Bridging the inherent bioactivity of botanical precursors with the tunable physicochemical properties of nanoscale particles, traditional Chinese medicine-derived carbon dots (TCM-CDs) have emerged as a promising innovative platform in sustainable nanotheranostics. Synthesized via eco-friendly routes such as hydrothermal or microwave-assisted carbonization, these nanomaterials exhibit “pharmacological inheritance,” a notable phenomenon wherein bioactive moieties from precursors (e.g., polyphenols, alkaloids, and polysaccharides) are partially retained or structurally transformed, contributing to their observed therapeutic activities. By combining structural features including ultrasmall size (<10 nm) and tunable optical properties with intrinsic bioactivity, TCM-CDs show promising preclinical potential in imaging-guided therapy, anti-inflammation, oncology, hemostasis, and tissue regeneration. Despite this considerable preclinical promise, the transition of TCM-CDs from bench to bedside remains impeded by critical bottlenecks, including batch-to-batch heterogeneity, lack of standardized quality control (QC) protocols, suboptimal delivery precision, and incomplete understanding of their long-term biosafety and underlying molecular mechanisms. Herein, we systematically review the latest advancements in the synthesis, purification, and biomedical applications of TCM-CDs. To facilitate future clinical translation, we highlight that integrating multi-omics technologies with precisely controlled manufacturing processes represents a critical path toward advancing the next generation of herbal-derived nanomedicines.
Tyrosine phosphorylation (pTyr) plays critical roles in diverse physiological and pathological processes. Mass spectrometry (MS)-based strategies have been developed for mapping pTyr proteome. Nonetheless, ultradeep characterization of pTyr signaling is still hindered by inherently low abundance and highly dynamic range. Here, we designed a sensitive and cost-efficient workflow combining Src homology 2 (SH2) superbinder-based enrichment with data independent acquisition (DIA)-based MS analysis for tyrosine phosphoproteomics. By applying the hybrid DIA (hybDIA) search strategy, this approach identified over 7000 high-confidence pTyr sites, achieving the deepest coverage of the tyrosine phosphoproteome in a single measurement reported thus far. Moreover, this approach showed superior performance on detection sensitivity, data completeness, quantitative robustness, and accuracy. Applied to gefitinib resistance in non-small cell lung cancers (NSCLCs), multiple canonical resistance pathways, including phosphatidylinositol 3-kinase/protein kinase B (PI3K-AKT), epidermal growth factor (EGF) receptor-tyrosine kinase inhibitor (EGFR-TKI) resistance, and erythroblastic leukemia viral oncogene B (ERBB) pathways, were significantly enriched, confirming the reliability and applicability of our analytical workflow. In addition, glycolysis/gluconeogenesis and neurotrophin signaling pathways showed high potential to participate in gefitinib resistance. In light of these findings, drug combination experiments further demonstrated that inhibitors of acetyl-CoA carboxylase (ACC) or tropomyosin receptor kinase (TRK) significantly enhanced the therapeutic efficacy of gefitinib. Phosphorylation at nuclear envelope membrane protein 1 (NEMP1) Ser368, ribosomal protein S6 kinase b1 (RPS6KB1) Ser441/Thr444, and cortactin (CTTN) Tyr154 showed potentially important roles in gefitinib resistance. Functional rescue assay of NEMP1 Ser368 and RPS6KB1 Ser441/Thr444 further demonstrated that phosphorylation-defective mutant, compared to the phosphomimetic mutant, significantly enhanced the sensitivity of resistant cell to gefitinib. Collectively, this study provides a powerful tool for Tyr phosphoproteomics, facilitating mechanistic insights into the EGFR-TKI resistance in NSCLC.
Immunological agents are vital for treating cancer and autoimmune diseases, yet their therapeutic potential is limited by complex multi-organ adverse drug reactions (ADRs). Existing databases lack comprehensive integration of real-world ADR patterns, quantitative risk metrics, and mechanistic insights. To address this, the comprehensive atlas of immunological agents and adverse drug reactions (IMADRD) was developed. IMADRD provides three core data components: (1) ADR occurrence features, leveraging 194,035 clinical and real-world datasets; (2) quantitative metrics, featuring 18,305 unique ADRs across 27 system organ classes (SOCs), including 75,425 incidence records, 15,248 severity annotations, and 131,494 signal intensities; and (3) Drug-ADR-protein interaction networks, connecting 315 drugs, 2586 ADRs, 2361 proteins, and 22,892 associations. Then, targeted analysis of immune checkpoint inhibitors (ICIs) further validates IMADRD’s practical utility. Through systematic evaluation of approximately 100 ICIs, IMADRD accurately identifies high-risk SOCs and uncovers rare but severe neuropsychiatric risks. More notably, IMADRD enables quantitative assessment of drug similarity through ADR profile comparisons, providing clinicians with evidence-based guidance for selecting alternative therapies or avoiding known high-risk agents. Overall, by integrating and analyzing multidimensional data, IMADRD advances our understanding of the biological mechanisms of ADRs and supports the development of personalized immunotherapy regimens. IMADRD is freely accessible at http://imadrd.bipslab.com.
Glycosylation is crucial for the effector functions of therapeutic monoclonal antibodies (mAbs). However, the specific glycoprofile of tocilizumab, a humanized anti-interleukin-6 (IL-6) receptor mAb, remains inadequately characterized in the current literature. To address this, we designed and synthesized a novel pair of stable isotope-labeled mass spectrometry (MS) probes: d0/d15-[4-(1-carboxyethyl)phenyl]methyl-triphenylphosphonium (d0/d15-CEBTPP). Featuring a permanent positive charge, these probes not only significantly enhance the ionization efficiency and detection sensitivity of glycans in MS positive-ion mode, but also effectively neutralize the intrinsic negative charge of sialic acid residues, thereby substantially improving the structural stability of acidic glycans during analysis. Consequently, a novel relative quantification strategy for glycans was established by coupling d0/d15-CEBTPP isotopic labeling with ultra-high performance liquid chromatography-high-resolution MS (UHPLC-HRMS). Methodological validation using a sialoglycopeptide (SGP) model demonstrated excellent linearity (R2 = 0.9990), reproducibility (coefficient of variation (CV) = 3.50%), and accuracy (relative error (RE) = 0.76%) across a 10-fold molar mixing range. Application of this strategy to the comprehensive characterization of tocilizumab successfully identified 23 N-glycans, including 14 fucosylated and 7 sialylated glycoforms. Furthermore, the method was successfully applied in a rat model, precisely tracking the dynamic fluctuations of targeted serum glycan levels from 2 min to 36 h post-administration. Overall, this study establishes a robust strategy for the efficient and reliable relative quantification of trace glycans in complex biological matrices without relying on glycan standards. This work offers a powerful analytical tool for comprehensive glycosylation evaluation in biopharmaceuticals and disease-related comparative glycomics.
This comprehensive narrative review examines how artificial intelligence (AI) is being applied in drug discovery and pharmacy practice. In pharmaceutical research, methods such as machine learning (ML) and neural networks help predict key physicochemical properties, optimize pharmacokinetic and pharmacodynamic profiles, and support early toxicity assessment. These applications can accelerate the identification of promising drug candidates and make the design and conduct of clinical trials more efficient and cost-conscious. In clinical practice, AI supports medication management by automating prescription verification, assisting in dose selection, and detecting adverse drug reactions (ADRs) close to real time. AI based clinical decision support systems are increasingly used to reduce medication errors and to monitor patient adherence to treatment. High income countries are also deploying AI driven platforms in digital therapeutics, pharmaceutical supply chain management, and real time drug performance monitoring. Despite this progress, several barriers limit wider adoption. Biased or unrepresentative training data, the opaque “black box” nature of many models, and fragmented regulatory requirements remain major challenges. Ethical concerns related to patient privacy, data security, and algorithmic fairness require robust governance frameworks. Furthermore, the inherent complexity of biological systems and the shortage of professionals trained to interpret AI outputs constrain effective implementation. Overall, the potential of AI to reshape pharmaceutical science is substantial, but its long-term impact will depend on addressing regulatory and ethical issues and strengthening collaboration among researchers, healthcare professionals, and policy makers.
Chronic kidney disease (CKD) characterized by the progressive loss of renal, represents a significant global health challenge. Central to CKD progression is kidney fibrosis, an irreversible process marked by the accumulation of extracellular matrix proteins. The development of effective antifibrotic therapies is thus crucial for improving patient outcomes. We conducted a comprehensive analysis of bulk RNA sequencing data from unilateral ureteral obstruction (UUO) and folic acid (FA)-induced nephropathy mice models, combined with single-cell RNA sequencing (scRNA-seq) to explore cellular heterogeneity and molecular mechanisms of kidney fibrosis. Differential gene expression analysis, gene co-expression network (CN) analysis, time-series clustering, and cell marker analysis were employed to identify core fibrosis-related genes. Our analyses revealed a set of 37 core fibrosis-related genes which are largely associated with inflammation and immune response and contributed to the production of extracellular matrix (ECM), with Ckap4 standing out as a key marker of kidney fibrosis. In vivo experiments demonstrated that knockdown of Ckap4 significantly reduced kidney fibrosis in UUO mice, as evidenced by decreased collagen deposition and improved renal function. Additionally, the repurposed small molecule drugs, especially CGP-60474, showed promising antifibrotic effects, further highlighting the potential of Ckap4 as a therapeutic target.
Myocardial ischemia/reperfusion (I/R) injury persists as a pivotal unresolved bottleneck in treating acute myocardial infarction. Post-reperfusion inflammation, driven by macrophage metabolic and functional reprogramming, exacerbates this injury. Tectorigenin (TEC), an active component of Belamcanda chinensis, exerts marked cardioprotection against myocardial I/R injury by targeting macrophage-mediated inflammation. TEC significantly improved cardiac function, reduced infarct size, and suppressed cardiomyocyte apoptosis. RNA sequencing (RNA-seq) identified the type I interferon (IFN-I) signaling pathway as a key target. Mechanistically, TEC reduced macrophage reactive oxygen species (ROS) levels and upregulated the transcriptional repressor basic helix-loop-helix family member e41 (BHLHE41). BHLHE41 subsequently suppressed expression of the cytosolic RNA sensor melanoma differentiation-associated protein 5 (MDA5), encoded by IFN induced with helicase C domain 1 (Ifih1)), thereby blocking downstream activation of TANK-binding kinase 1 (TBK1) and IFN regulatory factor 3 (IRF3) to reduce IFN-β production. Moreover, multiple orthogonal assays identified 3-oxoacid CoA-transferase 1 (OXCT1), a crucial enzyme involved in ketone body metabolism, as a direct target of TEC, with TRP413 serving as the critical binding residue. This interaction promotes lysosomal degradation of OXCT1 and elevates intracellular ketone body levels, which enhance Bhlhe41 messenger RNA (mRNA) stability and inhibit MDA5-dependent IFN-I signaling. Exogenous ketone body supplementation mimicked these effects, while genetic knockdown of Oxct1 or Bhlhe41 confirmed their pivotal roles in regulating macrophage activation. Collectively, this study identifies a novel macrophage OXCT1-BHLHE41-MDA5 axis modulating IFN-I-driven inflammation during I/R injury, establishes TEC as a promising therapeutic agent, and highlights ketone metabolism as a potential cardioprotective target.
Apoptotic bodies (ApoBDs) are membrane-bound vesicular structures generated during the late stages of programmed cell death. They were traditionally regarded as inert cellular debris that is removed by phagocytes, but increasing evidence suggests that ApoBDs are not simply passive byproducts of apoptosis. Instead, they can participate in intercellular communication and contribute to the maintenance of tissue homeostasis. Their formation involves coordinated cytoskeletal remodeling, membrane blebbing, and vesicle fragmentation, which together package diverse nuclear and cytoplasmic components into membrane-enclosed structures. Recent studies further indicate that the composition and biological effects of ApoBDs are closely related to their cellular origin. ApoBDs derived from stem cells, immune cells, and tumor cells may carry different cargos and exhibit distinct functional tendencies, leading to different effects on recipient cells in different biological contexts. These source-related differences not only reflect the heterogeneity of ApoBDs, but also distinguish them from other extracellular vesicle subtypes. In this review, we summarize recent progress in the mechanisms of apoptotic body formation and discuss how cellular origin influences their cargo composition, recognition, uptake, and downstream biological effects. We also discuss their potential relevance in tissue repair, immune regulation, disease intervention, and biomarker-related applications, while highlighting current limitations in this field, including methodological inconsistency, insufficient standardization, and the challenges that still restrict their further translational application.
Black soybean oil (BSO) is a traditional Chinese medicine (TCM) for inflammatory skin disorders, yet its active basis remains unclear. Inspired by BSO preparation's similarity to carbon dot synthesis, we isolated fluorescent carbon-based nanoparticles (BSO-CNPs) from BSO. Analyses confirmed their characteristic nanostructure and fluorescence. Functionally, BSO-CNPs showed potent antioxidant activity, scavenging reactive oxygen species (ROS) and restoring cell viability under in vitro oxidative stress. In murine models of atopic dermatitis, topical BSO-CNPs alleviated clinical symptoms, reduced epidermal hyperplasia, suppressed mast cell infiltration, and downregulated inflammatory cytokines. Mechanistically, BSO-CNPs attenuate the NOD-like receptor family pyrin domain containing 3 (NLRP3)/caspase-1/gasdermin D (GSDMD) pyroptosis pathway by suppressing mitochondrial ROS and stabilizing lysosomal function. Furthermore, BSO-CNPs demonstrated high biosafety in vitro and in vivo. These findings elucidate the material basis of BSO’s therapeutic effects and highlight BSO-CNPs as a novel nanotherapeutic candidate for atopic dermatitis, bridging traditional herbal medicine with modern nanomedicine.
Traditional Chinese medicine (TCM) has long served as a valuable source for modern therapeutics. However, elucidating their precise mechanisms of action remains a major challenge. The inherent complexity of TCM, particularly the unclear relationship between its active substances and their molecular targets has significantly hindered its modernization and global acceptance. This review highlights recent advances in direct protein target identification technologies, including affinity-based and label-free strategies, and discusses their applicability in both monomeric compounds and complex TCM extracts. Importantly, we emphasize that while global targetome profiling can reveal extract-level protein interactions, it does not inherently resolve specific component-target relationships. To address this critical bottleneck, we outline a sequential analytical strategy that integrates global target identification with target-guided reverse ligand fishing technologies, offering a promising strategy to establish direct component-target correspondence within TCM complex mixtures. We further analyze the principles, representative applications, analytical strengths, and limitations of these approaches, and discuss future directions, including high-throughput interactomics and physiologically relevant models. By clarifying methodological integration strategies for mixture deconvolution, this review provides a practical roadmap for transitioning TCM research from empirical observations toward precise mechanistic decoding.
Epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2) are key oncogenic drivers in gastric and other solid tumors, yet the clinical benefit of conventional tyrosine kinase inhibitors such as lapatinib is limited by acquired resistance and incomplete pathway suppression. Proteolysis-targeting chimeras (PROTACs) provide an event-driven strategy for simultaneous target elimination, but their translation is often constrained by on-target, off-tumor toxicity in normal tissues expressing basal levels of EGFR/HER2. Here, we developed a series of lapatinib-derived PROTACs that recruit the von Hippel-Lindau (VHL) E3 ligase to induce dual degradation of EGFR and HER2. The lead compound, PROTAC-B7, showed superior antitumor activity to lapatinib, a selective EGFR degrader (PROTAC-A1), and a selective HER2 degrader (PROTAC-A2) in vitro and in vivo, but caused dose-limiting systemic toxicity. Mechanistically, we found that the fatty acid translocase CD36 mediates cellular uptake of fatty acid-conjugated PROTACs and is highly expressed in gastric cancer cells. Guided by this finding, we generated a second-generation molecule, PROTAC-B7.7, by conjugating PROTAC-B7 with a long-chain fatty acid. This modification enhanced CD36-dependent tumor cell uptake and intracellular accumulation, while reducing toxicity in normal cells with low CD36 expression. Consequently, PROTAC-B7.7 achieved more selective EGFR/HER2 degradation, stronger antitumor efficacy, and an improved therapeutic window in preclinical models. These findings establish CD36-mediated targeted delivery as a modular strategy to enhance the tumor selectivity and translational potential of PROTAC therapeutics.
Peroxynitrite (ONOO−) is a highly reactive species with potent oxidative properties that plays a crucial role in signal transduction. Abnormal levels of ONOO− can lead to a variety of diseases, including acute liver injury. Hence, ONOO− scavengers are emerging as potential bioactive molecules for combating acute liver injuries. However, the development of simultaneously water-soluble, selective and sensitive probes for subcellular ONOO− levels remains challenging, and there is also a lack of reliable and informative screening methods for ONOO− scavengers. In this work, we reported a luminescent iridium(III)-based probe with a 1,4:3,6-dianhydro-α-D-glucopyranose (DGP)-derived hydrazone as the recognition unit of ONOO−. The incorporation of DGP improved the aqueous solubility of the iridium(III) complex compared to the parent specie, from 7.80 × 10−3 g/L to 1.44 × 10−2 g/L. Moreover, the probe showed high sensitivity for ONOO−, with a detection limit (LOD) of as low as 0.14 μM. Finally, this probe was capable of detecting both exogenous and endogenous ONOO− in mitochondria in liver cells, and also demonstrated good penetration (approximately 75.6 μm) in 3D tumor spheroids. Leveraging its sensing merits, the probe was successfully applied for high-content screening (HCS) of the Food and Drug Administration (FDA)-approved drugs, identifying fezolinetant as an ONOO− scavenger. Taken together, this work provides a useful imaging tool for probing ONOO− functions and diagnosing liver injury, and also offers the first HCS platform for ONOO− scavengers, holding great promise for the diagnosis and therapy of ONOO−-related diseases in the future.
Knee osteoarthritis (KOA) is characterized by progressive osteochondral destruction driven by inflammatory and catabolic signaling, yet no approved structure-modifying pharmacotherapy currently exists. Duhuo Jisheng Decoction (DHJSD), a classic traditional Chinese medicine (TCM) formula widely used for KOA, requires a clearer understanding of its mechanism of action. To address this, we integrated multiple approaches: a destabilization of the medial meniscus (DMM) mouse model for initial therapeutic efficacy (n = 3 per group), bulk articular cartilage RNA-sequencing (RNA-seq) (Gene Expression Omnibus (GEO): GSE319157), network pharmacology, focused in vivo dose validation, in vitro dose-bridging assays, primary chondrocyte phospho-signaling assays, ex vivo complement activation, ultra-high-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS) serum metabolomics, molecular docking and dynamics simulations, cellular target engagement assays cellular thermal shift assay (CETSA), isothermal dose–response fingerprint CETSA (ITDRF-CETSA), and drug affinity responsive target stability (DARTS), and an in vivo C3a challenge-rescue experiment to interrogate the complement–IκB kinase beta (IKKβ)–nuclear factor kappa-B (NF-κB) axis. DHJSD treatment was associated with preservation of osteochondral integrity and cartilage matrix homeostasis in DMM mice. The initial dose cohort (2, 4, and 8 g/kg/day) showed a non-monotonic efficacy pattern; an independent focused validation cohort centered on 4 g/kg/day confirmed this dose as the most consistent for mechanistic interrogation within the refined range tested. RNA-seq and network pharmacology identified a complement–NF-κB program as a DHJSD-responsive network. Bulk-cartilage complement C3a receptor 1 (C3AR1) transcript behavior including anchor-cohort reverse transcription quantitative PCR (RT-qPCR) confirmation of the RNA-seq direction served as tissue-level contextual information, whereas mechanistic inference regarding C3AR1 was based on chondrocyte-level RT-qPCR, cartilage-localized protein expression, and downstream IKKβ–NF-κB signaling readouts. In primary chondrocytes, interleukin-1 beta (IL-1β) induced C3 and C3AR1 transcripts and activated the IKKβ–inhibitor of nuclear factor kappa B alpha (IκBα)–v-rel reticuloendotheliosis viral oncogene homolog A (RelA, P65) cascade; DHJSD-medicated serum derived from the 4 g/kg dose most consistently attenuated these responses. Intra-articular C3a challenge partially blunted DHJSD-mediated protection, whereas IKKβ inhibition with BMS-345541 partially restored structural and signaling endpoints. UHPLC-Q-TOF-MS prioritized geniposidic acid (GA) as a blood-exposed candidate constituent, and CETSA, ITDRF-CETSA, and DARTS provided convergent cellular evidence supporting GA–IKKβ interaction, although direct biochemical affinity remains to be determined. Collectively, these data support a restrained mechanistic model in which DHJSD attenuates a complement-amplified IKKβ–NF-κB inflammatory circuit in experimental KOA. GA emerges as a prioritized candidate requiring pharmacokinetic (PK) and biochemical validation, and adequately powered dose-optimization studies are still warranted.
Ulcerative colitis (UC) is a chronic and recurrent inflammatory bowel disease. Current therapeutic strategies achieve clinical remission in only 30%–60% of patients, indicating the crucial need for safer and more effective treatment options. Mitochondrial dysfunction, oxidative stress, and immune dysregulation are critically involved in UC pathogenesis. The NOD-like receptor pyrin domain-containing protein 3 (NLRP3) inflammasome, a key regulator of innate immunity, disrupts immune homeostasis disruption when aberrantly activated. In this study, we identified a berberine (BBR) derivative B4, which demonstrated markedly superior efficacy over the parent compound BBR in ameliorating dextran sulfate sodium (DSS)-induced UC mouse model, along with a favorable safety profile. Mechanistic studies revealed that B4 exerts its effects through dual targeting. B4 directly binds to heat shock protein 60 (HSP60), effectively reversing mitochondrial redox and metabolic imbalance, reducing reactive oxygen species (ROS) burst and HSP60 translocation, thereby suppressing NLRP3 activation at the source on one hand. Furthermore, B4 directly targets the NLRP3 protein to inhibit inflammasome assembly, which significantly reducing the secretion of interleukin-1beta (IL-1β) and interleukin-18 (IL-18), and suppressing pyroptosis. This dual-targeting synergy effectively interrupts the vicious positive feedback loop of “mitochondrial dysfunction-NLRP3 inflammasome activation-exacerbated inflammation-aggravated mitochondrial damage,” indicating that B4 is a promising therapeutic agent for UC. Moreover, this novel strategy of co-targeting the “mitochondria-NLRP3 inflammasome axis” provides a new direction for UC drug development.
