A-770041 reverses paclitaxel and doxorubicin resistance in osteosarcoma cells

BMC Cancer

Zhenfeng Duan⁵  Jianming Zhang¹  Shunan Ye⁵  Jacson Shen⁵  Edwin Choy⁵  Gregory Cote⁵  David Harmon⁵  Henry Mankin⁵  Yingqi Hua⁴  Yu Zhang³  Nathanael S Gray²  Francis J Hornicek⁵ 

[1] Cutaneous Biology Research Center, Massachusetts General Hospital, Boston 02114, MA, USA

[2] Dana Farber Cancer Institute, Harvard Medical School, Boston, MA 02115, USA

[3] Department of Orthopedic Surgery, Liu Hua Qiao Hospital, Guangzhou 510010, China

[4] Shanghai First People’s Hospital, Shanghai Jiaotong University, Shanghai 200080, China

[5] Center for Sarcoma and Connective Tissue Oncology, Massachusetts General Hospital, 100 Blossom St., Jackson 1115, Boston 02114, MA, USA

关键词: Doxorubicin;    Src kinase;    MDR;    Osteosarcoma;   

Others  :  1121060
DOI  :  10.1186/1471-2407-14-681

 received in 2014-06-04, accepted in 2014-09-16,  发布年份 2014

【 摘 要 】

Background

Reversing multidrug resistance (MDR) has been an important goal for clinical and investigational oncologists. In the last few decades, significant effort has been made to search for inhibitors to reverse MDR by targeting ATP-binding cassette (ABC) transporters (Pgp, MRP) directly, but these efforts have achieved little clinical success. Protein kinases play important roles in many aspects of tumor cell growth and survival. Combinations of kinase inhibitors and chemotherapeutics have been observed to overcome cancer drug resistance in certain circumstances.

Methods

We screened a kinase specific inhibitor compound library in human osteosarcoma MDR cell lines to identify inhibitors that were capable of reversing chemoresistance to doxorubicin and paclitaxel.

Results

We identified 18 small molecules that significantly increase chemotherapy drug-induced cell death in human osteosarcoma MDR cell lines U-2OS_MR and KHOSR_2. We identified A-770041 as one of the most effective MDR reversing agents when combined with doxorubicin or paclitaxel. A-770041 is a potent Src family kinase (Lck and Src) inhibitor. Western blot analysis revealed A-770041 inhibits both Src and Lck activation and expression. Inhibition of Src expression in U-2OS_MR and KHOSR₂ cell lines using lentiviral shRNA also resulted in increased doxorubicin and paclitaxel drug sensitivity. A-770041 increases the intracellular drug accumulation as demonstrated by calcein AM assay.

Conclusions

These results indicate that small molecule inhibitor A-770041 may function to reverse ABCB1/Pgp-mediated chemotherapy drug resistance. Combination of Src family kinase inhibitor with regular chemotherapy drug could be clinically effective in MDR osteosarcoma.

【 授权许可】

   
2014 Duan et al.; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]Bielack SS, Marina N, Ferrari S, Helman LJ, Smeland S, Whelan JS, Reaman GH: Osteosarcoma: the same old drugs or more? J Clin Oncol 2008, 26(18):3102-3103. author reply 3104–3105
• [2]Gill J, Ahluwalia MK, Geller D, Gorlick R: New targets and approaches in osteosarcoma. Pharmacol Ther 2013, 137(1):89-99.
• [3]Geller DS, Gorlick R: Osteosarcoma: a review of diagnosis, management, and treatment strategies. Clin Adv Hematol Oncol 2010, 8(10):705-718.
• [4]Chou AJ, Geller DS, Gorlick R: Therapy for osteosarcoma: where do we go from here? Paediatr Drugs 2008, 10(5):315-327.
• [5]Chou AJ, Gorlick R: Chemotherapy resistance in osteosarcoma: current challenges and future directions. Expert Rev Anticancer Ther 2006, 6(7):1075-1085.
• [6]Susa M, Iyer AK, Ryu K, Choy E, Hornicek FJ, Mankin H, Milane L, Amiji MM, Duan Z: Inhibition of ABCB1 (MDR1) expression by an siRNA nanoparticulate delivery system to overcome drug resistance in osteosarcoma. PLoS One 2010, 5(5):e10764.
• [7]Kobayashi E, Iyer AK, Hornicek FJ, Amiji MM, Duan Z: Lipid-functionalized dextran nanosystems to overcome multidrug resistance in cancer: a pilot study. Clin Orthop Relat Res 2013, 471(3):915-925.
• [8]Brambilla D, Zamboni S, Federici C, Lugini L, Lozupone F, De Milito A, Cecchetti S, Cianfriglia M, Fais S: P-glycoprotein binds to ezrin at amino acid residues 149–242 in the FERM domain and plays a key role in the multidrug resistance of human osteosarcoma. Int J Cancer 2012, 130(12):2824-2834.
• [9]Cagliero E, Ferracini R, Morello E, Scotlandi K, Manara MC, Buracco P, Comandone A, Baroetto Parisi R, Baldini N: Reversal of multidrug-resistance using Valspodar (PSC 833) and doxorubicin in osteosarcoma. Oncol Rep 2004, 12(5):1023-1031.
• [10]Pakos EE, Ioannidis JP: The association of P-glycoprotein with response to chemotherapy and clinical outcome in patients with osteosarcoma. A meta-analysis. Cancer 2003, 98(3):581-589.
• [11]Hotta T, Suzuki H, Nagai S, Yamamoto K, Imakiire A, Takada E, Itoh M, Mizuguchi J: Chemotherapeutic agents sensitize sarcoma cell lines to tumor necrosis factor-related apoptosis-inducing ligand-induced caspase-8 activation, apoptosis and loss of mitochondrial membrane potential. J Orthop Res 2003, 21(5):949-957.
• [12]Zhao Y, Zhang CL, Zeng BF, Wu XS, Gao TT, Oda Y: Enhanced chemosensitivity of drug-resistant osteosarcoma cells by lentivirus-mediated Bcl-2 silencing. Biochem Biophys Res Commun 2009, 390(3):642-647.
• [13]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(6):591-600.
• [14]Du J, Bernasconi P, Clauser KR, Mani DR, Finn SP, Beroukhim R, Burns M, Julian B, Peng XP, Hieronymus H, Maglathlin RL, Lewis TA, Liau LM, Nghiemphu P, Mellinghoff IK, Louis DN, Loda M, Carr SA, Kung AL, Golub TR: Bead-based profiling of tyrosine kinase phosphorylation identifies SRC as a potential target for glioblastoma therapy. Nat Biotechnol 2009, 27(1):77-83.
• [15]Scotlandi K, Picci P: Targeting insulin-like growth factor 1 receptor in sarcomas. Curr Opin Oncol 2008, 20(4):419-427.
• [16]Wan X, Helman LJ: The biology behind mTOR inhibition in sarcoma. Oncologist 2007, 12(8):1007-1018.
• [17]Kim SY, Helman LJ: Strategies to explore new approaches in the investigation and treatment of osteosarcoma. Cancer Treat Res 2009, 152:517-528.
• [18]Hirai H, Sootome H, Nakatsuru Y, Miyama K, Taguchi S, Tsujioka K, Ueno Y, Hatch H, Majumder PK, Pan BS, Kotani H: MK-2206, an allosteric Akt inhibitor, enhances antitumor efficacy by standard chemotherapeutic agents or molecular targeted drugs in vitro and in vivo. Mol Cancer Ther 2010, 9(7):1956-1967.
• [19]Wang YH, Xiong J, Wang SF, Yu Y, Wang B, Chen YX, Shi HF, Qiu Y: Lentivirus-mediated shRNA targeting insulin-like growth factor-1 receptor (IGF-1R) enhances chemosensitivity of osteosarcoma cells in vitro and in vivo. Mol Cell Biochem 2010, 341(1–2):225-233.
• [20]Duan Z, Choy E, Harmon D, Yang C, Ryu K, Schwab J, Mankin H, Hornicek FJ: Insulin-like growth factor-I receptor tyrosine kinase inhibitor cyclolignan picropodophyllin inhibits proliferation and induces apoptosis in multidrug resistant osteosarcoma cell lines. Mol Cancer Ther 2009, 8(8):2122-2130.
• [21]George JA, Chen T, Taylor CC: SRC tyrosine kinase and multidrug resistance protein-1 inhibitions act independently but cooperatively to restore paclitaxel sensitivity to paclitaxel-resistant ovarian cancer cells. Cancer Res 2005, 65(22):10381-10388.
• [22]Shukla S, Chen ZS, Ambudkar SV: Tyrosine kinase inhibitors as modulators of ABC transporter-mediated drug resistance. Drug Resist Updat 2012, 15(1–2):70-80.
• [23]Sodani K, Tiwari AK, Singh S, Patel A, Xiao ZJ, Chen JJ, Sun YL, Talele TT, Chen ZS: GW583340 and GW2974, human EGFR and HER-2 inhibitors, reverse ABCG2- and ABCB1-mediated drug resistance. Biochem Pharmacol 2012, 83(12):1613-1622.
• [24]Liu Y, Gray NS: Rational design of inhibitors that bind to inactive kinase conformations. Nat Chem Biol 2006, 2(7):358-364.
• [25]Bamborough P, Morse MA, Ray KP: Targeting IKKbeta for the treatment of rheumatoid arthritis. Drug News Perspect 2010, 23(8):483-490.
• [26]Patricelli MP, Nomanbhoy TK, Wu J, Brown H, Zhou D, Zhang J, Jagannathan S, Aban A, Okerberg E, Herring C, Nordin B, Weissig H, Yang Q, Lee JD, Gray NS, Kozarich JW: In situ kinase profiling reveals functionally relevant properties of native kinases. Chem Biol 2011, 18(6):699-710.
• [27]Weisberg E, Liu Q, Zhang X, Nelson E, Sattler M, Liu F, Nicolais M, Zhang J, Mitsiades C, Smith RW, Stone R, Galinsky I, Nonami A, Griffin JD, Gray N: Selective Akt inhibitors synergize with tyrosine kinase inhibitors and effectively override stroma-associated cytoprotection of mutant FLT3-positive AML cells. PLoS One 2013, 8(2):e56473.
• [28]Liu Y, Marks K, Cowley GS, Carretero J, Liu Q, Nieland TJ, Xu C, Cohoon TJ, Gao P, Zhang Y, Chen Z, Altabef AB, Tchaicha JH, Wang X, Choe S, Driggers EM, Zhang J, Bailey ST, Sharpless NE, Hayes DN, Patel NM, Janne PA, Bardeesy N, Engelman JA, Manning BD, Shaw RJ, Asara JM, Scully R, Kimmelman A, Byers LA, Gibbons DL, Wistuba II, Heymach JV, Kwiatkowski DJ, Kim WY, Kung AL, Gray NS, Root DE, Cantley LC, Wong KK: Metabolic and functional genomic studies identify deoxythymidylate Kinase as a target in LKB1 mutant lung cancer. Cancer Discov 2013, 3(8):870-879.
• [29]Lourda M, Trougakos IP, Gonos ES: Development of resistance to chemotherapeutic drugs in human osteosarcoma cell lines largely depends on up-regulation of Clusterin/Apolipoprotein J. Int J Cancer 2007, 120(3):611-622.
• [30]Duan Z, Ji D, Weinstein EJ, Liu X, Susa M, Choy E, Yang C, Mankin H, Hornicek FJ: Lentiviral shRNA screen of human kinases identifies PLK1 as a potential therapeutic target for osteosarcoma. Cancer Lett 2010, 293(2):220-229.
• [31]Burchat A, Borhani DW, Calderwood DJ, Hirst GC, Li B, Stachlewitz RF: Discovery of A-770041, a src-family selective orally active lck inhibitor that prevents organ allograft rejection. Bioorg Med Chem Lett 2006, 16(1):118-122.
• [32]Stachlewitz RF, Hart MA, Bettencourt B, Kebede T, Schwartz A, Ratnofsky SE, Calderwood DJ, Waegell WO, Hirst GC: A-770041, a novel and selective small-molecule inhibitor of Lck, prevents heart allograft rejection. J Pharmacol Exp Ther 2005, 315(1):36-41.
• [33]Meyn MA 3rd, Smithgall TE: Small molecule inhibitors of Lck: the search for specificity within a kinase family. Mini Rev Med Chem 2008, 8(6):628-637.
• [34]Cantiani L, Manara MC, Zucchini C, De Sanctis P, Zuntini M, Valvassori L, Serra M, Olivero M, Di Renzo MF, Colombo MP, Picci P, Scotlandi K: Caveolin-1 reduces osteosarcoma metastases by inhibiting c-Src activity and met signaling. Cancer Res 2007, 67(16):7675-7685.
• [35]Shor AC, Keschman EA, Lee FY, Muro-Cacho C, Letson GD, Trent JC, Pledger WJ, Jove R: Dasatinib inhibits migration and invasion in diverse human sarcoma cell lines and induces apoptosis in bone sarcoma cells dependent on SRC kinase for survival. Cancer Res 2007, 67(6):2800-2808.
• [36]Patel A, Tiwari AK, Chufan EE, Sodani K, Anreddy N, Singh S, Ambudkar SV, Stephani R, Chen ZS: PD173074, a selective FGFR inhibitor, reverses ABCB1-mediated drug resistance in cancer cells. Cancer Chemother Pharmacol 2013, 2013:2013.
• [37]Posthumadeboer J, van Egmond PW, Helder MN, de Menezes RX, Cleton-Jansen AM, Belien JA, Verheul HM, van Royen BJ, Kaspers GJ, van Beusechem VW: Targeting JNK-interacting-protein-1 (JIP1) sensitises osteosarcoma to doxorubicin. Oncotarget 2012, 3(10):1169-1181.
• [38]Spina A, Sorvillo L, Chiosi E, Esposito A, Di Maiolo F, Sapio L, Caraglia M, Naviglio S: Synergistic cytotoxic effects of inorganic phosphate and chemotherapeutic drugs on human osteosarcoma cells. Oncol Rep 2013, 29(5):1689-1696.
• [39]Naviglio S: The possible use of inorganic phosphate in osteosarcoma therapy. Future Oncol 2013, 9(9):1249-1251.
• [40]Michels S, Trautmann M, Sievers E, Kindler D, Huss S, Renner M, Friedrichs N, Kirfel J, Steiner S, Endl E, Wurst P, Heukamp L, Penzel R, Larsson O, Kawai A, Tanaka S, Sonobe H, Schirmacher P, Mechtersheimer G, Wardelmann E, Büttner R, Hartmann W: SRC signaling is crucial in the growth of synovial sarcoma cells. Cancer Res 2013, 73(8):2518-2528.
• [41]Heiska L, Melikova M, Zhao F, Saotome I, McClatchey AI, Carpen O: Ezrin is key regulator of Src-induced malignant phenotype in three-dimensional environment. Oncogene 2011, 30(50):4953-4962.
• [42]Le XF, Bast RC Jr: Src family kinases and paclitaxel sensitivity. Cancer Biol Ther 2011, 12(4):260-269.
• [43]Kitazaki T, Oka M, Nakamura Y, Tsurutani J, Doi S, Yasunaga M, Takemura M, Yabuuchi H, Soda H, Kohno S: Gefitinib, an EGFR tyrosine kinase inhibitor, directly inhibits the function of P-glycoprotein in multidrug resistant cancer cells. Lung Cancer 2005, 49(3):337-343.
• [44]Tsai CM, Chiu CH, Chang KT, Chen JT, Lai CL, Chen YM, Hsiao SY: Gefitinib enhances cytotoxicities of antimicrotubule agents in non-small-cell lung cancer cells exhibiting no sensitizing epidermal growth factor receptor mutation. J Thorac Oncol 2012, 7(8):1218-1227.