Developing and Testing Methylated Nano-Structured Dipeptides that Inhibit Src Kinase Activity in vitro for Anti-Cancer Applications
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Abstract
Here, with the aim of developing a novel anti-cancer treatment, seven dipeptides were designed that contained methylated tryptophan and/or methylated arginine and were produced using Fmoc solid-phase peptide synthesis. Overexpression of the Src tyrosine kinase enzyme has been implicated in the development of different cancers. Dipeptides containing unnatural amino acids such as methylated arginine (RCH3), dimethylated arginine (R(CH3)2), and/or methylated tryptophan (WCH3) residues have been earlier shown to inhibit the Src kinase enzyme. In this study, three such dipeptides W-RCH3, WCH3-RCH3, and W-R(CH3)2 were tested and were found to have IC50 values (the concentration at which 50% inhibition occurs) of 510 nM, 916 nM, and 1 μM, respectively, in acellular assays. The IC50 values of these dipeptides are considered low as in nanomolar concentrations compared to the cyclic penta to nano W-R ([W-R]5-[W-R]9) that are in the micromolar concentration scale for Src kinase.1 The IC50 values of these cyclic peptides are 0.81, 0.57, 0.35, 0.33, and 0.21 µM, respectively. 1 However, the unmethylated versions of these peptides did not show any inhibitory activity against Src kinase. All of these dipeptides (50 mM) did not show any cytotoxicity upon incubation up to 72 h with three different cancer cell lines, including leukemia (CCRF-CEM), breast adenocarcinoma(MDA-MB-231), and ovarian adenocarcinoma (SK-OV-3) cell lines, indicating the limited permeability of the peptides through the cell membrane. Further study is therefore needed to improve the permeability of these peptides for anticancer applications such as using a peptide carrier or additional peptide functionalization. In summary, this study provides a protocol to synthesize and test peptides that inhibit Src kinase activity and, thus, possess promising anticancer ability as demonstrated using acellular and cellular assays.
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