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| Establishment and Application of High-throughput Screening Model for SHP Inhibitors |
| GAO Wei1, SHAO Wei2, SU Gang2, LI Jia3, XIE Xiaodong2 |
1. Department of General Thoracic Surgery, Lanzhou General Hospital of Lanzhou Military Command, Lanzhou 730050, China;
2. Key Lab of Preclinical Study for New Drugs of Gansu Province, School of Basic Medical Science of Lanzhou University, Lanzhou 730020, China;
3. National Center for Drug Screening, Shanghai Institute of Materia Medica, Chinese Academy of Science, Shanghai 201203, China |
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Abstract: The aims of this study is to set up a high throughput screening model for SHP2 inhibitors and screen the potential inhibitors. By the E. coil expression system GST-SHP2 fusion protein is cloned and over expressed. High-purity SHP2 protein is purified though GSTSepharose column. After establishing the high through screening system with the colorimetric assay of SHP2, a library of 48000 pure compounds is screened using the 384 micro- plate. Among them 75 compounds have inhibitory effects over 50%. Ultimately, three inhibitors are identified as SHP2 inhibitors with high activity. The high throughput screening is a highly sensitive, inexpensive, and operationally simple assay method in identifying SHP2 inhibitors.
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Received: 10 August 2013
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[1] Lai L A, Zhao C, Zhang E E, et al. The Shp-2 tyrosine phosphatase[J]. Protein Phosphatases. 2004, 5: 275-299.
[2] Neel B G, Gu H, Pao L. The 'Shp'ing news: SH2 domain-containing tyrosine phosphatases in cell signaling[J]. Trends in Biochemical Sciences, 2003, 28(6): 284-293.
[3] Cunnick J M, Meng S, Ren Y, et al. Regulation of the mitogen-activated protein kinase signaling pathway by SHP2[J]. Journal of Biological Chemistry, 2002, 277(11): 9498-9504.
[4] QU C K. The SHP-2 tyrosine phosphatase: Signaling mechanisms and biological functions[J]. Cell Research, 2000, 10(4): 279-288.
[5] Chan R J, Feng G S. PTPN11 is the first identified proto-oncogene that encodes a tyrosine phosphatase[J]. Blood, 2007, 109(3): 862-867.
[6] Tartaglia M, Zampino G, Gelb B D. Noonan syndrome: Clinical aspects and molecular pathogenesis[J]. Molecular Syndromology, 2010, 1(1): 2- 26.
[7] Edouard T, Combier J P, Nédélec A, et al. Functional effects of PTPN11 (SHP2) mutations causing LEOPARD syndrome on epidermal growth factor-induced phosphoinositide 3-kinase/AKT/glycogen synthase kinase 3β signaling[J]. Molecular and Cellular Biology, 2010, 30(10): 2498-2507.
[8] Grossmann K S, Rosário M, Birchmeier C, et al. The tyrosine phosphatase SHP2 in development and cancer[J]. Advances in Cancer Research, 2010, 106: 53-89.
[9] Bentires-Alj M, Paez J G, David F S, et al. Activating mutations of the noonan syndrome-associated SHP2/PTPN11 gene in human solid tumors and adult acute myelogenous leukemia[J]. Cancer Research, 2004, 64 (24): 8816-8820.
[10] Christiansen D H, Desta F, Andersen M K, et al. Mutations of the PTPN11 gene in therapy-related MDS and AML with rare balanced chromosome translocations[J]. Genes, Chromosomes and Cancer, 2007, 46(6): 517-521.
[11] Yu Z H, Chen L, Wu L, et al. Small molecule inhibitors of SHP2 tyrosine phosphatase discovered by virtual screening[J]. Bioorganic & Medicinal Chemistry Letters, 2011, 21(14): 4238-4242.
[12] Mayr L M, Bojanic D. Novel trends in high-throughput screening[J]. Current Opinion in Pharmacology, 2009, 9(5): 580-588.
[13] Dechert U, Adam M, Harder K W, et al. Characterization of protein tyrosine phosphatase SH - PTP2. Study of phosphopeptide substrates and possible regulatory role of SH2 domains[J]. Journal of Biological Chemistry, 1994, 269(8): 5602-5611.
[14] ZHANG J, Chung T D Y, Oldenburg K R. Validation of high throughput screening assays[J]. Journal of Biomolecular Screening, 1999, 4(2): 67-73. |
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2014, Vol. 32 
