Mode of action of methotrexate-albumin in a human T-cell leukemia line and activity against an MTX-resistant clone

Anticancer Drug Des. Aug-Oct 2001;16(4-5):227-37.


Limitations of low mol. wt anticancer drugs are short tumor exposure times and toxicity to normal tissue. Methotrexate (MTX) covalently linked to human serum albumin (HSA) as a macromolecular carrier caused tumor regressions concomitant with a favorable toxicity profile in a clinical phase I trial (Hartung et aL, Clin. Cancer Res., 1999, 5, 753). We examined the uptake, intracellular degradation, metabolism and thymidylate synthase (TS) inhibition of MTX-HSA in the T-cell leukemia line CCRF-CEM and the MTX transport resistant clone CCRF-CEM/MTX. The number of MTX molecules per albumin molecule was determined by electrospray mass spectrometry. A loading ratio (LR) of approximately 1.4 mol MTX/albumin revealed intact complexes with one and two MTX molecules/albumin. In the complex with an LR of 5.7, albumin with up to 16 MTX molecules was seen. MTX-HSA was taken up by CCRF-CEM cells via endocytosis and cleaved by lysosomal enzymes. Liberated MTX was a poor substrate of folylpolyglutamate synthetase and was exported into the medium. TS was inhibited and cell survival was impaired by MTX-HSA in a time- and concentration-dependent manner. CCRF-CEM/MTX cells exhibited a growth inhibition of 30-40% after MTX-HSA treatment, but no TS inhibition. The alternative uptake route via endocytosis enables MTX-HSA to overcome transport resistance to MTX.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antimetabolites, Antineoplastic / metabolism
  • Antimetabolites, Antineoplastic / pharmacology*
  • Carrier Proteins / genetics
  • Carrier Proteins / metabolism
  • Clone Cells
  • Drug Resistance, Neoplasm
  • Humans
  • Immunochemistry
  • Leukemia, T-Cell / drug therapy*
  • Leukemia, T-Cell / pathology
  • Lysosomes / metabolism
  • Methotrexate / metabolism
  • Methotrexate / pharmacology*
  • Molecular Weight
  • Polyglutamic Acid / metabolism
  • Protein Binding
  • Serum Albumin / pharmacology*
  • Spectrometry, Mass, Electrospray Ionization
  • Survival Analysis
  • Thymidylate Synthase / metabolism
  • Tumor Cells, Cultured


  • Antimetabolites, Antineoplastic
  • Carrier Proteins
  • Serum Albumin
  • Polyglutamic Acid
  • Thymidylate Synthase
  • Methotrexate