Yu-Qian Zhao1, Yi-Qiong Yin2, Jie Liu2, Gui-Hua Wang1, Jian Huang1, Ling-Juan Zhu1, Jin-Hui Wang1,3. 1. School of Traditional Chinese Materia Medica, Shenyang Pharmaceutical University, Shenyang 110016, China. 2. Department of Gastrointestinal Surgery, State Key Laboratory of Biotherapy, Collaborative Innovation Center of Biotherapy, West China Hospital, Sichuan University, Chengdu 610041, China. 3. School of Pharmacy, Shihezi University, Shihezi 832002, China.
Abstract
AIM: Pim-2 is a short-lived serine/threonine kinase, which plays a key role in metastasis of breast cancer through persistent activation of STAT3. Although the crystal structure of Pim-2 has been reported, but thus far no specific Pim-2-targeted compounds have been reported. In this study, we identified a novel Pim-2 inhibitor, HJ-PI01, by in silico analysis and experimental validation. METHODS: The protein-protein interaction (PPI) network, chemical synthesis, molecular docking, and molecular dynamics (MD) simulations were used to design and discover the new Pim-2 inhibitor HJ-PI01. The anti-tumor effects of HJ-PI01 were evaluated in human breast MDA-MB-231, MDA-MB-468, MDA-MB-436, MCF-7 cells in vitro and in MDA-MB-231 xenograft mice, which were treated with HJ-PI01 (40 mg·kg(-1)·d(-1), ig) with or without lienal polypeptide (50 mg·kg(-1)·d(-1), ip) for 10 d. The apoptosis/autophage-inducing mechanisms of HJ-PI01 were elucidated using Western blots, immunoblots, flow cytometry, transmission electron microscopy and fluorescence microscopy. RESULTS: Based on the PrePPI network, the potential partners interacting with Pim-2 in regulating apoptosis (160 protein pairs) and autophagy (47 protein pairs) were identified. Based on the structural characteristics of Pim-2, a total of 15 compounds (HJ-PI01 to HJ-P015) were synthesized, which showed moderate or remarkable anti-proliferative potency in the human breast cancer cell lines tested. The most effective compound HJ-PI01 exerted a robust inhibition on MDA-MB-231 cells compared with chlorpromazine and the pan-Pim inhibitor PI003. Molecular dynamics (MD) simulation revealed that HJ-PI01 had a good binding score with Pim-2. Moreover, HJ-PI01 (300 nmol/L) induced death receptor-dependent and mitochondrial apoptosis as well as autophagic death in MDA-MB-231 cells. In MDA-MB-231 xenograft mice, administration of HJ-PI01 remarkably inhibited the tumor growth and induced tumor cell apoptosis in vivo. Co-administration of HJ-PI01 with lienal polypeptide could improve the anti-tumor activity of HJ-PI01 and reduce its toxicity. CONCLUSION: The newly synthesized compound, HJ-PI01, can induce death receptor/mitochondrial apoptosis and autophagic cell death by targeting Pim-2 in human breast cancer cells in vitro and in vivo.
AIM: Pim-2 is a short-lived serine/threonine kinase, which plays a key role in metastasis of breast cancer through persistent activation of STAT3. Although the crystal structure of Pim-2 has been reported, but thus far no specific Pim-2-targeted compounds have been reported. In this study, we identified a novel Pim-2 inhibitor, HJ-PI01, by in silico analysis and experimental validation. METHODS: The protein-protein interaction (PPI) network, chemical synthesis, molecular docking, and molecular dynamics (MD) simulations were used to design and discover the new Pim-2 inhibitor HJ-PI01. The anti-tumor effects of HJ-PI01 were evaluated in human breast MDA-MB-231, MDA-MB-468, MDA-MB-436, MCF-7 cells in vitro and in MDA-MB-231 xenograft mice, which were treated with HJ-PI01 (40 mg·kg(-1)·d(-1), ig) with or without lienal polypeptide (50 mg·kg(-1)·d(-1), ip) for 10 d. The apoptosis/autophage-inducing mechanisms of HJ-PI01 were elucidated using Western blots, immunoblots, flow cytometry, transmission electron microscopy and fluorescence microscopy. RESULTS: Based on the PrePPI network, the potential partners interacting with Pim-2 in regulating apoptosis (160 protein pairs) and autophagy (47 protein pairs) were identified. Based on the structural characteristics of Pim-2, a total of 15 compounds (HJ-PI01 to HJ-P015) were synthesized, which showed moderate or remarkable anti-proliferative potency in the humanbreast cancer cell lines tested. The most effective compound HJ-PI01 exerted a robust inhibition on MDA-MB-231 cells compared with chlorpromazine and the pan-Pim inhibitor PI003. Molecular dynamics (MD) simulation revealed that HJ-PI01 had a good binding score with Pim-2. Moreover, HJ-PI01 (300 nmol/L) induced death receptor-dependent and mitochondrial apoptosis as well as autophagic death in MDA-MB-231 cells. In MDA-MB-231 xenograft mice, administration of HJ-PI01 remarkably inhibited the tumor growth and induced tumor cell apoptosis in vivo. Co-administration of HJ-PI01 with lienal polypeptide could improve the anti-tumor activity of HJ-PI01 and reduce its toxicity. CONCLUSION: The newly synthesized compound, HJ-PI01, can induce death receptor/mitochondrial apoptosis and autophagic cell death by targeting Pim-2 in humanbreast cancer cells in vitro and in vivo.
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