Top

Published in:

01-12-2010 | PRECLINICAL STUDIES

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

Authors: Subhasree Roy Choudhury, Surajit Karmakar, Naren L. Banik, Swapan K. Ray

Published in: Investigational New Drugs | Issue 6/2010

Summary

Neuroblastoma is an extracranial, solid, and heterogeneous malignancy in children. The conventional therapeutic modalities are mostly ineffective and thus new therapeutic strategies for malignant neuroblastoma are urgently warranted. We examined the synergistic efficacy of combination of sorafenib (SF) and genistein (GST) in human malignant neuroblastoma SK-N-DZ (N-Myc amplified) and SH-SY5Y (N-Myc non-amplified) cell lines. MTT assay showed dose-dependent decrease in cell viability and the combination therapy more prominently inhibited the cell proliferation in both cell lines than either treatment alone. Apoptosis was confirmed morphologically by Wright staining. Flow cytometric analysis of cell cycle phase distribution and Annexin V-FITC/PI staining showed increase in subG1 DNA content and early apoptosis, respectively, after treatment with the combination of drugs. Apoptosis was further confirmed by scanning electron microscopy. Combination therapy showed activation of caspase-8, cleavage of Bid to tBid, increase in p53 and p21 expression, down regulation of anti-apoptotic Mcl-1, and increase in Bax:Bcl-2 ratio to trigger apoptosis. Down regulation of MDR, hTERT, N-Myc, VEGF, FGF-2, NF-κB, p-Akt, and c-IAP2 indicated suppression of angiogenic and survival pathways. Mitochondrial release of cytochrome c and Smac into cytosol indicated involvement of mitochondia in apoptosis. Increases in proteolytic activities of calpain and caspase-3 were also confirmed. Our results suggested that combination of SF and GST inhibited angiogenic and survival factors and increased apoptosis via receptor and mitochondria mediated pathways in both neuroblastoma SK-N-DZ and SH-SY5Y cell lines. Thus, this combination of drugs could be a potential therapeutic strategy against human malignant neuroblastoma cells having N-Myc amplification or non-amplification.

Hiyama E, Iehara T, Sugimoto T, Fukuzawa M, Hayashi Y, Sasaki F, Sugiyama M, Kondo S, Yoneda A, Yamaoka H, Tajiri T, Akazawa K, Ohtaki M (2008) Effectiveness of screening for neuroblastoma at 6 months of age: a retrospective population-based cohort study. Lancet 371:1173–1180CrossRefPubMed

Brown A, Jolly P, Wei H (1998) Genistein modulates neuroblastoma cell proliferation and differentiation through induction of apoptosis and regulation of tyrosine kinase activity and N-myc expression. Carcinogenesis 19:991–997CrossRefPubMed

Marcus K, Johnson M, Adam RM, O'Reilly MS, Donovan M, Atala A, Freeman MR, Soker S (2005) Tumor cell-associated neuropilin-1 and vascular endothelial growth factor expression as determinants of tumor growth in neuroblastoma. Neuropathology 25:178–187CrossRefPubMed

Peet AC, McConville C, Wilson M, Levine BA, Reed M, Dyer SA, Edwards EC, Strachan MC, McMullan DJ, Wilkes TM, Grundy RG (2007) 1H MRS identifies specific metabolite profiles associated with MYCN-amplified and non-amplified tumour subtypes of neuroblastoma cell lines. NMR Biomed 20:692–700CrossRefPubMed

Huynh H, Lee JW, Chow PK, Ngo VC, Lew GB, Lam IW, Ong HS, Chung A, Soo KC (2009) Sorafenib induces growth suppression in mouse models of gastrointestinal stromal tumor. Mol Cancer Ther 8:152–159CrossRefPubMed

Jane EP, Premkumar DR, Pollack IF (2006) Coadministration of sorafenib with rottlerin potently inhibits cell proliferation and migration in human malignant glioma cells. J Pharmacol Exp Ther 319:1070–1080CrossRefPubMed

Yang F, Van Meter TE, Buettner R, Hedvat M, Liang W, Kowolik CM, Mepani N, Mirosevich J, Nam S, Chen MY, Tye G, Kirschbaum M, Jove R (2008) Sorafenib inhibits signal transducer and activator of transcription 3 signaling associated with growth arrest and apoptosis of medulloblastomas. Mol Cancer Ther 7:3519–3526CrossRefPubMed

Sarkar FH, Adsule S, Padhye S, Kulkarni S, Li Y (2006) The role of genistein and synthetic derivatives of isoflavone in cancer prevention and therapy. Mini Rev Med Chem 6:401–407CrossRefPubMed

Li Z, Li J, Mo B, Hu C, Liu H, Qi H, Wang X, Xu J (2008) Genistein induces cell apoptosis in MDA-MB-231 breast cancer cells via the mitogen-activated protein kinase pathway. Toxicol In Vitro 22:1749–1753CrossRefPubMed

10.

Sergeev IN (2004) Genistein induces Ca²⁺-mediated, calpain/caspase-12-dependent apoptosis in breast cancer cells. Biochem Biophys Res Commun 321:462–467CrossRefPubMed

11.

Das A, Banik NL, Ray SK (2006) Mechanism of apoptosis with the involvement of calpain and caspase cascades in human malignant neuroblastoma SH-SY5Y cells exposed to flavonoids. Int J Cancer 119:2575–2585CrossRefPubMed

12.

Purnanam KL, Boustany RM (1999) In Assessment of cell viability and histochemical methods in apoptosis. In: Hannun YA, Boustany RM (eds) Apoptosis in neurobiology. CRC, New York, pp 129–152

13.

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 141:1265–1280CrossRefPubMed

14.

Karmakar S, Banik NL, Patel SJ, Ray SK (2007) 5-Aminolevulinic acid-based photodynamic therapy suppressed survival factors and activated proteases for apoptosis in human glioblastoma U87MG cells. Neurosci Lett 415:242–247CrossRefPubMed

15.

Ashkenazi A, Dixit VM (1998) Death receptors: signaling and modulation. Science 281:1305–1308CrossRefPubMed

16.

Choi YH, Choi BT, Lee WH, Rhee SH, Park KY (2001) Doenjang hexane fraction-induced G1 arrest is associated with the inhibition of pRB phosphorylation and induction of Cdk inhibitor p21 in human breast carcinoma MCF-7 cells. Oncol Rep 8:1091–1096PubMed

17.

Shusterman S, Maris JM (2005) Prospects for therapeutic inhibition of neuroblastoma angiogenesis. Cancer Lett 228:171–179CrossRefPubMed

18.

Zaghloul N, Hernandez SL, Bae JO, Huang J, Fisher JC, Lee A, Kadenhe-Chiweshe A, Kandel JJ, Yamashiro DJ (2009) Vascular endothelial growth factor blockade rapidly elicits alternative proangiogenic pathways in neuroblastoma. Int J Oncol 34:401–407PubMed

19.

Ribatti D, Marimpietri D, Pastorino F, Brignole C, Nico B, Vacca A, Ponzonim M (2004) Angiogenesis in neuroblastoma. Ann N Y Acad Sci USA 1028:133–142CrossRef

20.

Ray SK, Karmakar S, Nowak MW, Banik NL (2006) Inhibition of calpain and caspase-3 prevented apoptosis and preserved electrophysiological properties of voltage-gated and ligand-gated ion channels in rat primary cortical neurons exposed to glutamate. Neuroscience 139:577–595CrossRefPubMed

21.

Tran MA, Smith CD, Kester M, Robertson GP (2008) Combining nanoliposomal ceramide with sorafenib synergistically inhibits melanoma and breast cancer cell survival to decrease tumor development. Clin Cancer Res 14:3571–3581CrossRefPubMed

22.

Lasithiotakis KG, Sinnberg TW, Schittek B, Flaherty KT, Kulms D, Maczey E, Garbe C, Meier FE (2008) Combined inhibition of MAPK and mTOR signaling inhibits growth, induces cell death, and abrogates invasive growth of melanoma cells. J Invest Dermatol 128:2013–2023CrossRefPubMed

23.

Das A, Banik NL, Ray SK (2009) Retinoids induce differentiation and downregulate telomerase activity and N-Myc to increase sensitivity to flavonoids for apoptosis in human malignant neuroblastoma SH-SY5Y cells. Int J Oncol 34:757–765PubMed

24.

Ismail IA, Kang KS, Lee HA, Kim JW, Sohn YK (2007) Genistein-induced neuronal apoptosis and G2/M cell cycle arrest is associated with MDC1 up-regulation and PLK1 down-regulation. Eur J Pharmacol 575:12–20CrossRefPubMed

25.

Thasni KA, Rojini G, Rakesh SN, Ratheeshkumar T, Babu MS, Srinivas G, Banerji A, Srinivas P (2008) Genistein induces apoptosis in ovarian cancer cells via different molecular pathways depending on Breast Cancer Susceptibility gene-1 (BRCA1) status. Eur J Pharmacol 588:158–164CrossRefPubMed

26.

Di Francesco AM, Meco D, Torella AR, Barone G, D'Incalci M, Pisano C, Carminati P, Riccardi R (2007) The novel atypical retinoid ST1926 is active in ATRA resistant neuroblastoma cells acting by a different mechanism. Biochem Pharmacol 73:643–655CrossRefPubMed

27.

Goldschneider D, Blanc E, Raguénez G, Barrois M, Legrand A, Le Roux G, Haddada H, Bénard J, Douc-Rasy S (2004) Differential response of p53 target genes to p73 overexpression in SH-SY5Y neuroblastoma cell line. J Cell Sci 117:293–301CrossRefPubMed

28.

Moll UM, LaQuaglia M, Bénard J, Riou G (1995) Wild-type p53 protein undergoes cytoplasmic sequestration in undifferentiated neuroblastomas but not in differentiated tumors. Proc Natl Acad Sci USA 92:4407–4411CrossRefPubMed

29.

Zhang W, Konopleva M, Ruvolo VR, McQueen T, Evans RL, Bornmann WG, McCubrey J, Cortes J, Andreeff M (2008) Sorafenib induces apoptosis of AML cells via Bim-mediated activation of the intrinsic apoptotic pathway. Leukemia 22:808–818CrossRefPubMed

30.

Yeh TC, Chiang PC, Li TK, Hsu JL, Lin CJ, Wang SW, Peng CY, Guh JH (2007) Genistein induces apoptosis in human hepatocellular carcinomas via interaction of endoplasmic reticulum stress and mitochondrial insult. Biochem Pharmacol 73:782–792CrossRefPubMed

31.

Panka DJ, Wang W, Atkins MB, Mier JW (2006) The Raf inhibitor BAY 43–9006 (Sorafenib) induces caspase-independent apoptosis in melanoma cells. Cancer Res 66:1611–1619CrossRefPubMed

32.

Castro AF, Altenberg GA (1997) Inhibition of drug transport by genistein in multidrug-resistant cells expressing P-glycoprotein. Biochem Pharmacol 53:89–93CrossRefPubMed

33.

Krams M, Rudolph P, Harms D (2004) Proliferation and hTERT expression in neuroblastoma. Pathologe 25:317–323CrossRefPubMed

34.

Matsumoto M (1991) Inhibitors for protein tyrosine kinases, erbstatin, genistein and herbimycin A, induce differentiation of human neural tumor cell lines. Nippon Geka Hokan 60:113–121PubMed

35.

Jagadeesh S, Kyo S, Banerjee PP (2006) Genistein represses telomerase activity via both transcriptional and posttranslational mechanisms in human prostate cancer cells. Cancer Res 66:2107–2115CrossRefPubMed

36.

Eggert A, Ikegaki N, Kwiatkowski J, Zhao H, Brodeur GM, Himelstein BP (2000) High-level expression of angiogenic factors is associated with advanced tumor stage in human neuroblastomas. Clin Can Res 6:1900–1908

37.

Schomber T, Zumsteg A, Strittmatter K, Crnic I, Antoniadis H, Littlewood-Evans A, Wood J, Christofori G (2009) Differential effects of the vascular endothelial growth factor receptor inhibitor PTK787/ZK222584 on tumor angiogenesis and tumor lymphangiogenesis. Mol Cancer Ther 8:55–63CrossRefPubMed

38.

Murphy DA, Makonnen S, Lassoued W, Feldman MD, Carter C, Lee WM (2006) Inhibition of tumor endothelial ERK activation, angiogenesis, and tumor growth by sorafenib (BAY43–9006). Am J Pathol 69:1875–1885CrossRef

39.

Beg AA, Baltimore B (1996) An essential role for NF-κB in preventing TNF-α-induced cell death. Science 274:782–784CrossRefPubMed

40.

Li Y, Ahmed F, Ali S, Philip PA, Kucuk O, Sarkar FH (2005) Inactivation of nuclear factor kappaB by soy isoflavone genistein contributes to increased apoptosis induced by chemotherapeutic agents in human cancer cells. Cancer Res 65:6934–6942CrossRefPubMed

41.

Pelloski CE, Lin E, Zhang L, Yung WK, Colman H, Liu JL, Woo SY, Heimberger AB, Suki D, Prados M, Chang S, Barker FG III, Fuller GN, Aldape KD (2006) Prognostic associations of activated mitogen-activated protein kinase and Akt pathways in glioblastoma. Clin Cancer Res 12:3935–3941CrossRefPubMed

42.

Wang CY, Mayo MW, Korneluk RG, Goeddel DV, Baldwin AS Jr (1998) NF-κB antiapoptosis: induction of TRAF1 and TRAF2 and c-IAP1 and c-IAP2 to suppress caspase-8 activation. Science 281:1680–1683CrossRefPubMed

43.

Du C, Fang M, Li Y, Li L, Wang X (2000) Smac, a mitochondrial protein that promotes cytochrome c-dependent caspase activation by eliminating IAP inhibition. Cell 102:33–42CrossRefPubMed

44.

Makin GW, Corfe BM, Griffiths GJ, Thistlethwaite A, Hickman JA, Dive C (2001) Damage-induced Bax N-terminal change, translocation to mitochondria and formation of Bax dimers/complexes occur regardless of cell fate. EMBO J 20:6306–6315CrossRefPubMed

45.

Rahmani M, Davis EM, Bauer C, Dent P, Grant S (2005) Apoptosis induced by the kinase inhibitor BAY 43–9006 in human leukemia cells involves down-regulation of Mcl-1 through inhibition of translation. J Biol Chem 280:35217–35227CrossRefPubMed

Title: 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
Authors: Subhasree Roy Choudhury
Surajit Karmakar
Naren L. Banik
Swapan K. Ray
Publication date: 01-12-2010
Publisher: Springer US
Published in: Investigational New Drugs / Issue 6/2010
Print ISSN: 0167-6997
Electronic ISSN: 1573-0646
DOI: https://doi.org/10.1007/s10637-009-9324-7

Webinar | 19-02-2024 | 17:30 (CET)

Keynote webinar | Spotlight on antibody–drug conjugates in cancer

Watch now

Antibody–drug conjugates (ADCs) are novel agents that have shown promise across multiple tumor types. Explore the current landscape of ADCs in breast and lung cancer with our experts, and gain insights into the mechanism of action, key clinical trials data, existing challenges, and future directions.

Dr. Véronique Diéras

Prof. Fabrice Barlesi

Developed by: Springer Medicine

Keynote webinar | Spotlight on medication adherence

Springer Medicine

Summary

Please log in to get access to this content

Other articles of this Issue 6/2010

TAE226-mediated inhibition of focal adhesion kinase interferes with tumor angiogenesis and vasculogenesis

DH9, a novel PPARγ agonist suppresses the proliferation of ADPKD epithelial cells: An association with an inhibition of β-catenin signaling

Lovastatin overcomes gefitinib resistance in human non-small cell lung cancer cells with K-Ras mutations

New SRC/ABL inhibitors for chronic myeloid leukemia therapy show selectivity for T315I ABL mutant CD34+ cells

The epothilone B analogue ixabepilone in patients with advanced hepatobiliary cancers: a trial of the University of Chicago Phase II Consortium

Scaling the time-course of myelosuppression from rats to patients with a semi-physiological model

Keynote webinar | Spotlight on antibody–drug conjugates in cancer