# Functional Effects of Cyclization on Li1 Peptide Activity against Metacyclic Leishmania amazonensis Internalization

## TL;DR

A cyclic peptide called Li1 was found to reduce Leishmania infection in macrophages and mice without causing toxicity, suggesting it could be used to prevent the spread of the parasite.

## Contribution

This study provides the first direct comparison between a cyclic peptide (Li1) and its linear analogue (Li1nc) in inhibiting Leishmania amazonensis infection.

## Key findings

- Li1 significantly reduced infection rates in macrophages and parasite burden in mice compared to Li1nc.
- Li1 showed stronger binding to the parasite surface, particularly along the flagellum, indicating structure-dependent interaction.
- Neither peptide caused cytotoxicity or antipromastigote activity, and their effects were limited to pre-infection interactions.

## Abstract

Leishmaniases are caused by protozoa of the genus Leishmania, whose metacyclic promastigote forms initiate
infection in the mammalian host. Building upon previous work with Leishmania infantum, this study evaluated the cyclic
peptide Li1 and its linear analogue Li1nc regarding their capacity
to interfere with Leishmania amazonensis infection. Both peptides were synthesized by solid-phase peptide
synthesis (SPPS–Fmoc) and tested after pre-exposure of metacyclic
promastigotes prior to infection of murine peritoneal and THP-1–derived
macrophages. In contrast to the previous study conducted by our group,
which evaluated only Li1 against L. infantum, the present work introduces the first direct comparison between
the cyclic peptide and its linear analogue, allowing a structure–activity
assessment not previously available. Pre-exposure to Li1 (0.5 mg mL–1 ≅ 0.318 mmol L–1) significantly
reduced the infection rate and infection index in both macrophage
models, while Li1nc (0.324 mmol L–1) exhibited moderate
inhibition. Neither peptide displayed cytotoxicity toward host cells
(CC50 > 0.5 mg mL–1) nor direct antipromastigote
activity (IC50 > 0.5 mg mL–1). Confocal
microscopy revealed stronger and more defined binding of Li1 to the
parasite surface than Li1nc, particularly along the flagellum, supporting
structure-dependent interaction with surface molecules involved in
parasite internalization. In a murine model of cutaneous leishmaniasis,
preincubation of metacyclic promastigotes with Li1 (0.2 mg mL–1 ≅ 0.127 mmol L–1) reduced
parasite burden by 35.2%, whereas Li1nc (0.130 mmol L–1) achieved a 20.4% reduction relative to untreated controls. No significant
alterations in hepatic or renal biochemical parameters were observed,
indicating the absence of systemic toxicity. Notably, neither peptide
showed activity under a postinfection treatment regimen (2 mg Kg–1), suggesting that their effects are restricted to
early host–parasite interactions rather than therapeutic clearance.
Collectively, these findings demonstrate that Li1 acts through a structure-dependent
mechanism that interferes with host–parasite recognition and
reduces infectivity without inducing detectable toxicity. The results
support the translational potential of Li1 as a safe peptide scaffold
for prophylactic or paratransgenic strategies aimed at preventing Leishmania transmission.