Design, Synthesis And Activity Evaluation Of Penetrating Peptide-drug

As biological penetration enhancers, penetrating peptides are an effective way to promote the drug through the skin or cells by covalent binding or physical mixing with drugs, this can increase efficacy, obtain new physical and chemical properties as well as synergy. Because of its advantages, such as excellent biocompatibility, penetration effect, high transfer efficiency, no limitations, and so on, it offers a new direction and idea for the development of the field of drug delivery, so penetration peptides have important significance in the field of cancer therapy, biology and medicine and so on.Penetration peptide TD-1 was discovered through phage display technology, it is a cationic cyclic peptides with 11 amino acid, when physically mixed insulin and other protein molecules, can promote them through the skin into the systemic circulation and thus exert biological effects. Our group design and synthesis peptide TD-34 based on TD-1 by double lysine replacing peptide backbone, showing better transmembrane and transdermal penetration activity. In this study, penetration peptide TD-34 as the research object, select two anti-cancer drugs, doxorubicin (DOX) and methotrexate (MTX) as model medicine. Firstly, penetration peptide TD-34 was synthesized by solid-phase synthesis, covalently connect TD-34 and two small molecule compounds by chemical means, analysis purity by reversed-phase high-performance liquid chromatography, use electrospray ionization mass spectrometry to identify the molecular weight, results shown it’s consistent with the calculated molecular weight, proving that penetration peptide TD-34 as well as two conjugates are successful synthesized.Cytotoxicity assays were used to detect the cytotoxicity of TD-34, mixtures and covalent conjugates on three tumors cell lines (MCF-7, MDA-MB-231, Caco-2). Results show that two modes are more toxic than individual drug, but did not show significant differences between the two modes. After increasing mixture ratio of penetration peptide in mixtures, significantly enhanced cytotoxicity and overcome resistance of cancer cells, and produce more effects than sensitive cell.Tumor cells’uptake and retention of doxorubicin is obsered and measured by inverted fluorescence microscopy and flow cytometry, qualitative and quantitative analysis and comparison respectively. It was found that TD-34 promote doxorubicin’cellular uptake and retention in MCF-7 (human breast cancer cell line) and Caco-2(human colonic cancer cells), mainly through the interaction with the cell membraneand. In Caco-2 cells, the integrity of the cell monolayer is damaged, increasing the surface roughness was observed, explain that TD-34 may open the connection structure between Caco-2 cells, promote drug absorption through the bypass route.