Methemoglobin incites primaquine toxicity through single-electron oxidation and modification

Area of Science:

• Biochemistry
• Pharmacology
• Toxicology

Background:

• Primaquine (Pq) usage is limited by drug-associated hemotoxicity.
• Metabolic products of Pq are implicated in its toxicity.

Purpose of the Study:

• To investigate the mechanism by which methemoglobin (MetHb) contributes to primaquine (Pq) hemotoxicity.
• To characterize the oxidized form of Pq (Pqox) and its effects on red blood cells (RBCs).

Main Methods:

• Molecular modeling to identify the Pq binding pocket in MetHb.
• UPLC and mass spectrometry to predict Pq metabolites.
• Characterization of Pqox hemotoxicity using MetHb generation, oxidative stress assays, antioxidant enzyme inhibition studies, and SEM.

Main Results:

• MetHb binds Pq at the heme site (KD = 6.4 μM) and oxidizes it to Pqox via single electron transfer.
• Pqox administration to RBCs significantly increased MetHb formation, reactive oxygen species (ROS) generation, and inhibited antioxidant enzymes.
• SEM analysis revealed visible plasma membrane damage (holes) in Pqox-exposed RBCs.

Conclusions:

• The MetHb-H2O2 system converts Pq into a highly reactive oxidative form (Pqox), leading to severe hemolysis.
• The involvement of Pq radical species in MetHb-H2O2-mediated Pq hemolysis is indicated.
• MetHb appears to activate Pq's toxic properties, suggesting peroxidase inhibitors as a potential therapeutic strategy.