Structure-Activity Relationship of 3-Methylcytidine-5'-α,β-methylenediphosphates as CD73 Inhibitors

J Med Chem. 2022 Feb 10;65(3):2409-2433. doi: 10.1021/acs.jmedchem.1c01852. Epub 2022 Jan 26.

Abstract

We recently reported N4-substituted 3-methylcytidine-5'-α,β-methylenediphosphates as CD73 inhibitors, potentially useful in cancer immunotherapy. We now expand the structure-activity relationship of pyrimidine nucleotides as human CD73 inhibitors. 4-Chloro (MRS4598 16; Ki = 0.673 nM) and 4-iodo (MRS4620 18; Ki = 0.436 nM) substitution of the N4-benzyloxy group decreased Ki by ∼20-fold. Primary alkylamine derivatives coupled through a p-amido group with a varying methylene chain length (24 and 25) were functionalized congeners, for subsequent conjugation to carrier or reporter moieties. X-ray structures of hCD73 with two inhibitors indicated a ribose ring conformational adaptation, and the benzyloxyimino group (E configuration) binds to the same region (between the C-terminal and N-terminal domains) as N4-benzyl groups in adenine inhibitors. Molecular dynamics identified stabilizing interactions and predicted conformational diversity. Thus, by N4-benzyloxy substitution, we have greatly enhanced the inhibitory potency and added functionality enabling molecular probes. Their potential as anticancer drugs was confirmed by blocking CD73 activity in tumor tissues in situ.

Publication types

  • Research Support, N.I.H., Intramural
  • Research Support, Non-U.S. Gov't
  • Video-Audio Media

MeSH terms

  • 5'-Nucleotidase / antagonists & inhibitors*
  • 5'-Nucleotidase / metabolism
  • Adult
  • Cytosine Nucleotides / chemical synthesis
  • Cytosine Nucleotides / metabolism
  • Cytosine Nucleotides / pharmacology*
  • Diphosphonates / chemical synthesis
  • Diphosphonates / metabolism
  • Diphosphonates / pharmacology*
  • Enzyme Inhibitors / chemical synthesis
  • Enzyme Inhibitors / metabolism
  • Enzyme Inhibitors / pharmacology*
  • GPI-Linked Proteins / antagonists & inhibitors
  • GPI-Linked Proteins / metabolism
  • Humans
  • Male
  • Molecular Docking Simulation
  • Molecular Dynamics Simulation
  • Molecular Structure
  • Neoplasms / enzymology
  • Palatine Tonsil / enzymology
  • Protein Binding
  • Structure-Activity Relationship

Substances

  • Cytosine Nucleotides
  • Diphosphonates
  • Enzyme Inhibitors
  • GPI-Linked Proteins
  • 5'-Nucleotidase
  • NT5E protein, human