Radiolabeled 6-(2, 3-Dichlorophenyl)-N4-methylpyrimidine-2, 4-diamine (TH287): A Potential Radiotracer for Measuring and Imaging MTH1

Int J Mol Sci. 2020 Nov 23;21(22):8860. doi: 10.3390/ijms21228860.


MTH1 (MutT homolog 1) or NUDT1 (Nudix Hydrolase 1), also known as oxidized purine nucleoside triphosphatase, has potential as a biomarker for monitoring cancer progression and quantifying target engagement for relevant therapies. In this study, we validate one MTH1 inhibitor TH287 as a PET MTH1 radiotracer. TH287 was radiolabeled with tritium and the binding of [3H]TH287 to MTH1 was evaluated in live glioblastoma cells (U251MG) through saturation and competitive binding assays, together with in vitro enzymatic assays. Furthermore, TH287 was radiolabeled with carbon-11 for in vivo microPET studies. Saturation binding assays show that [3H]TH287 has a dissociation constant (Kd) of 1.97 ± 0.18 nM, Bmax of 2676 ± 122 fmol/mg protein for U251MG cells, and nH of 0.98 ± 0.02. Competitive binding assays show that TH287 (Ki: 3.04 ± 0.14 nM) has a higher affinity for MTH1 in U251MG cells compared to another well studied MTH1 inhibitor: (S)-crizotinib (Ki: 153.90 ± 20.48 nM). In vitro enzymatic assays show that TH287 has an IC50 of 2.2 nM in inhibiting MTH1 hydrolase activity and a Ki of 1.3 nM from kinetics assays, these results are consistent with our radioligand binding assays. Furthermore, MicroPET imaging shows that [11C]TH287 gets into the brain with rapid clearance from the brain, kidney, and heart. The results presented here indicate that radiolabeled TH287 has favorable properties to be a useful tool for measuring MTH1 in vitro and for further evaluation for in vivo PET imaging MTH1 of brain tumors and other central nervous system disorders.

Keywords: MTH1; PET; TH287; binding assay; radiotracer.

MeSH terms

  • Animals
  • Biomarkers, Tumor / isolation & purification*
  • Biomarkers, Tumor / metabolism
  • Brain / diagnostic imaging
  • Brain / metabolism
  • Cell Line, Tumor
  • Crizotinib / pharmacology
  • DNA Repair Enzymes / antagonists & inhibitors
  • DNA Repair Enzymes / genetics*
  • DNA Repair Enzymes / isolation & purification
  • Glioblastoma / diagnostic imaging*
  • Glioblastoma / genetics
  • Glioblastoma / pathology
  • Heart / diagnostic imaging
  • Humans
  • Kidney / diagnostic imaging
  • Kidney / metabolism
  • Mice
  • Phosphoric Monoester Hydrolases / antagonists & inhibitors
  • Phosphoric Monoester Hydrolases / genetics*
  • Phosphoric Monoester Hydrolases / isolation & purification
  • Positron Emission Tomography Computed Tomography
  • Pyrimidines / chemistry
  • Pyrimidines / pharmacology*


  • Biomarkers, Tumor
  • Pyrimidines
  • TH287 compound
  • Crizotinib
  • Phosphoric Monoester Hydrolases
  • 8-oxodGTPase
  • DNA Repair Enzymes