Mechanistic insights into the anti-inflammatory effects of isocoronarin D associated with reduced PKCδ and PI3K/Akt signalling in LPS-activated macrophages.

Curcuma comosa Roxb., a Thai medicinal plant, has long been used to relieve postmenopausal symptoms and is known to exhibit anti-inflammatory properties. Isocoronarin D, a labdane diterpene and the main bioactive compound in the aerial parts of C. comosa, has previously been reported to induce foetal haemoglobin synthesis. However, its anti-inflammatory activity, predicted molecular targets, pharmacokinetic characteristics, and toxicity remain insufficiently characterised. Therefore, this study integrates in silico predictions of its molecular targets, pharmacokinetic behaviour and toxicity with in vitro validation of its anti-inflammatory effects and underlying mechanisms to address these knowledge gaps. Initially, network pharmacology analysis identified protein kinase Cδ (PKCδ), phosphoinositide 3-kinase (PI3K), and cyclooxygenase-2 (COX-2) as inflammation-related targets computationally associated with isocoronarin D. These predicted targets were subsequently validated in vitro, where isocoronarin D significantly reduced the production of inflammatory mediators, including inducible nitric oxide synthase (iNOS), COX-2, tumour necrosis factor-α, interleukin (IL)-1β, and IL-6, in lipopolysaccharide (LPS)-activated RAW264.7 macrophages. Moreover, isocoronarin D inhibited the phosphorylation of PKCδ, PI3K, and Akt, indicating suppression of key inflammatory signalling pathways. The observed inhibition of these inflammatory proteins, together with the predicted interactions revealed by molecular docking, supports a mechanistically relevant role for these proteins as inflammatory targets of isocoronarin D. In addition, in silico SwissADME, pkCSM, and GUSAR analyses suggested drug-like characteristics, favourable pharmacokinetic properties, and a non-toxic safety profile. Collectively, these findings indicate that isocoronarin D exerts multi-target anti-inflammatory activity and warrants further in vivo investigation to better elucidate its therapeutic relevance in inflammation-related diseases.