Dual carbonic anhydrase ix/xii inhibitors and carbon monoxide releasing molecules modulate LPS-mediated inflammation in mouse macrophages

Low concentrations of carbon monoxide (CO) were reported to exhibit anti-inflammatory effects when administered in cells by suitable chemotypes such as CO releasing molecules (CO-RMs). In addition, the pH-modulating abilities of specific carbonic anhydrase isoforms played a crucial role in different models of inflammation and neuropathic pain. Herein, we report a series of chemical hybrids consisting of a Carbonic Anhydrase (CA) inhibitor linked to a CO-RM tail (CAI/CO-RMs). All compounds and their precursors were first tested in vitro for their inhibition activity against the human CA I, II, IX, and XII isoforms as well their CO releasing properties, aiming at corroborating the data by means of molecular modelling techniques. Then, their impact on metabolic activity modulation of RAW 264.7 mouse macrophages for 24 and 48 h was assessed with or without lipopolysaccharide (LPS) stimulation. The compounds were shown to counteract the inflammatory stimulus as also indicated by the reduced tumor necrosis factor alpha (TNF-α) release after treatment. All the biological results were compared to those of N-acetylcysteine (NAC) as a reference antioxidant compound. Within the series, two CAI/CO-RM hybrids (1 and 2), bearing both the well-known scaffold able to inhibit CAs (acesulfame) and the cobalt-based CO releasing portion, induced a higher anti-inflammatory effect up to 48 h at concentrations lower than NAC.