title: Transduction of Recombinant M3-p53-R12 Protein Enhances Human Leukemia Cell Apoptosis.
abstract: Tumor suppressor protein p53 plays important roles in initiating cell cycle arrest and promoting tumor cell apoptosis. Previous studies have shown that p53 is either mutated or defective in approximately 50% of human cancers; therefore restoring normal p53 activity in cancer cells might be an effective anticancer therapeutic approach. Herein, we designed a chimeric p53 protein flanked with the MyoD N-terminal transcriptional activation domain (amino acids 1-62, called M3) and a poly-arginine (R12) cell penetrating signal in its N-and C-termini respectively. This chimeric protein, M3-p53-R12, can be expressed in E. coli and purified using immobilized metal ion chromatography followed by serial refolding dialysis. The purified M3-p53-R12 protein retains DNA-binding activity and gains of cell penetrating ability. Using MTT assay, we demonstrated that M3-p53-R12 inhibited the growth of K562, Jurkat as well as HL-60 leukemia cells carrying mutant p53 genes. Results from FACS analysis also demonstrated that transduction of M3-p53-R12 protein induced cell cycle arrest of these leukemia cells. Of special note, M3-p53-R12 has no apoptotic effect on normal mesenchymal stem cells (MSC) and leukocytes, highlighting its differential effects on normal and tumor cells. To sum up, our results reveal that purified recombinant M3-p53-R12 protein has functions of suppressing the leukemia cell lines' proliferation and launching cell apoptosis, suggesting the feasibility of using M3-p53-R12 protein as an anticancer drug. In the future we will test whether this chimeric protein can preferentially trigger the death of malignant cancer cells without affecting normal cells in animals carrying endogenous or xenographic tumors.
authors.affiliation: 1. Department of Life Sciences, National Central University, Jhongli 32001, Taiwan;; 2. Taiwan Advance Bio-Pharma Inc., New Taipei City 22180, Taiwan.
authors.forename: Tsung Chi
authors.initials: TC
authors.lastname: Lu
authors.affiliation: 1. Department of Life Sciences, National Central University, Jhongli 32001, Taiwan;
authors.forename: Guan-Hao
authors.initials: GH
authors.lastname: Zhao
authors.affiliation: 1. Department of Life Sciences, National Central University, Jhongli 32001, Taiwan;; 3. The Affiliated Jhongli Senior High School of National Central University, Jhongli 32047, Taiwan;
authors.forename: Yao Yun
authors.initials: YY
authors.lastname: Chen
authors.affiliation: 1. Department of Life Sciences, National Central University, Jhongli 32001, Taiwan;
authors.forename: Chia-Ying
authors.initials: CY
authors.lastname: Chien
authors.affiliation: 2. Taiwan Advance Bio-Pharma Inc., New Taipei City 22180, Taiwan.
authors.forename: Chi-Hung
authors.initials: CH
authors.lastname: Huang
authors.affiliation: 4. Department of Biochemistry, Chang Gung University, Taoyuan 333, Taiwan;; 5. Liver Research Center, Chang Gung Memorial Hospital, Linko, Taoyuan 333, Taiwan.
authors.forename: Kwang Hui
authors.initials: KH
authors.lastname: Lin
authors.affiliation: 1. Department of Life Sciences, National Central University, Jhongli 32001, Taiwan;
authors.forename: Shen Liang
authors.initials: SL
authors.lastname: Chen
keywords: MyoD; and protein drug.; apoptosis; cancer; leukemia; p53