Skip to main content
SpringerLink
Log in

Enzymatic and non-enzymatic mechanisms of dimesna metabolism

  • Original Article
  • Published:
Amino Acids Aims and scope Submit manuscript

Abstract

The chemical reduction of the disulfide homodimer dimesna to its constituent mesna moieties is essential for its mitigation of nephrotoxicity associated with cisplatin and ifosfamide anticancer therapies and enhancement of dialytic clearance of the cardiovascular risk factor homocysteine. The objective of this study was to investigate potential enzymatic and non-enzymatic mechanisms of intracellular dimesna reduction. Similar to endogenous intracellular disulfides, dimesna undergoes thiol–disulfide exchange with thiolate anion-forming sulfhydryl groups via the two-step SN2 reaction. Determination of equilibrium constants of dimesna reduction when mixed with cysteine or glutathione provided a mechanistic explanation for dramatic cysteine and homocysteine depletion, but sparing of the endogenous antioxidant glutathione, previously observed during mesna therapy. Dimesna was reduced by recombinant enzymes of the thioredoxin system; however, oxidation of NADPH by the glutaredoxin system was only observed in the presence of combined dimesna and reduced glutathione, suggesting formation of oxidized glutathione following an initial non-enzymatic reduction of dimesna. Production of mesna by enzymatic and non-enzymatic mechanisms in HeLa cell lysate following dimesna incubation was demonstrated by a loss in mesna production following protein denaturation and prediction of residual non-enzymatic mesna production by mathematical modeling of thiol–disulfide exchange reactions. Reaction modeling also revealed that mixed disulfides make up a significant proportion of intracellular thiols, supporting their role in providing additional nephroprotection, independent of direct platinum conjugation.

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
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

Abbreviations

APN:

Aminopeptidase N

DTNB:

5,5′-Dithio-bis-2-nitrobenzoic acid

E°′MSSM/MSH :

Half-cell potential of the dimesna/mesna redox system

EDTA:

Ethylenediaminetetraacetic acid

ESRD:

End-stage renal disease

CCBL:

Cysteine-S-conjugate-β-lyase

Cys:

Cysteine

Cys34 :

Cysteine-34

CySSyC:

Cystine

F :

Faraday constant

FBS:

Fetal bovine serum

GGT:

Γ-Glutamyltranspeptidase

GR:

Glutathione reductase

GRX1:

Human glutaredoxin

GS-mesna:

Mesna-glutathione disulfide

GSH:

Glutathione

GSSG:

Glutathione disulfide

HPLC-FD:

High-performance liquid chromatography with fluorescence detection

k1–4:

Micro-rate constants

K eq :

Equilibrium constant

k obs :

Pseudo-first-order rate constants

M :

Mesna moiety

MBB:

Monobromobimane

MM :

Dimesna

MM 0 :

Dimesna, starting concentration

NADPH:

Nicotinamide adenine dinucleotide phosphate

pKa :

Acid dissociation constant

\({\hat R}\) :

Gas constant

R :

Thiol moiety

RM :

Oxidized thiol and mesna species

RR :

Oxidized thiol species

RSSM :

Mixed disulfide intermediate

T:

Absolute temperature

TNB:

3-Thio-6-nitrobenzoate

Trxr1:

Rat thioredoxin reductase 1

TRX1:

Human thioredoxin

References

  • Arner ES, Zhong L, Holmgren A (1999) Preparation and assay of mammalian thioredoxin and thioredoxin reductase. Methods Enzymol 300:226–239

    Article  CAS  PubMed  Google Scholar 

  • Benesch RE, Benesch R (1955) The acid stength of the –SH group in cysteine and related compounds. J Am Chem Soc 77:5877–5881

    Article  CAS  Google Scholar 

  • Bostom AG, Lathrop L (1997) Hyperhomocysteinemia in end-stage renal disease: prevalence, etiology, and potential relationship to arteriosclerotic outcomes. Kidney Int 52(1):10–20

    Article  CAS  PubMed  Google Scholar 

  • Boven E, Verschraagen M, Hulscher TM, Erkelens CA, Hausheer FH, Pinedo HM, van der Vijgh WJ (2002) BNP7787, a novel protector against platinum-related toxicities, does not affect the efficacy of cisplatin or carboplatin in human tumour xenografts. Eur J Cancer 38(8):1148–1156

    Article  CAS  PubMed  Google Scholar 

  • Brock N, Pohl J, Stekar J (1981a) Detoxification of urotoxic oxazaphosphorines by sulfhydryl compounds. J Cancer Res Clin Oncol 100(3):311–320

    Article  CAS  PubMed  Google Scholar 

  • Brock N, Pohl J, Stekar J (1981b) Studies on the urotoxicity of oxazaphosphorine cytostatics and its prevention. 2. Comparative study on the uroprotective efficacy of thiols and other sulfur compounds. Eur J Cancer Clin Oncol 17(11):1155–1163

    Article  CAS  PubMed  Google Scholar 

  • Brock N, Pohl J, Stekar J, Scheef W (1982) Studies on the urotoxicity of oxazaphosphorine cytostatics and its prevention–III. Profile of action of sodium 2-mercaptoethane sulfonate (mesna). Eur J Cancer Clin Oncol 18(12):1377–1387

    Article  CAS  PubMed  Google Scholar 

  • Cutler MJ, Urquhart BL, Velenosi TJ, Meyer Zu, Schwabedissen HE, Dresser GK, Leake BF, Tirona RG, Kim RB, Freeman DJ (2012) In vitro and in vivo assessment of renal drug transporters in the disposition of mesna and dimesna. J Clin Pharmacol 52(4):530–542

    Article  CAS  PubMed  Google Scholar 

  • Di Giuseppe D, Priora R, Coppo L, Ulivelli M, Bartalini S, Summa D, Margaritis A, Frosali S, Di Simplicio P (2014) The control of hyperhomocysteinemia through thiol exchange mechanisms by mesna. Amino Acids 46(2):429–439

    Article  PubMed  Google Scholar 

  • Finkelstein JD (1998) The metabolism of homocysteine: pathways and regulation. Eur J Pediatr 157(Suppl 2):S40–S44

    Article  CAS  PubMed  Google Scholar 

  • Gilbert HF (1995) Thiol/disulfide exchange equilibria and disulfide bond stability. Methods Enzymol 251:8–28

    Article  CAS  PubMed  Google Scholar 

  • Goren MP, Hsu LC, Li JT (1998) Reduction of dimesna to mesna by the isolated perfused rat liver. Cancer Res 58(19):4358–4362

    CAS  PubMed  Google Scholar 

  • Hanigan MH, Gallagher BC, Taylor PT Jr, Large MK (1994) Inhibition of gamma-glutamyl transpeptidase activity by acivicin in vivo protects the kidney from cisplatin-induced toxicity. Cancer Res 54(22):5925–5929

    CAS  PubMed  Google Scholar 

  • Hanigan MH, Gallagher BC, Taylor PT Jr (1996) Cisplatin nephrotoxicity: inhibition of gamma-glutamyl transpeptidase blocks the nephrotoxicity of cisplatin without reducing platinum concentrations in the kidney. Am J Obstet Gynecol 175(2):270–273

    Article  CAS  PubMed  Google Scholar 

  • Hanigan MH, Lykissa ED, Townsend DM, Ou CN, Barrios R, Lieberman MW (2001) Gamma-glutamyl transpeptidase-deficient mice are resistant to the nephrotoxic effects of cisplatin. Am J Pathol 159(5):1889–1894

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Hausheer FH, Shanmugarajah D, Leverett BD, Chen X, Huang Q, Kochat H, Petluru PN, Parker AR (2010) Mechanistic study of BNP7787-mediated cisplatin nephroprotection: modulation of gamma-glutamyl transpeptidase. Cancer Chemother Pharmacol 65(5):941–951

    Article  CAS  PubMed  Google Scholar 

  • Hausheer FH, Parker AR, Petluru PN, Jair KW, Chen S, Huang Q, Chen X, Ayala PY, Shanmugarajah D, Kochat H (2011) Mechanistic study of BNP7787-mediated cisplatin nephroprotection: modulation of human aminopeptidase N. Cancer Chemother Pharmacol 67(2):381–391

    Article  CAS  PubMed  Google Scholar 

  • Hensley ML, Hagerty KL, Kewalramani T, Green DM, Meropol NJ, Wasserman TH, Cohen GI, Emami B, Gradishar WJ, Mitchell RB, Thigpen JT, Trotti A, Trotti A III, von Hoff D, Schuchter LM (2008) American Society of Clinical Oncology 2008 clinical practice guideline update: use of chemotherapy and radiation therapy protectants. J Clin Oncol 27(1):127–145

    Article  PubMed  Google Scholar 

  • Holmgren A (1989) Thioredoxin and glutaredoxin systems. J Biol Chem 264(24):13963–13966

    CAS  PubMed  Google Scholar 

  • Jocelyn PC (1972) Biochemistry of the SH group. Academic Press, New York

    Google Scholar 

  • Kurowski V, Wagner T (1997) Urinary excretion of ifosfamide, 4-hydroxyifosfamide, 3- and 2-dechloroethylifosfamide, mesna, and dimesna in patients on fractionated intravenous ifosfamide and concomitant mesna therapy. Cancer Chemother Pharmacol 39(5):431–439

    Article  CAS  PubMed  Google Scholar 

  • Lauterburg BH, Nguyen T, Hartmann B, Junker E, Kupfer A, Cerny T (1994) Depletion of total cysteine, glutathione, and homocysteine in plasma by ifosfamide/mesna therapy. Cancer Chemother Pharmacol 35(2):132–136

    Article  CAS  PubMed  Google Scholar 

  • Mallamaci F, Zoccali C, Tripepi G, Fermo I, Benedetto FA, Cataliotti A, Bellanuova I, Malatino LS, Soldarini A (2002) Hyperhomocysteinemia predicts cardiovascular outcomes in hemodialysis patients. Kidney Int 61(2):609–614

    Article  PubMed  Google Scholar 

  • Masuda N, Negoro S, Hausheer F, Nakagawa K, Matsui K, Kudoh S, Takeda K, Yamamoto N, Yoshimura N, Ohashi Y, Fukuoka M (2011) Phase I and pharmacologic study of BNP7787, a novel chemoprotector in patients with advanced non-small cell lung cancer. Cancer Chemother Pharmacol 67(3):533–542

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Meister A, Anderson ME (1983) Glutathione. Annu Rev Biochem 52:711–760

    Article  CAS  PubMed  Google Scholar 

  • Millis KK, Weaver KH, Rabenstein DL (1993) Oxidation/reduction potential of glutathione. J Org Chem 58:4144–4146

    Article  CAS  Google Scholar 

  • Moustapha A, Naso A, Nahlawi M, Gupta A, Arheart KL, Jacobsen DW, Robinson K, Dennis VW (1998) Prospective study of hyperhomocysteinemia as an adverse cardiovascular risk factor in end-stage renal disease. Circulation 97(2):138–141

    Article  CAS  PubMed  Google Scholar 

  • Mudd SH, Finkelstein JD, Refsum H, Ueland PM, Malinow MR, Lentz SR, Jacobsen DW, Brattstrom L, Wilcken B, Wilcken DE, Blom HJ, Stabler SP, Allen RH, Selhub J, Rosenberg IH (2000) Homocysteine and its disulfide derivatives: a suggested consensus terminology. Arterioscler Thromb Vasc Biol 20(7):1704–1706

    Article  CAS  PubMed  Google Scholar 

  • Ormstad K, Uehara N (1982) Renal transport and disposition of Na-2-mercaptoethane sulfonate disulfide (dimesna) in the rat. FEBS Lett 150(2):354–358

    Article  CAS  PubMed  Google Scholar 

  • Ormstad K, Orrenius S, Lastbom T, Uehara N, Pohl J, Stekar J, Brock N (1983) Pharmacokinetics and metabolism of sodium 2-mercaptoethanesulfonate in the rat. Cancer Res 43(1):333–338

    CAS  PubMed  Google Scholar 

  • Pendyala L, Creaven PJ, Schwartz G, Meropol NJ, Bolanowska-Higdon W, Zdanowicz J, Murphy M, Perez R (2000) Intravenous ifosfamide/mesna is associated with depletion of plasma thiols without depletion of leukocyte glutathione. Clin Cancer Res 6(4):1314–1321

    CAS  PubMed  Google Scholar 

  • Pendyala L, Schwartz G, Smith P, Zdanowicz J, Murphy M, Hausheer F (2003) Modulation of plasma thiols and mixed disulfides by BNP7787 in patients receiving paclitaxel/cisplatin therapy. Cancer Chemother Pharmacol 51(5):376–384

    CAS  PubMed  Google Scholar 

  • Robinson K, Gupta A, Dennis V, Arheart K, Chaudhary D, Green R, Vigo P, Mayer EL, Selhub J, Kutner M, Jacobsen DW (1996) Hyperhomocysteinemia confers an independent increased risk of atherosclerosis in end-stage renal disease and is closely linked to plasma folate and pyridoxine concentrations. Circulation 94(11):2743–2748

    Article  CAS  PubMed  Google Scholar 

  • Selhub J (1999) Homocysteine metabolism. Annu Rev Nutr 19:217–246

    Article  CAS  PubMed  Google Scholar 

  • Shanmugarajah D, Ding D, Huang Q, Chen X, Kochat H, Petluru PN, Ayala PY, Parker AR, Hausheer FH (2009) Analysis of BNP7787 thiol-disulfide exchange reactions in phosphate buffer and human plasma using microscale electrochemical high performance liquid chromatography. J Chromatogr B Analyt Technol Biomed Life Sci 877(10):857–866

    Article  CAS  PubMed  Google Scholar 

  • Shemin D, Lapane KL, Bausserman L, Kanaan E, Kahn S, Dworkin L, Bostom AG (1999) Plasma total homocysteine and hemodialysis access thrombosis: a prospective study. J Am Soc Nephrol 10(5):1095–1099

    CAS  PubMed  Google Scholar 

  • Smith PF, Booker BM, Creaven P, Perez R, Pendyala L (2003) Pharmacokinetics and pharmacodynamics of mesna-mediated plasma cysteine depletion. J Clin Pharmacol 43(12):1324–1328

    Article  CAS  PubMed  Google Scholar 

  • Srinivasan U, Mieyal PA, Mieyal JJ (1997) pH profiles indicative of rate-limiting nucleophilic displacement in thioltransferase catalysis. Biochemistry 36(11):3199–3206

    Article  CAS  PubMed  Google Scholar 

  • Stofer-Vogel B, Cerny T, Kupfer A, Junker E, Lauterburg BH (1993) Depletion of circulating cyst(e)ine by oral and intravenous mesna. Br J Cancer 68(3):590–593

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Townsend DM, Deng M, Zhang L, Lapus MG, Hanigan MH (2003) Metabolism of cisplatin to a nephrotoxin in proximal tubule cells. J Am Soc Nephrol 14(1):1–10

    Article  CAS  PubMed  Google Scholar 

  • Urquhart BL, House AA, Cutler MJ, Spence JD, Freeman DJ (2006) Thiol exchange: an in vitro assay that predicts the efficacy of novel homocysteine lowering therapies. J Pharm Sci 95(8):1742–1750

    Article  CAS  PubMed  Google Scholar 

  • Urquhart BL, Freeman DJ, Spence JD, House AA (2007a) The effect of mesna on plasma total homocysteine concentration in hemodialysis patients. Am J Kidney Dis 49(1):109–117

    Article  CAS  PubMed  Google Scholar 

  • Urquhart BL, Freeman DJ, Spence JD, House AA (2007b) Mesna as a nonvitamin intervention to lower plasma total homocysteine concentration: implications for assessment of the homocysteine theory of atherosclerosis. J Clin Pharmacol 47(8):991–997

    Article  CAS  PubMed  Google Scholar 

  • Verschraagen M, Boven E, Ruijter R, van der Born K, Berkhof J, Hausheer FH, van der Vijgh WJ (2003) Pharmacokinetics and preliminary clinical data of the novel chemoprotectant BNP7787 and cisplatin and their metabolites. Clin Pharmacol Ther 74(2):157–169

    Article  CAS  PubMed  Google Scholar 

  • Verschraagen M, Boven E, Torun E, Hausheer FH, Bast A, van der Vijgh WJ (2004) Possible (enzymatic) routes and biological sites for metabolic reduction of BNP7787, a new protector against cisplatin-induced side-effects. Biochem Pharmacol 68(3):493–502

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

The authors are grateful to Dr. Jack Bend for his expert advice in preparation of this manuscript, Dr. Richard Kim for use of the Kim Lab spectrophotometer and Sara LeMay for her skilled animal work.

Conflict of interest

A. A. House, B. L. Urquhart, and D. J. Freeman have applied for a patent for the use of mesna to lower homocysteine in patients with ESRD.

Author information

Authors and Affiliations

  1. School of Pharmacy, University of Waterloo, 200 University Ave W., Waterloo, ON, Canada

    Murray J. Cutler

  2. Department of Physiology and Pharmacology, Western University, London, ON, Canada

    Thomas J. Velenosi, Bradley L. Urquhart & David J. Freeman

  3. Department of Physiology, University of Toronto, Toronto, ON, Canada

    Ankur Bodalia

  4. Division of Nephrology, Department of Medicine, University Hospital, London, ON, Canada

    Andrew A. House

  5. Division of Clinical Pharmacology, Department of Medicine, University Hospital, London, ON, Canada

    Bradley L. Urquhart & David J. Freeman

  6. Lawson Health Research Institute, London, ON, Canada

    Bradley L. Urquhart & David J. Freeman

Authors
  1. Murray J. Cutler
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Thomas J. Velenosi
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ankur Bodalia
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Andrew A. House
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Bradley L. Urquhart
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. David J. Freeman
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Murray J. Cutler.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Cutler, M.J., Velenosi, T.J., Bodalia, A. et al. Enzymatic and non-enzymatic mechanisms of dimesna metabolism. Amino Acids 47, 511–523 (2015). https://doi.org/10.1007/s00726-014-1882-0

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00726-014-1882-0

Keywords

Search

Navigation