Determination of the cytoprotective agent WR-2721 (Amifostine, Ethyol) and its metabolites in human blood using monobromobimane fluorescent labeling and high-performance liquid chromatography

Cancer Chemother Pharmacol. 1998;42(5):400-6. doi: 10.1007/s002800050836.


Purpose: WR-2721 [S-2-(3-aminopropylamino)ethylphosphorothioic acid] is a chemoprotective agent that is currently in pediatric clinical trials. It is a prodrug that is dephosphorylated by alkaline phosphatase to the active free thiol form, WR-1065 [S-2-(3-aminopropylamino)ethanethiol]. It is likely that adequate and sustained cellular levels of the drug are necessary for optimum cytoprotection. To date, a method to measure both plasma and cellular levels of WR-2721 and its metabolites in clinical samples has not been available.

Methods: In the study reported here the monobromobimane (mBBr) fluorescent labeling method was used to measure these levels when drug was added in vitro to blood samples from normal volunteers. In addition, we present pharmacokinetic data from a pediatric patient receiving WR-2721 (825 mg/m2 x 2).

Results: The results can be summarized as follows: (1) WR-2721 was detected in the patient's plasma with a half-life of about 10 min; (2) the WR-1065 concentration in the blood cellular fraction was similar to that of plasma; (3) both WR-1065 and WR-SS-low molecular weight (WR-SS-LMW) metabolites disappeared from plasma and the cellular fraction by 3.6 h after WR-2721 infusion; (4) a large proportion of WR-1065 was oxidized in plasma to WR-SS protein and WR-SS-LMW; (5) a large proportion of WR-1065 in the cellular fraction was oxidized to WR-SS-protein; (6) the WR-SS-LMW concentration in the cellular fraction was low; and (7) saturation of plasma and cellular protein binding sites was possible.

Conclusions: The pharmacokinetic data that were generated with this technique could guide clinical trials using WR-2721.

Publication types

  • Case Reports
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amifostine / analysis*
  • Amifostine / metabolism
  • Bridged Bicyclo Compounds
  • Child
  • Chromatography, High Pressure Liquid / methods*
  • Female
  • Fluorescent Dyes
  • Half-Life
  • Humans
  • Mercaptoethylamines / analysis
  • Prodrugs / analysis*
  • Prodrugs / metabolism
  • Radiation-Protective Agents / analysis*
  • Radiation-Protective Agents / metabolism


  • Bridged Bicyclo Compounds
  • Fluorescent Dyes
  • Mercaptoethylamines
  • Prodrugs
  • Radiation-Protective Agents
  • WR 1065
  • Amifostine
  • monobromobimane