Background/Objectives: Cancer is one of the leading causes of death worldwide, and skin cancer is especially prevalent and lethal in Brazil. Despite advancements in treatment, there is still a need for new anticancer agents that are effective, selective, and less toxic. This study aimed to evaluate the cytotoxic and therapeutic potential of the peptide PEPAD. Methods: The cytotoxicity of PEPAD was assessed by MTT assay in murine melanoma (B16F10-Nex2), human melanoma (SK-MEL-28), breast (MCF-7), and cervical (HeLa) cancer cell lines. Selectivity was evaluated in healthy cells (RAW 264.7 and FN1). Morphological changes were analyzed by microscopy. Cell migration was assessed using scratch assays. Apoptotic features were evaluated using MitoTracker Deep Red, NucBlue, CaspACETM labeling, and flow cytometry. Immunogenic cell death was investigated by calreticulin and HMGB1 release. Molecular dynamics simulations explored peptide structure and interaction with lipid membranes. Results: PEPAD showed IC₅₀ values of 7.4 µM and 18 µM in B16F10-Nex2 and SK-MEL-28 cells, respectively, and >60 µM in MCF-7 and HeLa cells. Low toxicity was observed in healthy cells (IC₅₀ > 56 µM), indicating high selectivity. Apoptotic morphology and reduced cell migration were observed. Flow cytometry and fluorescence probes confirmed apoptosis and mitochondrial swelling. Calreticulin and HMGB1 release indicated immunogenic cell death. Simulations showed that PEPAD maintains a stable α-helical conformation and interacts with membranes. Conclusions: These findings highlight PEPAD’s selective cytotoxicity and its potential as an anticancer agent with apoptotic and immunogenic properties, making it a promising candidate for therapeutic development.

Inga cylindrica, a tropical fruit tree of the Fabaceae family (subfamily Mimosoideae), is native to South America. The seeds from this family are an essential source of trypsin inhibitors, which display promising bioactivity for increasing host defense against pathogens. The aim of the present study was to characterize the antimicrobial and antibiofilm activities of the trypsin inhibitor extracted from I. cylindrica seeds, ICTI. ICTI demonstrated antifungal activity with a minimum inhibitory concentration (MIC) of 32.11 μmol.L^-1, and a minimum fungicidal concentration (MFC) of 32.1 μmol.L^-1, against Cryptococcus gattii, Candida albicans, Candida glabrata and Candida guilliermondii. Combining ICTI with Amphotericin B had a significant synergistic effect, reducing the concentration of the antibiotic by 75% for C. albicans and 94% for C. gatti. The significant increase (16 x) in activity observed with ergosterol (1.01 mol.L^-1) for C. albicans and C. gatti, and the lack of activity against bacterial strains, suggests that ICTI interferes with the integrity of the fungal cell membrane. Treatment with ICTI at 10 x MIC resulted in a 51% reduction in biofilm formation and a 56% decrease in mature biofilm colonies for C. albicans. Finally, ICTI displayed no toxicity in the in vivo Galleria mellonella model. Given its antifungal and antibiofilm properties, ICTI could be a promising candidate for the development of new antimicrobial drug prototypes.

Background/Objectives: The rise in cancer cases is a major global public health problem. Treating the disease and maintaining health generate high economic and social costs. In recent years, treatment resistance has become a challenge, reinforcing the need for new anticancer agents. In this context, anticancer peptides (ACPs) stand out for their selective interaction with cancer cell membranes. This study aimed to evaluate the activity of the RQ18 peptide against cancer cells. Methods: The cytotoxicity of RQ18 was determined by the MTT assay in murine melanoma (B16F10-Nex2), human melanoma (Sk-mel-28 and A375), breast cancer (MCF-7), cervical cancer (HeLa), murine macrophage (RAW 264.7), and human fibroblast (FN1). Caspase-3/7 activity was further evaluated in B16F10-Nex2, Sk-mel-28, and A375 melanoma cells to investigate the involvement of apoptotic pathways. In addition, intracellular oxidative stress was assessed using the DCFH-DA probe in these three melanoma cell lines. In B16F10-Nex2 cells, cellular morphological changes were examined by microscopy, while apoptosis-associated events were characterized using the markers MitoTracker Deep Red, and NucBlue, complemented by flow cytometry analyses. In addition, molecular dynamics simulations were employed to investigate its interactions with lipid membranes. Results: The peptide exhibited anticancer activity, with IC₅₀ values of 4.4, 11.58 and 59.99 µM·mL⁻¹, against cells B16F10-Nex2, Sk-mel-28 and A375, respectively. Compared with non-cancerous cells, RQ18 preferentially affected cancer cells, especially B16F10-Nex2. In contrast, the peptide was nontoxic to healthy cells, thus indicating selectivity for cancer cells. Morphological analysis revealed alterations compatible with apoptosis, including cell rounding and chromatin condensation. RQ18 increased caspase-3/7 activity in B16F10-Nex2, Sk-mel-28, and A375 cells and induced intracellular oxidative stress, as indicated by increased DCFH-DA fluorescence. Flow cytometry data confirmed the induction of apoptotic cell death in B16F10-Nex2 cells. Caspase activation, detected by a caspase-3/7 assay in three melanoma cell lines, together with flow cytometry data, supported the involvement of apoptotic cell death. Molecular dynamics simulations suggest that the peptide interacts stably with cancer cell-like membranes and retains an α-helical conformation during membrane association. Conclusions: In summary, the RQ18 peptide demonstrates preferential anticancer activity, particularly against melanoma cells, supporting further investigation of its therapeutic potential.