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Anti-proliferative effect on a colon adenocarcinoma cell line exerted by a membrane disrupting antimicrobial peptide
Yu-Ching Chen1, Tsung-Lin Tsai, Xin-Hong Ye
1a Institute of Molecular Medicine and Department of Life Science ; National Tsing Hua University ; HsinChu , Taiwan, R.O.C.
Abstract:
The antimicrobial and anticancer activities of an antimicrobial peptide (AMP) KL15 obtained through in silico modification on the sequences of 2 previously identified bacteriocins m2163 and m2386 from Lactobacillus casei ATCC 334 by us have been studied. While significant bactericidal effect on the pathogenic bacteria Listeria, Escherichia, Bacillus, Staphylococcus, Enterococcus is exerted by KL15, the AMP can also kill 2 human adenocarcinoma cells SW480 and Caco-2 with measured IC50 as 50 μg/ml or 26.3 μM. However, the IC50 determined for KL15 on killing the normal human mammary epithelial cell H184B5F5/M10 is 150 μg/ml. The conformation of KL15 dissolved in 50% 2,2,2-trifluroroethanol or in 2 large unilamellar vesicle systems determined by circular dichroism spectroscopy appears to be helical. Further, the cell membrane permeability of treated SW480 cells by KL15 appears to be significantly enhanced as studied by both flow cytometry and confocal microscopy. As observed under a scanning electron microscope, the morphology of treated SW480 cells is also significantly changed as treating time by 80 μg/ml KL15 is increased. KL15 appears to be able to pierce the cell membrane of treated SW480 cells so that numerous porous structures are generated and observable. Therefore, KL15 is likely to kill the treated SW480 cells through the necrotic pathway similar to some recently identified AMPs by others.
Insights
The novel antimicrobial peptide (AMP) KL15 exhibits potent bactericidal and anticancer activities. KL15 effectively targets pathogenic bacteria and human cancer cells, inducing cell death through membrane permeabilization and necrosis.
Area of Science:
- Biochemistry
- Molecular Biology
- Antimicrobial Research
Background:
- Antimicrobial peptides (AMPs) are crucial components of innate immunity.
- Bacteriocins, a class of AMPs, show potential for therapeutic applications.
- In silico modification offers a method to enhance peptide properties.
Purpose of the Study:
- To investigate the antimicrobial and anticancer activities of a novel AMP, KL15.
- To characterize the mechanism of action of KL15 against bacterial and cancer cells.
- To evaluate the selectivity of KL15 towards cancer cells versus normal cells.
Main Methods:
- In silico design and modification of antimicrobial peptide KL15.
- Antimicrobial activity assays against pathogenic bacteria (Listeria, Escherichia, Bacillus, Staphylococcus, Enterococcus).
- Anticancer activity assays against human adenocarcinoma cells (SW480, Caco-2) and normal mammary epithelial cells (H184B5F5/M10) using IC50 determination.
- Circular dichroism spectroscopy to determine peptide conformation.
- Flow cytometry and confocal microscopy to assess cell membrane permeability.
- Scanning electron microscopy to observe cellular morphology changes.
Main Results:
- KL15 demonstrated significant bactericidal effects against multiple pathogenic bacteria.
- KL15 exhibited potent cytotoxicity against SW480 and Caco-2 cancer cells with IC50 values of 50 μg/ml (26.3 μM).
- KL15 showed a higher IC50 (150 μg/ml) for normal human mammary epithelial cells, indicating selectivity.
- Circular dichroism spectroscopy revealed a helical conformation of KL15 in solution and lipid environments.
- Flow cytometry, confocal microscopy, and scanning electron microscopy indicated that KL15 enhances cell membrane permeability, leading to cell membrane pore formation and altered cell morphology.
Conclusions:
- The novel antimicrobial peptide KL15 possesses significant antimicrobial and anticancer properties.
- KL15's mechanism of action involves inducing cell membrane permeabilization and pore formation, leading to cell death via necrosis.
- KL15 demonstrates selectivity towards cancer cells over normal human cells, suggesting therapeutic potential.
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