title: Inhibition of cell proliferation, migration and invasion by a glioma-targeted fusion protein combining the p53 C terminus and MDM2-binding domain.
abstract: OBJECTIVE
  The aim of this study was to develop multifunctional fusion proteins for targeting and delivering therapy elements into glioma cells.
  
  
  METHODS
  Multifunctional fusion proteins were expressed in Escherichia coli and purified using Ni-NTA resin affinity chromatography. Human glioma cells and primary astrocytes were used to analyse their functions. Targeting proteins location to glioma cells was observed by confocal microscopy. Effects of cell viability and proliferation were evaluated using the Cell Counting Kit 8 and colony formation assays. Glioma cell migration and invasion were assessed using transwell assays, and apoptosis was analysed by flow cytometry. In addition, changes in expression of proteins related to the cell cycle and apoptosis were determined by Western blotting.
  
  
  RESULTS
  The protein with highest bioactivity was GL1-riHA2-p53c+m-TAT (GHPc+mT), which combines glioma-targeting peptide GL1 (G), and C terminus (Pc) and mouse double minute domains (Pm) of p53, with the destabilizing lipid membrane peptide riHA2 (H) and cell-penetrating peptide TAT (T). The purified fusion protein was stable in cell culture medium and specifically targeted, and was internalized by, epidermal growth factor receptor (EGFR)-overexpressing glioma cells (U87ΔEGFR). It inhibited cell proliferation, migration and invasion, while flow cytometric analysis showed increased apoptosis. In addition, GHPc+mT caused significant changes in expression of proteins related to the cell cycle and apoptosis.
  
  
  CONCLUSIONS
  GHPc+mT is a multifunctional protein combining targeting, inhibition of glioma cell proliferation and induction of apoptosis, providing some potential to be developed into an effective protein drug delivery system for glioma therapy.
authors: Department of Biotechnology, Dalian Medical University, Dalian, 116044, China.|Department of Hematology, The First Affiliated Hospital of Dalian Medical University, Dalian, 116011, China.
authors: Jiawen
authors: J
authors: Yu
authors: Department of Biotechnology, Dalian Medical University, Dalian, 116044, China.
authors: Meihua
authors: M
authors: Guo
authors: Department of Biotechnology, Dalian Medical University, Dalian, 116044, China.
authors: Ting
authors: T
authors: Wang
authors: Department of Biotechnology, Dalian Medical University, Dalian, 116044, China.
authors: Xiang
authors: X
authors: Li
authors: Department of Biotechnology, Dalian Medical University, Dalian, 116044, China.
authors: Dan
authors: D
authors: Wang
authors: Department of Biotechnology, Dalian Medical University, Dalian, 116044, China.
authors: Xinying
authors: X
authors: Wang
authors: Department of Biotechnology, Dalian Medical University, Dalian, 116044, China.
authors: Qian
authors: Q
authors: Zhang
authors: Department of Biotechnology, Dalian Medical University, Dalian, 116044, China.
authors: Liang
authors: L
authors: Wang
authors: Department of Biotechnology, Dalian Medical University, Dalian, 116044, China.
authors: Yang
authors: Y
authors: Zhang
authors: College of Life Sciences, Liaoning Normal University, Dalian, 116029, China.
authors: Chunhui
authors: C
authors: Zhao
authors: Department of Biotechnology, Dalian Medical University, Dalian, 116044, China.
authors: Bin
authors: B
authors: Feng
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D005910:Glioma / Q000473:pathology*; D006801:Humans; D009361:Neoplasm Invasiveness; D011485:Protein Binding / Q000187:drug effects; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000494:pharmacology*; D014410:Tumor Stem Cell Assay; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D011993:Recombinant Fusion Proteins; D016159:Tumor Suppressor Protein p53