Synthesis and biological activity of methotrexate analogues with two acid groups and a hydrophobic aromatic ring in the side chain

J Med Chem. 1991 Feb;34(2):574-9. doi: 10.1021/jm00106a016.


The heretofore unknown gamma-(m-carboxyanilide) and gamma-(m-boronoanillide) derivatives of methotrexate (MTX) and the gamma-(m-carboxyanilide) derivatives of aminopterin (AMT) were prepared and tested as inhibitors of dihydrofolate reductase (DHFR) and as inhibitors of cell growth in culture with the aim of comparing their activity with that of N alpha-(4-amino-4-deoxypteroyl)-N delta-hemiphthaloyl-L-ornithine, a potent antifolate whose side chain likewise contains a hydrophobic aromatic ring with an acid group on the ring. All three anilides were potent DHFR inhibitors, with activity comparable to MTX and AMT. The gamma-(m-boronoanilide) displayed growth inhibitory potency similar to that of the hemiphthaloylornithine analogue, with an IC50 of only 0.7 nM. This compound, which is the most potent of the gamma-amides of MTX tested to date, is also the first reported example of an antifolate with a B(OH)2 group in the side chain and is especially novel because of its potential to form a stable tetrahedral boronate complex by reaction with electron rich OH or NH2 groups in the active site of DHFR or other folate enzymes. In antitumor assays against L1210 leukemia in mice, N alpha-(4-amino-4-deoxypteroyl)-N delta-hemiphthaloyl-L-ornithine gave a T/C of greater than 263% at 20 mg/kg (qdx9) and 300% at 16 mg/kg (bidx10), whereas maximally tolerated doses of MTX of 8 mg/kg (qdx9) and 1 mg/kg (bidx10) gave T/C values of 213 and 188%, respectively. MTX gamma-(m-boronoanilide) was also active, with a T/C of 175% at 32 mg/kg (qdx9), the highest dose tested.

Publication types

  • Comparative Study
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Aminopterin / analogs & derivatives*
  • Animals
  • Antineoplastic Agents / chemical synthesis*
  • Antineoplastic Agents / pharmacology
  • Antineoplastic Agents / therapeutic use
  • Cell Line
  • Folic Acid Antagonists
  • Leukemia L1210 / drug therapy
  • Methotrexate / analogs & derivatives*
  • Mice
  • Structure-Activity Relationship


  • Antineoplastic Agents
  • Folic Acid Antagonists
  • Aminopterin
  • Methotrexate