Neurochemical Research

, 36:2292 | Cite as

Valproic Acid Induced Differentiation and Potentiated Efficacy of Taxol and Nanotaxol for Controlling Growth of Human Glioblastoma LN18 and T98G Cells

  • Subhasree Roy Choudhury
  • Surajit Karmakar
  • Naren L. Banik
  • Swapan K. Ray
Original Paper


Glioblastoma shows poor response to current therapies and warrants new therapeutic strategies. We examined the efficacy of combination of valproic acid (VPA) and taxol (TX) or nanotaxol (NTX) in human glioblastoma LN18 and T98G cell lines. Cell differentiation was manifested in changes in morphological features and biochemical markers. Cell growth was controlled with down regulation of vascular endothelial growth factor (VEGF), epidermal growth factor receptor (EGFR), nuclear factor-kappa B (NF-κB), phospho-Akt (p-Akt), and multi-drug resistance (MDR) marker, indicating suppression of angiogenic, survival, and multi-drug resistance pathways. Cell cycle analysis showed that combination therapy (VPA and TX or NTX) increased the apoptotic sub G1 population and apoptosis was further confirmed by Annexin V-FITC/PI binding assay and scanning electron microscopy. Combination therapy caused activation of caspase-8 and cleavage of Bid to tBid and increased Bax:Bcl-2 ratio and mitochondrial release of cytochrome c and apoptosis-inducing factor (AIF). Upregulation of calpain and caspases (caspase-9 and caspase-3) and substrate degradation were also detected in course of apoptosis. The combination of VPA and NTX most effectively controlled the growth of LN18 and T98G cells. Therefore, this combination of drugs can be used as an effective treatment for controlling growth of human glioblastoma cells.


Apoptosis Glioblastoma Nanotaxol Taxol Valproic acid 



This investigation was supported in part by the NS-57811 and NS-62327 grants from the National Institutes of Health and the SCIRF-11-002 grant from the State of South Carolina.


  1. 1.
    Narayana A, Leibel SA (2004) Primary and metastatic brain tumors in adults. In: Leibel SA, Phillips TL (eds) Textbook of radiation oncology. Elsevier, Philadelphia, pp 463–495Google Scholar
  2. 2.
    Bähr O, Rieger J, Duffner F, Meyermann R, Weller M, Wick W (2003) P-glycoprotein and multidrug resistance-associated protein mediate specific patterns of multidrug resistance in malignant glioma cell lines, but not in primary glioma cells. Brain Pathol 13:482–494PubMedCrossRefGoogle Scholar
  3. 3.
    Tang R, Faussat AM, Majdak P, Perrot JY, Chaoui D, Legrand O, Marie JP (2004) Valproic acid inhibits proliferation and induces apoptosis in acute myeloid leukemia cells expressing P-gp and MRP1. Leukemia 18:1246–1255PubMedCrossRefGoogle Scholar
  4. 4.
    Rowinsky EK, Wright M, Monsarrat B, Lesser GJ, Donehower RC (1993) Taxol: pharmacology, metabolism and clinical implications. Cancer Surv 17:283–304PubMedGoogle Scholar
  5. 5.
    Karmakar S, Banik NL, Patel SJ, Ray SK (2007) Combination of all-trans retinoic acid and taxol regressed glioblastoma T98G xenografts in nude mice. Apoptosis 12:2077–2087PubMedCrossRefGoogle Scholar
  6. 6.
    Karmakar S, Banik NL, Ray SK (2008) Combination of all-trans retinoic acid and taxol-induced differentiation and apoptosis in human glioblastoma U87MG xenografts in nude mice. Cancer 112:596–607PubMedCrossRefGoogle Scholar
  7. 7.
    Prados MD, Schold SC, Spence AM, Berger MS, McAllister LD, Mehta MP, Gilbert MR, Fulton D, Kuhn J, Lamborn K, Rector DJ, Chang SM (1996) Phase II study of paclitaxel in patients with recurrent malignant glioma. J Clin Oncol 14:2316–2321PubMedGoogle Scholar
  8. 8.
    Weiss RB, Donehower RC, Wiernik PH, Ohnuma T, Gralla RJ, Trump DL et al (1990) Hypersensitivity reactions from taxol. J Clin Oncol 8:1263–1268PubMedGoogle Scholar
  9. 9.
    Gallo JM, Li S, Guo P, Reed K, Ma J (2003) The effect of P-glycoprotein on Taxol brain and brain tumor distribution in mice. Cancer Res 63:5114–5117PubMedGoogle Scholar
  10. 10.
    Fellner S, Bauer B, Miller DS, Schaffri M, Fankhänel M, Spruss T et al (2002) Transport of paclitaxel (Taxol) across the blood-brain barrier in vitro and in vivo. J Clin Invest 110:1309–1318PubMedGoogle Scholar
  11. 11.
    Joo KM, Park K, Kong DS, Song SY, Kim MH, Lee GS et al (2008) Oral taxol chemotherapy for brain tumors: ideal combination treatment of taxol and P-glycoprotein inhibitor. Oncol Rep 19:17–23PubMedGoogle Scholar
  12. 12.
    Koziara JM, Whisman TR, Tseng MT, Mumper RJ (2006) In vivo efficacy of novel taxol nanoparticles in taxol-resistant human colorectal tumors. J Cont Rel 112:312–319CrossRefGoogle Scholar
  13. 13.
    Desai N, Trieu V, Yao Z, Louie L, Ci S, Yang A et al (2006) Increased antitumor activity, intratumor taxol concentrations, and endothelial cell transport of cremophor-free, albumin-bound taxol, ABI-007, compared with cremophor-based taxol. Clin Cancer Res 12:1317–1324PubMedCrossRefGoogle Scholar
  14. 14.
    Green MR, Manikhas GM, Orlov S, Afanasyev B, Makhson AM, Bhar P, Hawkins MJ (2006) Abraxane, a novel Cremophor-free, albumin-bound particle form of taxol for the treatment of advanced non-small-cell lung cancer. Ann Oncol 17:1263–1268PubMedCrossRefGoogle Scholar
  15. 15.
    Gradishar WJ, Tjulandin S, Davidson N, Shaw H, Desai N, Bhar P, Hawkins M, O’Shaughnessy J (2005) Phase III trial of nanoparticle albumin-bound taxol compared with polyethylated castor oil-based taxol in women with breast cancer. J Clin Oncol 23:7794–7803PubMedCrossRefGoogle Scholar
  16. 16.
    Abe T, Hasegawa S, Taniguchi K, Yokomizo A, Kuwano T, Ono M et al (1994) Possible involvement of multidrug-resistance-associated protein (MRP) gene expression in spontaneous drug resistance to vincristine, etoposide and adriamycin in human glioma cells. Int J Cancer 58:860–864PubMedCrossRefGoogle Scholar
  17. 17.
    Rieger L, Rieger J, Winter S, Streffer J, Esser P, Dichgans J, Meyermann R, Weller M (2000) Evidence for a constitutive, verapamil-sensitive, non-P-glycoprotein multidrug resistance phenotype in malignant glioma that is unaltered by radiochemotherapy in vivo. Acta Neuropathol 99:555–562PubMedCrossRefGoogle Scholar
  18. 18.
    Karmakar S, Weinberg MS, Banik NL, Patel SJ, Ray SK (2006) Activation of multiple molecular mechanisms for apoptosis in human malignant glioblastoma T98G and U87MG cells treated with sulforaphane. Neuroscience 141:1265–1280PubMedCrossRefGoogle Scholar
  19. 19.
    Dhandapani KM, Mahesh VB, Brann DW (2007) Curcumin suppresses growth and chemoresistance of human glioblastoma cells via AP-1 and NF-κB transcription factors. J Neurochem 102:522–538PubMedCrossRefGoogle Scholar
  20. 20.
    Karmakar S, Davis KA, Choudhury SR, Deeconda A, Banik NL, Ray SK (2009) Bcl-2 inhibitor and apigenin worked synergistically in human malignant neuroblastoma cell lines and increased apoptosis with activation of extrinsic and intrinsic pathways. Biochem Biophys Res Commun 388:705–710PubMedCrossRefGoogle Scholar
  21. 21.
    Roy Choudhury S, Karmakar S, Banik NL, Ray SK (2010) Synergistic efficacy of sorafenib and genistein in growth inhibition by down regulating angiogenic and survival factors and increasing apoptosis through upregulation of p53 and p21 in malignant neuroblastoma cells having N-Myc amplification or non-amplification. Invest New Drugs 28:812–824PubMedCrossRefGoogle Scholar
  22. 22.
    Karmakar S, Choudhury SR, Banik NL, Ray SK (2010) Activation of multiple molecular mechanisms for increasing apoptosis in human glioblastoma T98G xenograft. J Cancer Sci Ther 2:107–113PubMedCrossRefGoogle Scholar
  23. 23.
    Knizetova P, Ehrmann J, Hlobilkova A, Vancova I, Kalita O, Kolar Z, Bartek J (2008) Autocrine regulation of glioblastoma cell cycle progression, viability and radioresistance through the VEGF-VEGFR2 (KDR) interplay. Cell Cycle 7:2553–2561PubMedCrossRefGoogle Scholar
  24. 24.
    Aldape KD, Ballman K, Furth A, Buckner JC, Giannini C, Burger PC et al (2004) Immunohistochemical detection of EGFRvIII in high malignancy grade astrocytomas and evaluation of prognostic significance. J Neuropathol Exp Neurol 63:700–707PubMedGoogle Scholar
  25. 25.
    Sakahira H, Enari M, Nagata S (1998) Cleavage of CAD inhibitor in CAD activation and DNA degradation during apoptosis. Nature 391:96–99PubMedCrossRefGoogle Scholar
  26. 26.
    Das CM, Aguilera D, Vasquez H, Prasad P, Zhang M, Wolff JE, Gopalakrishnan V (2007) Valproic acid induces p21 and topoisomerase-II (α/β) expression and synergistically enhances etoposide cytotoxicity in human glioblastoma cell lines. J Neurooncol 85:159–170PubMedCrossRefGoogle Scholar
  27. 27.
    Selesniemi K, Reedy M, Gultice A, Guilbert LJ, Brown TLb (2005) Transforming growth factor-β induces differentiation of the labyrinthine trophoblast stem cell line SM10. Stem Cells Dev 14:697–711PubMedCrossRefGoogle Scholar
  28. 28.
    Jones J, Bentas W, Blaheta RA, Makarevic J, Hudak L, Wedel S et al (2008) Modulation of adhesion and growth of colon and pancreatic cancer cells by the histone deacetylase inhibitor valproic acid. Int J Mol Med 22:293–299PubMedGoogle Scholar
  29. 29.
    Zhang R, Banik NL, Ray SK (2008) Differential sensitivity of human glioblastoma LN18 (PTEN-positive) and A172 (PTEN-negative) cells to Taxol for apoptosis. Brain Res 1239:216–225PubMedCrossRefGoogle Scholar
  30. 30.
    George J, Banik NL, Ray SK (2009) Bcl-2 siRNA augments taxol mediated apoptotic death in human glioblastoma U138MG and U251MG cells. Neurochem Res 34:66–78PubMedCrossRefGoogle Scholar
  31. 31.
    Son MJ, Song HS, Kim MH, Kim JT, Kang CM, Jeon JW et al (2006) Synergistic effect and condition of pegylated interferon alpha with paclitaxel on glioblastoma. Int J Oncol 28:1385–1392PubMedGoogle Scholar
  32. 32.
    Pipas JM, Meyer LP, Rhodes CH, Cromwell LD, McDonnell CE, Kingman LS et al (2005) A Phase II trial of paclitaxel and topotecan with filgrastim in patients with recurrent or refractory glioblastoma multiforme or anaplastic astrocytoma. J Neurooncol 71:301–305PubMedCrossRefGoogle Scholar
  33. 33.
    Pelloski CE, Lin E, Zhang L, Yung WK, Colman H, Liu JL et al (2006) Prognostic associations of activated mitogen-activated protein kinase and Akt pathways in glioblastoma. Clin Cancer Res 12:3935–3941PubMedCrossRefGoogle Scholar
  34. 34.
    Beg AA, Baltimore D (1996) An essential role for NFκB in preventing TNF-α induced cell death. Science 274:782–784PubMedCrossRefGoogle Scholar
  35. 35.
    Kemper EM, Boogerd W, Thuis I, Beijnen JH, van Tellingen O (2004) Modulation of the blood-brain barrier in oncology: therapeutic opportunities for the treatment of brain tumours? Cancer Treat Rev 30:415–423PubMedCrossRefGoogle Scholar
  36. 36.
    Roth W, Wagenknecht B, Grimmel C, Dichgans J, Weller M (1998) Taxol-mediated augmentation of CD95 ligand-induced apoptosis of human malignant glioma cells: association with bcl-2 phosphorylation but neither activation of p53 nor G2/M cell cycle arrest. Br J Cancer 77:404–411PubMedCrossRefGoogle Scholar
  37. 37.
    Desai A, Vyas T, Amiji M (2008) Cytotoxicity and apoptosis enhancement in brain tumor cells upon coadministration of taxol and ceramide in nanoemulsion formulations. J Pharm Sci 97:2745–2756PubMedCrossRefGoogle Scholar
  38. 38.
    Bokelmann I, Mahlknecht U (2008) Valproic acid sensitizes chronic lymphocytic leukemia cells to apoptosis and restores the balance between pro- and antiapoptotic proteins. Mol Med 14:20–27PubMedCrossRefGoogle Scholar
  39. 39.
    Häcker S, Dittrich A, Mohr A, Schweitzer T, Rutkowski S, Krauss J et al (2009) Histone deacetylase inhibitors cooperate with IFN-gamma to restore caspase-8 expression and overcome TRAIL resistance in cancers with silencing of caspase-8. Oncogene 28:3097–3110PubMedCrossRefGoogle Scholar
  40. 40.
    Mielgo A, Torres VA, Clair K, Barbero S, Stupack DG (2009) Paclitaxel promotes a caspase 8-mediated apoptosis through death effector domain association with microtubules. Oncogene 28:3551–3562PubMedCrossRefGoogle Scholar
  41. 41.
    Park SJ, Wu CH, Gordon JD, Zhong X, Emami A, Safa AR (2004) Taxol induces caspase-10-dependent apoptosis. J Biol Chem 279:51057–51067PubMedCrossRefGoogle Scholar
  42. 42.
    Haldar S, Jena N, Croce CM (1995) Inactivation of Bcl-2 by phosphorylation. Proc Natl Acad Sci USA 92:4507–4511PubMedCrossRefGoogle Scholar
  43. 43.
    Kim R (2005) Recent advances in understanding the cell death pathways activated by anticancer therapy. Cancer 103:1551–1560PubMedCrossRefGoogle Scholar
  44. 44.
    Cregan SP, Dawson VL, Slack RS (2004) Role of AIF in caspase dependent and caspase-independent cell death. Oncogene 23:2785–2796PubMedCrossRefGoogle Scholar
  45. 45.
    Polster BM, Basañez G, Etxebarria A, Hardwick JM, Nicholls DG (2005) Calpain I induces cleavage and release of apoptosis-inducing factor from isolated mitochondria. J Biol Chem 280:6447–6454PubMedCrossRefGoogle Scholar
  46. 46.
    Catalano MG, Poli R, Pugliese M, Fortunati N, Boccuzzi G (2007) Valproic acid enhances tubulin acetylation and apoptotic activity of taxol on anaplastic thyroid cancer cell lines. Endocr Relat Cancer 14:839–845PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Subhasree Roy Choudhury
    • 1
  • Surajit Karmakar
    • 1
  • Naren L. Banik
    • 2
  • Swapan K. Ray
    • 1
  1. 1.Department of Pathology, Microbiology, and ImmunologyUniversity of South Carolina School of MedicineColumbiaUSA
  2. 2.Department of NeurosciencesMedical University of South CarolinaCharlestonUSA

Personalised recommendations