Fragment-Derived Nicotinic Acid Analogues Inhibit hCA III and Downregulate CA3 Expression in HepG2 Cells

Journal article

Chronic oxidative stress and lipid imbalance drive metabolic disorders such as obesity and non-alcoholic fatty liver disease, yet few therapies target the upstream redox imbalance in key tissues. Human carbonic anhydrase III (hCA III), a redox-associated enzyme enriched in liver and adipose tissue, has long remained pharmacologically elusive due to its low catalytic activity and lack of modulators. Here, we identify fragment-like nicotinic acid derivatives as non-sulfonamide hCA III modulators and evaluate their associated cellular effects. Using an esterase activity assay, we screened 25 analogues and identified two fragment-like hits, compound 17 (2-thioethyl) and compound 22 (6-morpholino), with IC50 values of 487 and 361 µM, respectively. Orthogonal thermal shift analysis supported compound-protein interaction, and selected hits were subsequently evaluated in HepG2 cells. Both compounds were associated with reduced CA3 mRNA expression after treatment at 1 µM, while their cellular phenotypes diverged, with compound 22 increasing ROS under oxidative stress conditions and compound 17 affecting mitochondrial membrane potential. Taken together, these findings identify tractable nicotinic acid-derived fragment hits and associated cellular phenotypes that warrant further mechanistic investigation. These fragment-like hits provide a practical starting point for studying the redox-linked biology of hCA III.

Authors: Abuhammad, A; Sabri, T; Ababneh, NA; Ali, RA; Ismail, MA

• Publication status: Published
• Peer review status: Peer reviewed
• Publisher: MDPI
• Journal: Biomolecules
• Volume: 16
• Issue: 4
• Pages: 599
• Article number: 599
• Publication date: 2026-04-17
• Acceptance date: 2026-04-15
• DOI: 10.3390/biom16040599
• EISSN: 2218-273X
• ISSN: 2218-273X
• Language: English
• Keywords: oxidative stress; carbonic anhydrase III; fragment-based drug discovery; nicotinic acid derivatives; metabolic disease; gene expression modulation