Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

Effects of cucurbitacins on cell morphology are associated with sensitization of renal carcinoma cells to TRAIL-induced apoptosis

Abstract

Cucurbitacins B and D were among the compounds identified as sensitizers of cancer cells to TRAIL-mediated apoptosis in a high-throughput screen. Therefore a series of cucurbitacins was further investigated for TRAIL sensitization and possible mechanisms of action. A total of six cucurbitacins promoted TRAIL-induced apoptosis (B, I, E, C, D, and K) and one (P) was inactive. Sensitization of renal adenocarcinoma cells to TRAIL was apparent after as little as 1–4 h pretreatment and did not require continued presence of cucurbitacin. Active cucurbitacins induced caspase-8 activation only after subsequent TRAIL addition and caspase activation was required for apoptosis suggesting amplified proximal signaling from TRAIL death receptors. Cucurbitacin-sensitized TRAIL-induced cytotoxicity was inhibited by N-acetyl cysteine. Structure–activity relationship analysis in comparison to published studies suggests that TRAIL-sensitizing and general cytotoxic activities of cucurbitacins may be decoupled. Cucurbitacins are reported to be inhibitors of STAT3 activation. However, their TRAIL-sensitizing activity is STAT3-independent. Treatment of renal carcinoma cells with active cucurbitacins produced rapid and dramatic changes in cell morphology and cytoskeletal organization (also prevented by NAC). Therefore, cucurbitacins may be useful as tools for investigating the molecular mechanism(s) of action of TRAIL sensitizers, particularly with regard to temporal aspects of sensitization and modulation of TRAIL signaling by cell morphology, and could form the basis for future therapeutic development in combination with TRAIL death receptor agonists.

This is a preview of subscription content, log in to check access.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

References

  1. 1.

    Pavet V, Portal MM, Moulin JC, Herbrecht R, Gronemeyer H (2010) Towards novel paradigms for cancer therapy. Oncogene 30:1–20

  2. 2.

    Holoch PA, Griffith TS (2009) TNF-related apoptosis-inducing ligand (TRAIL): a new path to anti-cancer therapies. Eur J Pharmacol 625:63–72

  3. 3.

    Mahmood Z, Shukla Y (2010) Death receptors: targets for cancer therapy. Exp Cell Res 316:887–899

  4. 4.

    Gonzalvez F, Ashkenazi A (2010) New insights into apoptosis signaling by Apo2L/TRAIL. Oncogene 29:4752–4765

  5. 5.

    Sayers TJ (2011) Targeting the extrinsic apoptosis signaling pathway for cancer therapy. Cancer Immunol Immunother 60:1173–1180

  6. 6.

    Ivanov VN, Bhoumik A, Ronai Z (2003) Death receptors and melanoma resistance to apoptosis. Oncogene 22:3152–3161

  7. 7.

    Rahman M, Pumphrey JG, Lipkowitz S (2009) The TRAIL to targeted therapy of breast cancer. Adv Cancer Res 103:43–73

  8. 8.

    Testa U (2010) TRAIL/TRAIL-R in hematologic malignancies. J Cell Biochem 110:21–34

  9. 9.

    Stegehuis JH, de Witt LH, de Vries EG, Groen HJ, de Jong S, Kruyt FA (2010) TRAIL receptor targeting therapies for non-small cell lung cancer: current status and perspectives. Drug Resist Updates 13:2–15

  10. 10.

    Booth NL, Sayers TJ, Brooks AD, Thomas CL, Jacobsen K, Goncharova EI, McMahon JB, Henrich CJ (2009) A cell-based high-throughput screen to identify synergistic TRAIL sensitizers. Cancer Immunol Immunother 58:1229–1244

  11. 11.

    Lee DH, Iwanski GB, Thoennissen NH (2010) Cucurbitacin: ancient compound shedding new light on cancer treatment. ScientificWorldJournal 5:413–418

  12. 12.

    Deng J, Grande F, Neamati N (2007) Small molecule inhibitors of Stat3 signaling pathway. Curr Cancer Drug Targets 7:91–107

  13. 13.

    Aggarwal BB, Sethi G, Ahn KS, Sandur SK, Pandey MK, Kunnumakkara AB, Sung B, Ichikawa H (2006) Targeting signal-transducer-and-activator-of-transcription-3 for prevention and therapy of cancer: modern target but ancient solution. Ann NY Acad Sci 1091:151–169

  14. 14.

    Yasuda S, Yogosawa S, Izutani Y, Nakamura Y, Watanabe H, Sakai T (2010) Cucurbitacin B induces G2 arrest and apoptosis via a reactive oxygen species-dependent mechanism in human colon adenocarcinoma SW480 cells. Mol Nutr Food Res 54:559–565

  15. 15.

    Finnberg N, Klein-Szanto AJ, El-Deiry WS (2008) TRAIL-R deficiency in mice promotes susceptibility to chronic inflammation and tumorigenesis. J Clin Invest 118:111–123

  16. 16.

    Huang S, Sinicrope FA (2010) Sorafenib inhibits STAT3 activation to enhance TRAIL-mediated apoptosis in human pancreatic cancer cells. Mol Cancer Ther 9:742–750

  17. 17.

    Lanuti P, Bertagnolo V, Pierdomenico L, Bascelli A, Santavenere E, Alinari L, Capitani S, Miscia S, Marchisio M (2009) Enhancement of TRAIL cytotoxicity by AG-490 in human ALL cells is characterized by downregulation of cIAP-1 and cIAP-2 through inhibition of JAK2/STAT3. Cell Res 19:1079–1089

  18. 18.

    Kusuba M, Nakao K, Goto T, Nishimura D, Kawashimo H, Shibata H, Motoyoshi Y, Taura N, Ichikawa T, Hamasaki K, Eguchi K (2007) Abrogation of constitutive STAT3 activity sensitizes human hepatoma cells to TRAIL-mediated apoptosis. J Hepatol 47:546–555

  19. 19.

    Kannappan R, Ravindran J, Prasad S, Sung B, Yadav VR, Reuter S, Chaturvedi MM, Aggarwal BB (2010) Gamma-tocotrienol promotes TRAIL-induced apoptosis through reactive oxygen species/extracellular signal-regulated kinase/p53-mediated upregulation of death receptors. Mol Cancer Ther 9:2196–2207

  20. 20.

    Lee TJ, Um HJ, Min do S, Park JW, Choi KS, Kwon TK (2009) Withaferin A sensitizes TRAIL-induced apoptosis through reactive oxygen species-mediated up-regulation of death receptor 5 and down-regulation of c-FLIP. Free Radic Biol Med 46:1639–1649

  21. 21.

    Sung B, Ravindran J, Prasad S, Pandey MK, Aggarwal BB (2010) Gossypol induces death receptor-5 through activation of the ROS-ERK-CHOP pathway and sensitizes colon cancer cells to TRAIL. J Biol Chem 285:35418–35427

  22. 22.

    Brooks AD, Jacobsen KM, Li W, Shanker A, Sayers TJ (2010) Bortezomib sensitizes human renal cell carcinomas to TRAIL apoptosis through increased activation of caspase-8 in the death-inducing signaling complex. Mol Cancer Res 8:729–738

  23. 23.

    Li L, Cheung S-h, Evans EL, Shaw PE (2010) Modulation of gene expression and tumor cell growth by redox modification of STAT3. Cancer Res 70:8222–8232

  24. 24.

    Ishidorj G, Johnston JB, Gibson SB (2010) Inhibition of constitutive activation of STAT3 by cucurbitacin-I (JSI-124) sensitized human B-leukemia cells to apoptosis. Mol Cancer Ther 9:3302–3314

  25. 25.

    Ganten TM, Koschny R, Haas TL, Sykora J, Li-Weber M, Herzer K, Walczak H (2005) Proteasome inhibition sensitizes hepatocellular carcinoma cells, but not human hepatocytes, to TRAIL. Hepatology 42:588–597

  26. 26.

    Hellwig CT, Kohler BF, Lehtivarjo A-K, Dussmann H, Courtney MJ, Prehn JHM, Rehm M (2008) Real time analysis of tumor necrosis factor-related apoptosis-inducing ligand/cycloheximide-induced caspase activities during apoptosis initiation. J Biol Chem 283:21676–21685

  27. 27.

    Oberst A, Pop C, Tremblay AG, Blais V, Denault J-B, Salvesen GS, Green DR (2010) Inducible dimerization and inducible cleavage reveal a requirement for both processes in caspase-8 activation. J Biol Chem 285:16632–16642

  28. 28.

    Pennarun B, Meijer A, de Vries EGE, Kleibeuker JH, Kruyt F, de Jong S (2010) Playing the DISC: turning on TRAIL death receptor-mediated apoptosis in cancer. Biochim Biophys Acta 1805:123–140

  29. 29.

    Zafarullah M, Li WQ, Sylvester J, Ahmad M (2003) Molecular mechanisms of N-acetylcysteine actions. Cell Mol Life Sci 60:6–20

  30. 30.

    Yin D, Wakimoto N, Xing H, Lu D, Huynh T, Wang X, Black KL, Koeffler HP (2008) Cucurbitacin B markedly inhibits growth and rapidly affects the cytoskeleton in glioblastoma multiforme. Int J Cancer 123:1364–1375

  31. 31.

    Haritunians T, Gueller S, Zhang L, Badr R, Yin D, Xing H, Fung MC, Koeffler HP (2008) Cucurbitacin B induces differentiation, cell cycle arrest, and actin cytoskeletal alterations in myeloid leukemia cells. Leuk Res 32:1366–1373

  32. 32.

    Phipps LE, Hino S, Muschel RJ (2011) Targeting cell spreading: a method of sensitizing metastatic tumor cells to TRAIL-induced apoptosis. Mol Cancer Res 9:249–258

  33. 33.

    Afifi MS, Ross SA, ElSohly MA, Naeem ZE, Halaweish FT (1999) Cucurbitacins of Cucumis prophetarum. J Chem Ecol 25:847–859

  34. 34.

    Bartalis J, Halaweish FT (2005) Relationship between cucurbitacins reversed-phase high-performance liquid chromatography hydrophobicity index and basal cytotoxicity on HepG2 cells. J Chromatogr B 818:159–166

  35. 35.

    van Dang G, Rode BM, Stuppner H (1994) Quantitative electronic structure–activity relationship (QESAR) of natural cytotoxic compounds: maytansinoids, quassinoids and cucurbitacins. Eur J Pharm Sci 2:331–350

  36. 36.

    Sun J, Baskovich MA, Jove R, Livingston SK, Coppola D, Sebti SM (2005) Cucurbitacin Q: a selective STAT3 activation inhibitor with potent antitumor activity. Oncogene 24:3236–3245

  37. 37.

    Gamero AM, Young HA, Wiltrout RH (2004) Inactivation of Stat3 in tumor cells: releasing a brake on immune responses against cancer? Cancer Cell 5:111–112

  38. 38.

    Aggarwal BB, Kunnumakkara AB, Harikumar KB, Gupta SR, Tharakan ST, Koca C, Day S, Sung B (2009) Signal transducer and activator of transcription-3, inflammation, and cancer: how intimate is the relationship? Ann NY Acad Sci 1171:59–76

  39. 39.

    Boykin C, Zhang G, Chen YH, Zhang RW, Fan XE, Yang WM, Lu Q (2011) Cucurbitacin IIa: a novel class of anti-cancer drug inducing non-reversible actin aggregation and inhibiting survivin independent of JAK2/STAT3 phosphorylation. Br J Cancer 104:781–789

  40. 40.

    Iwanski GB, Lee DH, En-Gal S, Doan NB, Castor B, Vogt M, Toh M, Bokemeyer C, Said JW, Thoennissen NH, Koeffler HP (2010) Cucurbitacin B, a novel in vivo potentiator of gemcitabine with low toxicity in the treatment of pancreatic cancer. Br J Pharmacol 160:998–1007

Download references

Acknowledgments

Thanks to Heidi Bokesh and Kirk Gustafson for performing and interpreting chemical analysis to confirm identities of the compounds obtained from the DTP repository. Thanks also to Anna Maciag for assistance with ROS assays and to Tommy Turbyville for help with actin staining and confocal microscopy. This project has been funded in whole or in part with Federal funds from the national Cancer Institute, National Institutes of Health, under contract HHSN261200800001E. The content of this publication does not necessarily reflect the views or policies of the Department of Health and Human Services, nor does mention of trade names, commercial products or organizations imply endorsement by the US Government. This research was supported (in part) by the Intramural Research Program of NIH, National Cancer Institute, Center for Cancer Research.

Conflict of interest

The authors declare that they have no conflict of interest.

Author information

Correspondence to Curtis J. Henrich.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (DOCX 333 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Henrich, C.J., Thomas, C.L., Brooks, A.D. et al. Effects of cucurbitacins on cell morphology are associated with sensitization of renal carcinoma cells to TRAIL-induced apoptosis. Apoptosis 17, 79–89 (2012). https://doi.org/10.1007/s10495-011-0652-7

Download citation

Keywords

  • Cucurbitacins
  • TRAIL
  • TRAIL sensitizers
  • Apoptosis
  • STAT3
  • Cell morphology