Development of New Leishmanicidal Compounds via Bioconjugation of Antimicrobial Peptides and Antileishmanial Guanidines.

Leishmaniasis refers to a collection of diseases caused by protozoa from the Leishmania genus. These diseases, along with other parasitic afflictions, pose a significant public health issue, particularly given the escalating number of at-risk patients. This group includes immunocompromised individuals and those residing in impoverished conditions. The treatment of leishmaniasis is crucial, particularly in light of the mortality rate associated with nontreatment, which stands at 20-30,000 deaths per year globally. However, the therapeutic options currently available are limited, often ineffective, and potentially toxic. Consequently, the pursuit of new therapeutic alternatives is warranted. This study aims to design, synthesize, and evaluate the leishmanicidal activity of antimicrobial peptides functionalized with guanidine compounds and identify those with enhanced potency and selectivity against the parasite. Accordingly, three bioconjugates were obtained by using the solid-phase peptide synthesis protocol. Each proved to be more potent against intracellular amastigotes than their respective peptide or guanidine compounds alone and demonstrated higher selectivity to the parasites than to the host cells. Thus, the conjugation strategy employed with these compounds effectively contributes to the development of new molecules with leishmanicidal activity.

Leishmanicidal Activity of Guanidine Derivatives against Leishmania infantum.

Leishmaniasis is a neglected tropical infectious disease with thousands of cases annually; it is of great concern to global health, particularly the most severe form, visceral leishmaniasis. Visceral leishmaniasis treatments are minimal and have severe adverse effects. As guanidine-bearing compounds have shown antimicrobial activity, we analyzed the cytotoxic effects of several guanidine-bearing compounds on Leishmania infantum in their promastigote and amastigote forms in vitro, their cytotoxicity in human cells, and their impact on reactive nitrogen species production. LQOFG-2, LQOFG-6, and LQOFG-7 had IC50 values of 12.7, 24.4, and 23.6 µM, respectively, in promastigotes. These compounds exhibited cytotoxicity in axenic amastigotes at 26.1, 21.1, and 18.6 µM, respectively. The compounds showed no apparent cytotoxicity in cells from healthy donors. To identify mechanisms of action, we evaluated cell death processes by annexin V and propidium iodide staining and nitrite production. Guanidine-containing compounds caused a significant percentage of death by apoptosis in amastigotes. Independent of L. infantum infection, LQOFG-7 increased nitrite production in peripheral blood mononuclear cells, which suggests a potential mechanism of action for this compound. Therefore, these data suggest that guanidine derivatives are potential anti-microbial molecules, and further research is needed to fully understand their mechanism of action, especially in anti-leishmanial studies.

Novel Selective and Low-Toxic Inhibitor of LmCPB2.8ΔCTE (CPB) One Important Cysteine Protease for Leishmania Virulence.

Leishmaniasis is a highly prevalent, yet neglected disease caused by protozoan parasites of the genus Leishmania. In the search for newer, safer, and more effective antileishmanial compounds, we herein present a study of the mode of action in addition to a detailed structural and biological characterization of LQOF-G6 [N-benzoyl-N'-benzyl-N″-(4-tertbutylphenyl)guanidine]. X-ray crystallography and extensive NMR experiments revealed that LQOF-G6 nearly exclusively adopts the Z conformation stabilized by an intramolecular hydrogen bond. The investigated guanidine showed selective inhibitory activity on Leishmania major cysteine protease LmCPB2.8ΔCTE (CPB) with ~73% inhibition and an IC50-CPB of 6.0 µM. This compound did not show any activity against the mammalian homologues cathepsin L and B. LQOF-G6 has been found to be nontoxic toward both organs and several cell lines, and no signs of hepatotoxicity or nephrotoxicity were observed from the analysis of biochemical clinical plasma markers in the treated mice. Docking simulations and experimental NMR measurements showed a clear contribution of the conformational parameters to the strength of the binding in the active site of the enzyme, and thus fit the differences in the inhibition values of LQOF-G6 compared to the other guanidines. Furthermore, the resulting data render LQOF-G6 suitable for further development as an antileishmanial drug.