Inflammatory bowel disease (IBD) is an umbrella term for chronic diseases that produce inflammation in the digestive system, particularly Crohn's disease (CD) and ulcerative colitis (UC). UC is more limited in location than CD and is characterized by hemorrhagic diarrhea, a condition with serious ramifications on patients’ quality of life (Kobayashi et al., 2020). Early UC is typically treated with corticosteroids, but adverse reactions to these drugs can be severe and can lead to drug dependence and drug resistance (Matsuoka et al., 2018). At present, the drugs commonly used in clinics include aminosalicylate and immunosuppressive agents, but these drugs have low efficacy and are also accompanied by serious side effects (Wan et al., 2014).Thus, there is an urgent need to find more effective drugs with less toxicity and side effects. Previous research has indicated that inhibition of the NF-κB pathway, the PI3K/Akt pathway, the JAK/STAT pathway, the MAPK pathway, and the Notch pathway can all be effective in relieving UC (Chen et al., 2021; Huang et al., 2022; Lin et al., 2022; Yang et al., 2022; Zheng et al., 2022).

Rabdosia rubescens (Hemsl.) H.Hara is a kind of natural plant with homologous medicinal and edible properties (We have checked with The World Flora Online website and verified. 20120323 http://www.worldfloraonline.org/tpl/kew-103019). It is commonly used as Chinese herbal medicine and can also be used as herbal tea to drink. In traditional Chinese medicine, Rabdosia rubescens has the effects of clearing heat and removing toxins, activating blood circulation and relieving pain, it is mainly used for treating inflammatory diseases such as sore throats and tonsillitis (Chen et al., 2022). Recent studies have revealed that its active components possess antitumor activity (Yang et al., 2018). In current clinical applications, Rabdosia rubescens is mainly used for the treatment of inflammatory diseases such as pharyngitis and tonsillitis (Leung et al., 2005; Zhang et al., 2020), pharmacochemical and pharmacological studies have shown that its active constituents are mainly diterpenoids, the most abundant of which are oridonin and ponicidin (Lu et al., 2007). Oridonin has been shown to have in anti-inflammatory and anti-tumor properties. However, oridonin has also been shown to have certain hepatotoxicity in animal experiments (Li et al., 2021) and to affect the expression levels of the drug metabolism enzymes CYP2C and CYP3A (Yi-Wen et al., 2018), potentially resulting in adverse drug interactions (Zhang et al., 2014). In contrast, there have been few similar studies into the anti-inflammation activity of ponicidin.

Heat shock protein 90 (HSP90) is the most abundant molecular chaperone protein in cells (Hartl et al., 2011), and it controls the function and activity of client by promoting the correct folding of proteins or the binding between proteins (Schopf et al., 2017). There are hundreds of client proteins of HSP90, including protein kinase B (AKT), mitogen-activated protein kinase (MAPK), extracellular regulated protein kinases (ERKs), and other transcription factors such as notch1 and nuclear factor kappa-B (NF-κB), peroxisome-proliferators-activated receptors (PPARs) and epidermal growth factor receptor (EGFR), and transforming growth factor-β (TGF-β). Under stress conditions, HSP90 will is highly expressed to maintain cell homeostasis, increasing up to 5% (Fujita et al., 2017), and thus promoting the correct folding and maturation of misfolded clients, MAPK and NF-κB inflammation-related signal pathways are then activated, and the release of inflammatory factors increases (Costa et al., 2020). Therefore, inhibition of HSP90 can negatively regulate inflammatory diseases such as arthritis, colitis, and septicemia (Chatterjee et al., 2007; Collins et al., 2013; Conte et al., 2015). HSP90 inhibitors exert anti-inflammatory activity mainly by inhibiting inflammatory factors and increasing the expression of heat shock protein 70 (HSP70), which can also inhibit the synthesis of inflammatory factors such as TNF, IL-1β, and IL-6 (Costa et al., 2020; Tanaka et al., 2007). Based on the powerful biological role of HSP90, targeting HSP90 to treat inflammatory diseases is a promising treatment approach.

In this study, we firstly verified the anti-inflammatory activity of ponicidin using cells and animal models and explored the effects on signaling pathways. Meanwhile, we identified the target of ponicidin using molecular target fishing, and the interaction site of ponicidin with the target was further characterized using molecular dynamics simulations, molecular interaction assays and kinase activity assay in vitro. Taken together, this work provides a theoretical basis for future studies of the anti-inflammatory activity of ponidin and a structural basis for the development of targeted HSP90 inhibitors.