Killing Effects Of FOXM1c Targeted Lead Peptide P201 To Liver Cancer Cells And Rational Design Of The Peptide Sequences

Liver cancer is one of malignant tumors that threaten human life and health seriously. In China, about 130 thousand people die from liver cancer every year, which makes it as one of the most affected countries. FoxMl, as a special member of Fox family, is overexpressed in human cancer cells, and closely related to advanced cancers, metastasis, proliferation, drug resistance and poor prognosis. Also, multiple oncogenic pathways have been reported to crosstalk with FoxM1. Thus, FoxMl has become an important marker and target for cancer therapy and intervention. P201 peptide, high affinity to FOXM1c-DBD selected from phage display peptide library, indeed could become a potential approach for targeted anticancer drugs.In this study, firstly, killing effects and death pathways of targeted FoxM1 dodecapeptide P201 to human liver cancer cells were investigated primarily. Secondly, rational design of the peptide sequences was conducted as well as key residues. Finally, a optimized peptide on amino acid level was tested fot its killing effects to liver cancer cells.MTT assay and morphological changes revealed that the inhibition rate of HepG-2 cells could increase to 96% at the concentration of 60μg/mL of P201 peptide after 48h, as IC50 value of 25.16μg/mL. For MCF-7 and 293T cells, the inhibition rate could also increase to 64.1% and 61.9% at the concentration of 60μg/mL of P201 peptide after 48h, in a time- and dose-dependent manner within a certain range. Furthermore, AO-EB assay, DNA Ladder and FCM showed that the lethal effect of P201 peptide to HepG-2 cells was related to apoptosis, with possible multiple death pathways for further study.In addition, the 10th residue of P201 peptide was considered as the site for optimization and Asparagine was selected to be new residue in the 10th site through virtual alanine mutagenesis, LibDock, CDOCKER and MVD. Also, five amino acids (1st,2nd,4th,7th and 9th residues) were defined by a combination of motif search, antisense peptide amino acid analysis, virtual alanine mutagenesis and molecular docking.The killing effects of M1Oaa-D peptide to HepG2 cells was significantly below P201 peptide by CCK-8 assay, with the inhibition rate of 26.3% at the concentration of 100 μg/mL after 48h. At low concentration, M10aa-D peptide could even promote cell proliferation. The killing effects of peptides were related to their affinity to FOXM1.All these results provided insights for development of P201 peptide to a novel targeted cancer therapy, in the meantime, indicated that more studies on molecular mechanism and optimization should go on and meet requirements of peptide research for anticancer drugs.