Novel production method of innovative antiangiogenic and antitumor small peptides in Escherichia coli.
Background
Developing innovative drugs with potent efficacy, specificity, and high safety remains an ongoing task in antitumor therapy development. In the last few years, peptide drugs have become attractive agents in cancer therapy. HM-3, mainly with antiangiogenic effect, and AP25, with an additional antiproliferative effect, are two peptides designed in our laboratory targeting αvβ3 and α5β1 integrins, respectively. The low molecular weight of the two peptides renders their recombinant expression very difficult, and the complicated structure of AP25 makes its chemical synthesis restricted, which presents a big challenge for its development.


Methods
Bifunctional peptides designed by the ligation of HM-3 and AP25, using linkers with different flexibility, were prepared using recombinant DNA technology in Escherichia coli. The fusion peptides were expressed in a modified auto-induction medium based on a mixture of glucose, glycerol, and lactose as carbon substrates and NH4+ as nitrogen source without any amino acid or other elements. Subsequently, the antiangiogenic, antiproliferative, and cell adhesion assays were conducted to evaluate the bioactivity of the two fusion peptides.


Results
The peptides were successfully expressed in a soluble form without any induction, which allows the culture to reach higher cell density before protein expression occurs. Human umbilical vein endothelial cell migration assay and chick embryo chorioallantoic membrane assay showed, at low doses, a significantly increased antiangiogenic effect (>75%) of the purified products compared with the single molecules. Meanwhile, MTT assay confirmed their enhanced antitumor activity against gastric cancer cell line MGC-803; however, no significant effect was observed on hepatoma HepG2 cells and no cytotoxicity on normal human lens epithelial cell SRA01/04 and human epithelial esophageal cells.


Conclusion
Bifunctional molecules with antiangiogenic and antiproliferative effects were obtained by using this technique, which presents an alternative for small peptide production, instead of the conventional chemical method. The increased molecular weight facilitates the peptide expression with a simultaneous improvement in their stability and biological activity.
['Setrerrahmane|Sarra|S|', 'Yu|Jian|J|', 'Hao|Jingchao|J|', 'Zheng|Heng|H|', 'Xu|Hanmei|H|']
D020533:Angiogenesis Inhibitors D000970:Antineoplastic Agents D002448:Cell Adhesion D002460:Cell Line D002465:Cell Movement D049109:Cell Proliferation D004274:DNA, Recombinant D004305:Dose-Response Relationship, Drug D004354:Drug Screening Assays, Antitumor D004926:Escherichia coli D005818:Genetic Engineering D006801:Humans D010455:Peptides D013329:Structure-Activity Relationship
antiangiogenic antitumor innovative drugs bifunctional peptides linkers auto-induction