Skip to main content
SpringerLink
Log in

Myeloid-derived suppressor cell function is reduced by Withaferin A, a potent and abundant component of Withania somnifera root extract

  • Original Article
  • Published:
Cancer Immunology, Immunotherapy Aims and scope Submit manuscript

Abstract

Myeloid cells play a crucial role in tumor progression. The most common tumor-infiltrating myeloid cells are myeloid-derived suppressor cells (MDSC) and tumor-associated macrophages (TAMs). These cells promote tumor growth by their inherent immune suppressive activity which is enhanced by their cross-talk. The root extract of the plant Withania somnifera (Ashwagandha) (WRE) has been reported to reduce tumor growth. HPLC analysis identified Withaferin A (WA) as the most abundant constituent of WRE and led us to determine whether the anti-tumor effects of WRE and WA involve modulating MDSC and TAM activity. A prominent effect of MDSC is their production of IL-10 which increases upon cross-talk with macrophages, thus polarizing immunity to a pro-tumor type 2 phenotype. In vitro treatment with WA decreased MDSC production of IL-10 and prevented additional MDSC production of IL-10 generated by MDSC–macrophage cross-talk. Macrophage secretion of IL-6 and TNFα, cytokines that increase MDSC accumulation and function, was also reduced by in vitro treatment with WA. Much of the T-cell suppressive activity of MDSC is due to MDSC production of reactive oxygen species (ROS), and WA significantly reduced MDSC production of ROS through a STAT3-dependent mechanism. In vivo treatment of tumor-bearing mice with WA decreased tumor weight, reduced the quantity of granulocytic MDSC, and reduced the ability of MDSC to suppress antigen-driven activation of CD4+ and CD8+ T cells. Thus, adjunctive treatment with WA reduced myeloid cell-mediated immune suppression, polarized immunity toward a tumor-rejecting type 1 phenotype, and may facilitate the development of anti-tumor immunity.

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

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  1. Nagaraj S, Gabrilovich DI (2010) Myeloid-derived suppressor cells in human cancer. Cancer J 16:348–353. doi:10.1097/PPO.0b013e3181eb3358

    Article  PubMed  CAS  Google Scholar 

  2. Kodumudi KN, Weber A, Sarnaik AA, Pilon-Thomas S (2012) Blockade of myeloid-derived suppressor cells after induction of lymphopenia improves adoptive T cell therapy in a murine model of melanoma. J Immunol 189:5147–5154. doi:10.4049/jimmunol.1200274

    Article  PubMed  CAS  Google Scholar 

  3. Ostrand-Rosenberg S (2010) Myeloid-derived suppressor cells: more mechanisms for inhibiting antitumor immunity. Cancer Immunol Immunother 59:1593–1600. doi:10.1007/s00262-010-0855-8

    Article  PubMed  Google Scholar 

  4. Sinha P, Clements VK, Ostrand-Rosenberg S (2005) Interleukin-13-regulated M2 macrophages in combination with myeloid suppressor cells block immune surveillance against metastasis. Cancer Res 65:11743–11751. doi:10.1158/0008-5472.CAN-05-0045

    Article  PubMed  CAS  Google Scholar 

  5. Sinha P, Clements VK, Ostrand-Rosenberg S (2005) Reduction of myeloid-derived suppressor cells and induction of M1 macrophages facilitate the rejection of established metastatic disease. J Immunol 174:636–645

    PubMed  CAS  Google Scholar 

  6. Sinha P, Clements VK, Bunt SK, Albelda SM, Ostrand-Rosenberg S (2007) Cross-talk between myeloid-derived suppressor cells and macrophages subverts tumor immunity toward a type 2 response. J Immunol 179:977–983

    PubMed  CAS  Google Scholar 

  7. Christina AJ, Joseph DG, Packialakshmi M, Kothai R, Robert SJ, Chidambaranathan N, Ramasamy M (2004) Anticarcinogenic activity of Withania somnifera Dunal against Dalton’s ascitic lymphoma. J Ethnopharmacol 93:359–361. doi:10.1016/j.jep.2004.04.004

    Article  PubMed  CAS  Google Scholar 

  8. Muralikrishnan G, Dinda AK, Shakeel F (2010) Immunomodulatory effects of Withania somnifera on azoxymethane induced experimental colon cancer in mice. Immunol Invest 39:688–698. doi:10.3109/08820139.2010.487083

    Article  PubMed  Google Scholar 

  9. Padmavathi B, Rath PC, Rao AR, Singh RP (2005) Roots of Withania somnifera inhibit forestomach and skin carcinogenesis in mice. Evid Based Complement Alternat Med 2:99–105. doi:10.1093/ecam/neh064

    Article  PubMed  Google Scholar 

  10. Senthilnathan P, Padmavathi R, Magesh V, Sakthisekaran D (2006) Chemotherapeutic efficacy of paclitaxel in combination with Withania somnifera on benzo(a)pyrene-induced experimental lung cancer. Cancer Sci 97:658–664. doi:10.1111/j.1349-7006.2006.00224.x

    Article  PubMed  CAS  Google Scholar 

  11. Leyon PV, Kuttan G (2004) Effect of Withania somnifera on B16F-10 melanoma induced metastasis in mice. Phytother Res 18:118–122. doi:10.1002/ptr.1378

    Article  PubMed  CAS  Google Scholar 

  12. Davis L, Kuttan G (2002) Effect of Withania somnifera on CTL activity. J Exp Clin Cancer Res 21:115–118

    PubMed  CAS  Google Scholar 

  13. Srinivasan S, Ranga RS, Burikhanov R, Han SS, Chendil D (2007) Par-4-dependent apoptosis by the dietary compound Withaferin A in prostate cancer cells. Cancer Res 67:246–253. doi:10.1158/0008-5472.CAN-06-2430

    Article  PubMed  CAS  Google Scholar 

  14. Oh JH, Lee TJ, Kim SH, Choi YH, Lee SH, Lee JM, Kim YH, Park JW, Kwon TK (2008) Induction of apoptosis by Withaferin A in human leukemia U937 cells through down-regulation of Akt phosphorylation. Apoptosis 13:1494–1504. doi:10.1007/s10495-008-0273-y

    Article  PubMed  CAS  Google Scholar 

  15. Stan SD, Hahm ER, Warin R, Singh SV (2008) Withaferin A causes FOXO3a- and Bim-dependent apoptosis and inhibits growth of human breast cancer cells in vivo. Cancer Res 68:7661–7669. doi:10.1158/0008-5472.CAN-08-1510

    Article  PubMed  CAS  Google Scholar 

  16. Mandal C, Dutta A, Mallick A, Chandra S, Misra L, Sangwan RS (2008) Withaferin A induces apoptosis by activating p38 mitogen-activated protein kinase signaling cascade in leukemic cells of lymphoid and myeloid origin through mitochondrial death cascade. Apoptosis 13:1450–1464. doi:10.1007/s10495-008-0271-0

    Article  PubMed  CAS  Google Scholar 

  17. Davis L, Kuttan G (2001) Effect of Withania somnifera on DMBA induced carcinogenesis. J Ethnopharmacol 75:165–168

    Article  PubMed  CAS  Google Scholar 

  18. Koduru S, Kumar R, Srinivasan S, Evers MB, Damodaran C (2010) Notch-1 inhibition by Withaferin-A: a therapeutic target against colon carcinogenesis. Mol Cancer Ther 9:202–210. doi:10.1158/1535-7163.MCT-09-0771

    Article  PubMed  CAS  Google Scholar 

  19. Lee J, Sehrawat A, Singh SV (2012) Withaferin A causes activation of Notch2 and Notch4 in human breast cancer cells. Breast Cancer Res Treat 136:45–56. doi:10.1007/s10549-012-2239-6

    Article  PubMed  CAS  Google Scholar 

  20. Ichikawa H, Takada Y, Shishodia S, Jayaprakasam B, Nair MG, Aggarwal BB (2006) Withanolides potentiate apoptosis, inhibit invasion, and abolish osteoclastogenesis through suppression of nuclear factor-kappaB (NF-kappaB) activation and NF-kappaB-regulated gene expression. Mol Cancer Ther 5:1434–1445. doi:10.1158/1535-7163.MCT-06-0096

    Article  PubMed  CAS  Google Scholar 

  21. Kaileh M, Vanden Berghe W, Heyerick A, Horion J, Piette J, Libert C, De Keukeleire D, Essawi T, Haegeman G (2007) Withaferin a strongly elicits IkappaB kinase beta hyperphosphorylation concomitant with potent inhibition of its kinase activity. J Biol Chem 282:4253–4264. doi:10.1074/jbc.M606728200

    Article  PubMed  CAS  Google Scholar 

  22. Falsey RR, Marron MT, Gunaherath GM, Shirahatti N, Mahadevan D, Gunatilaka AA, Whitesell L (2006) Actin microfilament aggregation induced by Withaferin A is mediated by annexin II. Nat Chem Biol 2:33–38. doi:10.1038/nchembio755

    Article  PubMed  CAS  Google Scholar 

  23. Mohan R, Hammers HJ, Bargagna-Mohan P, Zhan XH, Herbstritt CJ, Ruiz A, Zhang L, Hanson AD, Conner BP, Rougas J, Pribluda VS (2004) Withaferin A is a potent inhibitor of angiogenesis. Angiogenesis 7:115–122. doi:10.1007/s10456-004-1026-3

    Article  PubMed  CAS  Google Scholar 

  24. Sinha P, Clements VK, Fulton AM, Ostrand-Rosenberg S (2007) Prostaglandin E2 promotes tumor progression by inducing myeloid-derived suppressor cells. Cancer Res 67:4507–4513. doi:10.1158/0008-5472.CAN-06-4174

    Article  PubMed  CAS  Google Scholar 

  25. Pulaski BA, Ostrand-Rosenberg S (1998) Reduction of established spontaneous mammary carcinoma metastases following immunotherapy with major histocompatibility complex class II and B7.1 cell-based tumor vaccines. Cancer Res 58:1486–1493

    PubMed  CAS  Google Scholar 

  26. Sinha P, Chornoguz O, Clements VK, Artemenko KA, Zubarev RA, Ostrand-Rosenberg S (2011) Myeloid-derived suppressor cells express the death receptor Fas and apoptose in response to T cell-expressed FasL. Blood 117:5381–5390. doi:10.1182/blood-2010-11-321752

    Article  PubMed  CAS  Google Scholar 

  27. Kusmartsev S, Nefedova Y, Yoder D, Gabrilovich DI (2004) Antigen-specific inhibition of CD8+ T cell response by immature myeloid cells in cancer is mediated by reactive oxygen species. J Immunol 172:989–999

    PubMed  CAS  Google Scholar 

  28. Corzo CA, Cotter MJ, Cheng P, Cheng F, Kusmartsev S, Sotomayor E, Padhya T, McCaffrey TV, McCaffrey JC, Gabrilovich DI (2009) Mechanism regulating reactive oxygen species in tumor-induced myeloid-derived suppressor cells. J Immunol 182:5693–5701. doi:10.4049/jimmunol.0900092

    Article  PubMed  CAS  Google Scholar 

  29. Ostrand-Rosenberg S, Sinha P (2009) Myeloid-derived suppressor cells: linking inflammation and cancer. J Immunol 182:4499–4506. doi:10.4049/jimmunol.0802740

    Article  PubMed  CAS  Google Scholar 

  30. Bronte V, Serafini P, De Santo C, Marigo I, Tosello V, Mazzoni A, Segal DM, Staib C, Lowel M, Sutter G, Colombo MP, Zanovello P (2003) IL-4-induced arginase 1 suppresses alloreactive T cells in tumor-bearing mice. J Immunol 170:270–278

    PubMed  CAS  Google Scholar 

  31. Thaiparambil JT, Bender L, Ganesh T, Kline E, Patel P, Liu Y, Tighiouart M, Vertino PM, Harvey RD, Garcia A, Marcus AI (2011) Withaferin A inhibits breast cancer invasion and metastasis at sub-cytotoxic doses by inducing vimentin disassembly and serine 56 phosphorylation. Int J Cancer 129:2744–2755. doi:10.1002/ijc.25938

    Article  PubMed  CAS  Google Scholar 

  32. Davis L, Kuttan G (2002) Effect of Withania somnifera on cell mediated immune responses in mice. J Exp Clin Cancer Res 21:585–590

    PubMed  CAS  Google Scholar 

  33. Corzo CA, Condamine T, Lu L, Cotter MJ, Youn JI, Cheng P, Cho HI, Celis E, Quiceno DG, Padhya T, McCaffrey TV, McCaffrey JC, Gabrilovich DI (2010) HIF-1alpha regulates function and differentiation of myeloid-derived suppressor cells in the tumor microenvironment. J Exp Med 207:2439–2453. doi:10.1084/jem.20100587

    Article  PubMed  CAS  Google Scholar 

  34. Yang L, DeBusk LM, Fukuda K, Fingleton B, Green-Jarvis B, Shyr Y, Matrisian LM, Carbone DP, Lin PC (2004) Expansion of myeloid immune suppressor Gr+ CD11b+ cells in tumor-bearing host directly promotes tumor angiogenesis. Cancer Cell 6:409–421. doi:10.1016/j.ccr.2004.08.031

    Article  PubMed  CAS  Google Scholar 

  35. Gabrilovich DI, Nagaraj S (2009) Myeloid-derived suppressor cells as regulators of the immune system. Nat Rev Immunol 9:162–174. doi:10.1038/nri2506

    Article  PubMed  CAS  Google Scholar 

  36. Tacke RS, Lee HC, Goh C, Courtney J, Polyak SJ, Rosen HR, Hahn YS (2012) Myeloid suppressor cells induced by hepatitis C virus suppress T-cell responses through the production of reactive oxygen species. Hepatology 55:343–353. doi:10.1002/hep.24700

    Article  PubMed  CAS  Google Scholar 

  37. Nagaraj S, Gupta K, Pisarev V, Kinarsky L, Sherman S, Kang L, Herber DL, Schneck J, Gabrilovich DI (2007) Altered recognition of antigen is a mechanism of CD8+ T cell tolerance in cancer. Nat Med 13:828–835. doi:10.1038/nm1609

    Article  PubMed  CAS  Google Scholar 

  38. Hahm ER, Moura MB, Kelley EE, Van Houten B, Shiva S, Singh SV (2011) Withaferin A-induced apoptosis in human breast cancer cells is mediated by reactive oxygen species. PLoS One 6:e23354. doi:10.1371/journal.pone.0023354

    Article  PubMed  CAS  Google Scholar 

  39. Mayola E, Gallerne C, Esposti DD, Martel C, Pervaiz S, Larue L, Debuire B, Lemoine A, Brenner C, Lemaire C (2011) Withaferin A induces apoptosis in human melanoma cells through generation of reactive oxygen species and down-regulation of Bcl-2. Apoptosis 16:1014–1027. doi:10.1007/s10495-011-0625-x

    Article  PubMed  CAS  Google Scholar 

  40. Malik F, Kumar A, Bhushan S, Khan S, Bhatia A, Suri KA, Qazi GN, Singh J (2007) Reactive oxygen species generation and mitochondrial dysfunction in the apoptotic cell death of human myeloid leukemia HL-60 cells by a dietary compound withaferin A with concomitant protection by N-acetyl cysteine. Apoptosis 12:2115–2133. doi:10.1007/s10495-007-0129-x

    Article  PubMed  CAS  Google Scholar 

  41. Um HJ, Min KJ, Kim DE, Kwon TK (2012) Withaferin A inhibits JAK/STAT3 signaling and induces apoptosis of human renal carcinoma Caki cells. Biochem Biophys Res Commun 427:24–29. doi:10.1016/j.bbrc.2012.08.133

    Article  PubMed  CAS  Google Scholar 

  42. Lee J, Hahm ER, Singh SV (2010) Withaferin A inhibits activation of signal transducer and activator of transcription 3 in human breast cancer cells. Carcinogenesis 31:1991–1998. doi:10.1093/carcin/bgq175

    Article  PubMed  CAS  Google Scholar 

  43. Malara N, Foca D, Casadonte F, Sesto MF, Macrina L, Santoro L, Scaramuzzino M, Terracciano R, Savino R (2008) Simultaneous inhibition of the constitutively activated nuclear factor kappaB and of the interleukin-6 pathways is necessary and sufficient to completely overcome apoptosis resistance of human U266 myeloma cells. Cell Cycle 7:3235–3245

    Article  PubMed  CAS  Google Scholar 

  44. Min KJ, Choi K, Kwon TK (2011) Withaferin A down-regulates lipopolysaccharide-induced cyclooxygenase-2 expression and PGE2 production through the inhibition of STAT1/3 activation in microglia cells. Int Immunopharmacol 11:1137–1142. doi:10.1016/j.intimp.2011.02.029

    Article  PubMed  CAS  Google Scholar 

  45. Munagala R, Kausar H, Munjal C, Gupta RC (2011) Withaferin A induces p53-dependent apoptosis by repression of HPV oncogenes and upregulation of tumor suppressor proteins in human cervical cancer cells. Carcinogenesis 32:1697–1705. doi:10.1093/carcin/bgr192

    Article  PubMed  CAS  Google Scholar 

  46. Karin M, Cao Y, Greten FR, Li ZW (2002) NF-kappaB in cancer: from innocent bystander to major culprit. Nat Rev Cancer 2:301–310. doi:10.1038/nrc780

    Article  PubMed  CAS  Google Scholar 

  47. Karin M, Greten FR (2005) NF-kappaB: linking inflammation and immunity to cancer development and progression. Nat Rev Immunol 5:749–759. doi:10.1038/nri1703

    Article  PubMed  CAS  Google Scholar 

  48. DiDonato JA, Mercurio F, Karin M (2012) NF-kappaB and the link between inflammation and cancer. Immunol Rev 246:379–400. doi:10.1111/j.1600-065X.2012.01099.x

    Article  PubMed  Google Scholar 

  49. Sinha P, Okoro C, Foell D, Freeze HH, Ostrand-Rosenberg S, Srikrishna G (2008) Proinflammatory S100 proteins regulate the accumulation of myeloid-derived suppressor cells. J Immunol 181:4666–4675

    PubMed  CAS  Google Scholar 

  50. Ndlovu MN, Van Lint C, Van Wesemael K, Callebert P, Chalbos D, Haegeman G, Vanden Berghe W (2009) Hyperactivated NF-{kappa}B and AP-1 transcription factors promote highly accessible chromatin and constitutive transcription across the interleukin-6 gene promoter in metastatic breast cancer cells. Mol Cell Biol 29:5488–5504. doi:10.1128/MCB.01657-08

    Article  PubMed  CAS  Google Scholar 

  51. Oh JH, Kwon TK (2009) Withaferin A inhibits tumor necrosis factor-alpha-induced expression of cell adhesion molecules by inactivation of Akt and NF-kappaB in human pulmonary epithelial cells. Int Immunopharmacol 9:614–619. doi:10.1016/j.intimp.2009.02.002

    Article  PubMed  CAS  Google Scholar 

  52. Mocellin S, Marincola FM, Young HA (2005) Interleukin-10 and the immune response against cancer: a counterpoint. J Leukoc Biol 78:1043–1051. doi:10.1189/jlb.0705358

    Article  PubMed  CAS  Google Scholar 

  53. Tanikawa T, Wilke CM, Kryczek I, Chen GY, Kao J, Nunez G, Zou W (2012) Interleukin-10 ablation promotes tumor development, growth, and metastasis. Cancer Res 72:420–429. doi:10.1158/0008-5472.CAN-10-4627

    Article  PubMed  CAS  Google Scholar 

  54. Murai M, Turovskaya O, Kim G, Madan R, Karp CL, Cheroutre H, Kronenberg M (2009) Interleukin 10 acts on regulatory T cells to maintain expression of the transcription factor Foxp3 and suppressive function in mice with colitis. Nat Immunol 10:1178–1184. doi:10.1038/ni.1791

    Article  PubMed  CAS  Google Scholar 

  55. Qian B, Deng Y, Im JH, Muschel RJ, Zou Y, Li J, Lang RA, Pollard JW (2009) A distinct macrophage population mediates metastatic breast cancer cell extravasation, establishment and growth. PLoS One 4:e6562. doi:10.1371/journal.pone.0006562

    Article  PubMed  Google Scholar 

  56. Lin EY, Li JF, Gnatovskiy L, Deng Y, Zhu L, Grzesik DA, Qian H, Xue XN, Pollard JW (2006) Macrophages regulate the angiogenic switch in a mouse model of breast cancer. Cancer Res 66:11238–11246. doi:10.1158/0008-5472.CAN-06-1278

    Article  PubMed  CAS  Google Scholar 

  57. Zheng Y, Cai Z, Wang S, Zhang X, Qian J, Hong S, Li H, Wang M, Yang J, Yi Q (2009) Macrophages are an abundant component of myeloma microenvironment and protect myeloma cells from chemotherapy drug-induced apoptosis. Blood 114:3625–3628. doi:10.1182/blood-2009-05-220285

    Article  PubMed  CAS  Google Scholar 

  58. Gabrilovich DI, Ostrand-Rosenberg S, Bronte V (2012) Coordinated regulation of myeloid cells by tumours. Nat Rev Immunol 12:253–268. doi:10.1038/nri3175

    Article  PubMed  CAS  Google Scholar 

  59. Huang B, Pan PY, Li Q, Sato AI, Levy DE, Bromberg J, Divino CM, Chen SH (2006) Gr-1+ CD115+ immature myeloid suppressor cells mediate the development of tumor-induced T regulatory cells and T-cell anergy in tumor-bearing host. Cancer Res 66:1123–1131. doi:10.1158/0008-5472.CAN-05-1299

    Article  PubMed  CAS  Google Scholar 

  60. Serafini P, Mgebroff S, Noonan K, Borrello I (2008) Myeloid-derived suppressor cells promote cross-tolerance in B-cell lymphoma by expanding regulatory T cells. Cancer Res 68:5439–5449. doi:10.1158/0008-5472.CAN-07-6621

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

We thank Drs. James B. McMahon, Kirk R. Gustafson, and Thomas J. Sayers (NIH) for helpful discussions, and Dr. Adam I. Marcus (Emory) for sharing his protocol for the preparation of the WRE extract. This study was supported by NIH RO1CA84232, NIH RO1CA115880, NIHRO1GM021248, and American Cancer Society IRG-97-153-07.

Conflict of interest

The authors declare no financial or commercial conflict of interest.

Author information

Authors and Affiliations

  1. Department of Biological Sciences, University of Maryland, Baltimore County, 1000 Hilltop Circle, Baltimore, MD, 21250, USA

    Pratima Sinha & Suzanne Ostrand-Rosenberg

Authors
  1. Pratima Sinha
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Suzanne Ostrand-Rosenberg
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Pratima Sinha.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (PDF 424 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Sinha, P., Ostrand-Rosenberg, S. Myeloid-derived suppressor cell function is reduced by Withaferin A, a potent and abundant component of Withania somnifera root extract. Cancer Immunol Immunother 62, 1663–1673 (2013). https://doi.org/10.1007/s00262-013-1470-2

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00262-013-1470-2

Keywords

Search

Navigation