10.6084/m9.figshare.5632279 Xiufeng Liu Xiufeng Liu Xintong Liu Xintong Liu Suwen Sunchen Suwen Sunchen Meixia Liu Meixia Liu Chen Shen Chen Shen Juanjuan Wu Juanjuan Wu Wanli Zhao Wanli Zhao Boyang Yu Boyang Yu Jihua Liu Jihua Liu A novel tumor-activated ALA fusion protein for specific inhibition on the growth and invasion of breast cancer cells MDA-MB-231 Taylor & Francis Group 2017 Fusion protein tumor activate uPA AGAP apoptosis 2017-11-25 04:53:48 Journal contribution https://tandf.figshare.com/articles/journal_contribution/A_novel_tumor-activated_ALA_fusion_protein_for_specific_inhibition_on_the_growth_and_invasion_of_breast_cancer_cells_MDA-MB-231/5632279 <p><b>Objective:</b> The aim of this research was to develop a novel ALA fusion protein for target to the malignant cells surface with high uPAR expression and locally release of the scorpion toxin AGAP in an uPA-cleavable manner. It will provide an effective approach for controlled release of the peptide toxins to treat cancerous cells.</p> <p><b>Methods:</b> The ALA fusion proteins were expressed in pichia pastoris, and the recombinant proteins were purified by Ni-NTA affinity chromatography. The proteins were added to human breast cancer cells (MDA-MB-231) and human embryonic kidney cells (HEK-293) in order to investigate the characteristic of selective targeting and releasing of scorpion toxin AGAP in cancer cells with high uPAR expression. The inhibitory effect of ALA on MDA-MB-231, MCF7, LO2 and HEK-293 was evaluated by MTT assay. Moreover, the antiproliferation mechanism of ALA was determined by flow cytometric and western blot analysis.</p> <p><b>Results:</b> The results showed that ALA could target MDA-MB-231 cells and the scorpion toxin AGAP could be released with high efficiency and selectivity. ALA inhibited the growth and invasion of breast cancer cells MDA-MB231. Also, cell apoptosis pathway was found to be associated with the inhibition mechanism of ALA according to the data of flow cytometric and western blot analysis. Therefore, ALA could be a novel antitumor candidate for targeting treatment of malignant cell.</p> <p><b>Conclusions:</b> This study successfully demonstrated that fusion of biotoxins with tumor target domain could provide a simple yet effective way to delivery of peptide or protein drugs.</p>