, Volume 8, Issue 6, pp 1026–1036 | Cite as

Androgen receptor activation results in metabolite signatures of an aggressive prostate cancer phenotype: an NMR-based metabonomics study

  • Neil MacKinnon
  • Amjad P. Khan
  • Arul M. Chinnaiyan
  • Thekkelnaycke M. Rajendiran
  • Ayyalusamy Ramamoorthy
Original Article


Metabolomic studies have proven to provide a unique perspective of the cellular dysfunction developing as a result of prostate cancer (PCa) onset and progression, facilitated primarily by mass spectrometry (MS) and nuclear magnetic resonance (NMR) techniques. PCa develops as an androgen-dependent disease with the expression of the androgen receptor (AR), where patient treatment typically involves androgen ablation therapy. In response, it is theorized that PCa transforms to an androgen-hypersensitive or androgen-independent state, where treatment options are severely reduced. Under the hypothesis that AR stimulation increases the aggressivity of pre-existing PCa, NMR spectroscopy was utilized in the delineation of the metabonomic response of an androgen-dependent PCa cell line (LnCAP) as a result of AR activation. Metabolite profiles were determined after 12, 24, and 48 h of exposure to methyltrienolone (R1881), an AR agonist. Principal components analysis revealed the relative myo-inositol and phosphocholine levels were severely altered after androgen treatment. Furthermore, univariate analysis revealed multiple metabolic perturbations in response to R1881 exposure, including amino acid, choline, the phosphocholine/glycerophosphocholine ratio, and UDP-coupled sugar metabolism, which are consistent with reported measurements between normal and PCa samples. These results suggest that androgen-sensitive PCa may transform to an aggressive phenotype upon AR activation.


NMR Metabonomic Prostate cancer LnCAP Androgen R1881 

Supplementary material

11306_2012_398_MOESM1_ESM.pdf (345 kb)
Supplementary material 1 (PDF 344 kb)
11306_2012_398_MOESM2_ESM.xls (66 kb)
Supplementary material 2 (XLS 66 kb)


  1. Ackerstaff, E., Pfug, B. R., Nelson, J. B., & Bhujwalla, Z. M. (2001). Detection of increased choline compounds with proton nuclear magnetic resonance spectroscopy subsequent to malignant transformation of human prostatic epithelial cells. Cancer Research, 61, 3599–3603.PubMedGoogle Scholar
  2. Albers, M. J., Butler, T. N., Rahwa, I., Bao, N., Keshari, K. R., Swanson, M. G., et al. (2009). Evaluation of the ERETIC method as an improved quantitative reference for 1H HR-MAS spectroscopy of prostate tissue. Magnetic Resonance in Medicine, 61, 525–532.PubMedCrossRefGoogle Scholar
  3. Baek, S. H., Ohgi, K. A., Nelson, C. A., Welsbie, D., Chen, C., Sawyers, C. L., et al. (2006). Ligand-specific allosteric regulation of coactivator functions of androgen receptor in prostate cancer cells. Proceedings of the National Academy of Sciences of the USA, 103, 3100–3105.PubMedCrossRefGoogle Scholar
  4. Bao, B. Y., Chuang, B. F., Wang, Q., Sartor, O., Balk, S. P., Brown, M., et al. (2008). Androgen receptor mediates the expression of UDP-glucuronosyltransferase 2 BI5 and BI7 genes. The Prostate, 68, 839–848.PubMedCrossRefGoogle Scholar
  5. Beckonert, O., Monnerjahn, J., Bonk, U., & Leibfritz, D. (2003). Visualizing metabolic changes in breast-cancer tissue using 1H-NMR spectroscopy and self-organizing maps. NMR in Biomedicine, 16, 1–11.PubMedCrossRefGoogle Scholar
  6. Butcher, N. J., Tetlow, N. L., Cheung, C., Broadhurst, G. M., & Minchin, R. F. (2007). Induction of human arylamine N-acetyltransferase type i by androgens in human prostate cancer cells. Cancer Research, 67, 85–92.PubMedCrossRefGoogle Scholar
  7. Cai, C., Wang, H., Xu, Y., Chen, S., & Balk, S. P. (2009). Reactivation of androgen receptor-regulated TMPRSS2:ERG gene expression in castration-resistant prostate cancer. Cancer Research, 69, 6027–6032.PubMedCrossRefGoogle Scholar
  8. Cai, J., Kandagatla, P., Singareddy, R., Kropinski, A., Sheng, S., Cher, M. L., et al. (2010). Androgens induce functional CXCR4 through ERG factor expression in TMRPRSS2-ERG fusion-positive prostate cancer cells. Translational Oncology, 3, 195–203.PubMedGoogle Scholar
  9. Chen, M., Tanner, M., Levine, A. C., Levina, E., Ohouo, P., & Buttyan, R. (2009). Androgenic regulation of hedgehog signaling pathway components in prostate cancer cells. Cell Cycle, 8, 149–157.PubMedCrossRefGoogle Scholar
  10. Cheng, L. L., Wu, C., Smith, M. R., & Gonzalez, R. G. (2001). Non-destructive quantitation of spermine in human prostate tissue samples using HRMAS 1H NMR spectroscopy at 9.4 T. FEBS Letters, 494(1–2), 112–116.PubMedCrossRefGoogle Scholar
  11. Cheng, L. L., Burns, M. A., Taylor, J. L., He, W., Halperin, E. F., McDougal, W. S., et al. (2005). Metabolic characterization of human prostate cancer with tissue magnetic resonance spectroscopy. Cancer Research, 65, 3030–3034.PubMedGoogle Scholar
  12. Chipuk, J. E., Cornelius, S. C., Pultz, N. J., Jorgensen, J. S., Bonham, M. J., Kim, S. J., et al. (2002). The androgen receptor represses transforming growth factor-β signaling through interaction with Smad3. The Journal of Biological Chemistry, 277, 1240–1248.PubMedCrossRefGoogle Scholar
  13. Church, D. R., Lee, E., Thompson, T. A., Basu, H. S., Ripple, M. O., Ariazi, R. A., et al. (2005). Induction of AP-I activity by androgen activation of the androgen receptor in LNCaP human prostate carcinoma cells. The Prostate, 63, 155–168.PubMedCrossRefGoogle Scholar
  14. Costello, L. C., Franklin, R. B., & Narayan, P. (1999). Citrate in the diagnosis of prostate cancer. The Prostate, 38, 237–245.PubMedCrossRefGoogle Scholar
  15. Damber, J. E., & Aus, G. (2008). Prostate cancer. Lancet, 37, 1710–1721.CrossRefGoogle Scholar
  16. Dieterle, F., Ross, A., Schlotterbeck, G., & Senn, H. (2006). Probabilistic quotient normalization as robust method to account for dilution of complex biological mixtures. Application in 1H NMR metabonomics. Analytical Chemistry, 78, 4281–4290.PubMedCrossRefGoogle Scholar
  17. Feldman, B. J., & Feldman, D. (2001). The development of androgen-independent prostate cancer. Nature Reviews Cancer, 1, 34–45.PubMedCrossRefGoogle Scholar
  18. Flammand, V., Zhao, H., & Peehl, D. M. (2010). Targeting monoamine oxidase A in advanced prostate cancer. Journal of Cancer Research and Clinical Oncology, 136, 1761–1771.CrossRefGoogle Scholar
  19. Gannon, P. O., Godin-Ethier, J., Hassler, M., Delvoye, N., Aversa, M., Poisson, A. O., et al. (2010). Androgen-regulated expression of arginase 1, arginase 2 and interleukin-8 in human prostate cancer. PLoS ONE, 5, doi:10.1371/journal.pone.0012107.
  20. Gavrielides, M. V., Gonzalez-Guerrico, A. M., Riobo, N. A., & Kazanietz, M. G. (2006). Androgens regulate protein kinase Cδ transcription and modulate its apoptotic function in prostate cancer cells. Cancer Research, 66, 11792–11801.PubMedCrossRefGoogle Scholar
  21. Hobisch, A., Eder, I. E., Putz, T., Horninger, W., Bartsch, G., Klocker, H., et al. (1998). Interleukin-6 regulates prostate-specific protein expression in prostate carcinoma cells by activation of the androgen receptor. Cancer Research, 58, 4640–4645.PubMedGoogle Scholar
  22. Jackson, J. E. (1991). A user’s guide to principal components. New York: Wiley.CrossRefGoogle Scholar
  23. Jennbacken, K., Gustavsson, H., Tešan, T., Horn, M., Vallbo, C., Welén, K., et al. (2009). The prostatic environment suppresses growth of androgen-independent prostate cancer xenografts: An effect influenced by testosterone. The Prostate, 69, 1164–1175.PubMedCrossRefGoogle Scholar
  24. Kimura, K., Markowski, M., Bowen, C., & Gelmann, E. P. (2001). Androgen blocks apoptosis of hormone-dependent prostate cancer cells. Cancer Research, 61, 5611–5618.PubMedGoogle Scholar
  25. Lee, Y. C., Cheng, C. J., Huang, M., Bilen, M. A., Ye, X., Navone, N. M., et al. (2010). Androgen depletion up-regulates cadherin-11 expression in prostate cancer. Journal of Pathology, 221, 68–76.PubMedCrossRefGoogle Scholar
  26. Levin, Y. S., Albers, M. J., Butler, T. N., Spielman, D., Peehl, D. M., & Kurhanewicz, J. (2009). Methods for metabolic evaluation of prostate cancer cells using proton and 13C HR-MAS spectroscopy and [3-13C] pyruvate as a metabolic substrate. Magnetic Resonance in Medicine, 62, 1091–1098.PubMedCrossRefGoogle Scholar
  27. Loberg, R. D., McGregor, N., Ying, C., Sargent, E., & Pienta, K. J. (2007). In vivo evaluation of AT-101 (R-(-)-gossypol acetic acid) in androgen-independent growth of VCaP prostate cancer cells in combination with surgical castration. Neoplasia, 9, 1030–1037.PubMedCrossRefGoogle Scholar
  28. Makkonen, H., Kauhanen, M., Jääskeläinen, T., & Palvimo, J. J. (2011). Androgen receptor amplification is reflected in the transcriptional responses of vertebral-cancer of the prostate cells. Molecular and Cellular Endocrinology, 331, 57–65.PubMedCrossRefGoogle Scholar
  29. Mani, R. S., Tomlins, S. A., Callahan, K., Ghosh, A., Nyati, M. K., Varambally, S., et al. (2009). Induced chromosomal proximity and gene fusions in prostate cancer. Science, 326, 1230.PubMedCrossRefGoogle Scholar
  30. Massie, C. E., Lynch, A., Ramos-Montoya, A., Boren, J., Stark, R., Fazli, L., et al. (2011). The androgen receptor fuels prostate cancer by regulating central metabolism and biosynthesis. The EMBO Journal, 1–15, doi:10.1038/emboj.2011.158.
  31. Migliaccio, A., Castoria, G., Di Domenico, M., de Falco, A., Bilancio, A., Lombardi, M., et al. (2000). Steroid-induced androgen receptor-oestradiol receptor β-Src complex triggers prostate cancer cell proliferation. The EMBO Journal, 19, 5406–5417.PubMedCrossRefGoogle Scholar
  32. Nicholson, J. K., Focall, P. J. D., Spraul, M., Farrant, R. D., & Lindon, J. C. (1995). 750 MHz 1H and 1H–13C NMR spectroscopy of human blood plasma. Analytical Chemistry, 67, 793–811.PubMedCrossRefGoogle Scholar
  33. Park, S. Y., Kim, Y. J., Gao, A. C., Mohler, J. L., Onate, S. A., Hidalgo, A. A., et al. (2006). Hypoxia increases androgen receptor activity in prostate cancer cells. Cancer Research, 66, 5121–5129.PubMedCrossRefGoogle Scholar
  34. Putluri, N., Shojaie, A., Vasu, V. T., Nalluri, S., Vareed, S. K., Putluir, V., et al. (2011). Metabolomic profiling reveals a role for androgen in activating amino acid metabolism and methylation in prostate cancer cells. PLoS ONE, 6, doi:10.1371/journal.pone.0021417.
  35. Qi, H., Fillion, C., Labrie, Y., Grenier, J., Fournier, A., Berger, L., et al. (2002). AlbZIP, a novel bZIP gene located on chromosome 1q21.3 that is highly expressed in prostate tumors and of which the expression is up-regulated by androgens in LNCaP human prostate cancer cells. Cancer Research, 62, 721–733.PubMedGoogle Scholar
  36. Rantalainen, M., Cloarec, O., Beckonert, O., Wilson, I. D., Jackson, D., Tonge, R., et al. (2006). Statistically integrate metabonomic-proteomic studies on a human prostate cancer xenograft model in mice. Journal of Proteome Research, 5, 2642–2655.PubMedCrossRefGoogle Scholar
  37. Ross, B. D. (1991). Biochemical considerations in 1H spectroscopy. Glutamate and glutamine; Myo-inositol and related metabolites. NMR in Biomedicine, 4, 59–63.PubMedCrossRefGoogle Scholar
  38. Serkova, N. J., Gamito, E. J., Jones, R. H., O’Donnell, C., Brown, J. L., Green, S., et al. (2008). The metabolites citrate, myo-inositol, and spermine are potential age-independent markers of prostate cancer in human expressed prostatic secretions. The Prostate, 68, 620–628.PubMedCrossRefGoogle Scholar
  39. Sitter, B., Sonnewald, U., Spraul, M., Fjösne, H. E., & Gribbestad, I. S. (2002). High-resolution magic angle spinning MRS of breast cancer tissue. NMR in Biomedicine, 15, 327–337.PubMedCrossRefGoogle Scholar
  40. Song, K., Wang, H., Krebs, T. L., Wang, B., Kelley, T. J., & Danielpour, D. (2010). DHT selectively reverses Smad3-mediated/TGF-2-induced responses through transcriptional down-regulation of Smad3 in prostate epithelial cells. Molecular Endocrinology, 24, 2019–2029.PubMedCrossRefGoogle Scholar
  41. Sreekumar, A., Poisson, L. M., Rajendiran, T. M., Khan, A. P., Cao, Q., Yu, J., et al. (2009). Metabolomic profiles delineate potential role for sarcosine in prostate cancer progression. Nature, 457, 910–915.PubMedCrossRefGoogle Scholar
  42. Swanson, M. G., Vigneron, D. B., Tabatabai, Z. L., Males, R. G., Schmitt, L., Carroll, P. R., et al. (2003). Proton HR-MAS spectroscopy and quantitative pathologic analysis of MRI/3D-MRSI-targeted postsurgical prostate tissues. Magnetic Resonance in Medicine, 50, 944–954.PubMedCrossRefGoogle Scholar
  43. Swanson, M. G., Zektzer, A. S., Tabatabia, Z. L., Simko, J., Jarso, S., Keshari, K. R., et al. (2006). Quantitative analysis of prostate metabolites using 1H HR-MAS spectroscopy. Magnetic Resonance in Medicine, 55, 1257–1264.PubMedCrossRefGoogle Scholar
  44. Swanson, M. G., Keshari, K. R., Tabatabai, Z. L., Simko, J. P., Shinohara, K., Carroll, P. R., et al. (2008). Quantification of choline- and ethanolamine-containing metabolites in human prostate tissues using 1H HR-MAS total correlation spectroscopy. Magnetic Resonance in Medicine, 60, 33–40.PubMedCrossRefGoogle Scholar
  45. Teahan, O., Bevan, C. L., Waxman, J., & Keun, H. C. (2010). Metabolic signatures of malignant progression in prostate epithelial cells. International Journal of Biochemistry and Cell Biology,. doi:10.1016/j.biocel.2010.07.003.PubMedGoogle Scholar
  46. Tessem, M. B., Swanson, M. G., Keshari, K. R., Albers, M. J., Joun, D., Tabatabai, Z. L., et al. (2008). Evaluation of lactate and alanine as metabolic biomarkers of prostate cancer using 1H HR-MAS spectroscopy of biopsy tissues. Magnetic Resonance in Medicine, 60, 510–516.PubMedCrossRefGoogle Scholar
  47. Tiechert, F., Verschoyle, R. D., Greaves, P., Edwards, R. E., Teahan, O., Jones, D. J. L., et al. (2008). Metabolic profiling of transgenic adenocarcinoma of mouse prostate (TRAMP) tissue by 1H-NMR analysis: Evidence for unusual phospholipid metabolism. The Prostate, 68, 1035–1047.CrossRefGoogle Scholar
  48. Tomlins, A. M., Foxall, P. J. D., Lindon, J. C., Lynch, M. J., Spraul, M., Everett, J. R., et al. (1998). High resolution magic angle spinning 1H nuclear magnetic resonance analysis of intact prostatic hyperplastic and tumour tissues. Analytical Communications, 35, 113–115.CrossRefGoogle Scholar
  49. Tomlins, S. A., Rhodes, D. R., Perner, S., Dhanasekaran, S. M., Mehra, R., Sun, X. W., et al. (2005). Recurrent fusion of TMPRSS2 and ETS transcription factor genes in prostate cancer. Science, 310, 644–648.PubMedCrossRefGoogle Scholar
  50. Tomlins, S. A., Laxman, B., Varambally, S., Cao, X., Yu, J., Helgeson, B. E., et al. (2008). Role of the TMPRSS2-ERG gene fusion in prostate cancer. Neoplasia, 10, 177–188.PubMedCrossRefGoogle Scholar
  51. Van Asten, J. J. A., Cuijpers, V., van de Hulsbergen-Kaa, C., Soede-Huijbregts, C., Witjes, J. A., Verhofstad, A., et al. (2008). High resolution magic angle spinning NMR spectroscopy for metabolic assessment of cancer presence and Gleason score in human prostate needle biopsies. Magnetic Resonance Materials in Physics Biology and Medicine, 21, 435–442.CrossRefGoogle Scholar
  52. Veselkov, K. A., London, J. C., Ebbels, T. M. D., Crockford, D., Volynkin, V. V., Holmes, E., et al. (2009). Recursive segment-wise peak alignment of biological 1H NMR spectra for improved metabolic biomarker recovery. Analytical Chemistry, 81, 56–66.PubMedCrossRefGoogle Scholar
  53. Wang, Q., Li, W., Zhang, Y., Yuan, X., Xu, K., Yu, J., et al. (2009). Androgen receptor regulates a distinct transcription program in androgen-independent prostate cancer. Cell, 138, 245–256.PubMedCrossRefGoogle Scholar
  54. Wong, J. W. H., Cagney, G., & Cartwright, H. M. (2005a). SpecAlign-processing and alignment of mass spectra datasets. Bioinformatics, 21, 2088–2090.PubMedCrossRefGoogle Scholar
  55. Wong, J. W. H., Durante, C., & Cartwright, H. M. (2005b). Application of fast fourier transform cross-correlation for the alignment of large chromatographic and spectral datasets. Analytical Chemistry, 77, 5655–5661.PubMedCrossRefGoogle Scholar
  56. Wu, C., Taylor, J. L., He, W., Zepeda, A. G., Halperin, E. F., Bielecki, A., et al. (2003). Proton high-resolution magic angle spinning NMR analysis of fresh and previously frozen tissue of human prostate. Magnetic Resonance in Medicine, 50, 1307–1311.PubMedCrossRefGoogle Scholar
  57. Yu, J., Yu, J., Mani, R. S., Cao, Q., Brenner, C. J., Cao, X., et al. (2010). An integrated network of androgen receptor, polycomb, and TMPRSS2-ERG gene fusions in prostate cancer progression. Cancer Cell, 17, 443–454.PubMedCrossRefGoogle Scholar
  58. Zhu, H., Mazor, M., Kawano, Y., Walker, M. M., Leung, H. Y., Armstrong, K., et al. (2004). Analysis of Wnt gene expression in prostate cancer: Mutual inhibition of WNT11 and the androgen receptor. Cancer Research, 2004(64), 7918–7926.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  • Neil MacKinnon
    • 1
  • Amjad P. Khan
    • 2
  • Arul M. Chinnaiyan
    • 2
    • 3
  • Thekkelnaycke M. Rajendiran
    • 2
  • Ayyalusamy Ramamoorthy
    • 1
  1. 1.Biophysics and Department of ChemistryUniversity of MichiganAnn ArborUSA
  2. 2.Michigan Center for Translational Pathology, Department of PathologyUniversity of Michigan, Medical SchoolAnn ArborUSA
  3. 3.Department of UrologyComprehensive Cancer Center and Howard Hughes Medical Institute, University of Michigan, Medical SchoolAnn ArborUSA

Personalised recommendations