Pretreatment with serine protease inhibitors impairs Leishmania amazonensis survival on macrophages
Patrícia de Almeida Machado1,2,3, Pollyanna Stephanie Gomes1,2, Elaine Soares Coimbra3
1Laboratório de Imunologia Clínica, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz-Fiocruz, Rio de Janeiro, RJ, 21040-360, Brazil.
Background:
Leishmaniases are neglected tropical diseases with great clinical and epidemiological importance. The current chemotherapy available for the treatment of leishmaniasis presents several problems, such as adverse effects, toxicity, long treatment time, and parasite resistance. The discovery of new therapeutic alternatives is extremely essential, and the discovery of cellular targets is a tool that helps in the development of new drugs. Serine proteases emerge as important virulence factors in the Leishmania genus, as they participate in important processes involved in their infectivity, virulence, and survival. In this work, we evaluated the leishmanicidal effect of different serine protease inhibitors (Benzamidine, PF-429242, PMSF, TLCK, and TPCK). Additionally, we determined the implication of pretreatment with these inhibitors on the entry and survival of parasites within macrophages, as well as the conversion of promastigotes into amastigotes, to discover the importance of serine proteases in the establishment of infection and, consequently, as targets for new drugs for Leishmania.
Results:
In general, the inhibitors had low toxicity in host macrophages, and three showed some effect in promastigote and amastigote forms of L. amazonensis (PF-429242, TLCK, and TPCK). Using a short incubation interval, we pretreated L. amazonensis promastigotes with these five compounds before in vitro infection. Pretreatment with PF-429242, TLCK, and TPCK considerably compromised the survival of these parasites inside host macrophages, without altering the entry of promastigotes into these cells and differentiation into amastigotes. In addition, treatment with PF-429242 and TPCK was able to reduce the serine proteases' enzymatic activity using subtilisin substrate on L. amazonensis promastigote lysate.
Conclusions:
This work highlights the importance of serine proteases in L. amazonensis as a possible target for new therapeutic alternatives in Leishmania spp.
Insights
New drugs targeting serine proteases show promise for treating leishmaniasis. Inhibitors like PF-429242 and TPCK reduced parasite survival in macrophages, indicating potential therapeutic strategies for this neglected tropical disease.
Area of Science:
- Parasitology
- Drug Discovery
- Molecular Biology
Background:
- Leishmaniases are neglected tropical diseases requiring new treatments due to issues with current chemotherapy, including resistance and toxicity.
- Serine proteases are key virulence factors in Leishmania, essential for parasite infectivity, virulence, and survival.
- Identifying cellular targets is crucial for developing novel anti-leishmanial drugs.
Purpose of the Study:
- To evaluate the leishmanicidal effects of various serine protease inhibitors.
- To determine the impact of these inhibitors on parasite entry and survival within macrophages.
- To assess the role of serine proteases in Leishmania infection establishment and their potential as drug targets.
Main Methods:
- Screening of five serine protease inhibitors (Benzamidine, PF-429242, PMSF, TLCK, TPCK) for leishmanicidal activity.
- Assessing inhibitor toxicity in host macrophages.
- Pretreating Leishmania amazonensis promastigotes with inhibitors before in vitro infection of macrophages.
- Measuring parasite survival, entry, and differentiation within macrophages.
- Evaluating the effect of inhibitors on serine protease enzymatic activity in parasite lysates.
Main Results:
- Three inhibitors (PF-429242, TLCK, TPCK) demonstrated activity against promastigote and amastigote forms of L. amazonensis with low host cell toxicity.
- Pretreatment with PF-429242, TLCK, and TPCK significantly reduced parasite survival within macrophages.
- Inhibitor pretreatment did not affect parasite entry into macrophages or their differentiation into amastigotes.
- PF-429242 and TPCK treatments reduced serine protease activity in L. amazonensis lysates.
Conclusions:
- Serine proteases are critical for L. amazonensis virulence and survival within host cells.
- Inhibitors targeting serine proteases, such as PF-429242 and TPCK, show potential as novel therapeutic agents against leishmaniasis.
- This study underscores the importance of serine proteases as viable targets for developing new drugs to combat Leishmania infections.
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