Skip to main content
SpringerLink
Log in

Detection of Total UDP-Glucuronosyltransferase (UGT) Activity in Melanoma Cells

  • Protocol
  • First Online:
Methods in Molecular Biology

Part of the book series: Methods in Molecular Biology

  • 81 Accesses

Abstract

The UDP-glucuronosyltransferases (UGTs) are integrally involved in the clearance of a wide range of drugs used to combat human diseases. UGT expression levels and activity can be induced by drug addition to cells and has been proposed as a potential intratumoral drug resistance mechanism. Traditional methods of assaying UGT activity are drug-centric and require HPLCs with multiple detectors (dependent on individual drug). Here, we describe a generalized method to detect total UGT activity (intrinsic or induced) via the UGT-Glo assay which utilizes a general UGT substrate with luminescence as the readout eliminating the need for multiple HPLC detectors to detect total UGT activity in a given sample. The method detailed here can be applied for any UGT containing sample, allowing for the efficient detection of total UGT activity to be the functional endpoint using a plate reader. In this manner, global changes in UGT activity can be monitored in response to a wide variety of stimuli.

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

Access this chapter

Log in via an institution

Institutional subscriptions

References

  1. Nagar S, Remmel RP (2006) Uridine diphosphoglucuronosyltransferase pharmacogenetics and cancer. Oncogene 25:1659–1672

    Article  PubMed  CAS  Google Scholar 

  2. Tukey RH, Strassburg CP (2000) Human UDP-glucuronosyltransferases: metabolism, expression, and disease. Annu Rev Pharmacol Toxicol 40:581–616

    Article  PubMed  CAS  Google Scholar 

  3. Chouinard S, Yueh MF, Tukey RH et al (2008) Inactivation by UDP-glucuronosyltransferase enzymes: the end of androgen signaling. J Steroid Biochem Mol Biol 109:247–253

    Article  PubMed  CAS  Google Scholar 

  4. Starlard-Davenport A, Lyn-Cook B, Radominska-Pandya A (2008) Identification of UDP-glucuronosyltransferase 1A10 in non-malignant and malignant human breast tissues. Steroids 73:611–620

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  5. Gregory PA, Lewinsky RH, Gardner-Stephen DA, Mackenzie PI (2004) Regulation of UDP glucuronosyltransferases in the gastrointestinal tract. Toxicol Appl Pharmacol 199:354–363

    Article  PubMed  CAS  Google Scholar 

  6. Giuliani L, Ciotti M, Stoppacciaro A et al (2005) UDP-glucuronosyltransferases 1A expression in human urinary bladder and colon cancer by immunohistochemistry. Oncol Rep 13:185–191

    PubMed  CAS  Google Scholar 

  7. King CD, Rios GR, Green MD, Tephly TR (2000) UDP-glucuronosyltransferases. Curr Drug Metab 1:143–161

    Article  PubMed  CAS  Google Scholar 

  8. Zheng Z, Fang JL, Lazarus P (2002) Glucuronidation: an important mechanism for detoxification of benzo[a]pyrene metabolites in aerodigestive tract tissues. Drug Metab Dispos 30:397–403

    Article  PubMed  CAS  Google Scholar 

  9. Dellinger RW, Fang JL, Chen G, Weinberg R, Lazarus P (2006) Importance of UDP-glucuronosyltransferase 1A10 (UGT1A10) in the detoxification of polycyclic aromatic hydrocarbons: decreased glucuronidative activity of the UGT1A10139Lys isoform. Drug Metab Dispos 34:943–949

    PubMed  CAS  Google Scholar 

  10. Dellinger RW, Matundan HH, Ahmed AS, Duong PH, Meyskens FL Jr (2012) Anti-cancer drugs elicit re-expression of UDP-glucuronosyltransferases in melanoma cells. PLoS One 7, e47696

    Article  PubMed  ADS  PubMed Central  CAS  Google Scholar 

  11. Bansal SK, Gessner T (1980) A unified method for the assay of uridine diphosphoglucuronyltransferase activities toward various aglycones using uridine diphospho[U-14C]glucuronic acid. Anal Biochem 109:321–329

    Article  PubMed  CAS  Google Scholar 

  12. Fisher MB, Campanale K, Ackermann BL, VandenBranden M, Wrighton SA (2000) In vitro glucuronidation using human liver microsomes and the pore-forming peptide alamethicin. Drug Metab Dispos 28:560–566

    PubMed  CAS  Google Scholar 

  13. Dellinger RW, Chen G, Blevins-Primeau AS, Krzeminski J, Amin S, Lazarus P (2007) Glucuronidation of PhIP and N-OH-PhIP by UDP-glucuronosyltransferase 1A10. Carcinogenesis 28:2412–2418

    Article  PubMed  CAS  Google Scholar 

  14. Radominska-Pandya A, Pokrovskaya ID, Xu J et al (2002) Nuclear UDP-glucuronosyltransferases: identification of UGT2B7 and UGT1A6 in human liver nuclear membranes. Arch Biochem Biophys 399:37–48

    Article  PubMed  CAS  Google Scholar 

  15. Chen G, Dellinger RW, Gallagher CJ, Sun D, Lazarus P (2008) Identification of a prevalent functional missense polymorphism in the UGT2B10 gene and its association with UGT2B10 inactivation against tobacco-specific nitrosamines. Pharmacogenet Genomics 18:181–191

    Article  PubMed  CAS  Google Scholar 

  16. Dellinger RW, Garcia AM, Meyskens FL Jr (2014) Differences in the glucuronidation of resveratrol and pterostilbene; altered enzyme specificity and potential gender differences. Drug Metab Pharmacokinet 29(2):112–119

    Article  PubMed  PubMed Central  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Chao Family Comprehensive Cancer Center and Departments of Medicine (RWD, FLM), Biological Chemistry (FLM), Epidemiology (RWD) and Public Health (FLM), University of California Irvine, Sprague Hall B200, Irvine, CA, 92697, USA

    Ryan W. Dellinger Ph.D. & Frank L. Meyskens Jr.

Authors
  1. Ryan W. Dellinger Ph.D.
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Frank L. Meyskens Jr.
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Ryan W. Dellinger Ph.D. .

Rights and permissions

Reprints and permissions

Copyright information

© 2015 Springer Science+Business Media New York

About this protocol

Cite this protocol

Dellinger, R.W., Meyskens, F.L. (2015). Detection of Total UDP-Glucuronosyltransferase (UGT) Activity in Melanoma Cells. In: Methods in Molecular Biology. Humana Press. https://doi.org/10.1007/7651_2015_298

Download citation

  • DOI: https://doi.org/10.1007/7651_2015_298

  • Published:

  • Publisher Name: Humana Press

Publish with us

Policies and ethics

Search

Navigation