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Molecular Cancer
Published in: Molecular Cancer 1/2010

Open Access 01-12-2010 | Research

RNAi phenotype profiling of kinases identifies potential therapeutic targets in Ewing's sarcoma

Authors: Shilpi Arora, Irma M Gonzales, R Tanner Hagelstrom, Christian Beaudry, Ashish Choudhary, Chao Sima, Raoul Tibes, Spyro Mousses, David O Azorsa

Published in: Molecular Cancer | Issue 1/2010

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Abstract

Background

Ewing's sarcomas are aggressive musculoskeletal tumors occurring most frequently in the long and flat bones as a solitary lesion mostly during the teen-age years of life. With current treatments, significant number of patients relapse and survival is poor for those with metastatic disease. As part of novel target discovery in Ewing's sarcoma, we applied RNAi mediated phenotypic profiling to identify kinase targets involved in growth and survival of Ewing's sarcoma cells.

Results

Four Ewing's sarcoma cell lines TC-32, TC-71, SK-ES-1 and RD-ES were tested in high throughput-RNAi screens using a siRNA library targeting 572 kinases. Knockdown of 25 siRNAs reduced the growth of all four Ewing's sarcoma cell lines in replicate screens. Of these, 16 siRNA were specific and reduced proliferation of Ewing's sarcoma cells as compared to normal fibroblasts. Secondary validation and preliminary mechanistic studies highlighted the kinases STK10 and TNK2 as having important roles in growth and survival of Ewing's sarcoma cells. Furthermore, knockdown of STK10 and TNK2 by siRNA showed increased apoptosis.

Conclusion

In summary, RNAi-based phenotypic profiling proved to be a powerful gene target discovery strategy, leading to successful identification and validation of STK10 and TNK2 as two novel potential therapeutic targets for Ewing's sarcoma.
Appendix
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Literature
1.
Khoury JD: Ewing sarcoma family of tumors. Adv Anat Pathol. 2005, 12: 212-220. 10.1097/01.pap.0000175114.55541.52CrossRefPubMed
2.
Ludwig JA: Ewing sarcoma: historical perspectives, current state-of-the-art, and opportunities for targeted therapy in the future. Curr Opin Oncol. 2008, 20: 412-418. 10.1097/CCO.0b013e328303ba1dCrossRefPubMed
3.
Riggi N, Stamenkovic I: The Biology of Ewing sarcoma. Cancer Lett. 2007, 254: 1-10. 10.1016/j.canlet.2006.12.009CrossRefPubMed
4.
Ginsberg JP, de Alava E, Ladanyi M, Wexler LH, Kovar H, Paulussen M, Zoubek A, Dockhorn-Dworniczak B, Juergens H, Wunder JS: EWS-FLI1 and EWS-ERG gene fusions are associated with similar clinical phenotypes in Ewing's sarcoma. J Clin Oncol. 1999, 17: 1809-1814.PubMed
5.
Llombart-Bosch A, Navarro S: Immunohistochemical detection of EWS and FLI-1 proteinss in Ewing sarcoma and primitive neuroectodermal tumors: comparative analysis with CD99 (MIC-2) expression. Appl Immunohistochem Mol Morphol. 2001, 9: 255-260. 10.1097/00022744-200109000-00010PubMed
6.
Scotlandi K, Baldini N, Cerisano V, Manara MC, Benini S, Serra M, Lollini PL, Nanni P, Nicoletti G, Bernard G: CD99 engagement: an effective therapeutic strategy for Ewing tumors. Cancer Res. 2000, 60: 5134-5142.PubMed
7.
Grier HE, Krailo MD, Tarbell NJ, Link MP, Fryer CJ, Pritchard DJ, Gebhardt MC, Dickman PS, Perlman EJ, Meyers PA: Addition of ifosfamide and etoposide to standard chemotherapy for Ewing's sarcoma and primitive neuroectodermal tumor of bone. N Engl J Med. 2003, 348: 694-701. 10.1056/NEJMoa020890CrossRefPubMed
8.
Miser JS, Krailo MD, Tarbell NJ, Link MP, Fryer CJ, Pritchard DJ, Gebhardt MC, Dickman PS, Perlman EJ, Meyers PA: Treatment of metastatic Ewing's sarcoma or primitive neuroectodermal tumor of bone: evaluation of combination ifosfamide and etoposide--a Children's Cancer Group and Pediatric Oncology Group study. J Clin Oncol. 2004, 22: 2873-2876. 10.1200/JCO.2004.01.041CrossRefPubMed
9.
Joo J, Christensen L, Warner K, States L, Kang HG, Vo K, Lawlor ER, May WA: GLI1 is a central mediator of EWS/FLI1 signaling in Ewing tumors. PLoS One. 2009, 4: e7608- 10.1371/journal.pone.0007608PubMedCentralCrossRefPubMed
10.
Herrero-Martin D, Osuna D, Ordonez JL, Sevillano V, Martins AS, Mackintosh C, Campos M, Madoz-Gurpide J, Otero-Motta AP, Caballero G: Stable interference of EWS-FLI1 in an Ewing sarcoma cell line impairs IGF-1/IGF-1R signalling and reveals TOPK as a new target. Br J Cancer. 2009, 101: 80-90. 10.1038/sj.bjc.6605104PubMedCentralCrossRefPubMed
11.
Ordonez JL, Osuna D, Herrero D, de Alava E, Madoz-Gurpide J: Advances in Ewing's sarcoma research: where are we now and what lies ahead?. Cancer Res. 2009, 69: 7140-7150. 10.1158/0008-5472.CAN-08-4041CrossRefPubMed
12.
Luo W, Gangwal K, Sankar S, Boucher KM, Thomas D, Lessnick SL: GSTM4 is a microsatellite-containing EWS/FLI target involved in Ewing's sarcoma oncogenesis and therapeutic resistance. Oncogene. 2009, 28: 4126-4132. 10.1038/onc.2009.262CrossRefPubMed
13.
Anderson PM, Pearson M: Novel therapeutic approaches in pediatric and young adult sarcomas. Curr Oncol Rep. 2006, 8: 310-315. 10.1007/s11912-006-0038-0CrossRefPubMed
14.
Nencioni A, Sandy P, Dillon C, Kissler S, Blume-Jensen P, Van Parijs L: RNA interference for the identification of disease-associated genes. Curr Opin Mol Ther. 2004, 6: 136-140.PubMed
15.
MacKeigan JP, Murphy LO, Blenis J: Sensitized RNAi screen of human kinases and phosphatases identifies new regulators of apoptosis and chemoresistance. Nat Cell Biol. 2005, 7: 591-600. 10.1038/ncb1258CrossRefPubMed
16.
Aza-Blanc P, Cooper CL, Wagner K, Batalov S, Deveraux QL, Cooke MP: Identification of modulators of TRAIL-induced apoptosis via RNAi-based phenotypic screening. Mol Cell. 2003, 12: 627-637. 10.1016/S1097-2765(03)00348-4CrossRefPubMed
17.
Azorsa DO, Gonzales IM, Basu GD, Choudhary A, Arora S, Bisanz KM, Kiefer JA, Henderson MC, Trent JM, Von Hoff DD, Mousses S: Synthetic lethal RNAi screening identifies sensitizing targets for gemcitabine therapy in pancreatic cancer. J Transl Med. 2009, 7: 43- 10.1186/1479-5876-7-43PubMedCentralCrossRefPubMed
18.
Whitehurst AW, Bodemann BO, Cardenas J, Ferguson D, Girard L, Peyton M, Minna JD, Michnoff C, Hao W, Roth MG: Synthetic lethal screen identification of chemosensitizer loci in cancer cells. Nature. 2007, 446: 815-819. 10.1038/nature05697CrossRefPubMed
19.
Hu K, Lee C, Qiu D, Fotovati A, Davies A, Abu-Ali S, Wai D, Lawlor ER, Triche TJ, Pallen CJ, Dunn SE: Small interfering RNA library screen of human kinases and phosphatases identifies polo-like kinase 1 as a promising new target for the treatment of pediatric rhabdomyosarcomas. Mol Cancer Ther. 2009, 8: 3024-3035. 10.1158/1535-7163.MCT-09-0365PubMedCentralCrossRefPubMed
20.
Birmingham A, Selfors LM, Forster T, Wrobel D, Kennedy CJ, Shanks E, Santoyo-Lopez J, Dunican DJ, Long A, Kelleher D: Statistical methods for analysis of high-throughput RNA interference screens. Nat Methods. 2009, 6: 569-575. 10.1038/nmeth.1351PubMedCentralCrossRefPubMed
21.
Solly K, Wang X, Xu X, Strulovici B, Zheng W: Application of real-time cell electronic sensing (RT-CES) technology to cell-based assays. Assay Drug Dev Technol. 2004, 2: 363-372. 10.1089/adt.2004.2.363CrossRefPubMed
22.
Subbiah V, Anderson P, Lazar AJ, Burdett E, Raymond K, Ludwig JA: Ewing's sarcoma: standard and experimental treatment options. Curr Treat Options Oncol. 2009, 10: 126-140. 10.1007/s11864-009-0104-6CrossRefPubMed
23.
Andersson MK, Aman P: Proliferation of Ewing sarcoma cell lines is suppressed by the receptor tyrosine kinase inhibitors gefitinib and vandetanib. Cancer Cell Int. 2008, 8: 1- 10.1186/1475-2867-8-1PubMedCentralCrossRefPubMed
24.
Sabbatini P, Korenchuk S, Rowand JL, Groy A, Liu Q, Leperi D, Atkins C, Dumble M, Yang J, Anderson K: GSK1838705A inhibits the insulin-like growth factor-1 receptor and anaplastic lymphoma kinase and shows antitumor activity in experimental models of human cancers. Mol Cancer Ther. 2009, 8: 2811-2820. 10.1158/1535-7163.MCT-09-0423CrossRefPubMed
25.
Zenali MJ, Zhang PL, Bendel AE, Brown RE: Morphoproteomic confirmation of constitutively activated mTOR, ERK, and NF-kappaB pathways in Ewing family of tumors. Ann Clin Lab Sci. 2009, 39: 160-166.PubMed
26.
Wakahara K, Ohno T, Kimura M, Masuda T, Nozawa S, Dohjima T, Yamamoto T, Nagano A, Kawai G, Matsuhashi A: EWS-Fli1 up-regulates expression of the Aurora A and Aurora B kinases. Mol Cancer Res. 2008, 6: 1937-1945. 10.1158/1541-7786.MCR-08-0054CrossRefPubMed
27.
Javelaud D, Besancon F: NF-kappa B activation results in rapid inactivation of JNK in TNF alpha-treated Ewing sarcoma cells: a mechanism for the anti-apoptotic effect of NF-kappa B. Oncogene. 2001, 20: 4365-4372. 10.1038/sj.onc.1204570CrossRefPubMed
28.
Hotfilder M, Sondermann P, Senss A, van Valen F, Jurgens H, Vormoor J: PI3K/AKT is involved in mediating survival signals that rescue Ewing tumour cells from fibroblast growth factor 2-induced cell death. Br J Cancer. 2005, 92: 705-710. 10.1038/sj.bjc.6602384PubMedCentralCrossRefPubMed
29.
Wishart MJ, Dixon JE: Gathering STYX: phosphatase-like form predicts functions for unique protein-interaction domains. Trends Biochem Sci. 1998, 23: 301-306. 10.1016/S0968-0004(98)01241-9CrossRefPubMed
30.
Siligan C, Ban J, Bachmaier R, Spahn L, Kreppel M, Schaefer KL, Poremba C, Aryee DN, Kovar H: EWS-FLI1 target genes recovered from Ewing's sarcoma chromatin. Oncogene. 2005, 24: 2512-2524. 10.1038/sj.onc.1208455CrossRefPubMed
31.
Uren A, Toretsky JA: Pediatric malignancies provide unique cancer therapy targets. Curr Opin Pediatr. 2005, 17: 14-19. 10.1097/01.mop.0000147904.84978.aeCrossRefPubMed
32.
Yap TA, Papadopoulos K, Fearen I, Carpenter C, Delgado L, Taylor A, Domsch J, Olmos D, Brunetto AT, Tan DS, Decordova S, Heaton SP, Garrett MD, Patnaik A, Sullivan D, Yan L, de Bono JS, Tolcher AW: First-in-man, first-in-class phase I study of two schedules of MK-2206, a novel oral Akt inhibitor, in patients (pts) with advanced solid tumors, including ovarian and castration resistant prostate cancer (CRPC). Molecular Cancer Therapeutics (Meeting abstract supplement). 2009, 8: B135-CrossRef
33.
Rhodes N, Heerding DA, Duckett DR, Eberwein DJ, Knick VB, Lansing TJ, McConnell RT, Gilmer TM, Zhang SY, Robell K: Characterization of an Akt kinase inhibitor with potent pharmacodynamic and antitumor activity. Cancer Res. 2008, 68: 2366-2374. 10.1158/0008-5472.CAN-07-5783CrossRefPubMed
34.
Kawamoto T, Akisue T, Kishimoto K, Hara H, Imabori M, Fujimoto T, Kurosaka M, Hitora T, Kawaguchi Y, Yamamoto T: Inhibition of PKCalpha activation in human bone and soft tissue sarcoma cells by the selective PKC inhibitor PKC412. Anticancer Res. 2008, 28: 825-832.PubMed
35.
Martiny-Baron G, Kazanietz MG, Mischak H, Blumberg PM, Kochs G, Hug H, Marme D, Schachtele C: Selective inhibition of protein kinase C isozymes by the indolocarbazole Go 6976. J Biol Chem. 1993, 268: 9194-9197.PubMed
36.
Tibes R, Jimeno A, Von Hoff DD, Walker R, Pacciarini MA, Scaburri A, Fiorentini F, Borad MJ, Jameson GS, Hidalgo M: Phase I dose escalation study of the oral multi-CDK inhibitor PHA-848125. J Clin Oncol. 2008, 26: 15S#3531-
37.
Bu Y, Yang Z, Li Q, Song F: Silencing of polo-like kinase (Plk) 1 via siRNA causes inhibition of growth and induction of apoptosis in human esophageal cancer cells. Oncology. 2008, 74: 198-206. 10.1159/000151367CrossRefPubMed
38.
Spankuch-Schmitt B, Wolf G, Solbach C, Loibl S, Knecht R, Stegmuller M, von Minckwitz G, Kaufmann M, Strebhardt K: Downregulation of human polo-like kinase activity by antisense oligonucleotides induces growth inhibition in cancer cells. Oncogene. 2002, 21: 3162-3171. 10.1038/sj.onc.1205412CrossRefPubMed
39.
Takai N, Hamanaka R, Yoshimatsu J, Miyakawa I: Polo-like kinases (Plks) and cancer. Oncogene. 2005, 24: 287-291. 10.1038/sj.onc.1208272CrossRefPubMed
40.
Yamaguchi U, Honda K, Satow R, Kobayashi E, Nakayama R, Ichikawa H, Shoji A, Shitashige M, Masuda M, Kawai A: Functional genome screen for therapeutic targets of osteosarcoma. Cancer Sci. 2009,
41.
He ZL, Zheng H, Lin H, Miao XY, Zhong DW: Overexpression of polo-like kinase1 predicts a poor prognosis in hepatocellular carcinoma patients. World J Gastroenterol. 2009, 15: 4177-4182. 10.3748/wjg.15.4177PubMedCentralCrossRefPubMed
42.
Ikezoe T, Yang J, Nishioka C, Takezaki Y, Tasaka T, Togitani K, Koeffler HP, Yokoyama A: A novel treatment strategy targeting polo-like kinase 1 in hematological malignancies. Leukemia. 2009, 23: 1564-1576. 10.1038/leu.2009.94CrossRefPubMed
43.
Li JJ, Li SA: Mitotic kinases: the key to duplication, segregation, and cytokinesis errors, chromosomal instability, and oncogenesis. Pharmacol Ther. 2006, 111: 974-984. 10.1016/j.pharmthera.2006.02.006CrossRefPubMed
44.
Baldwin A, Li W, Grace M, Pearlberg J, Harlow E, Munger K, Grueneberg DA: Kinase requirements in human cells: II. Genetic interaction screens identify kinase requirements following HPV16 E7 expression in cancer cells. Proc Natl Acad Sci USA. 2008, 105: 16478-16483. 10.1073/pnas.0806195105PubMedCentralCrossRefPubMed
45.
Bommi-Reddy A, Almeciga I, Sawyer J, Geisen C, Li W, Harlow E, Kaelin WG, Grueneberg DA: Kinase requirements in human cells: III. Altered kinase requirements in VHL-/- cancer cells detected in a pilot synthetic lethal screen. Proc Natl Acad Sci USA. 2008, 105: 16484-16489. 10.1073/pnas.0806574105PubMedCentralCrossRefPubMed
46.
Grueneberg DA, Degot S, Pearlberg J, Li W, Davies JE, Baldwin A, Endege W, Doench J, Sawyer J, Hu Y: Kinase requirements in human cells: I. Comparing kinase requirements across various cell types. Proc Natl Acad Sci USA. 2008, 105: 16472-16477. 10.1073/pnas.0808019105PubMedCentralCrossRefPubMed
47.
Grueneberg DA, Li W, Davies JE, Sawyer J, Pearlberg J, Harlow E: Kinase requirements in human cells: IV. Differential kinase requirements in cervical and renal human tumor cell lines. Proc Natl Acad Sci USA. 2008, 105: 16490-16495. 10.1073/pnas.0806578105PubMedCentralCrossRefPubMed
48.
Baird K, Davis S, Antonescu CR, Harper UL, Walker RL, Chen Y, Glatfelter AA, Duray PH, Meltzer PS: Gene expression profiling of human sarcomas: insights into sarcoma biology. Cancer Res. 2005, 65: 9226-9235. 10.1158/0008-5472.CAN-05-1699CrossRefPubMed
49.
Ellinger-Ziegelbauer H, Karasuyama H, Yamada E, Tsujikawa K, Todokoro K, Nishida E: Ste20-like kinase (SLK), a regulatory kinase for polo-like kinase (Plk) during the G2/M transition in somatic cells. Genes Cells. 2000, 5: 491-498. 10.1046/j.1365-2443.2000.00337.xCrossRefPubMed
50.
Walter SA, Cutler RE, Martinez R, Gishizky M, Hill RJ: Stk10, a new member of the polo-like kinase kinase family highly expressed in hematopoietic tissue. J Biol Chem. 2003, 278: 18221-18228. 10.1074/jbc.M212556200CrossRefPubMed
51.
Kuramochi S, Matsuda Y, Okamoto M, Kitamura F, Yonekawa H, Karasuyama H: Molecular cloning of the human gene STK10 encoding lymphocyte-oriented kinase, and comparative chromosomal mapping of the human, mouse, and rat homologues. Immunogenetics. 1999, 49: 369-375. 10.1007/s002510050509CrossRefPubMed
52.
Kuramochi S, Moriguchi T, Kuida K, Endo J, Semba K, Nishida E, Karasuyama H: LOK is a novel mouse STE20-like protein kinase that is expressed predominantly in lymphocytes. J Biol Chem. 1997, 272: 22679-22684. 10.1074/jbc.272.36.22679CrossRefPubMed
53.
Eisenmann KM, McCarthy JB, Simpson MA, Keely PJ, Guan JL, Tachibana K, Lim L, Manser E, Furcht LT, Iida J: Melanoma chondroitin sulphate proteoglycan regulates cell spreading through Cdc42, Ack-1 and p130cas. Nat Cell Biol. 1999, 1: 507-513. 10.1038/70302CrossRefPubMed
54.
Satoh T, Kato J, Nishida K, Kaziro Y: Tyrosine phosphorylation of ACK in response to temperature shift-down, hyperosmotic shock, and epidermal growth factor stimulation. FEBS Lett. 1996, 386: 230-234. 10.1016/0014-5793(96)00449-8CrossRefPubMed
55.
Kato-Stankiewicz J, Ueda S, Kataoka T, Kaziro Y, Satoh T: Epidermal growth factor stimulation of the ACK1/Dbl pathway in a Cdc42 and Grb2-dependent manner. Biochem Biophys Res Commun. 2001, 284: 470-477. 10.1006/bbrc.2001.5004CrossRefPubMed
56.
Howlin J, Rosenkvist J, Andersson T: TNK2 preserves epidermal growth factor receptor expression on the cell surface and enhances migration and invasion of human breast cancer cells. Breast Cancer Res. 2008, 10: R36- 10.1186/bcr2087PubMedCentralCrossRefPubMed
Metadata
Title
RNAi phenotype profiling of kinases identifies potential therapeutic targets in Ewing's sarcoma
Authors
Shilpi Arora
Irma M Gonzales
R Tanner Hagelstrom
Christian Beaudry
Ashish Choudhary
Chao Sima
Raoul Tibes
Spyro Mousses
David O Azorsa
Publication date
01-12-2010
Publisher
BioMed Central
Published in
Molecular Cancer / Issue 1/2010
Electronic ISSN: 1476-4598
DOI
https://doi.org/10.1186/1476-4598-9-218

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