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STAT3 knockdown by siRNA induces apoptosis in human cutaneous T-cell lymphoma line Hut78 via downregulation of Bcl-xL

Abstract

Cutaneous T-cell lymphomas (CTCLs) are non-Hodgkin’s lymphomas resulting from clonal expansion and localization of malignant T-lymphocytes to the skin. CTCL cells have defective apoptosis. Signal transducers and activators of transcription (STAT) are a family of transcription factors known to play important roles in the development and progression of several human cancers by promoting cell proliferation and protecting against apoptosis. In this study, we investigated the specific role of STAT3, a major component of the STAT family, in growth and survival of human CTCL cell line Hut78. Western immunoblot analysis showed elevated expression of STAT3 and phospho-STAT3(Y705) in human CTCL cells as compared to freshly isolated peripheral blood lymphocytes (PBLs). Specific knockdown of STAT3 expression in Hut78 cells by RNA interference induced morphological and biochemical changes indicating apoptotic cell death. Moreover, STAT3 inhibition downregulated the expression of Bcl2 family of anti-apoptotic gene Bcl-xL. These observations suggest that STAT3 is required for the survival of CTCL cells and strongly indicate that targeting STAT3 using siRNA techniques may serve a novel therapeutic strategy for the treatment of CTCL.

Abbreviations

CTCL:

cutaneous T-cell lymphoma

MF:

mycosis fungoides

PBL:

peripheral blood lymphocyte

siRNA:

small interfering RNA

SS:

Sézary syndrome

STAT:

signal transducers and activators of transcription

TUNEL:

terminal deoxynucleotidyl transferase dUTP nick end labeling

References

  1. Siegel, R.S. and Kuzel, T.M. Cutaneous T-cell lymphoma/leukemia. Curr. Trends Options Oncol. 1 (2000) 43–50.

    Article  CAS  Google Scholar 

  2. Kotz, E.A., Anderson, D. and Thiers, B.H. Cutaneous T-cell lymphoma. J. Eur. Acad. Dermatol. Venereol. 17 (2003) 131–137.

    Article  CAS  PubMed  Google Scholar 

  3. Willemze, R., Jaffe, E.S., Burg, G., Burg, G., Cerroni, L., Berti, E., Swerdlow, S.H., Ralfkiaer, E., Chimenti, S., Diaz-Perez, J.L., Duncan, L.M., Grange, F., Harris, N.L., Kempf, W., Kerl, H., Kurrer, M., Knobler, R., Pimpinelli, N., Sander, C., Santucci, M., Sterry, W., Vermeer, M.H., Wechsler, J., Whittaker, S. and Meijer, C.J.L.M. WHO-EORTC classification for cutaneous lymphomas. Blood 105 (2005) 3768–3785.

    Article  CAS  PubMed  Google Scholar 

  4. Neelis, K.J., Schimmel, E.C., Vermeer, M.H., Senff, N.J., Willemze, R. and Noordijk, E.M. Low-dose palliative radiotherapy for cutaneous B- and T-cell lymphomas. Int. J. Rad. Oncol. Biol. Phys. 74 (2009) 154–158.

    Google Scholar 

  5. Kim, E.J., Hess, S., Richardson, S.K., Newton, S., Showe, L.C., Benoit, B.M., Ubriani, R., Vittorio, C.C., Junkins-Hopkins, J.M., Wysocka, M. and Rook, A.H. Immunopathogenesis and therapy of cutaneous T cell lymphoma. J. Clin. Invest. 115 (2005) 798–812.

    CAS  PubMed  Google Scholar 

  6. Diamandidou, E., Cohen, R.R. and Kurzrock, R. Mycosis fungoides and Sezary syndrome. Blood 88 (1996) 2385–2409.

    CAS  PubMed  Google Scholar 

  7. Qin, J-Z., Dummer, R., Burg, G. and Dobbeling, U. Constitutive and interleukin-7/interleukin-15 stimulated DNA binding of Myc, Jun, and novel Myc-like proteins in cutaneous T-cell lymphoma cells. Blood 93 (1999) 260–267.

    CAS  PubMed  Google Scholar 

  8. Kari, L., Loboda, A., Nebozhyn, M., Rook, A.H., Vonderheid, E.C., Nichols, C., Virok, D., Chang, C., Horng, W.H., Johnston, J., Wysocka, M., Showe, M.K. and Showe, L.C. Classification and prediction of survival in patients with the leukemic phase of cutaneous T cell lymphoma. J. Exp. Med. 197 (2003) 1477–1488.

    Article  CAS  PubMed  Google Scholar 

  9. Duvic, M. and Cather, J. Emerging new therapies for cutaneous T-cell lymphoma. Dermatol. Clin. 18 (2000) 147–156.

    CAS  PubMed  Google Scholar 

  10. Querfeld, C., Rizvi, M.A., Kuzel, T.M., Guitart, J., Rademaker, A., Sabharwal, S.S., Krett, N.L. and Rosen, S.T. The selective protein kinase C beta inhibitor enzastaurin induces apoptosis in cutaneous T-cell lymphoma cell lines through the AKT pathway. J. Invest. Dermatol. 126 (2006) 1641–1647.

    Article  CAS  PubMed  Google Scholar 

  11. Contassot, E. and French, L.E. Targeting apoptosis defects in cutaneous T-cell lymphomas. J. Invest. Dermatol. 129 (2009) 1059–1061.

    Article  CAS  PubMed  Google Scholar 

  12. Ni, X., Zhang, C., Talpur, R. and Duvic, M. Resistance to activation-induced cell death and bystander cytotoxicity via the Fas/Fas ligand pathway are implicated in the pathogenesis of cutaneous T cell lymphomas. J. Invest. Dermatol. 124 (2005) 741–750.

    Article  CAS  PubMed  Google Scholar 

  13. Wu, J., Nihal, M., Siddiqui, J., Vonderheid, E.C. and Wood, G.S. Low FAS/CD95 expression by CTCL correlates with reduced sensitivity to apoptosis that can be restored by FAS upregulation. J. Invest. Dermatol. 129 (2009) 1165–1173.

    Article  CAS  PubMed  Google Scholar 

  14. Zhang, C.L., Kamarasher, J., Qin, J.Z., Burg, G., Dummer, R. and Dobbeling, U. Expression of apoptosis regulators in cutaneous T-cell lymphomas. J. Pathol. 200 (2003) 249–254.

    Article  CAS  PubMed  Google Scholar 

  15. Takeda, K. and Akira, S. STAT family of transcription factors in cytokinemediated biological responses. Cytokine Growth Factor Rev. 11 (2000) 199–207.

    Article  CAS  PubMed  Google Scholar 

  16. Bromberg, J. and Darnell, J.E.Jr. The role of STAT3 in transcriptional control and their impact on cellular function. Oncogene 19 (2000) 2468–2473.

    Article  CAS  PubMed  Google Scholar 

  17. Levy, D.E. and Lee, C.K. What does Stat3 do? J. Clin. Invest. 109 (2002) 1143–1148.

    CAS  PubMed  Google Scholar 

  18. Kortylewski, M., Jove, R. and Yu, H. Targeting STAT3 affects melanoma on multiple fronts. Cancer Metastasis Rev. 24 (2005) 315–327.

    Article  CAS  PubMed  Google Scholar 

  19. Turkson, J. STAT proteins as novel targets for cancer drug discovery. Expert Opin. Ther. Targets 8 (2004) 409–422.

    Article  CAS  PubMed  Google Scholar 

  20. Sommer, V.H., Clemmensen, O.J., Nielsen, O., Wasik, M., Lovato, P., Brender, C., Eriksen, K.W., Woetmann, A., Kaestel, C.G., Nissen, M.H., Ropke, C., Skov, S. and Ødum, N. In vivo activation of STAT3 in cutaneous T-cell lymphoma. Evidence for an antiapoptotic function of STAT3. Leukemia 18 (2004) 1288–1295.

    Article  CAS  PubMed  Google Scholar 

  21. Nielsen, M., Kaltoft, K., Nordahl, M., Ropke, C., Geisler, C., Mustelin, T., Dobson, P., Svejgaard, A. and Ødum, N. Constitutive activation of a slowly migrating isoform of Stat3 in mycosis fungoides: tyrphostin AG490 inhibits Stat3 activation and growth of mycosis fungoides tumor cell lines. Proc. Natl. Acad. Sci. USA 94 (1997) 6764–6769.

    Article  CAS  PubMed  Google Scholar 

  22. Zhang, Q., Raghunath, P.N., Xue, L., Majewski, M., Carpentieri, D.F., Ødum, N., Morris, S., Skorski, T. and Wasik, M.A. Multilevel dysregulation of STAT3 activation in anaplastic lymphoma kinase-positive T/null-cell lymphoma. J. Immunol. 168 (2002) 466–474.

    CAS  PubMed  Google Scholar 

  23. Eriksen, K.W., Kaltoft, K., Mikkelsen, G., Nielsen, M., Zhang, Q., Geisler, C., Nissen, M.H., Ropke, C., Wasik, M.A. and Ødum, N. Constitutive STAT3-activation in Sezary syndrome: tyrphostin AG490 inhibits STAT3-activation, interleukin-2 receptor expression and growth of leukemic Sezary cells. Leukemia 15 (2001) 787–793.

    Article  CAS  PubMed  Google Scholar 

  24. Michel, L., Dupuy, A., Jean-Louis, F., Sors, A., Poupon, J., Viguier, M., Musette, P., Dubertret, L., Degos, L., Dombret, H. and Bachelez, H. Arsenic trioxide induces apoptosis of cutaneous T cell lymphoma cells: evidence for a partially caspase-independent pathway and potentiation by ascorbic acid (vitamin C). J. Invest. Dermatol. 121 (2003) 881–893.

    Article  CAS  PubMed  Google Scholar 

  25. Volkov, Y., Long, A., McGrath, S., Eidhin, D.Ni. and Kelleher, D. Crucial importance of PKC-β(I) in LFA-1-mediated locomotion of activated T cells. Nat. Immunol. 2 (2001) 508–514.

    Article  CAS  PubMed  Google Scholar 

  26. Verma, N.K., Dempsey, E., Conroy, J., Olwell, P., Mcelligott, A.M., Davies, A.M., Kelleher, D., Butini, S., Campiani, G., Williams, D.C., Zisterer, D.M., Lawler, M. and Volkov, Y. A new microtubule targeting compound PBOX-15 inhibits T-cell migration via post-translational modifications of tubulin. J. Mol. Med. 86 (2008) 457–469.

    Article  CAS  PubMed  Google Scholar 

  27. Verma, N.K., Dourlat, J., Davies, A.M., Long, A., Liu, W.Q., Garbay, C., Kelleher, D. and Volkov, Y. STAT3-stathmin interactions control microtubule dynamics in migrating T-cells. J. Biol. Chem. 284 (2009) 12349–12362.

    Article  CAS  PubMed  Google Scholar 

  28. Elbashir, S.M., Harborth, J., Lendeckel, W., Yalcin, A., Weber, K. and Tuschl, T. Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature 411 (2001) 494–498.

    Article  CAS  PubMed  Google Scholar 

  29. Konnikova, L., Kotecki, M., Kruger, M.M. and Cochran, B. Knockdown of STAT3 expression by RNAi induces apoptosis in astrocytoma cells. BMC Cancer 3 (2003) 23–32.

    Article  PubMed  Google Scholar 

  30. Gao, L.F., Xu, D.Q., Wen, L.J., Zhang, X.Y., Shao, Y.T. and Zhao, X.J. Inhibition of STAT3 expression by siRNA suppresses growth and induces apoptosis in laryngeal cancer cells. Acta Pharmacol. Sin. 26 (2005) 377–383.

    Article  CAS  PubMed  Google Scholar 

  31. Grandis, J.R., Drenning, S.D., Zeng, Q., Watkins, S.C., Melhem, M.F., Endo, S., Johnson, D.E., Huang, L., He, Y. and Kim, J.D. Constitutive activation of Stat3 signaling abrogates apoptosis in squamous cell carcinogenesis in vivo. Proc. Natl. Acad. Sci. USA 97 (2000) 4227–4232.

    Article  CAS  PubMed  Google Scholar 

  32. Mora, L.B., Buettner, R., Seigne, J., Diaz, J., Ahmad, N., Garcia, R., Bowman, T., Falcone, R., Fairclough, R., Cantor, A., Muro-Cacho, C., Livingston, S., Karras, J., Pow-Sang, J. and Jove, R. Constitutive activation of Stat3 in human prostate tumors and cell lines: direct inhibition of Stat3 signaling induces apoptosis of prostate cancer cells. Cancer Res. 62 (2002) 6659–6666.

    CAS  PubMed  Google Scholar 

  33. Elmore, S. Apoptosis: a review of programmed cell death. Toxicol. Pathol. 35 (2007) 495–516.

    Article  CAS  PubMed  Google Scholar 

  34. Reed, J.C. Bcl2 family proteins. Oncogene 17 (1998) 3225–3236.

    Article  PubMed  Google Scholar 

  35. Osella-Aate, S., Zaccagna, A., Savoia, P., Quaglino, P., Salomone, B. and Bernengo, M.G. Expression of apoptosis markers on peripheral blood lymphocytes from patients with cutaneous T-cell lymphoma during extracorporeal photochemotherapy. J. Am. Acad. Dermatol. 44 (2001) 40–47.

    Article  Google Scholar 

  36. Yu, H. and Jove, R. The STATs of cancer—new molecular targets come of age. Nat. Rev. Cancer 4 (2004) 97–105.

    Article  CAS  PubMed  Google Scholar 

  37. Alshamsan, A., Hamdya, S., Samuel, J., El-Kadi, A.O.S., Lavasanifar, A. and Uludag, H. The induction of tumor apoptosis in B16 melanoma following STAT3 siRNA delivery with a lipid-substituted polyethylenimine. Biomaterials 31 (2010) 1420–1428.

    Article  CAS  PubMed  Google Scholar 

  38. Yue, P. and Turkson, J. Targeting STAT3 in cancer: how successful are we? Expert Opin. Investig. Drugs 18 (2009) 45–56.

    Article  CAS  PubMed  Google Scholar 

  39. Cejka, D., Losert, D. and Wacheck, V. Short interfering RNA (siRNA): tool or therapeutic? Clin. Sci. (Lond.) 110 (2006) 47–58.

    Article  CAS  Google Scholar 

  40. Grivennikov, S.I. and Karin, M. Dangerous liaisons: STAT3 and NF-κB collaboration and crosstalk in cancer. Cytokine Growth Factor Rev. (2009) in press. DOI:10.1016/j.cytogfr.2009.11.005.

  41. Takeda, K., Noguchi, K., Shi, W., Tanaka, T., Matsumoto, M., Yoshida, N., Kishimoto, T. and Akira, S. Targeted disruption of the mouse Stat3 gene leads to early embryonic lethality. Proc. Natl. Acad. Sci. USA 94 (1997) 3801–3804.

    Article  CAS  PubMed  Google Scholar 

  42. de la Iglesia, N., Konopka, G., Lim, K.L., Nutt, C.L., Bromberg, J.F., Frank, D.A., Mischel, P.S., Louis, D.N. and Bonni, A. Deregulation of a STAT3-interleukin 8 signaling pathway promotes human glioblastoma cell proliferation and invasiveness. J. Neurosci. 28 (2008) 5870–5878.

    Article  PubMed  Google Scholar 

  43. Li, S.D., Chen, Y.C., Hackett, M.J. and Huang, L. Tumor-targeted delivery of siRNA by self-assembled nanoparticles. Mol. Ther. 1 (2008) 163–169.

    Article  Google Scholar 

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Correspondence to Navin K. Verma.

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Verma, N.K., Davies, A.M., Long, A. et al. STAT3 knockdown by siRNA induces apoptosis in human cutaneous T-cell lymphoma line Hut78 via downregulation of Bcl-xL . Cell Mol Biol Lett 15, 342–355 (2010). https://doi.org/10.2478/s11658-010-0008-2

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