Targeted antitumor therapy using peptide bioconjugation

Peptides, have being explored for their broad spectrum of action in various diseases, including about its anticancer activity. Anticancer peptides have cationic nature, which allow their interaction with the anionic components of the membranes of the target cells. For example, melittin, a molecule well described, shows activity in several tumor cell lines, however it exhibits non-specific toxicity against normal cells. Therefore, bioconjugation strategies with aptamer peptides, a term derived from the analogy with DNA and RNA aptamers, are molecules that show target protein recognition properties and can be used to overcome the limitations arising from the low specificity and high toxicity of melittin. Aptamers are molecules with affinity for membrane proteins, such as CD44, which are overexpressed in various tumor cell lines. Then, the objective of this work was to synthesize peptides bioconjugated between melittin and aptamer peptides; evaluate the cytotoxic activity of these peptides and bioconjugates and analyze the binding capacity of aptamers with membranes in breast adenocarcinoma (MCF-7, ATCC HTB-22), lung adenocarcinoma (A549, ATCC CCL-185) and non-tumor lung (MRC-5, ATCC CCL-171) cell lines. Peptides were obtained by Solid Phase Peptide Synthesis and the molecules were purified by High Performance Liquid Chromatography. Cell viability assays were performed using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) technique, and the affinity and quantification of the aptamers were observed using automated microscopy and flow cytometry assays. Three synthesis strategies were considered for the bioconjugate synthesis, with the aptamer linked to the N-terminus or C-terminus of melittin, and one synthesis in which the aptamer and melittin were linked by a spacer to analyze whether this spacing could improve the mobility of this molecule. Analysis by automated microscopy and flow cytometry showed that the aptamers have affinity to the membrane at different concentrations tested. In the cell viability assay, it was observed that the bioconjugates have selectivity, since they show cytotoxic activity only in tumor cell lines, while cell viability in the non-tumor cell line remains high. With the results obtained, it is possible to observe that the bioconjugation between antitumor peptides with aptamers can be an excellent strategy visualizing a greater specificity for the treatment of different types of cancer.