Discovery of a Mixed and Prodrug-Like Inhibition Mechanism for Phosphocoumarins and Phosphoquinolinones against Human Carbonic Anhydrases

  • J Med Chem. 2026 May 14;69(9):11638-11648. doi: 10.1021/acs.jmedchem.6c00915.
Alessio Nocentini  1 ,  Simone Giovannuzzi  1 ,  Vincenzo Alterio  2 ,  Alessandro Bonardi  1 ,  Rudolfs Barons  3  4 ,  Raivis Zalubovskis  3  4 ,  Wagdy M Eldehna  5 ,  Rossella Aronne  6 ,  Davide Esposito  2 ,  Enrico Luchinat  7  8 ,  Giuseppina De Simone  2 ,  Gianluca Bartolucci  1 ,  Paola Gratteri  1 ,  Mattia Mori  6 ,  Claudiu T Supuran  1
Affiliations
  • 1. NEUROFARBA Department, Section of Pharmaceutical Sciences, University of Florence, Sesto Fiorentino 50019, Italy.
  • 2. Institute of Biostructures and Bioimaging, National Research Council, Napoli 80145, Italy.
  • 3. Latvian Institute of Organic Synthesis, Riga LC-1006, Latvia.
  • 4. Institute of Chemistry and Chemical Technology, Riga Technical University, Riga LV-1048, Latvia.
  • 5. Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Kafrelsheikh University, Kafrelsheikh 33516, Egypt.
  • 6. Department of Biotechnology, Chemistry and Pharmacy, University of Siena, Siena 53100, Italy.
  • 7. CERM - Magnetic Resonance Center, University of Florence, Sesto Fiorentino 50019, Italy.
  • 8. Department of Chemistry "Ugo Schiff", University of Florence, Sesto Fiorentino 50019, Italy.
PMID: 42055938 DOI: 10.1021/acs.jmedchem.6c00915
Abstract

Phosphocoumarins and a first-in-class unsubstituted phosphoquinolinone are disclosed as previously unrecognized Carbonic Anhydrase (CA) inhibitors, displaying multimodal inhibition within a tunable coumarin-like scaffold. Acidic phosphocoumarins display inhibition of physiologically relevant human CAs, particularly tumor-associated isoforms IX and XII (KIs: 0.08-0.28 μM) through a composite, two-step mechanism: the ligand first anchors the zinc-bound water molecule before displacing it to directly coordinate the catalytic zinc ion, without CA-mediated hydrolysis. Conversely, a methyl-ester phosphocoumarin functions as an isoform-selective prodrug, undergoing CA-mediated cyclic phosphoester hydrolysis to selectively generate a potent hCA IX/XII inhibitor (KIs: 54-62 nM), whereas the phosphoquinolinone acts as a direct binder (KIs: 0.18-0.29 μM vs hCA IX/XII). The complementary mechanisms are supported by QM/MM and long-time scale MD simulations, crystallographic studies, 31P NMR, HRMS, and MS/MS. Selected derivatives exhibit low-micromolar antiproliferative activity and induce Apoptosis in Cancer cells, fostering phosphorus-heterocycles as a mechanistically rich platform for isoform-selective CA inhibition and targeted drug design.

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