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D-Amino Acid Analogues of the Antimicrobial Peptide CDT Exhibit Anti- Cancer Properties in A549, a Human Lung Adenocarcinoma Cell Line

[ Vol. 24 , Issue. 7 ]

Author(s):

Hedeel Guy Evans, Jeffrey W. Guthrie, Murali Jujjavarapu, Nathan Hendrickson, Anna Eitel, Yeji Park, Jennifer Garvey, Rebecca Newman, Daniel Esckilsen and Deborah L. Heyl*   Pages 590 - 598 ( 9 )

Abstract:


Introduction: The importance of the antitumor activity of some antimicrobial peptides (AMPs) is being increasingly recognized. The antimicrobial peptide, tachyplesin, has been shown to exhibit anticancer properties and a linear, cysteine deleted analogue (CDT), was found to retain its antibacterial function.

Objectives: The objective was to test CDT and related analogues against normal mammalian, bacterial, and cancer cells to determine their effectiveness and then utilize specific assays to determine a possible mechanism of action.

Methods: We used sequence reversal and D-amino acids to synthesize four CDT analogues by solid phase peptide synthesis. A number of assays were used including liposome dye-leakage, antibacterial activity against both Gram-positive and Gram-negative bacterial strains, hemolytic assays, methyl thiazolyl tetrazolium (MTT), and apoptosis to examine their effectiveness as both AMPs and anti-cancer peptides (ACPs). We then tested the analogues for their ability to inhibit proliferation of the human lung cancer cell line, A549.

Results: We found that D-CDT exhibited the best bactericidal properties of those tested and was not damaging to red blood cells. Both D-CDT and reverse D-CDT showed a dose-dependent reduction of cell viability. However, D-CDT was most effective with an IC50 of 9.814 μM, a value 9-fold lower than that calculated for reverse D-CDT (90.16 μM). Apoptosis does not appear to be a mechanism by which D-CDT exerts its anticancer properties since > 100 μM was required to increase activation of caspase 3. Moreover, the ERK1/2 pathway is also unlikely since only a modest (20%) decrease of activity was observed with > 100 μM D-CDT. However, D-CDT was found to operate via a hyaluronan (HA)-dependent mechanism as pretreatment of the cells with hyaluronidase decreased the cytotoxic effects of D-CDT on A549 cells and increased its IC50 29-fold to 283.9 μM.

Conclusion: D-CDT is both an effective AMP and ACP, and likely exerts its anticancer effects through both membranolytic as well as an HA-mediated mechanism.

Keywords:

Antimicrobial, anticancer, peptide, A549, hyaluronidase, MTT assay, hemolytic.

Affiliation:

Department of Chemistry, Eastern Michigan University, Ypsilanti, MI 48197, Department of Chemistry, Eastern Michigan University, Ypsilanti, MI 48197, Department of Chemistry, Eastern Michigan University, Ypsilanti, MI 48197, Department of Chemistry, Eastern Michigan University, Ypsilanti, MI 48197, Department of Chemistry, Eastern Michigan University, Ypsilanti, MI 48197, Department of Chemistry, Eastern Michigan University, Ypsilanti, MI 48197, Department of Chemistry, Eastern Michigan University, Ypsilanti, MI 48197, Department of Chemistry, Eastern Michigan University, Ypsilanti, MI 48197, Department of Chemistry, Eastern Michigan University, Ypsilanti, MI 48197, Department of Chemistry, 541 Science Complex, Eastern Michigan University, Ypsilanti, MI 48197

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