Discovery of cinnamoyl-flavonoid hybrid derivatives as inhibitors of SARS-CoV-2 Mpro and anti-inflammatory agents: Experimental and in silico insights into their efficacy against lipopolysaccharide-induced lung injury.

The chemical structures of the parental compounds of flavonoids from Boesenbergia rotunda were modified by conjugation with cinnamic acid to form cinnamoyl-flavonoid hybrid derivatives with enhanced anti-inflammatory and SARS-CoV-2 Mpro-inhibitory properties. Cinnamoyl-flavonoid hybrid derivatives 6 and 10 showed the potential to inhibit SARS-CoV-2 Mpro with IC50 values of 52.49 and 22.62 μM. Compounds 6 and 10 showed lower cytotoxicity in the human lung cell lines MRC-5 and A549 at concentrations greater than 50 μM. The effects of compounds 6 and 10 on cell viability were studied in a 3D co-culture model of A549 and MRC-5 treated with lipopolysaccharide (LPS) and observed through confocal microscopy. Compounds 6 and 10 downregulated p65 mRNA expression, resulting in a reduction of pro-inflammatory cytokines, including Interleukin 8 (IL-8) and Monocyte Chemoattractant Protein-1 (MCP-1/CCL2), leading to an anti-inflammatory response through Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signalling pathways. Compound 6 showed potential anti-inflammatory activity, downregulating Bcl-2 Associated X gene (BAX), which resulted in inhibition of apoptotic cell death when compared to compound 10. In silico molecular dynamic simulation shed light on how these cinnamoyl-flavonoid hybrid derivatives interact with myeloid differentiation factor 2 (MD-2), which is involved in the inflammatory response. Our findings suggest that cinnamoyl-flavonoid hybrid derivatives show potential as anti-inflammatory drugs and anti-SARS-CoV-2 drugs.