Synthesis, biological, and antitumor activity of a highly potent 6-substituted pyrrolo[2,3-d]pyrimidine thienoyl antifolate inhibitor with proton-coupled folate transporter and folate receptor selectivity over the reduced folate carrier that inhibits β-glycinamide ribonucleotide formyltransferase

J Med Chem. 2011 Oct 27;54(20):7150-64. doi: 10.1021/jm200739e. Epub 2011 Sep 22.

Abstract

2-Amino-4-oxo-6-substituted pyrrolo[2,3-d]pyrimidine antifolates with a thienoyl side chain (compounds 1-3, respectively) were synthesized for comparison with compound 4, the previous lead compound of this series. Conversion of hydroxyl acetylen-thiophene carboxylic esters to thiophenyl-α-bromomethylketones and condensation with 2,4-diamino-6-hydroxypyrimidine afforded the 6-substituted pyrrolo[2,3-d]pyrimidine compounds of type 18 and 19. Coupling with l-glutamate diethyl ester, followed by saponification, afforded 1-3. Compound 3 selectively inhibited the proliferation of cells expressing folate receptors (FRs) α or β, or the proton-coupled folate transporter (PCFT), including KB and IGROV1 human tumor cells, much more potently than 4. Compound 3 was more inhibitory than 4 toward β-glycinamide ribonucleotide formyltransferase (GARFTase). Both 3 and 4 depleted cellular ATP pools. In SCID mice with IGROV1 tumors, 3 was more efficacious than 4. Collectively, our results show potent antitumor activity for 3 in vitro and in vivo, associated with its selective membrane transport by FRs and PCFT over RFC and inhibition of GARFTase, clearly establishing the 3-atom bridge as superior to the 1-, 2-, and 4-atom bridge lengths for the activity of this series.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphate / metabolism
  • Animals
  • Antineoplastic Agents / chemical synthesis*
  • Antineoplastic Agents / chemistry
  • Antineoplastic Agents / pharmacology
  • Cell Line, Tumor
  • Drug Screening Assays, Antitumor
  • Female
  • Folate Receptor 1 / metabolism*
  • Folate Receptor 2 / metabolism*
  • Folic Acid Antagonists / chemical synthesis*
  • Folic Acid Antagonists / chemistry
  • Folic Acid Antagonists / pharmacology
  • Glutamates / chemical synthesis*
  • Glutamates / chemistry
  • Glutamates / pharmacology
  • Humans
  • Mice
  • Mice, SCID
  • Neoplasm Transplantation
  • Oocytes / drug effects
  • Oocytes / physiology
  • Patch-Clamp Techniques
  • Phosphoribosylglycinamide Formyltransferase / antagonists & inhibitors*
  • Proton-Coupled Folate Transporter / metabolism*
  • Pyrimidines / chemical synthesis*
  • Pyrimidines / chemistry
  • Pyrimidines / pharmacology
  • Pyrimidinones / chemical synthesis*
  • Pyrimidinones / chemistry
  • Pyrimidinones / pharmacology
  • Reduced Folate Carrier Protein / metabolism*
  • Stereoisomerism
  • Structure-Activity Relationship
  • Transplantation, Heterologous
  • Xenopus

Substances

  • Antineoplastic Agents
  • Folate Receptor 1
  • Folate Receptor 2
  • Folic Acid Antagonists
  • Glutamates
  • N-((5-((2-amino-4-oxo-4,7-dihydro-3H-pyrrolo(2,3-d)pyrimidin-6-yl)propyl)thiophen-2-yl)carbonyl)glutamic acid
  • Proton-Coupled Folate Transporter
  • Pyrimidines
  • Pyrimidinones
  • Reduced Folate Carrier Protein
  • Adenosine Triphosphate
  • Phosphoribosylglycinamide Formyltransferase