Liu Shiwei,Yuan Fukang,Dong Hui,Zhang Jiaqi,Mao Xinyu,Liu Yangsui,Li Huansong
Background Hepatocellular carcinoma (HCC) is the leading cause of cancer-related deaths worldwide, and the lack of effective biomarkers for early detection leads to poor therapeutic outcomes. Prostaglandin E Synthase 3 (PTGES3) is a putative prognostic marker in many solid tumors; however, its expression and biological functions in HCC have not been determined. The proteolysis-targeting chimera (PROTAC) is an established technology for targeted protein degradation. Compared to the small-molecule PROTAC, the peptide PROTAC (p-PROTAC) utilizes peptides bound to target proteins to mediate the ubiquitination and degradation of undruggable proteins. This study aimed to use the PROTAC technology to develop a PTGES3 degrader liposome complex containing a PTGES3-binding peptide and the E3 ubiquitin ligase ligand pomalidomide for regulating cell function and provide a novel pathway for treating HCC. Results In this study, we demonstrated that PTGES3 is highly expressed in HCC at the transcriptional and protein levels; furthermore, PTGES3 was identified as a novel drug target that could potentially treat HCC. Hence, we developed PTGES3-PROTACs by adjusting the ligand ratio to optimize the efficacy of degradation agents. The results revealed that PTGES3-PROTAC effectively degraded PTGES3 protein and strongly weakened the HCC malignant phenotype in vitro and in vivo. Conclusions Our findings revealed that the highly selective PTGES3 proteolysis is a potential therapeutic strategy for HCC, and PTGES3 degraders PTGES3-PROTACs can be developed as safe and effective drugs for HCC treatment. Graphical abstract
BackgroundSimiao Pill (SMP) has been demonstrated to suppress inflammation and modulate immune function, thereby influencing the onset and progression of rheumatoid arthritis (RA). Nonetheless, the specific molecular mechanisms and targets through which SMP mediates metabolic regulation and enhances immune function have yet to be fully elucidated.ObjectiveIn this study, we employed an integrated approach combining the analysis of dysregulated metabolites and proteins to identify, screen, and validate the metabolic regulatory targets of SMP in adjuvant-induced arthritis (AIA) rats by using pseudotargeted metabolomics and 4D-DIA quantitative proteomics methodologies.MethodsAn AIA rat model was developed, and SMP was administered to AIA rats. Subsequently, assessments were conducted on paw edema, arthritis scores, histopathological changes and IL-1 β content of inflammatory factors in AIA rats. UHPLC-QTOF-MS/MS was employed to analyze endogenous metabolites in the serum. Metabolic pathway and protein profile were performed on the biomarkers. The protein-lipid-phenotype map for the SMP-treated rats was constructed and the primary target closely related to the metabolic regulation of SMP was further screened and verified.ResultsPseudotargeted metabolomics analysis revealed that SMP can mitigate the down-regulation of lipid levels in AIA rats. Pathway enrichment analysis identified arachidonic acid metabolism as the most significantly affected metabolic pathway and SMP was found to substantially ameliorate the dysregulation of this pathway in AIA rats. Subsequent protein profiling led to the identification of five key proteins, with noteworthy obvious corrective effects observed on Ptges3 and Alox15 due to SMP treatment. A comprehensive protein-lipid-phenotypic landscape of SMP-treated rats was analyzed for the specific molecular expressions associated with the arachidonic acid pathway. According to the correlation matrix of dysregulated metabolite/protein, we found that Ptges3 was ranked as the primary target closely related to the metabolic regulation of SMP, a finding further validated through immunofluorescence staining in rat joint and synovial cells.ConclusionOur study confirmed that SMP exerts an anti-arthritic effect by modulating the arachidonic acid metabolic network via the Ptges3 protein in rat joints and human rheumatoid arthritis synovial fibroblasts. This finding offers a novel mechanistic insight into the pharmacological action of SMP in AIA rats. It informs future research on the therapeutic potential of SMP in RA.
Introduction Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal lung disease characterized by chronic inflammation and fibroblast activation, with limited treatment options. The bioactive diterpenoid andrographolide (AP) exhibits anti-inflammatory and antifibrotic properties, but its direct molecular targets and precise mechanism remain unclear. Objectives This study aimed to identify the primary functional target of AP and elucidate the molecular mechanism underlying its therapeutic effects against pulmonary fibrosis. Methods We employed activity-based protein profiling (ABPP) to identify covalent targets of AP in macrophages. Functional validation was performed using siRNA knockdown, enzymatic assays, molecular docking, and biophysical analyses. The therapeutic efficacy and target dependency of AP were evaluated in bleomycin-induced pulmonary fibrosis and LPS-induced acute lung injury mouse models, utilizing lung-specific Ptges3 knockdown. Results AP covalently bound to Cys58 of prostaglandin E synthase 3 (Ptges3), an allosteric site distinct from its catalytic and Hsp90-binding regions. This binding inhibited Ptges3 enzymatic activity, reduced prostaglandin E2 (PGE 2 ) production, and disrupted the Ptges3-Hsp90 chaperone complex, leading to suppressed NF-κB signaling. Genetic knockdown of Ptges3 significantly attenuated the anti-inflammatory and antifibrotic effects of AP both in vitro and in vivo . Conclusions Our findings establish Ptges3 as a critical functional target of AP. AP attenuates pulmonary fibrosis through a dual mechanism involving covalent inhibition of Ptges3 and disruption of the Ptges3-Hsp90-NF-κB axis, highlighting AP as a promising therapeutic agent and Ptges3 as a novel druggable target for IPF.
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