O-GlcNAcylation regulates the stability of transferrin receptor (TFRC) to control the ferroptosis in hepatocellular carcinoma cells.

Ferroptosis is an iron-dependent programmed cell death (PCD) enforced by lipid peroxidation accumulation. Transferrin receptor (TFRC), one of the signature proteins of ferroptosis, is abundantly expressed in hepatocellular carcinoma (HCC). However, post-translational modification (PTM) of TFRC and the underlying mechanisms for ferroptosis regulation remain less understood. In this study, we found that TFRC undergoes O-GlcNAcylation, influencing Erastin-induced ferroptosis sensitivity in hepatocytes. Further mechanistic studies found that Erastin can trigger de-O-GlcNAcylation of TFRC at serine 687 (Ser687), which diminishes the binding of ubiquitin E3 ligase membrane-associated RING-CH8 (MARCH8) and decreases polyubiquitination on lysine 665 (Lys665), thereby enhancing TFRC stability that favors labile iron accumulation. Therefore, our findings report O-GlcNAcylation on an important regulatory protein of ferroptosis and reveal an intriguing mechanism by which HCC ferroptosis is controlled by an iron metabolism pathway.

Drug repurposing-based nanoplatform via modulating autophagy to enhance chemo-phototherapy against colorectal cancer.

Multi-modal combination therapy is regarded as a promising approach to cancer treatment. Combining chemotherapy and phototherapy is an essential multi-modal combination therapy endeavor. Ivermectin (IVM) is a potent antiparasitic agent identified as having potential antitumor properties. However, the fact that it induces protective autophagy while killing tumor cells poses a challenge to its further application. IR780 iodide (IR780) is a near-infrared (NIR) dye with outstanding photothermal therapy (PTT) and photodynamic therapy (PDT) effects. However, the hydrophobicity, instability, and low tumor uptake of IR780 limit its clinical applications. Here, we have structurally modified IR780 with hydroxychloroquine, an autophagy inhibitor, to synthesize a novel compound H780. H780 and IVM can form H780-IVM nanoparticles (H-I NPs) via self-assembly. Using hyaluronic acid (HA) to modify the H-I NPs, a novel nano-delivery system HA/H780-IVM nanoparticles (HA/H-I NPs) was synthesized for chemotherapy-phototherapy of colorectal cancer (CRC). Under NIR laser irradiation, HA/H-I NPs effectively overcame the limitations of IR780 and IVM and exhibited potent cytotoxicity. In vitro and in vivo experiment results showed that HA/H-I NPs exhibited excellent anti-CRC effects. Therefore, our study provides a novel strategy for CRC treatment that could enhance chemo-phototherapy by modulating autophagy.

Mitochondrial-Targeted CS@KET/P780 Nanoplatform for Site-Specific Delivery and High-Efficiency Cancer Immunotherapy in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is a form of malignancy with limited curative options available. To improve therapeutic outcomes, it is imperative to develop novel, potent therapeutic modalities. Ketoconazole (KET) has shown excellent therapeutic efficacy against HCC by eliciting apoptosis. However, its limited water solubility hampers its application in clinical treatment. Herein, a mitochondria-targeted chemo-photodynamic nanoplatform, CS@KET/P780 NPs, is designed using a nanoprecipitation strategy by integrating a newly synthesized mitochondria-targeted photosensitizer (P780) and chemotherapeutic agent KET coated with chondroitin sulfate (CS) to amplify HCC therapy. In this nanoplatform, CS confers tumor-targeted and subsequently pH-responsive drug delivery behavior by binding to glycoprotein CD44, leading to the release of P780 and KET. Mechanistically, following laser irradiation, P780 targets and destroys mitochondrial integrity, thus inducing apoptosis through the enhancement of reactive oxygen species (ROS) buildup. Meanwhile, KET-induced apoptosis synergistically enhances the anticancer effect of P780. In addition, tumor cells undergoing apoptosis can trigger immunogenic cell death (ICD) and a longer-term antitumor response by releasing tumor-associated antigens (TAAs) and damage-associated molecular patterns (DAMPs), which together contribute to improved therapeutic outcomes in HCC. Taken together, CS@KET/P780 NPs improve the bioavailability of KET and exhibit excellent therapeutic efficacy against HCC by exerting chemophototherapy and antitumor immunity.

Identification of glycogene-based prognostic signature and validation of B3GNT7 as a potential biomarker and therapeutic target in breast cancer.

BACKGROUND: Breast cancer is the most common cancer worldwide, with the fifth highest mortality rate among all cancers and high risk of metastasis. However, potential biomarkers and molecular mechanisms underlying the stratification of breast cancer in terms of clinical outcomes remain to be investigated. Therefore, we aimed to find a novel prognostic biomarker and therapeutic target for breast cancer patients. METHODS: Unsupervised hierarchical clustering was used to perform comprehensive transcriptomic study of total 185 glycogenes in public datasets of breast cancer with clinicopathological and survival information. A glycogene-based signature for subtype classification was discovered using Limma packages, and relevance to four known molecular features was identified by GSVA. Experimental verification was performed and biological functions of B3GNT7 were characterized by quantitative RT-PCR, western blot, transwell assays, and lectin immunofluorescence staining in breast cancer cells. RESULTS: A 23-glycogene signature was identified for the classification of breast cancer. Among the 23 glycogenes, B3GNTs showed significantly positive associations with ER-/Her2- subtype in breast cancer patients (n = 2655). Overexpressed B3GNT7 were correlated with poor prognosis in breast cancer patients based on public datasets. B3GNT7 depletion inhibited cell proliferation, migration, and invasion, and decreased global fucosylation in MDA-MB-231 and HCC1937 breast cancer cells. CONCLUSIONS: Herein, we discovered a unique 23-gene signature for breast cancer patient glycogene-type classification. Among these genes, B3GNT7 was shown to be a potential biomarker for unfavorable outcomes and therapeutic target of breast cancer.

G-Quadruplex Structures as a "Switch" Regulate ATF4 Expression in Ferroptotic HepG2 Cells.

G-quadruplex (G4) is a noncanonical structure folded in a widespread manner by guanine-rich tandem repeated sequences. As a key response factor, activating transcription factor 4 (ATF4) has dual functions in managing iron-dependent ferroptosis by regulating amino acid synthesis and antioxidant-related gene expression. In our study, the activity of ATF4 expression was elevated in HepG2 cells induced by erastin. Based on preliminary bioinformatics analyses, the G-tract region, named WT, had high potential to form G4, and it was found that PDS could markedly weaken the increase of ATF4 expression by reducing the sensitivity of HepG2 cells toward erastin. In circular dichroism spectra, WT oligonucleotides showed characteristic molar ellipticity at specific wavelengths of parallel G4 structures, while corresponding single-base mutants possessed a weaker ability to form G4, which were consistent with immunostaining results. In addition, endogenous G4 formed by the WT motif was significantly destroyed in HepG2 cells treated with erastin. After being transfected with WT oligonucleotides, the levels of ATF4 mRNA decreased significantly regardless of being treated with erastin or not. Meanwhile, mutations of G-tracts could advantageously impact the luciferase expression downstream of an ATF4 promoter in reporter assays, manifesting that the decrease of endogenous G4 in the ATF4 promoter was positively associated with the expression enhanced by erastin in HepG2 cells.

Research Progress of Ferroptosis: A Bibliometrics and Visual Analysis Study.

Background: Ferroptosis is a type of cell death with major topic of debate under current research and plays an important role in disease regulation. Objective: In this study, the literature management software Bibexcel and knowledge graph tool VOSviewer were used to summarize and analyze the international research trends and hotspots about ferroptosis in recent years, which highlight the disease mechanism, diagnosis, and treatment related to ferroptosis. Material/Methods. The core collection database of Web of Science was used for retrieving ferroptosis research literature. The information such as the amount of text, the country, the period, the institution, the fund, and the keywords was extracted by the bibliometric tool Bibexcel. The cooccurrence and clustering function of VOSviewer were used to analyze the high-frequency keywords and the cooperative network of the author, institution, and country. Results: The research of ferroptosis started late and was formally proposed in 2012. It has developed rapidly and presented an "exponential" growth trend. China, the United States, Germany, Japan, and France are the main national forces of ferroptosis research development. The United States and China have a relatively high degree of support and attention to ferroptosis. Exploring oxidative stress, inducers/inhibitors, synergistic antitumor effect, relationships with other cell death types, GSH/GPX4 and iron metabolism imbalance related mechanisms of ferroptosis, and ferroptosis in the nervous system disease, ischemia-reperfusion injury, tumor, inflammation, and age-related diseases are the hot research directions. Conclusion: Ferroptosis has been a research hotspot in the field of biomedicine in recent years and has attracted the attention of scholars all over the world. The occurrence mechanism of ferroptosis and its application in neurological diseases, ischemia and reperfusion injury, tumors, inflammation, and aging are the hot directions of current research. In the future, ferroptosis can be appropriately considered for strengthening new approaches, new diseases, new inductors, new inhibitors, clinical transformation, and traditional medicine research.

Identification of Potential Inhibitors Against the TGF-ß/BMPs-Activin Receptor- like Kinase 1 Signal Pathway.

INTRODUCTION: In many diseased states, especially fibrosis and cancer, TGF-ß family members are overexpressed and the outcome of signaling is diverted toward disease progression. As the result of activin receptor-like kinase 1 (ALK1) plays a key role in TGF-ß signaling, discovering inhibitors of ALK1 to block TGF-ß signaling for a therapeutic benefit has become an effective strategy. METHODS: In this work, ZINC15894217 and ZINC12404282 were identified as potential ALK1 inhibitors using molecular docking, molecular dynamics simulation and MM/PBSA calculations studies. The analysis of energy decomposition found that Val208, Val216, Lys229, Gly283, Arg334 and Leu337 acted as crucial residues for ligand binding and system stabilizing. RESULTS: In addition, these compounds displayed excellent pharmacological and structural properties, which can be further evaluated through in vitro and in vivo experiments for the inhibition of ALK1 to be developed as drugs against fibrosis and tumor. CONCLUSION: Overall, our study illustrated a time- and cost-effective computer aided drug design procedure to identify potential ALK1 inhibitors. It would provide useful information for further development of ALK1 inhibitors to improve disease related to TGF-ß signal pathway.

Computer-aided screening for suppressor of variegation 4-20 homolog 1 inhibitors and their preliminary activity validation in human osteosarcoma.

Histone methylation/demethylation facilitate to maintain balanced histone methylation levels and underpin gene regulation, playing the key roles in epigenetic regulation. Suppressor of variegation 4-20 homolog 1 (SUV420H1), a member of class Histone Lysine Methyltransferase and a key enzyme in the epigenetic regulation of the pathways controlling metabolism and tumorigenesis, is crucial to maintain cell homeostasis. The inhibition of SUV420H1 has emerged as a promising candidate for drug development and cancer therapy. Herein, two potential and potent SUV420H1 inhibitors (ZINC08398384, ZINC08439608) were identified through in silico approach and in vitro biological experiments. In vitro biological tests demonstrated that these compounds can inhibit the proliferation of U2OS cells and restrict its migration ability. And the level of dimethylation of lysine 20 on histone H4 (H4K20me2) was markedly decreased by these compounds-treatment in a dose-dependent manner. These results indicated that ZINC08398384 and ZINC08439608 are potential SUV420H1 inhibitors and could be developed as promising drug candidates applied to cancer epigenetic therapy.Communicated by Ramaswamy H. Sarma.

A novel mannose-binding lectin from Liparis nervosa with anti-fungal and anti-tumor activities.

Plant lectins are carbohydrate-binding proteins with nonimmune origin, which can reversibly bind with carbohydrates, agglutinate cells, and precipitate polysaccharides and glycoconjugates. Plant lectins have attracted much attention for their anti-virus, anti-proliferation, and pro-apoptosis properties. Thus the exploration of new lectins has received special attention. Here we purified a mannose-binding lectin from the rhizomes of Liparis nervosa by ion exchange chromatography on DEAE-Sepharose, affinity chromatography on Mannose-Sepharose 4B, and gel filtration chromatography on Sephacryl S-100. The purified L. nervosa lectin (LNL) was identified to be a monomeric protein with a molecular mass of 13 kDa. LNL exhibited hemagglutinating activity towards rabbit erythrocytes, and its activity could be strongly inhibited by D-mannose, N-acetyl glucosamine and thyroglobulin. In vitro experiments showed that LNL exhibited a comparable anti-fungal activity against Piricularia oryzae (Cavara), Bipolaris maydis, Fusarium graminearum, and Sclerotium rolfsii, and anti-proliferation activity against tumor cells by inducing apoptosis. The full-length cDNA sequence of LNL is 715 bp in length and contains a 525 bp open reading frame (ORF) encoding a 110-residue mature protein. It was predicted to have three mannose-binding conserved motifs 'QXDXNXVXY'. The binding pattern of LNL was further revealed by homology modeling and molecular docking. We demonstrated that LNL is not only a potential therapeutic candidate against tumor but also a new anti-fungal agent.

The Prospects of Therapeutic Potential and Drug Development Targeting Autophagy in Cancer.

Autophagy is a self-protection mechanism of cells. Cells can degrade damaged organelles and macromolecules in this way to guarantee the growth and development of cells. In recent years, more and more researches have found that autophagy also plays a certain role in the occurrence and development of tumors. The dual role of autophagy in the development of tumors includes inhibiting the development of tumors; meanwhile, under the condition of insufficient nutrition, autophagy degrades organelles to reduce oxidative stress and provide nutrition and energy for tumor cells so as to protect tumor cells. The regulation of autophagy depends on the development of the tumor, and the corresponding autophagy inducers or inhibitors are constantly emerging, which provides a new direction for tumor treatment.

Drug Development and Treatment of Autophagy in Other Diseases.

In addition to tumors and aging that are associated with autophagy, many other diseases are also regulated by autophagy, including liver disease, myopathy, immune pathogen infection, cardiovascular disease, and so on. This chapter will detail the relationship between autophagy and these diseases and their underlying molecular mechanisms. We summarized the current research status of autophagy as a target for the treatment of related diseases, and prospected the development of related drugs and therapeutic strategies. We hope to provide new ideas for finding new therapeutic targets through the autophagic signaling pathways.

Systems Biology Approaches in Autophagy Research.

As a classical form of programmed cell death, autophagy is widely involved in cellular metabolism and vital for the maintenance of homeostasis in physiological and pathological states. With multiple levels of regulation and signaling integrated in, autography presents complicated relevance with various diseases, such as cancer and neurological diseases. The emerging subject, systems biology, along with multi-omics approaches, offers a new strategy to investigate these interactive processes from a holistic perspective. In this chapter, we focus on the systems biology method for autophagy research and introduce essential research skills and procedures. The critical step of systematic study is to explore interplay between biological molecules based on massive biological data, which requires construction of networks in different biological levels, modification, and identification of key pathways and targets via optimized algorithm and experimental verification. Guided by systems biology research, drug design can thus be strengthened by efficient screening and accurate evaluation. Overall, systems biology promises to act as a powerful tool which both helps to clarify the profound mechanism and to develop efficacious medicine.

FerrDb: a manually curated resource for regulators and markers of ferroptosis and ferroptosis-disease associations.

Ferroptosis is a mode of regulated cell death that depends on iron. Cells die from the toxic accumulation of lipid reactive oxygen species. Ferroptosis is tightly linked to a variety of human diseases, such as cancers and degenerative diseases. The ferroptotic process is complicated and consists of a wide range of metabolites and biomolecules. Although great progress has been achieved, the mechanism of ferroptosis remains enigmatic. We have currently entered an era of extensive knowledge advancement, and thus, it is important to find ways to organize and utilize data efficiently. We have observed a high-quality knowledge base of ferroptosis research is lacking. In this study, we downloaded 784 ferroptosis articles from the PubMed database. Ferroptosis regulators and markers and associated diseases were extracted from these articles and annotated. In summary, 253 regulators (including 108 drivers, 69 suppressors, 35 inducers and 41 inhibitors), 111 markers and 95 ferroptosis-disease associations were found. We then developed FerrDb, the first manually curated database for regulators and markers of ferroptosis and ferroptosis-disease associations. The database has a user-friendly interface, and it will be updated every 6 months to offer long-term service. FerrDb is expected to help researchers acquire insights into ferroptosis.Database URL: http://www.zhounan.org/ferrdb.

Multifunctional gold nanoparticles as smart nanovehicles with enhanced tumour-targeting abilities for intracellular pH mapping and in vivo MR/fluorescence imaging.

A number of multimodal agents have been developed for tumour imaging and diagnosis, but most of them cannot be used to study the detailed physiological or pathological changes in living cells at the same time. Herein, a series of pH-responsive magnetic resonance and fluorescence imaging (MRI/FI) dual-modal "nanovehicles" are developed and tested. These new dual-modal materials allow for intercellular pH sensing, and those with units that are dually sensitive towards both acidic and basic environments have the ability for intracellular pH mapping and can be used to quantify pH at the cellular level. In addition, detailed pH changes in organelles (including lysosomes and mitochondria) can be investigated at the same time. On the other hand, with the tumour-targeting peptide (cRGD)-modified dual-modal nanovehicles, in vivo tumour MR and fluorescence imaging, which is suitable for cancer diagnosis, can be achieved. Moreover, it has been proved that these materials can pass through the blood brain barrier (BBB). By combining the above mentioned promising properties, these novel multifunctional "nanovehicles" may provide a new method for studying the role of pH during cancer diagnosis and treatment.

Metabolic gene NR4A1 as a potential therapeutic target for non-smoking female non-small cell lung cancer patients.

BACKGROUND: Although cigarette smoking is considered one of the key risk factors for lung cancer, 15% of male patients and 53% of female patients with lung cancer are non-smokers. Metabolic changes are critical features of cancer. Therapeutic target identification from a metabolic perspective in non-small cell lung cancer (NSCLC) tissue of female non-smokers has long been ignored. RESULTS: Based on microarray data retrieved from Affymetrix expression arrays E-GEOD-19804, we found that the downregulated genes in non-smoking female NSCLC patients tended to participate in protein/amino acid and lipid metabolism, while upregulated genes were more involved in protein/amino acid and carbohydrate metabolism. Combining nutrient metabolic co-expression, protein-protein interaction network construction and overall survival assessment, we identified NR4A1 and TIE1 as potential therapeutic targets for NSCLC in female non-smokers. To accelerate the drug development for non-smoking female NSCLC patients, we identified nilotinib as a potential agonist targeting NR4A1 encoded protein by molecular docking and molecular dynamic stimulation. We also show that nilotinib inhibited proliferation and induced senescence of cells in non-smoking female NSCLC patients in vitro. CONCLUSIONS: These results not only uncover nutrient metabolic characteristics in non-smoking female NSCLC patients, but also provide a new paradigm for identifying new targets and drugs for novel therapy for such patients.

Preclinical assessment of histone deacetylase inhibitor quisinostat as a therapeutic agent against esophageal squamous cell carcinoma.

Esophageal squamous cell carcinoma (ESCC) is one of the most serious life-threatening malignancies. Although chemotherapeutic targets and agents for ESCC have made much progress recently, the efficacy is still unsatisfactory. Therefore, there is still an unmet medical need for patients with ESCC. Here, we report the expression status of HDAC1 in human ESCC and matched paracancerous tissues, and the results indicated that HDAC1 was generally upregulated in ESCC specimens. Furthermore, we comprehensively assessed the anti-ESCC activity of a highly active HDAC1 inhibitor quisinostat. Quisinostat could effectively suppress cellular viability and proliferation of ESCC cells, as well as induce cell cycle arrest and apoptosis even at low treatment concentrations. The effectiveness was also observed in KYSE150 xenograft model when quisinostat was administered at tolerated doses (3 mg/kg and 10 mg/kg). Meanwhile, quisinostat also had the ability to suppress the migration and invasion (pivotal steps of tumor metastasis) of ESCC cells. Western blot analysis indicated that quisinostat exerted its anti-ESCC effects mainly through blockade of Akt/mTOR and MAPK/ERK signaling cascades. Overall, HDAC1 may serve as a potential therapeutic target for ESCC, and quisinostat deserves to be further assessed as a promising drug candidate for the treatment of ESCC.

Screening of novel histone deacetylase 7 inhibitors through molecular docking followed by a combination of molecular dynamics simulations and ligand-based approach.

Histone acetylation/deacetylation is a key mechanism for transcription regulation which plays an important role in control of gene expression, tissue growth, and development. In particular, histone deacetylase 7 (HDAC7), a member of class IIa HDACs, is crucial to maintain cell homeostasis, and HDAC7 has emerged as a new target for cancer therapy. In this study, molecular docking was applied to screen candidate inhibitors and 21 compounds were found. Following the 50 ns molecular dynamics simulations and binding free energy calculation, ZINC00156160, ZINC01703144, ZINC04293665, and ZINC13900201 were identified as potential HDAC7 inhibitors, which would provide a sound starting point for further studies involving molecular modeling coupled with biochemical experiments. Meanwhile, similarity computation and substructure search were combined, and then we found that compounds sharing common backbone "CC(=O)N[C@@H](CSc1ccccc1)C(=O)O" could be efficient to inhibit the bioactivity of HDAC7. Then comparative molecular similarity indices analysis (CoMSIA) techniques were implemented to investigate the relationship between properties of the substituent group and bioactivities of small molecules. The CoMSIA model exhibited powerful predictivity, with satisfactory statistical parameters such as q2 of 0.659, R2 of 0.952, and F of 268.448. Contour maps of the CoMSIA model gave insight into the feature requirements of the common backbone for the HDAC7 inhibitory activity. Finally, details of designing novel HDAC7 inhibitors were confirmed by a combination of receptor-based docking and ligand-based structure-activity relationship. Communicated by Ramaswamy H. Sarma.

Computer-aided identification of a novel pyruvate kinase M2 activator compound.

OBJECTIVES: The aim of this study was to obtain antitumour molecules targeting to activate PKM2 through adequate computational methods combined with biological activity experiments. METHODS: The structure-based virtual screening was utilized to screen effective activator targeting PKM2 from ZINC database. Molecular dynamics simulations were performed to evaluate the stability of the small molecule-binding PKM2 complex systems. Then, cell survival experiments, glutaraldehyde crosslinking reaction, western blot, and qPCR experiments were used to detect the effects of top hits on various cancer cells and the targeting specificity of PKM2. RESULTS: Two small molecules in 1,5-2H-pyrrole-dione were obtained after virtual screening. In vitro experiments demonstrated that ZINC08383544 specifically activated PKM2 and affected the expression of upstream and downstream genes of PKM2 during glycolysis, leading to the inhibition of tumour cell growth. These results indicate that ZINC08383544 conforms to the characteristics of PKM2 activator and is potential to be a novel PKM2 activator as antitumour drug. DISCUSSION: This work proves that ZINC08383544 promotes the formation of PKM2 tetramer, effectively blocks PKM2 nuclear translocation, and inhibits the growth of tumour, and ZINC08383544 may be a novel activator of PKM2. This work may provide a good choice of drug or molecular fragments for the antitumour strategy targeting PKM2. Screening of targeted drugs by combination of virtual screening and bioactivity experiments is a rapid method for drug discovery.

Structure and energy based quantitative missense variant effect analysis provides insights into drug resistance mechanisms of anaplastic lymphoma kinase mutations.

Anaplastic lymphoma kinase (ALK) is considered as a validated molecular target in multiple malignancies, such as non-small cell lung cancer (NSCLC). However, the effectiveness of molecularly targeted therapies using ALK inhibitors is almost universally limited by drug resistance. Drug resistance to molecularly targeted therapies has now become a major obstacle to effective cancer treatment and personalized medicine. It is of particular importance to provide an improved understanding on the mechanisms of resistance of ALK inhibitors, thus rational new therapeutic strategies can be developed to combat resistance. We used state-of-the-art computational approaches to systematically explore the mutational effects of ALK mutations on drug resistance properties. We found the activation of ALK was increased by substitution with destabilizing mutations, creating the capacity to confer drug resistance to inhibitors. In addition, results implied that evolutionary constraints might affect the drug resistance properties. Moreover, an extensive profile of drugs against ALK mutations was constructed to give better understanding of the mechanism of drug resistance based on structural transitions and energetic variation. Our work hopes to provide an up-to-date mechanistic framework for understanding the mechanisms of drug resistance induced by ALK mutations, thus tailor treatment decisions after the emergence of resistance in ALK-dependent diseases.

Identification of two potential glycogen synthase kinase 3ß inhibitors for the treatment of osteosarcoma.

Osteosarcoma is the most common primary malignant bone tumor among adolescents worldwide with high mortality rate. Glycogen synthase kinase 3ß (GSK3ß) is a serine/threonine kinase and is considered as a validated target in osteosarcoma therapy. Therefore, the study of GSK3ß inhibitors is one of the most popular fields in anti-osteosarcoma drug development. Here, the tools of bioinformatics were used to screen novel effective inhibitors of GSK3ß from ZINC Drug Database. The molecular docking, molecular dynamic simulations, MM/GBSA, and energy decomposition analysis were performed to identify the inhibitors. Finally, ZINC08383479 and ZINC08441251 were selected as potential GSK3ß inhibitors. These two inhibitors were evaluated by GSK3ß kinase inhibition assay in vitro. The inhibition of cell proliferation was tested in osteosarcoma cell lines U2OS and MG63 in vitro. The result showed that ZINC08383479 and ZINC08441251 had high inhibition activity against GSK3ß. We found that CHIR99021 (a known GSK3ß inhibitor), ZINC08383479, and ZINC08441251 had significant inhibition activity in U2OS cells and MG63 cells. These findings may provide new ideas for the design of more potent GSK3ß inhibitors and therapeutic targets for osteosarcoma.