cause common side effects such as nephrotoxicity and drug rash, as well as rare side effects such as hypersensitivity reactions and peripheral neuropathy (von der Ahe et al. 2018). Melarsoprol is used for treating the second stage of T. b. rhodesiense HAT, but it has a life-threatening side effect known as encephalopathic syndrome (Seixas et al. 2020). Eflornithine, while equally effective, is less toxic than melarsoprol and is used to treat the second stage of T. b. gambiense HAT. Eflornithine is often combined with nifurtimox as a combination therapy, which has been shown to be more effective and less toxic than eflornithine monotherapy (Kansiime et al. 2018). Furthermore, fexinidazole is currently the only drug known to be effective against both the first and early second stages of T. b. gambiense (Lindner et al. 2020). Besides, it has found to be effective against acute mouse model infected with T. b. rhodesiense HAT (Bernhard et al. 2022). Then, fexinidazole was proven to be safe and effective against both stages of T. b. rhodesiense HAT in the clinical trial sponsored by Drugs for Neglected Diseases initiative (DNDi) (accessed 18 May 2024). In fact, the European Medicines Agency (EMA) adopted fexinidazole as the first oral treatment for the acute form of T. b. rhodesiense HAT in 2023. Apart from that, according to a report by DNDi, fexinidazole has also been involved in a phase II clinical trial to test its antiparasitic activity against T. cruzi (De Rycker et al. 2023). However, this project was halted due to the relapse of T. cruzi infection in patients after a 10-weeks post-treatment follow-up (Pinazo et al. 2024). Fexinidazole has some common side effects such as headache, vomiting, insomnia, and nausea, as well as occasional serious side effects such as breathing difficulty, chest pain, hives, and facial swelling (Kumesu et al. 2022; Mesu et al. 2021).

Currently, due to the availability of commercial drugs and preventive measures such as serological screening tests for detecting the T. b. gambiense parasite, insecticide-impregnated screens for vector control, and safe water supply, there has been a significant reduction in the incidence cases, which less than 800 reported cases of both T. b. gambiense HAT and less than 40 reported cases of T. b. rhodesiense HAT in 2022, as reported by WHO (accessed 27 November 2023). This decreased in cases of T. b. gambiense HAT has accelerated the elimination of HAT as a public health problem, which is the goal set by the WHO for 2020.

WHO has set new goals for 2030, which include the interruption of transmission for HAT (accessed 27 November 2023). This involves reducing the number of reported cases of T. b. gambiense HAT to less than 500 and T. b. rhodesiense HAT to less than 1 case per 10,000 people per year (Compaoré et al. 2022). However, it is important to note that some treatments such as melarsoprol, can have life-threatening side effects. Besides, there are ongoing problems with commercial drugs, such as resistant strains of T. brucei have been identified, demonstrating resistance to pentamidine, suramin, melarsoprol, eflornithine and nifurtimox (Kasozi et al. 2022; Vincent et al. 2010). In addition, there is currently no drug that effectively targets both stages of T. b rhodesiense HAT. Hence, the current drugs are not sufficient to address the changing epidemiological situation. In response to these challenges and the 2030 target achievements, studies have been conducted to evaluate the efficacy of APPs against both T. b rhodesiense and T. b gambiense, with the aim of uncovering the antiparasitic properties of naturally-derived APPs against T. brucei.

One such peptide, bacteriocin AS-48 (AP00929) a circular peptide was found in the bacterium Enterococcus faecalis, exhibiting antiparasitic effect against both forms of T. brucei (Table 6). This peptide is classified as α-helix cationic peptide with net charge of + 6 and is recognized as a medium-sized peptide with a length of 70 amino acid residues. It falls into the UCBB class in the database, indicating its status as a circular long peptide (Table 6; Maqueda et al. 2004). A recent study by Martínez-García et al. (2018) showed its efficacy, with an EC₅₀ value of 0.0017 µM against the bloodstream form (BSF) of T. b. rhodesiense HAT and an EC₅₀ value of 0.0026 µM against the BSF of T. b. gambiense HAT (Table 7). This highlights its potential as a treatment for both forms of the disease. Additionally, the peptide has been found to be non-toxic to human cell lines at a concentration of 12.5 µM (Table 7). The mechanism of action of bacteriocin AS-48 involves binding of the peptide to the variant surface glycoprotein (VSG) on the membrane of T. brucei parasites and entering the cell through clathrin-mediated endocytosis, ultimately leading to autophagic cell death (Martínez-García et al. 2018). Apart from peptides isolated from bacterial species, cupiennin 1a (AP00485) has also demonstrated significant parasitic activity, with an IC₅₀ value ranging from 0.055 to 0.061 µM against the BSF of T. b. rhodesiense (Table 7; Kuhn-Nentwig et al. 2011). This peptide not only demonstrated significant effect against T. cruzi, but also against T. b. rhodesiense, indicating that its potential as an antiparasitic candidate for both parasitic diseases. The cationic nature of this peptide allows for cytolytic activity to occur on the negatively charged membrane of the parasite, ultimately eliminating the BSF of T. b. rhodesiense. Additionally, temporin-SHd (AP02118), secreted from the frog skin of Pelophylax saharica, has also shown antiparasitic activity against T. brucei. Based on Abbassi et al. (2013), temporin-SHd not only demonstrated effectiveness against T. cruzi but also against the procyclic form of T. b. gambiense with an IC₅₀ value of 21.8 µM (Table 7). The mechanism of action of temporin-SHd likely involves the pore formation on the parasite membrane to kill T. b. gambiense.

Table 6 Summary of the characteristics of APPs mentioned in APD3 database

Table 7 Summary of APPs mentioned in APD3 database, exhibiting antitrypanosomal activity against T. b. gambiense & T. b. rhodesiense

Other than that, peptides derived from the frog Silurana tropicalis, such as CPF-St4 (AP02283), CPF-St5 (AP02284), CPF-St6 (AP02285), CPF-St7 (AP02286), Magainin-St1 (AP02287), XPF-St4 (AP02288), XPF-St5 (AP02289), XPF-St1(AP02290), XPF-St6 (AP02291), XPF-St7 (AP02292), XPF-St8 (AP02293), PFQa-St2 (AP02294) and PGLa-St2 (AP02295) peptides, have shown efficacy against T. brucei brucei (T. b. brucei) (Roelants et al. 2013). T. b. brucei is a subspecies of T. brucei that exclusively infects non-human vertebrates (Balmer et al. 2011). In Roelants’s study, the effectiveness of these peptides against T. b. brucei parasites was determined by the minimum concentration needed to kill 95% of the parasites within 30 min (Roelants et al. 2013). Among these, CPF-St5 showed the highest potency in killing T. b. brucei with an LC₉₅ value ranging from 1 to 2 µM (Table 6). The antiparasitic evaluations of the remaining peptides are shown in Table 6. They are cationic medium-sized peptides with a net charge ranging from + 3 to + 5 and the lengths ranging from 16 to 27, as shown in Table 7 (Roelants et al. 2013). In the database, they are classified as the UCLL class, indicating their linear structure (Table 7). Therefore, the predicted mechanism of action of these peptides involves the interaction between their cationic side and the negatively charged of phospholipid heads on membrane, whereas the hydrophobic sides of the peptides infiltrate the intermembrane region (Brogden 2005). This process potentially promoting membrane pore formation, and ultimately resulting in cell lysis. However, this study focused solely on testing the effectiveness of these 13 peptides against T. b. brucei and did not assess their activities against T. b. rhodesiense and T. b. gambiense, which are human pathogenic subspecies of T. brucei. Therefore, further research could evaluate their efficacy against these two human-infective forms, considering their demonstrated antiparasitic activities against T. b. brucei.

Apart from the APPs mentioned in APD3 Antimicrobial Peptide Database that showed antiparasitic activities against T. brucei, there are certain AMPs not documented in the database have been reported to exhibit antiparasitic effects against T. brucei. Although the majority of AMPs are considered ribosomally templated peptides, non-ribosomally templated peptides (NRPs) has also demonstrated its potential as antitrypanosomal agents against T. b. rhodesiense. One example of an NRP-AMP is photoditritide, a cyclic peptide derived from the entomopathogenic bacterium Photorhabdus temperata Meg1 strain (Table 8). This bacterium is commonly associated with the Heterorhabditis nematode and has a symbiotic relationship with them (Thanwisai et al. 2012). Photoditritide is composed of two D-form homoarginine amino acid residues and is produced by a non-ribosomal peptide synthetase (NRPS). A study conducted by Zhao et al. (2019) has shown that photoditritide possesses mediocre antitrypanosomal effects against the BSF of T. b. rhodesiense HAT, with an IC₅₀ value of 13 µM (Table 9). Although this IC₅₀ value is less potent compared to the control drug melarsoprol, which has an IC₅₀ value of 0.005 µM, photoditritide did not show any cytotoxicity effect against mammalian L6 cells in the study, distinguishing it from the highly toxic melarsoprol (Zhao et al. 2019). Therefore, further research could focus on the potential of using photoditritide as a template for synthetic modifications to enhance its antiparasitic activity against T. b. rhodesiense while maintaining low cytotoxicity against mammalian cells. Additional research is also needed to determine the mechanism of action of this peptide, which has yet to be determined.

Table 8 Summary of the characteristics of APPs not mentioned in APD3 database

Table 9 Summary of APPs not mentioned in APD3 database, exhibiting antitrypanosomal activity against T. b. gambiense & T. b. rhodesiense

Several NRP-AMPs that have been found to possess antitrypanosomal activity against T. b. rhodesiense, including szentiamide derived from the bacterium Xenorhabdus szentirmaii, and xenobactin derived from bacteria of the Xenorhabdus genus (Table 8). These bacteria are commonly associated with nematodes of the Steinernema genus and have a symbiotic relationship with them (Ohlendorf et al. 2011). Both szentiamide and xenobactin, are categorized as depsipeptides, and have shown moderate antitrypanosomal effects against T. b. rhodesiense. However, szentiamide, has been found to be more effective, with an IC₅₀ value of 10 µg/mL, compared to xenobactin, which has an IC₅₀ value of 31.64 µg/mL which showed in Table 9 (Grundmann et al. 2013; Nollmann et al. 2012). While xenobactin requires a higher concentration to exhibit antiparasitic activity against T. b. rhodesiense, it did not show any cytotoxicity effect towards mammalian L6 cells (Grundmann et al. 2013). Conversely, while szentiamide showed enhanced antitrypanosomal effects, it also demonstrated mild toxicity to mammalian L6 cells with a IC₅₀ of 57.4 µg/mL (Nollmann et al. 2012). However, the mechanisms of action for both peptides are still unclear, and further research is needed to determine their antiparasitic mechanism actions.

Furthermore, some peptide antibiotics are derived from fungi, such as leucinostatin A and leucinostatin B (both isolated from Paecilomyces lilacinus), alamethicin I (isolated from Trichoderma viride), and tsushimycin (isolated from Streptomyces) (Ishiyama et al. 2009). These peptides are unique from typical AMPs because they contain uncommon amino acids such as 2-aminoisobutyric acid (Aib), 4-methylproline (MePro), 2,4-diaminobutyric acid (Dab), β-alanine, 2-amino-6-hydroxy-4-methyl-8-oxodecanoic acid (AHMOD), threo-β-hydroxyleucine (HyLeu), pipecolinic acid (Pip), and β-methylaspartate (Map) (Abe et al. 2018; Bunkóczi et al. 2005; Harrington 2011). According to Ishiyama et al. (2009), leucinostatin A and leucinostatin B are well-characterized lipophilic peptide antibiotics that have been shown to have an antitrypanosomal effect against the bloodstream form of T. b. rhodesiense strain STIB900 with an IC₅₀ value of 3.4 ng/mL (leucinostatin A) and 4.4 ng/mL (leucinostatin B), respectively (Table 9). Notably, leucinostatins A and B exhibited a superior antitrypanosomal effect compared to the control drug suramin, with an IC₅₀ of 52 ng/mL. This suggests their potential as promising candidates for peptide-based antitrypanosomal therapeutics. Although the mechanisms of action for both peptides are still unclear, it has been hypothesized that they may function as voltage-gated channels on membrane phospholipids, thereby disrupting the homeostasis of the parasite (Ishiyama et al. 2009).

Alamethicin I also exhibited antitrypanosomal activity against the bloodstream form of T. b. rhodesiense strain STIB900, with an IC₅₀ value of 380 ng/mL, as shown in Table 9 (Ishiyama et al. 2009). In comparison, tsushimycin is a cyclic dodecamer lipopeptide derived from Streptomyces and demonstrated an IC₅₀ of 2490 ng/mL (Tables 8 and 9). It has been proposed that the mechanism of action for alamethicin I involves the formation of ionophores. This peptide orients itself perpendicularly to the lipid membrane. Upon contact with the membrane, the peptide oligomerizes, and develop cylindrical pores to allow ions and water to pass through (Ishiyama et al. 2009). The presence of α-aminoisobutyric acid (Aib) in alamethicin I contributes to the formation of ionophores, affecting its helical structure and leading to the creation of voltage-gated channels on the membrane(Mathew and Balaram 1983; Rahaman and Lazaridis 2014). In contrast, the mechanism of action for tsushimycin involves the suppression of dolichol-phosphate sugar (glucose and mannose) complex formation (Elbein 1981). These complexes play an important role in providing sugar moieties to glycosylphosphatidylinositol (GPI) anchors. GPI-anchors, such as VSG, typically act as a protective coat for T. brucei parasites, which are vital for their survival (Lillico et al. 2003). Overall, the suggested mechanisms of action for both peptides revolve around their interactions with trypanosome membranes.

In summary, the antiparasitic activities of APPs against T. b. gambiense and T. b. rhodesiense suggest that bacteriocin AS-48 has significant potential as an antitrypanosomal therapeutics. It has shown superior efficacy against the bloodstream form of both species (T. b. gambiense and T. b. rhodesiense) without causing cytotoxicity against mammalian cells. Its EC₅₀ value is lower than that of fexinidazole (0.7 to 3.3 µM), which is currently used as a treatment for HAT, indicating that bacteriocin AS-48 is more effective against T. brucei (Kaiser et al. 2011). Besides, bacteriocin AS-48 works by binding to VSG and inducing clathrin-mediated endocytosis, leading to autophagic cell death. In contrast, fexinidazole depends on electron reductions activated by NTR, which can potentially develop resistance to fexinidazole (Wyllie et al. 2016). Given its superior efficacy against two forms of HAT and its distinct mechanism of action, bacteriocin AS-48 could be a competitive alternative to current treatments for HAT. Additionally, Temporin-SHd and cupiennin 1a have also shown antiparasitic effects against T. cruzi and T. brucei parasites, suggesting their potential as treatments for both Chagas disease and HAT, where fexinidazole has limitations. Besides, it is noticeable that peptides such as CPF-St4, CPF-St5, CPF-St6, CPF-St7, Magainin-St1, XPF-St4, XPF-St5, XPF-St1, XPF-St6, XPF-St7, XPF-St8, PFQa-St2 and PGLa-St2 peptides, have shown efficacy against bloodstream form of T. b. brucei, a subspecies of T. brucei that exclusively infects non-human vertebrates. In addition, NRP-AMPs include photoditritide, szentiamide and xenobactin, have been effective against BSF of T. b. rhodesiense. Also, fungi-derived peptides such as leucinostatin A, leucinostatin B, alamethicin I as well as tsushimycin exhibited effectiveness against T. b. rhodesiense bloodstream form via mechanism include forming voltage-gated channels and ionophores on membrane phospholipids or suppressing dolichol-phosphate-sugar complexes formed. However, it is important to note that most evaluation of peptides has focused on the vitro study. To fully elucidate the potential of these peptides as antitrypanosomal therapeutics, further in vivo research is necessary.

Leishmaniasis

Apart from Chagas disease and human African trypanosomiasis, another neglected tropical disease caused by protozoan parasites is leishmaniasis. This disease is transmitted when the female phlebotomine sand flies bite an infected person, becoming infected with the Leishmania parasite. The infected sand flies then transmit the parasite to a healthy person when they bite them (Bettaieb et al. 2020). The main vector for this transmission are female phlebotomine sand flies from the Phlebotomus and Lutzomyia genera (Dostálová and Volf 2012). Some examples of pathogenic leishmania parasites that affect humans include L. donovani, L. chagasi, L. infantum, L. amazonensis, L. tropica, L. major, L. mexicana, L. pifanoi, L. tarentolae, and L. panamensis (Alemayehu and Alemayehu 2017; Blackwell et al. 2020; Steverding 2017). Leishmaniasis can manifest in three main forms: cutaneous leishmaniasis, mucocutaneous leishmaniasis, and visceral leishmaniasis (Torres-Guerrero et al. 2017). The most common form is cutaneous leishmaniasis, which is primarily caused by L. tropica, L. major, L. donovani, L. aethiopica, L. mexicana, L. amazonensis, L. venezuelensis, L. [Viannia] brazilliensis, L. [Viannia] guyanensis, L. [Viannia] panamensis and L. donovani chagasi and results in skin sores, such as ulcers. Mucocutaneous leishmaniasis is characterized by the destruction of mucous membranes and symptoms such as odynophagia, oral and pharyngeal lesions, due to the spread of parasites from the skin sores to the naso-oropharyngeal mucosa (Cobo et al. 2016; Ruas et al. 2014). This form is usually caused by L aethiopica, L. [Viannia] brazilliensis, L. [Viannia] panamensis, and L. [Viannia] guyanensis. In contrast, visceral leishmaniasis, also known as kala-azar, is characterized by symptoms such as hepatomegaly, splenomegaly, intermittent fever, anemia, and weight loss (Bern et al. 2005). It is considered the most severe form and is typically caused by L. infantum and L. donovani. If left untreated, it can continue to affect internal organs such as the liver, spleen, and bone marrow, ultimately resulting in the death of the patient (Zijlstra 2021). According to the WHO, there are approximately 1 million new cases of cutaneous leishmaniasis reported worldwide each year, underscoring its significant global spread (accessed 30 November 2023). Visceral leishmaniasis is particularly concerning due to its high fatality rate and symptoms such as irregular bouts of fever, anemia, and an enlargement of the spleen and liver. It is estimated that 50 to 90 thousand new cases of visceral leishmaniasis occurred each year in South America, East Africa, South-East Asia, and the Mediterranean region, including countries such as Iraq, Somalia, Yemen and Sudan, making it one of the top parasitic diseases with high mortality and outbreak potential in the world (Bi et al. 2018).

Based on the GBD study, the prevalence of cutaneous and mucocutaneous leishmaniasis has decreased in most countries from 1990 to 2019, with notable exceptions in certain countries such as the Syrian Arab Republic and Morocco, located in the Middle East and North Africa (accessed on 30 November 2023). In Syrian, the prevalence of cutaneous and mucocutaneous leishmaniasis increased from 133.64 cases per 100 000 in 1990 to 1519.33 cases per 100 000 in 2019. Similarly, in Morocco, the prevalence of cases increased from 76.58 cases per 100 000 in 1990 to about 208.19 prevalent cases per 100 000 in 2019, as reported by the GBD study (accessed on 30 November 2023). Another notable example reported by GBD study is Afghanistan, which is known as an endemic region for cutaneous leishmaniasis, the prevalence cases have decreased from 8414.24 cases per 100 000 to 7145.8 cases per 100 000 between 1990 and 2019. While the number cases of cutaneous leishmaniasis in Afghanistan has seem to have a minor decrease every year, the country still experiences approximately 400 cases per 100 000 every year, as reported by the GBD study. This suggests that leishmaniasis continues to pose a significant public health problem in Afghanistan.

The GBD study has analyzed the prevalence of visceral leishmaniasis in endemic countries, including South America, Africa, South-East Asia, and the Middle East. Overall, there has been a decrease in prevalent cases between 1990 and 2019 in most endemic countries. However, there are notable exceptions such as Brazil, Paraguay, Bangladesh, India, South Sudan, and Iraq, which have experienced the re-emergence of visceral leishmaniasis in specific years, as documented in the GBD study. Despite a consistent annual decrease in prevalence in Africa, it remains the most affected country, with an average of 50 new cases per 100,000 reported annually. This underscores the persistent challenge of eliminating visceral leishmaniasis in endemic countries such as, Africa, despite the implementation of vector control measures (such as indoor residual spraying), rK39 rapid diagnostic tests (RDTs), and treatment efforts (Alvar et al. 2021).

The life cycle of Leishmania begins when an infected female sand fly consumes a blood meal and injects promastigotes into the host’s skin, as shown in Fig. 11. These promastigotes are then engulfed by macrophages or the mononuclear phagocytic system, causing them to transform into amastigotes. These amastigotes reproduce through binary fission, persist within the cells, and subsequently infect other cells. During the sand fly stage, when the sand fly feeds on the blood of an infected host, they become infected by ingesting amastigotes within macrophages. These amastigotes then transform into promastigotes, multiply within the fly’s gut, and migrate to the proboscis (Damianou et al. 2020; Sunter and Gull 2017; Teixeira et al. 2013). Currently, there are several drugs available for treating cutaneous leishmaniasis (CL) and visceral leishmaniasis (VL), such as antimonials (glucantime, sodium stibogluconate), miltefosine, amphotericin B, paromomycin, and pentamidine (Singh et al. 2016). While these drugs have shown excellent efficacy against CL and VL, they have also been found to be toxic to mammalian cells with prolonged use which can cause a serious renal toxicity, cardiotoxicity and even death (Vikrant et al. 2015). Furthermore, some strains of Leishmania parasites, such as L. donovani, have developed resistance to sodium stibogluconate, miltefosine, and amphotericin-B (Garza-Tovar et al. 2020; Jain and Jain 2018; Moore and Lockwood 2010). Some researchers have found evidence of persister forms in Leishmania spp., which presents a significant challenge for current treatment methods in eradicating Leishmaniasis (Barrett et al. 2019; De Rycker et al. 2023). There are currently several promising chemical compounds in Phase I and II clinical trials for treating visceral leishmaniasis. These include DNDI-6899 (formerly GSK899/DDD853651; CRK12 inhibitor), GSK245 (formerly GSK3494245/DDD01305143), and LXE408, developed by GSK, DNDi, and Novartis, respectively. However, there is a lack of recent drug discovery efforts targeting cutaneous leishmaniasis and mucocutaneous leishmaniasis. Therefore, it is an urgent need to discover new compounds that can potentially serve as therapeutics for these forms of leishmaniasis. This is particularly crucial in addressing the issues associated with the currently available drugs, such as serious renal toxicity and the emergence of persister cells and drug resistance.

Fig. 11

The lifecycle Leishmania parasite in human stage and sandfly stage. Created with BioRender.com

The APD3 database has highlighted several APPs derived from insects, amphibians and mammals that have demonstrated promising antileishmanial effects against specific species of Leishmania, including L. major, L. mexicana, and L. amazonensis, which are responsible for causing cutaneous leishmaniasis (accessed on 10 January 2024). For example, melittin (AP00146) demonstrated an EC₅₀ value of 74.01 µg/mL against promastigotes. Decoralin (AP00723), a peptide from the Eumenes-like potter wasp Oreumenes decorates, demonstrated an IC₅₀ value of 72 µM against promastigotes. PduDef (AP01364), a defensin peptide from the sand fly Phlebotomus duboscqi, an insect vector of L. major, demonstrated an IC₅₀ value of 68–85 µM against promastigotes, as shown in Tables 10 and 11 (Boulanger et al. 2004; Konno et al. 2007; Pérez-Cordero et al. 2011). Both melittin and decoralin are grouped into the UCLL class, and have a helix structure with net charges of + 6 and + 2, respectively (Table 10). This structural classification highlights their high affinity towards the anionic membrane of the parasite, contributing to pore formation on the membrane. In contrast, PduDef peptide belongs to the UCSS class due to the presence of six cysteine residues connected by three disulfide bridges (Table 10). Based on Boulanger et al. (2004), PduDef peptide induces an immune response in the sand fly midgut, eliminating promastigotes upon detection of phosphoglycans and lipophosphoglycan (LPG) on the promastigotes’ surface. Although both melittin and PduDef peptides have shown antileishmanial activity against L. major species, melittin has lower specificity, and may potentially targeting normal mammalian cells. Furthermore, the cytotoxic effect of PduDef on mammalian host cells has not been tested, emphasizing the need for further investigations in this regard.

Table 10 Summary of the characteristics of APPs mentioned in APD3 database

Table 11 Summary of APPs mentioned in APD3 database, exhibiting antitrypanosomal activity against leishmania

The dermaseptin peptide family, including dermaseptin-S1 (AP00157), dermaseptin DPh-1 (AP00763), and dermaseptin 01 (dermaseptin-O1; AP01389) derived from amphibians, was also found to be effective against both L. major and L. amazonensis. Dermaseptin-S1, isolated from the tree frog Phyllomedusa sauvagii, showed an EC₅₀ value of 17.81 µg/mL against promastigotes and an EC₅₀ value of 1.31 µg/mL against amastigotes of L. major, indicating its superior efficacy against both forms, as described in Table 11 (Pérez-Cordero et al. 2011). Dermaseptin-S1was found to possess better antiparasitic activity against amastigotes than promastigotes. However, it showed toxicity to human dendritic cells with an LC₅₀ value of 10.66 µg/mL and its mechanisms of action remain unclear, which could potentially limit its use as an antileishmanial agent (Table 11). Two other peptides, dermaseptin DPh-1 and dermaseptin 01, derived from the tree frog Phyllomedusa hypochondrialis, displayed the ability to fully eliminate promastigotes of L. amazonensis at concentration of 64 µg/mL, as summarized in Tables 10 and 11 (Brand et al. 2006). While dermaseptin DPh-1 showed no toxicity to mammalian blood cells (white and red blood cells) up to a concentration of 53 µM, the toxicity of dermaseptin 01 was not addressed in Brand’s study (Brand et al. 2006). However, in a study by Brand et al. (2002), dermaseptin 01 did not show a haemolytic effect against human red blood cell at a concentration of 128 µg/mL (Table 11). Leite et al. (2008) also confirmed the non-toxic effect of dermaseptin 01 against mammalian peritoneal cells, with approximately 80% cell viability observed when treated with 46.7 µM of dermaseptin 01 (Table 11). Dermaseptin-S1 is a polycationic peptide with an α-helical structure peptide whereas dermaseptin DPh-1 and dermaseptin 01 are linear (UCLL class) cationic peptides with a net charge of + 4 (Table 10). Although they have shown antileishmanial activity against L. major and L. amazonensis, dermaseptin-S1 exhibited low specificity when targeting L. major, suggesting its potential effects against normal mammalian cells (Garza-Tovar et al. 2020). Additionally, the efficacy of dermaseptin DPh-1 and dermaseptin 01 against amastigotes of L. amazonensis has not been tested and their mechanisms of action remain unclear, highlighting the necessity for further investigation in this regard.

In addition to the dermaseptin peptide family, the phylloseptin peptide family has also been studied for its antiparasitic effects against L. major. This family includes phylloseptin-S1 (AP01581), phylloseptin-S2 (AP03220) and phylloseptin-S4 (AP03222), which have been isolated from the monkey frog Phyllomedusa sauvagii. Based on Raja et al. (2013), phyllopseptin-S1 has the highest potency against promastigotes of L. major with an IC₅₀ value of 12.6 µM. This is followed by phyllopseptin-S2 with an IC₅₀ value of 13.3 µM and phylloseptin-S4 with an IC₅₀ value of 18 µM, as summarized in Table 11. However, it should be noted that they have also been reported to be toxic to human monocytes THP-1, with an LC₅₀ value of 23 µM, which may limit their use as an antileishmanial agent, as summarized in Table 11. They are α-helix cationic peptides with a net charge of + 2 and they are classified as the UCLL class in the database, highlighting their α-helix linear cationic properties (Table 10). While their exact mechanisms of action are still unclear, it is believed that the cationic and amphipathic properties of these peptides are suggested to contribute to their enhanced activity by allowing them to bind to the anionic parasitic membrane surface. This, in turn, implement them to form a helical amphipathic structure, which can permeabilize the parasitic membrane and ultimately leading to cell collapse when the peptides reach a critical concentration threshold (Raja et al. 2013). However, it is noticeable that this study only focused on the peptides’ effectiveness against promastigotes of L. major and their efficacy against amastigotes has not been tested. Therefore, future research should consider evaluating these peptides against all forms of L. major and addressing potential toxicities to provide a more comprehensive understanding of their therapeutic applications.

Additionally, peptides from the temporin family, such as temporin A (AP00094), temporin B (AP00095), temporin-SHa (also known as temporin-1Sa; AP00898), Temporin F (AP00098) and Temporin L (AP00101), have shown antiparasitic activity against L. mexicana (Eggimann et al. 2015). Among these, temporin-SHa showed the highest antiparasitic effect against promastigote of L. mexicana (ED₅₀ = 4 µM), followed by temporin L (ED₅₀ = 5 µM), temporin A (ED₅₀ = 8 µM), temporin F (ED₅₀ = 14 µM) and temporin B (ED₅₀ = 38 µM), as shown in Table 11. Notably, only temporin-SHa and temporin L showed activities against amastigotes of L. mexicana with ED₅₀ = 42 µM (temporin-SHa) and 83 µM (temporin L) (Table 11). This highlights that temporin 1Sa and temporin L have the potential to be a promising antileishmanial candidates against both forms of L. mexicana. These peptides are α-helix cationic peptides with a net charge of + 2 and are classified into the UCLL class in the database, highlighting their α-helix linear cationic properties (Table 10). The mechanisms of actions of these peptides remain unclear. However, Eggimann et al. (2015) noted that the temporin peptides demonstrated better antiparasitic activities against promastigotes than amastigotes due to the presence of proteophosphoglycan (PPG) on the surface of promastigotes of L. mexicana. This presence of PPG forms a negatively charged membrane, which is more susceptible to temporin peptides with cationic properties, as confirmed by Eggimann et al. (2015). This indicates that the peptides likely to interact with PPG on the membrane to exert their activity against promastigotes. This study also lacks a cytotoxicity test of the peptides against human cells, emphasizing the need for further examination in this regard. In addition, there is one temporin peptide, which known as temporin-SHd, has been reported to possess efficacy against promastigotes of Leishmania parasites, including L. major (IC₅₀ = 14.6 µM), L. tropica (IC₅₀ = 13.9 µM) and L. amazonensis (IC₅₀ = 14.1 µM), with a low cytotoxicity effect against human monocytes (LC₅₀ = 66 µM), as summarized in Table 11 (Abbassi et al. 2013). Therefore, Temporin-SHd has the potential to be a promising antiparasitic candidate against T. cruzi, T. brucei, and Leishmania parasites due to its significant efficacy against three parasites and low cytotoxicity effect against human monocytes.

In addition to the APPs isolated from amphibians, APPs derived from mammals have also been found to have an antiparasitic effect against L. major. These include unmodified BMAP-28 (L-BMAP-28; AP00367), retro-inverso isoform of BMAP-28 (RI-BMAP-28), and D- amino acid isoform of BMAP-28 (D-BMAP-28) (Lynn et al. 2011). BMAP-28 is a cathelicidin peptide isolated from the cattle Bos taurus and it is a linear (UCLL class) cationic peptide with an α-helix structure (Table 10). Among these three peptides, D-BMAP-28 showed the highest potent in reducing approximately 95% of cell viability of promastigotes of L. major Seidman wild type strain, followed by RI-BMAP-28 (reducing 80% of promastigotes) and L-BMAP-28 (reducing 40% of promastigotes) at a concentration of 2 µM (Table 11). Lynn et al. (2011) mentioned that D-BMAP-28 and RI-BMAP-28 showed excellent efficacy against promastigotes compared to L-BMAP-28, primarily because both peptides were not influenced by the proteolytic effect of glycoprotein 63 (GP63) on the promastigotes surface. Notably, L-BMAP-28 showed enhanced antiparasitic effect against L. major Seidman strain GP63 knockout, indicating the crucial role of GP63 in the interaction between the peptide and promastigote of L. major. GP63, also known as leishmanolysin, is a glycoprotein present on the surface of Leishmania parasite (Lieke et al. 2008). GP63 is significant for Leishmania as it is involved in facilitating parasite entry into macrophages, modulating the host immune responses, and triggering proteolytic effects to degrade the host defense proteins to promote the parasite’s survival (Isnard et al. 2012). Hence, GP63 likely facilitates the proteolytic activity against AMPs, as proven by the reduced antiparasitic effect of L-BMAP-28 in Lynn’s study (Lynn et al. 2011). Additionally, all three peptides, such as L-BMAP-28, RI-BMAP-28, and D-BMAP-28 also showed efficacy against amastigotes of L. major Seidman wild type strain. At a concentration of 2 µM, these peptides were able to reduce amastigotes by 82%, 80%, and 67%, respectively (Table 11). L-BMAP-28 showed the highest activity against amastigotes due to its down-regulation of amastigotes. They also showed no-toxicity against murine-derived macrophages, with approximately 96% cell viability when treated with a 5 µM concentration, as summarized in Table 11 (Lynn et al. 2011). Their mechanism of actions against L. major involves membrane disruption and late-stage apoptotic cell death. Overall, this study showed that L-DMAP-28 and its two isomers exhibited notable antileishmanial effects against promastigotes and intracellular amastigotes, indicating their potential for the development of antileishmanial therapeutics.

Conversely, human neutrophil peptide-1 (HNP-1; AP00176) is a defensin peptide isolated from humans. It is a cationic defensin peptide with β-sheet structure and three disulfide bonds between side chains, classified as the UCSS class (Table 10). Based on Dabirian et al. (2013), HNP-1 has the ability to kill approximately 40% of promastigotes of L. major at a concentration of 40 µg/mL, as shown in Table 11. Besides, the study found that HNP-1 also showed antiparasitic effects against amastigotes. In this study, the effectiveness of HNP-1 against amastigotes was evaluated by measuring the DNApol gene in infected LM-1 cells using Realtime-PCR. The results showed a significant decrease in number of gene copies in the LM-1 cells treated with 20 µg/mL of HNP-1 (53,867 gene copies) compared to the untreated LM-1 cells (24,018 gene copies), indicating its potential to eliminate both promastigotes and amastigotes of L. major (Table 11). The possible mechanisms of action of HNP-1 against promastigotes may involve early apoptosis followed by necrosis, or induction of cell death through necrosis alone (Dabirian et al. 2013).

Moreover, a study conducted by Söbirk et al. (2013) found that human chemokines, specifically CXCL2 (AP02077), CXCL6 (AP02185), CXCL9 (AP02079), CXCL10 (AP02080), CCL20 (AP02075), and CCL28 (AP02186), play a role as AMPs, exhibit antiparasitic effects against L. mexicana. The expression of chemokines is dependent on the infection of the Leishmania parasite and upon detection, they play an important role in mobilizing the host’s defense mechanisms to hinder the parasite at inoculation site and induce an immune response against the parasites (Oghumu et al. 2010). In the Söbirk study, the cytotoxicity of chemokines against promastigotes of L. mexicana was measured using an MTT-based assay (Söbirk et al. 2013). Results showed that CXCL6, CXCL9, and CCL28 had the highest activities against L. mexicana promastigotes, resulting in over 80% cell death, followed by CXCL2, CXCL10, and CCL20, which resulted in approximately 50% cell death at a concentration of 10 µM, as summarized in Table 11 (Söbirk et al. 2013). Interestingly, CLL28, the most effective chemokine against L. mexicana, was found to be non-toxic to HaCaT cells at a concentration of 10–20 µM (Table 11). The majority of chemokines demonstrated membrane disruption, subsequently leading to cell lysis and cell aggregation, as observed through flow cytometry and scanning electron microscopy (Söbirk et al. 2013). Overall, these findings suggest that chemokines have the potential to be developed into immunomodulatory drugs against Leishmania parasites in the future.

In addition, certain peptides isolated from amphibians have shown potential as antileishmanial agents against L. panamensis and L. braziliensis, the two species responsible for causing mucocutaneous leishmaniasis. For example, dermaseptin-S1 (Pérez-Cordero et al. 2011; AP00157), temporin-SHe (André et al. 2020; AP03245), and temporin-SHd (Abbassi et al. 2013; AP02118) were initially recognized for their effectiveness against L. major, but have also shown similar effectiveness against L. panamensis and L. braziliensis. In a study conducted by Pérez-Cordero et al. (2011), it was established that dermaseptin-S1 exhibited antileishmanial effects against L. panamensis, with an EC₅₀ value of 63.75 µg/mL against promastigotes and an EC₅₀ of 12.41 µg/mL against amastigotes, as summarized in Table 11. This suggests that dermaseptin-S1 is more effective against amastigotes than promastigotes. However, there are concerns about its applicability due to its toxicity to human dendritic cells. The mechanisms of action for both forms of L. panamensis are also unclear, highlighting the need further research to address these concerns. In contrast, temporin-SHe and temporin-SHd have also shown antiparasitic activities against promastigote forms of L. braziliensis with an IC₅₀ value of 10.5 µM and IC₅₀ value of 17.9 µM, as conducted by André et al. (2020) and Abbassi et al. (2013). Despite temporin-SHe has a better antiparasitic activity than temporin-SHd, it showed a higher toxicity effect against human THP-1 monocytes, with an LC₅₀ value of 11.4 µM compared to temporin-SHd with LC₅₀ value of 66 µM (Table 11; André et al. 2020). This suggests that temporin-SHd may be a promising antileishmanial candidate due to its lower toxicity. The mechanisms of action for both peptides are still unknown, but André et al. (2020) proposed that both peptides may eliminate the promastigotes through membrane permeabilization, which is consistent with previous findings involving membrane permeabilization against bacteria membranes. Overall, dermaseptin-S1 and temporin-SHe pose challenges due to their toxicity effects on human cells, while temporin-SHd emerges as a promising antileishmanial peptide with lower toxicity and enhanced efficacy, indicating its potential for future development.

The phylloseptin peptides, including phylloseptin-S1 (AP01581), phylloseptin-S2 (AP03220), and phylloseptin-S4 (AP03222), not only showed efficacy against L. major but also demonstrated antileishmanial effect against L. braziliensis promastigotes with IC₅₀ values of 15.3 µM (phylloseptin-S1), 15 µM (phylloseptin-S2), and 17.2 µM (phylloseptin-S4) (Table 11, Raja et al. 2013). Their mechanisms of action against L. braziliensis are predicted to be similar to those against L. major, involving plasma membrane permeabilization due to their cationic amphipathic properties. Additionally, peptides derived from insects, such as cecropin A (AP00139) from the moth Hyalophora cecropia and andropin (AP00343) isolated from the fly Drosophila mauritiana have been found to possess antileishmanial activity against L. panamensis (Table 11). Cecropin A and andropin have shown effectiveness against the amastigote forms of L. panamensis, with cecropin A having an EC₅₀ value of 2.48 µg/mL and a SI value of 40, while andropin having an EC₅₀ value of 23.45 µg/mL and SI value of 4 (Table 11; Pérez-Cordero et al. 2011). The superior efficacy of cecropin A suggests that cecropin A is more potent and selective than andropin in eliminating the amastigote forms of L. panamensis, making it a promising candidate for further development as an antileishmanial agent. The mechanism of action of cecropin A and andropin may involve an indirect mechanism, including the activation of the parasiticidal efficacy of infected phagocytes (Pérez-Cordero et al. 2011). Overall, cecropin A and andropin stand out as potent and non-toxic antileishmanial peptide against intracellular amastigote forms of L. panamensis, highlighting their potential as an antileishmanial agent for further research and development in the field of antileishmanial peptides.

Furthermore, peptides such as dermaseptin, phylloseptin, and temporin peptides, which have been isolated from amphibians, have shown efficacy against L. infantum and L. donovani, the causative species of visceral leishmaniasis. For example, dermaseptin 01 (AP01389) showed an IC₅₀ value of 10.8 µg/mL against promastigotes of L. infantum (Table 11; Eaton et al. 2014). The mechanism actions of dermaseptin 01 suggests that this peptide induces parasite cell death through membrane disruption. Another dermaseptin peptide, dermaseptin-H10 (AP00951), is a linear (UCLL class) cationic peptide with a net charge of + 2 (Table 10). It showed efficacy against promastigotes of L. infantum with an IC₅₀ value of 8.1 µM and amastigotes of L. infantum with an IC₅₀ value of 64.6 µM (Table 11). It is noteworthy that dermaseptin-H10 is non-toxic to mammalian peritoneal cells, maintaining approximately 100% viability up to a concentration of 64 µM, highlighting its potential as a potent antileishmanial agents against L. infantum (Brand et al. 2013). However, the mechanism of actions of dermaseptin-H10 against L. infantum remains unclear in this study.

Certain phylloseptin peptides, such as phylloseptin-S1 (AP01581), phylloseptin-S2 (AP03220), and phylloseptin-S4 (AP03222), have been found to have antileishmanial effect against promastigotes of L. infantum. Among these, phylloseptin-S1 showed the highest potency against promastigotes of L. infantum with an IC₅₀ value of 16.5 µM, followed by phyllopsetin-S2 with an IC₅₀ value of 18.5 µM and phylloseptin-S4 with an IC₅₀ value of 22 µM (Table 11; Raja et al. 2013). However, these peptides have also been reported to exhibit cytotoxicity effects on human monocytes THP-1, with LC₅₀ value of 23 µM. Their mechanism of action is predicted to involve membrane permeabilization of the parasite (Raja et al. 2013). Furthermore, temporin peptides, such as temporin-SHa (AP00898), temporin-SHe (AP03245), and temporin-SHd (AP02118), have also shown antileishmanial effects against L. infantum. In a study conducted by Abbassi et al. (2008), temporin-SHa demonstrated an IC₅₀ value of 20 µM against promastigotes and axenic amastigotes of L. infantum without showing any toxic effects on macrophages at that concentration (Table 11; Abbassi et al. 2008). Although temporin-SHa showed potent antileishmanial effects against two forms of L. infantum, its mechanism of action against L. Infantum is still unclear. Two additional temporin peptides, temporin-SHd and temporin-SHe, have, shown a stronger antileishmanial activity against promastigotes of L. Infantum, with an IC₅₀ values of 16.5 µM (temporin-SHd) and 4.6 µM (temporin-SHe), as shown in Table 11 (Abbassi et al. 2013; André et al. 2020). Despite temporin-SHe showed superior antiparasitic activity, it also exhibited higher toxicity with LC₅₀ value of 11.4 µM compared to temporin-SHd (LC₅₀ of 66 µM) against human monocytes THP-1 (Table 11). Also, temporin-SHd demonstrated an antiparasitic activity against axenic amastigotes with an IC₅₀ value of 23.5 µM and intracellular amastigotes with an IC₅₀ value 6.7 µM for L. infantum (Abbassi et al. 2013). However, the effectiveness of temporin-SHe against amastigotes was not evaluated in André’s study (André et al. 2020). The mechanism of actions of both peptides are suggested to involve membrane permeabilization via pore formation. In summary, temporin-SHd shows more potential as an antileishmanial agents of L. infantum due to its lower toxicity and enhanced activity against promastigotes and intracellular amastigotes of L. infantum.

Apart from their efficacy against L. infantum, temporin peptides, such as temporin A (AP00094) and temporin B (AP00095), have also been shown to have antileishmanial effects against L. donovani. In Mangoni’s study, temporin A exhibited antileishmanial activity against promastigotes of L. donovani with LC₅₀ value of 8.4 µM whereas temporin B showed an LC₅₀ value of 8.6 µM, as shown in Table 11 (Mangoni et al. 2005). Besides, both temporin A and temporin B showed antiparasitic activities against amastigotes of L. pifanoi, with LC₅₀ values of 14.6 µM (temporin A) and 7.1 µM (temporin B) (Mangoni et al. 2005). It is noteworthy that neither of these temporin peptides showed any toxicity effects against murine macrophages at a concentration of 80 µM. Mangoni et al. (2005) suggested that the mechanisms for both temporin peptides involve membrane permeabilization, as evidenced by transmission electron microscopy, which displayed severe damage to the parasite membrane and a collapse of the membrane potential. On the other hand, there are such peptides isolated from the toad Bombina variegate, known as bombinin H2 (AP00793) and bombinin H4 (AP00056), which have shown efficacy against promastigotes of L. donovani in another study conducted by Mangoni et al. (2006). In this study, the efficacy of both peptides against leishmania parasites was measured by their ability to inhibit MTT reduction compared to the untreated parasites, denoted as ED₅₀. Bombinin H4 showed a better efficacy with an ED₅₀ value of 1.7 µM compared to bombinin H2 with an ED₅₀ value of 7.3 µM. Besides, both peptides also showed activity against amastigotes of L. pifanoi with ED₅₀ value of 5.6 µM (bombinin H4) and 11 µM (bombinin H2) (Mangoni et al. 2006). Notably, neither peptides showed any toxic effects on human erythrocytes. In the APD3 database, both peptides are classified as the UCLL class, indicating that they have linear properties. They are also considered to have an α-helix 3D structure and positively charged with net charged of + 3 (Table 10). The enhanced efficacy of Bombinin H4 compared to bombinin H2 is due to its higher binding affinity towards the parasite membrane. Mangoni et al. (2006) suggested that the lower binding affinity of Bombinin H2 may results from its lower hydrophobic interaction with the parasite membrane. Another possible reason is that bombinin H2 acquired a higher percentage of β-sheet aggregation compared to bombinin H4, which may contribute to its lower efficacy (Simmaco et al. 2009). Therefore, bombinin H4 exhibits a greater affinity for binding to the membrane, subsequently leading to enhanced plasma membrane permeation and membrane disruption.

In addition to the APPs mentioned in APD3 Antimicrobial Peptide Database for their antiparasitic activities against Leishmania, there are certain AMPs not listed in the database that have been reported to show antiparasitic effects against Leishmania. For instance, one such AMPs derived from aquatic animals is tachyplesin, a peptide isolated from the horseshoe crab Tachypleus tridentatus. This peptide has also demonstrated antileishmanial effects against the promastigote form of L. braziliensis. In a study conducted by Löfgren et al. (2008), tachyplesin showed an LD₅₀ value of 4.7 µM and a SI value of 8 against promastigotes of L. braziliensis, as shown in Table 12. However, it should be noted that tachyplesin was not assessed for its activity against the amastigote form of L. braziliensis in this study. Additionally, the mechanism of action of tachyplesin against L. braziliensis promastigotes remains unclear. Therefore, further investigations are necessary to elucidate the specific mechanism involved in inhibiting L. braziliensis and to explore the potential antileishmanial activity of tachyplesin against L. braziliensis amastigotes.

Table 12 Summary of the characteristics of APPs not mentioned in APD3 database

Furthermore, tachyplesin has also demonstrated antileishmanial activity against L. infantum and L. donovani, the causative species of visceral leishmaniasis. In a study conducted by Kumar and Chugh (2021), tachyplesin exhibited an IC₅₀ of 3.96 µM against L. donovani promastigotes and removed all intracellular amastigote forms with a concentration of 20 µM, as summarized in Table 13. The mechanism involves the formation of pores on the membrane of both promastigotes and amastigotes, leading to the discharge of cytoplasmic content and irregular cell structure. Kumar and Chugh (2021) suggested that tachyplesin may interact with receptors such as LPG or glycoinositolphospholipids (GIPLs), making it more susceptible to the membrane of both promastigotes and amastigotes. Other AMPs that have shown promising antileishmanial effects against L. infantum and L. donovani include bicarinalin, and thionins. Bicarinalin, isolated from the ant venom Tetramorium bicarinatum, has shown inhibitory activity against the growth of L. infantum at a concentration of 1.5 µmol/L, as shown in Tables 12 and 13 (Téné et al. 2016). Importantly, no toxic effects were observed on human lymphocytes at concentrations ranging from 0.066 to 8.5 µmol/L of bicarinalin, with LC₅₀ value of 67.8 µmol/L (Téné et al. 2016). Bicarinalin is a partially amphipathic peptide with an α-helix structure. This structure allows it to permeabilize the anionic membrane of the parasite without inducing cell death. This property positions bicarinalin as a promising candidate for the development of effective antileishmanial drugs using a peptide-based approach, due to its robust antiparasitic activity against L. infantum and non-toxicity to human cells within a safe concentration range.

Table 13 Summary of APPs not mentioned in APD3 database, exhibiting antitrypanosomal activity against leishmania

In a separate study, thionins, a peptide derived from the plant source wheat Triticum aestivum, has demonstrated excellent activity in inhibiting the proliferation of promastigotes of L. donovani with an IC₅₀ value of 0.2 µM (Tables 13 and 12; Berrocal-Lobo et al. 2009). Besides, thionins has also shown activity against amastigotes of L. donovani with LC₅₀ value of 46.3 µM (Berrocal-Lobo et al. 2009). Thionins is more effective against promastigotes compared to amastigotes due to the presence of LPG and GP63 on the promastigote membrane. The mechanism of action of thionins is typically related to the permeabilization of parasite membranes, resulting in the collapse of pH and calcium ionic gradients on the membrane, as well as a reduction in intracellular ATP levels, ultimately leading to the death of the parasite (Berrocal-Lobo et al. 2009; Petranka et al. 2016).

In summary, many of the reported studies have confirmed the notion that APPs have promising antiparasitic properties against Leishmania parasites, as evidenced by their IC₅₀ values. For instance, dermaseptin, phylloseptin and temporin peptides have demonstrated significant efficacy against both promastigotes and amastigotes of several Leishmania species, such as L. major, L. Mexicana, L. braziliensis, L. panamensis, L. infantum and L. donovani. Notably, phylloseptin-S1, S2 and S4 have shown efficacy against three different species, such as L. major, L. braziliensis and L. infantum, which contribute to three main forms of leishmaniasis: cutaneous leishmaniasis, mucocutaneous leishmaniasis, visceral leishmaniasis, respectively. Moreover, temporin-SHd has shown efficacy against five species of Leishmania, including L. major, L. tropica, L. amazonensis, L. braziliensis, and L. infantum. This peptide has been particularly effective against L. infantum intracellular amastigotes, which are the parasite forms that remain in the infected humans. Also, this peptide has demonstrated efficacy against T. cruzi and T. b. gambiense, highlighting its potential as a versatile therapeutic agent against Chagas disease, HAT and the three main forms of leishmaniasis. This is significant because there is currently no single drug that is effective for all three NTDs. Furthermore, mammal-derived peptides (BMAP-28 and HNP-1) are also been effective against L. major, while human chemokines have exhibited antileishmanial activity against L. mexicana. This highlights their potential as therapeutic agents for cutaneous leishmaniasis, which is important considering the lack of new therapeutic options for this form of the disease. These peptides and chemokines could offer an alternative treatment option that is effective and yet potentially less toxic than existing drugs. Insect-derived peptides, such as cecropin-A and andropin, have also shown effective antiparasitic activity against L. panamensis. In particular, cecropin-A has shown good efficacy (EC₅₀ = 2.48 µg/mL) against amastigotes of L. panamensis, underscoring their potential as therapeutic agents for mucocutaneous leishmaniasis, which also lacks of new treatment options. Hence, the peptides derived from both mammal and insect sources could offer promising new treatments for both cutaneous and mucocutaneous leishmaniasis, addressing critical gaps in the current therapeutic landscape. Additionally, aquatic-derived peptide such as tachyplesin has emerged as a potent AMP, displaying effective antiparasitic activity against L. brazilliensis promastigotes and L. donovani promastigotes and amastigotes. However, the mechanism of tachyplesin against L. brazilliensis is unclear, highlight the need of further research. Meanwhile, thionins are the only plant-derived peptides that have been shown to have antileishmanial effects against both forms of L. donovani.

Lymphatic Filariasis

Lymphatic filariasis (LF), also known as elephantiasis, is a NTD caused by infection with parasitic helminth such as Brugia timori (B. timori), Wuchereria bancrofti (W. bancrofti) and Brugia malayi (B. malayi) (McNulty et al. 2013).At least 90% of LF cases is attributed to W. bancrofti (Gordon et al. 2018). LF is transmitted through mosquito bites, with the species like Culex, Anopheles, Mansonia and Aedes. The mosquitoes become infected when they bite and ingest microfilariae from an infected person. The microfilariae then mature into larvae within the mosquito’s midgut and are transmit the larvae to another person during a subsequent bite, where they develop into adult worms and disrupt the lymphatic system resulting in swelling and blockage. These worms produce millions of microfilariae that circulate in the blood, enabling the cycle to continue when another mosquito bites the infected person (Fimbo et al. 2020).

LF disease has three distinct manifestations which are asymptomatic, acute, and chronic. The majority of infected patients do not show any external signs of the disease. However, even in the absence of external signs, the microfilariae can caused hidden damage to the lymphatic system. In the acute stage, the symptoms can be observed, including inflammation of the lymph glands and lymph channels (lymphadenitis and lymphangitis), along with episodic fever and pulmonary eosinophilia (Bizhani et al. 2021). In the chronic stage, the inflammation of the lymph glands can progress to lymphoedema of the arms or legs and hydrocele, resulting in body deformities and significant disability (Chandy et al. 2011).

In the 1990s, LF became a major public health problem, with 72 countries located in Africa, South-East Asia, Western Pacific regions, Eastern Mediterranean and South America reporting endemic cases. It was estimated that 120 million people in these endemic countries were infected (Hotez and Ehrenberg 2010; Khieu et al. 2018; Fimbo et al. 2020). In response, the World Health Assembly passed Resolution WHA 50.29 in 1997, calling for the elimination of LF as a global public health problem (Molyneux 2003). This led to the establishment of the Global Programme to Eliminate Lymphatic Filariasis (GPELF) by the WHO in 2000 (accessed 15 May 2024). The GPELF’s elimination strategy focused on two main objectives, which are interrupting transmission of LF and controlling morbidity (specifically hydrocele and lymphedema) in affected populations. To achieve the goal of interruption of LF transmission, the WHO has described mass drug administration (MDA) to the entire at-risk population annually for 5 to 6 years. This involved the use of a two-drug regimen, which include diethylcarbamazine (DEC) with albendazole (ALB), ivermectin (IVM) with ALB, or ALB alone. However, recent clinical trials have shown that a triple-drug therapy combining DEC, ALB, and IVM is more effective against microfilariae in LF compared to the previously recommended two-drug regimen (King et al. 2018; Thomsen et al. 2016). As a result, the WHO now recommends the use of triple-drug regimens for the treatment of LF.

The WHO delivered a total of 9 billion treatments to more than 900 million people in 72 endemic countries from 2000 to 2021. This has resulted in the decline of number of reported infections with only 51 million infected people reported in 2018. As a result, 17 countries have achieved the elimination of LF by 2020 (accessed 15 May 2024). However, there are still 44 countries that rely on the MDA and 9 countries that have not yet received MDA. In addition to providing treatment, the WHO recommends an essential package of care for affected individuals, which includes surgery for hydrocele, treatment for infection, and management of adenolymphangitis episodes, aiming to alleviate suffering of patients, but also prevents the progression and severity of the disease, ultimately achieving the goal of eliminating LF (accessed 15 May 2024). The implementation of mosquito control measures such as insecticide-treated nets and indoor residual spraying has also been suggested to reduce the transmission of LF by killing mosquitoes (van den berg et al. 2013).

While the current triple-drug therapy has been shown to effectively kill microfilariae and sterilize adult filarial worms but the therapy fail to kill worms which may allow the new microfilariae production (King et al. 2018). Therefore, research into natural products offers promising new avenues for eliminating LF. To the best of our knowledge, no natural-derived peptides have been discovered to be effective as antiparasitic agents targeting lymphatic filariasis. In contrast, certain plant-derived phytochemicals have been found to have promising antifilarial activities. For example, phytochemicals such as gedunin and photogedunin isolated from the Xylocarpus granatum fruit, have demonstrated antifilarial activities against both microfilariae and adult worms of Brugia malayi in vitro study. They also showed good adulticidal efficacy in killing 80% and 70% of transplanted adult worms of B. malayi in a Jird model after 5 days of treatment with 100 mg/kg (Misra et al. 2011). Other phytochemicals, such as asperoside and strebloside from Streblus asper, exhibited adulticidal properties by killing over 70% worms of B. malayi in a Mastomys model after 5 days of treatment with 50 mg/kg (Chatterjee et al. 1992). Additionally, oleanonic acid and oleanolic acid have shown efficacy against adult worm of B. malayi in an vitro study, with LC₁₀₀ values of 31.25 µg/mL and 62.5 µg/mL, respectively (Misra et al. 2007).

According to Das et al. (2023), the limited availability of drugs has led researchers to explore alternative methods for targeting filarial worms such as inducing apoptosis to kill filarial worms. Phytochemicals such as ursolic acid isolated from Nyctanthes arbortristis have shown promising antifilarial activity, killing 100% microfilariae of W. bancrofti at 10 µg/mL by activating the proapoptotic genes such as eg1-1, ced-3, and ced-4 via the increase in ROS (Saini et al. 2014). Although natural products have the potential to be effective antifilarial agents, research in this area faces few limitations including limited funding, a lack of a centralized natural product library, a lack of suitable model organisms for screening novel antifilarial compounds. Kamal et al. (2023) suggested using Caenorhabditis elegans and Oscheius tipulae as model organisms for high-efficiency screening of natural products. By utilizing appropriate screening models, researchers can efficiently identify the potential compounds to develop new antifilarial agents.

Conclusion and Future Perspectives

Despite being caused by different parasites, the NTDs mentioned in this review share a common vector-borne mode of transmission, facilitated by infected insect vector. This review has revealed that peptides isolated from animal sources have significant potential as antiparasitic agents especially those derived from amphibian sources. Moreover, peptides derived from plants, such as thionins, and fungi, such as leucinostatins, alamethicin I, and tsushimycin, have also shown promise in treating human African trypanosomiasis. Notably, thionins, derived from plants, may be particularly effective against leishmania, exhibiting superior antileishmanial activity against L. donovani compared to animal-derived peptides. Structural parameters play a crucial role in conferring antitrypanosomal activity to nature-derived peptides against Chagas disease, human African trypanosomiasis, and leishmaniasis. This review highlights the fact that AMPs with antitrypanosomal activity against T. cruzi and Leishmania parasites typically possess a linear, α-helical, amphipathic structure, with a predominance of positively charged amino acid residues and are often belong to the category of short and medium-length peptides. The cationic nature of these peptides allows them to selectively target parasitic infected cells, as they are able to interact more strongly with the negatively charged ions present on the membranes of these cells, compared to normal cells. For example, in the case of Chagas disease, AMPs exhibit heightened selectivity and potent antitrypanosomal activity against the trypomastigote form. This efficacy is primarily due to the higher concentration of negatively charged ions on the membrane surface of trypomastigotes, making them are more susceptible to action of peptides.

In contrast, for LF disease, no natural-derived peptides have currently been discovered to be effective as antiparasitic agents targeting LF. Hence, further research on natural-derived peptides should focus on exploring their antiparasitic activities against nematodes, including B. timori, W. bancrofti and B. malayi, which are the causative agents of LF.

Peptides derived from nature often use direct microbial interactions to induce cell death by releasing intracellular contents. In addition, these peptides can employ various mechanisms to cause parasite cell death, including inducing autophagy, apoptosis, and necrosis, inhibiting dolichol-phosphate-sugar complexes, promoting of cytolysis and endocytosis, and forming ionophores. For instance, certain nature-derived peptides induce autophagy to break down cellular components, while others inhibit the formation of sugar complexes that are crucial for parasite cell survival. Furthermore, some nature-derived peptides damage the cell membrane or form ionophores on the membrane, ultimately leading to parasite cell death. These mechanisms of action differ significantly from those mechanism of current drugs, which primarily cause DNA damage in parasites, target certain enzymes involved in protein synthesis, and interfere with metabolic processes. The unique modes of action of nature-derived peptides could provide a complementary approach to existing treatments, potentially overcoming issues related to side effects and drug resistance.

Besides discussing alternative therapeutics for Chagas disease, HAT, and leishmaniasis, and LF, the WHO and CDC also emphasize the importance of implementing preventive measures to reduce the spread of these diseases (accessed 22 May 2024). These measures include regular insecticide spraying around human settlements to eliminate bugs, tsetse flies, sandflies and mosquitoes and improving housing by using materials that are less conducive to insect infestation. In areas prone to HAT and leishmaniasis, setting up insecticide-treated traps and insecticidal nets can control the breeding of tsetse flies and sandflies. In areas prone to LF, removing standing water and using biological control strategy such as mosquitofish (Gambusia affinis), along with the use of mosquito nets and applying mosquito repellents. Regular screenings of at-risk populations include blood and organ donors and pregnant women are necessary to detect diseases early and prevent transmission through blood and organ transfusion, and congenital transmission. Community education is also necessary in raising awareness about the risks of these diseases, promoting preventive measures, and advising against consumption of raw food in endemic areas in endemic areas can reduce the risk of transmission.

Although the practical use of AMPs is still in early stages, they offer several benefits. For example, AMPs can selectively target specific receptors, reducing off-target effects that can lead to toxicity. Besides, AMPs often exhibited high potency at low concentration, ranging from micromolar to nanomolar. This has been demonstrated by bacteriocin AS-48, which showed potency against BSF of T. b. rhodesiense at a nanomolar concentration. Due to their specificity and high potency, peptides prone to have fewer side effects on mammalian cells. Besides, peptides are biodegradable and are normally metabolized by proteases into amino acids. These amino acids can easily pass through barriers and reach their target site (López-García et al. 2022). Hence, this predictable metabolism reduces the risk of accumulation of toxic metabolites. In addition to the advantages of AMPs, the increasing of prevalence of infectious diseases and the demand for effective and safe treatments are also driving the growth of the AMPs market. Currently, AMPs are emerging as promising new options for bacterial and fungal infections, which is likely to increase the public acceptance of these peptides. As acceptance of AMP-based medications and therapies continue to grow, the global market for these peptides is anticipated to grow substantially in future.

Despite the potential benefits of using specific peptides isolated from nature to combat diseases such as Chagas disease, Human African trypanosomiasis, and Leishmaniasis, it is important to acknowledge their limitations. AMPs have their own set of limitations, such as their short half-life due to their susceptibility to degradation by cytosolic proteases, which prevents them from reaching their target sites (Starr and Wimley 2017). Besides, most peptides are small in size and easily degraded, causing them easily excreted by the kidneys (Benincasa et al. 2010). Another challenge faced by peptides is their poor oral absorption when administered orally, as they are difficult to cross the gastrointestinal tract and are rapidly degraded by digestive enzymes (Deshayes et al. 2022). To address the issues of rapid excretion by the kidney, the introduction of polyethylene glycol (PEG) to peptides has been shown to reduce clearance and improve their (Benincasa et al. 2015). Similarly, to overcome the challenge of oral administration, encapsulating AMPs in drug delivery systems (DDS) such as liposomes, microparticles, and nanoparticles can aid in crossing the gastrointestinal tract and avoiding exposure to the stomach (Deshayes et al. 2022).

Currently, there is no research utilizing antimicrobial peptides to target persister cells present in T. cruzi and Leishmania spp. In addition to the emergence of drug resistance, these persister cells also pose a significant challenge for existing drug in achieving a complete eradication of the disease. Additionally, certain peptides such as dermaseptin-DI1, dermaseptin-DI2, temporin F, temporin L, chemokines, mastoparan, and decoralin, lack comprehensive evaluation of their cytotoxic effects on mammalian cells. Additionally, some peptides, including melittin, dermaseptin-S1, phylloseptin-S1, -S2, -S4, and thionins, have been found to be toxic to normal cells. To address these concerns, one potential solution is to synthesize synthetic peptides based on natural templates. This approach could potentially reduce toxicity without compromising effectiveness against parasites. Another innovative approach involves hybridizing peptides, aiming to combine two nature-derived peptides to create a more powerful and safer treatment option. However, in order for these advancements to be successfully applied in practical applications, further in vivo study are necessary to determine the bioavailability of these peptides. These considerations highlight the importance of taking a comprehensive approach to developing antiparasitic peptides that are both potent and safe for therapeutic use. The production of AMPs is also a concern. As the demand for AMPs increases, methods such as directly isolating them from natural products or using chemical synthesis through solid phase peptide synthesis (SPPS) may not be cost-effective. To produce AMPs on a large-scale in a cost-efficient manner, biological production systems are needed. Yeast and Escherichia coli (E. coli) are the promising options for producing recombinant AMPs (Mazurkiewicz-Pisarek et al. 2023).

This review analyses the effectiveness of nature-derived APPs, specifically from amphibians, insects, mammals, bacteria, marines and plants in combating Chagas disease, Human African trypanosomiasis, and Leishmaniasis. They have shown a substantial antiparasitic effects, as evidenced by their low IC₅₀ values. This indicates their potential as potent treatments against these diseases. Consequently, future research should focus on developing synthetic peptides based on these nature-derived AMPs as a base template. This approach aims to reduce the peptide toxicity to human cells, optimize their effectiveness against parasites, and improve their overall bioavailability. These nature-derived AMPs have the potential to be promising candidates for antiparasitic therapeutic applications, contributing a potential alternative to current drugs and dealing with the concerns associated with resistance issues in these diseases.