RESUMO
Background: This study examined the effect of calcium-sensing receptor (CaSR) antagonism on human osteosarcoma cells and investigated the underlying molecular mechanisms of this effect through transcriptome sequencing. Methods: Human osteosarcoma cell lines MG-63 and Saos-2 were treated with different concentrations (0.1-10 µM) of the CaSR antagonist NPS-2143. Cell Counting Kit-8 (CCK-8) assays were used to detect the effect of CaSR antagonism on the viability of the cells. RNA sequencing was performed on cells treated with five µM NPS-2143 for 24 hours, followed by bioinformatic analysis to identify differentially expressed genes and enriched pathways. qRT-PCR was conducted to validate key genes. Results: CCK-8 assays showed that at low concentrations (0.1 and one µM), NPS-2143 had no significant effect on MG-63 and Saos-2 cell viability. At higher concentrations (five µM and 10 µM), the viability of MG-63 and Saos-2 cells was significantly reduced. Five µM was therefore selected for subsequent experiments. RNA sequencing revealed distinct gene expression profiles in NPS-2143-treated cells compared to controls. A total of 927 differentially expressed genes (DEGs) were identified in Saos-2 cells (378 upregulated, 549 downregulated), and 59 DEGs were identified in MG-63 cells (33 upregulated, 26 downregulated). Reactome and KEGG pathway enrichment analyses indicated significant enrichment of cholesterol and steroid biosynthesis-related pathways. Transcriptome sequencing showed that NPS-2143 modulated the expression of genes in cholesterol and steroid synthesis pathways. Real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) confirmed that NPS-2143 promoted the expression of the cholesterol and steroid synthesis pathway genes, CYP51A1, DHCR24, LSS, and MSMO1 in MG-63 and Saos-2 cells. Discussion: The inhibitory effect of NPS-2143 on MG-63 and Saos-2 osteosarcoma tumor cell viability was confirmed. CaSR antagonism significantly up-regulated genes involved in cholesterol and steroid biosynthesis, including CYP51A1, DHCR24, LSS, and MSMO1. These genes encode key enzymes in the cholesterol synthesis pathway, and their upregulation may lead to cholesterol overproduction. This may, in turn, lead to the formation of oxysterols, which are known to induce inflammation and cytotoxicity. These findings suggest a potential metabolic mechanism through which CaSR antagonists influence osteosarcoma cell viability. Although further validation is warranted, our results provide preliminary evidence implicating cholesterol biosynthesis as a mechanistic target in osteosarcoma and underscore the exploratory value of CaSR antagonists as metabolic regulators in cancer research.
Assuntos
Neoplasias Ósseas , Colesterol , Osteossarcoma , Receptores de Detecção de Cálcio , Esteroides , Humanos , Osteossarcoma/metabolismo , Osteossarcoma/tratamento farmacológico , Osteossarcoma/patologia , Osteossarcoma/genética , Receptores de Detecção de Cálcio/antagonistas & inibidores , Sobrevivência Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Colesterol/biossíntese , Colesterol/metabolismo , Esteroides/biossíntese , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/patologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , NaftalenosRESUMO
This study investigates the expression of Yes-associated protein (YAP) and androgen receptor (AR) in osteosarcoma and assess their prognostic significance. A retrospective cohort of 100 osteosarcoma patients and 30 adjacent normal tissues was analyzed by immunohistochemistry. Expression levels were correlated with clinicopathological features and progression-free survival. High expression of YAP (65%) and AR (60%) was significantly more frequent in osteosarcoma than in normal tissues. Both markers were associated with advanced stage and distant metastasis, and their co-overexpression predicted the shortest median progression-free survival (9 months). Interaction analysis confirmed a synergistic effect of YAP and AR on poor prognosis. YAP and AR are frequently co-overexpressed in osteosarcoma and jointly contribute to tumor aggressiveness and unfavorable outcomes. Their combined evaluation may serve as a novel prognostic indicator and potential therapeutic target.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Neoplasias Ósseas , Osteossarcoma , Receptores Androgênicos , Fatores de Transcrição , Humanos , Osteossarcoma/patologia , Osteossarcoma/metabolismo , Osteossarcoma/mortalidade , Receptores Androgênicos/metabolismo , Estudos Retrospectivos , Masculino , Feminino , Proteínas de Sinalização YAP , Neoplasias Ósseas/patologia , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/mortalidade , Adulto , Adolescente , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Prognóstico , Adulto Jovem , Biomarcadores Tumorais/metabolismo , Criança , Pessoa de Meia-Idade , Imuno-Histoquímica , Relevância ClínicaRESUMO
OBJECTIVE: Osteosarcoma (OS), the most common primary malignant bone tumor, exhibits poor prognosis, underscoring the need for new therapies. Juglone, a natural naphthoquinone, shows therapeutic promise; however, the effects of juglone on OS and underlying mechanisms remain to be elucidated. MATERIALS AND METHODS: In vitro experiments were validated using U2OS and HOS cell lines. Network pharmacology predicted juglone-OS targets, followed by protein-protein interaction (PPI) network construction and GO/KEGG enrichment analysis. Transcriptomics analysis was further confirmed. Molecular docking was performed using AutoDock Tools and Discovery Studio. Molecular dynamics simulations (MD) using GROMACS confirmed the stability of the juglone-target complex. In vivo experiments clarified the effect of juglone in vivo. ADME/T was profiled using Swiss and Protox. RESULTS: Juglone inhibited OS cell viability, promoted apoptosis, suppressed proliferation and migration, and induced G2/M arrest. A total of 234 targets were identified; PPI highlighted key interactors (TP53, AKT1, BCL2, CTNNB1, STAT3). Enrichment analyses revealed associations with oxidative stress and PI3K/AKT pathways. Molecular docking showed high-affinity binding to PI3K and AKT1, confirmed stable by molecular dynamics. Mechanistically, juglone inhibits OS via the ROS/PI3K/AKT pathway. In vivo, it potently inhibited tumor growth in xenograft models with short-term biosafety. ADME/T profiling indicated favorable pharmacokinetics but potential toxicity risks. CONCLUSION: This study first demonstrates juglone's anti-OS efficacy via the ROS/PI3K/AKT pathway. In vivo studies confirm potent tumor suppression with short-term safety, supporting its clinical promise for OS treatment.
Assuntos
Antineoplásicos , Neoplasias Ósseas , Naftoquinonas , Osteossarcoma , Fosfatidilinositol 3-Quinases , Proteínas Proto-Oncogênicas c-akt , Espécies Reativas de Oxigênio , Transcriptoma , Naftoquinonas/farmacologia , Naftoquinonas/uso terapêutico , Humanos , Osteossarcoma/tratamento farmacológico , Osteossarcoma/patologia , Osteossarcoma/genética , Osteossarcoma/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Farmacologia em Rede , Espécies Reativas de Oxigênio/metabolismo , Animais , Linhagem Celular Tumoral , Simulação de Acoplamento Molecular , Transdução de Sinais/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/metabolismo , Proliferação de Células/efeitos dos fármacos , Camundongos , Apoptose/efeitos dos fármacos , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/patologia , Neoplasias Ósseas/genética , Neoplasias Ósseas/metabolismo , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Ensaios Antitumorais Modelo de Xenoenxerto , Sobrevivência Celular/efeitos dos fármacos , Transcriptoma/efeitos dos fármacos , Simulação de Dinâmica Molecular , Movimento Celular/efeitos dos fármacos , Mapas de Interação de Proteínas/efeitos dos fármacosRESUMO
Osteosarcoma is the most common primary malignant tumour of bone. It is highly malignant and invasive, posing a serious threat to human health. Neoadjuvant chemotherapy with cisplatin and doxorubicin combined with surgical resection is the standard treatment option, which can improve the 5-year survival rate of patients to approximately 60%. However, chemotherapy resistance, postoperative distant metastasis, and adverse reactions are key bottlenecks leading to poor prognosis, and new precision treatment strategies are urgently needed. SLC7A11, a core functional subunit of the cystine-glutamate antiporter System Xc-, is a key regulator of iron homeostasis and is closely associated with the infiltration and function of various immune cells in the tumour immune microenvironment, thereby regulating osteosarcoma progression and the antitumor immune response. Targeting SLC7A11 to synchronously regulate iron homeostasis and remodel the immune microenvironment is expected to improve osteosarcoma treatment. To systematically assess the regulatory mechanism and clinical application potential of SLC7A11, we retrieved the relevant literature from PubMed, Web of Science, Elsevier ScienceDirect, and other databases from April 2001 to July 2025. We screened and integrated high-quality studies related to SLC7A11, iron death, and the immune microenvironment of osteosarcoma. This review focuses on the role of SLC7A11 in iron death-related pathways (such as the GPX4-GSH, FSP1-CoQ10, and GCH1-BH4 pathways) and immune cells in the tumour immune microenvironment (such as TAMs, tans, and T cells), commenting on the molecular mechanisms of osteosarcoma development and summarising the regulatory effects of inhibitors targeting SLC7A11 (including small molecule compounds, active ingredients of traditional Chinese medicine, non-coding RNAs, exosomes, and nanomaterials) on osteosarcoma. The SLC7A11-targeting strategies summarised in this review provide a solid theoretical basis for clinical transformation, such as optimising combination chemotherapy regimens, developing personalised immunotherapy regimens, and constructing targeted drug delivery systems, which are expected to effectively improve the efficacy and prognosis of patients with osteosarcoma.
Assuntos
Sistema y+ de Transporte de Aminoácidos , Neoplasias Ósseas , Ferroptose , Osteossarcoma , Microambiente Tumoral , Humanos , Ferroptose/imunologia , Microambiente Tumoral/imunologia , Osteossarcoma/imunologia , Osteossarcoma/tratamento farmacológico , Osteossarcoma/metabolismo , Sistema y+ de Transporte de Aminoácidos/metabolismo , Sistema y+ de Transporte de Aminoácidos/imunologia , Neoplasias Ósseas/imunologia , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/metabolismo , AnimaisRESUMO
BACKGROUND: This study aimed to evaluate the diagnostic value of special AT-rich sequence binding protein 2 (SATB2) in distinguishing between osteogenic tumors and non-osteogenic tumors, providing reliable scientific evidence for its use as an adjunct diagnostic tool in clinical practice. METHODS: We conducted systematic searches of the PubMed, EMBASE, Cochrane Library, and Web of Science databases to identify all relevant literature published up to June 2025 that studied SATB2 in the differential diagnosis between osteogenic and non-osteogenic tumors. The QUADAS-2 tool was used to evaluate the methodological quality of each included study. Meta-analysis was performed using STATA SE-64 and RevMan 5.4 software. RESULTS: 10 studies involving a total of 1234 cases were included, comprising 494 patients with osteogenic tumors and 740 patients with non-osteogenic tumors. The pooled sensitivity and specificity of SATB2 for differentiating osteogenic from non-osteogenic tumors were 0.97 (95 % CI: 0.90-0.99) and 0.88 (95 % CI: 0.73-0.96), respectively. The PLR was 8.17 (95 % CI: 3.30-20.23), and the NLR was 0.03 (95 % CI: 0.01-0.12). The DOR was 252.82 (95 % CI: 41.85-1527.16). The AUC was 0.98 (95 % CI: 0.97-0.99). CONCLUSION: SATB2 demonstrates high sensitivity and robust specificity as an adjunct diagnostic marker for differentiating osteogenic from non-osteogenic tumors. However, it should be noted that this study excluded tumors with ambiguous definitions, such as giant cell tumors of bone. Future research should further validate the clinical utility of SATB2 in these and other challenging lesions.
Assuntos
Neoplasias Ósseas , Imuno-Histoquímica , Proteínas de Ligação à Região de Interação com a Matriz , Osteogênese , Osteossarcoma , Fatores de Transcrição , Humanos , Proteínas de Ligação à Região de Interação com a Matriz/metabolismo , Neoplasias Ósseas/diagnóstico , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/patologia , Diagnóstico Diferencial , Fatores de Transcrição/metabolismo , Osteossarcoma/diagnóstico , Osteossarcoma/metabolismo , Biomarcadores Tumorais/metabolismoRESUMO
Osteosarcoma, a prevalent primary malignant bone tumor, primarily affects adolescents and young adults. Current treatment strategies involve a combination of surgical intervention and chemotherapy. However, the effectiveness of chemotherapy is constrained by considerable challenges, such as drug resistance and insensitivity. Ferroptosis, a form of programmed cell death that is distinct from apoptosis, presents a promising alternative target for cancer therapy. Ferroptosis is characterized by irondependent lipid peroxidation, producing reactive oxygen species (ROS) and suppressing glutathione peroxidase 4 (GPX4). Notably, ferroptosis circumvents the conventional mechanisms associated with apoptosis. Inducing ferroptosis in cancer cells may help overcome drug resistance and enhance the effectiveness of existing treatments, including chemotherapy, radiotherapy and immunotherapy. Acetylshikonin is a derivative of naphthoquinone that possesses anticancer properties. However, the effects of acetylshikonin on the treatment of osteosarcoma and the mechanisms by which it induces cancer cell death remain unclear. The present study aimed to investigate the potential of acetylshikonin to induce apoptosis in osteosarcoma cells. Using cell viability assays, ROS detection, mitochondrial membrane potential analysis and ferroptosis inhibitor rescue experiments, the results demonstrated that acetylshikonin significantly reduced the viability of osteosarcoma cell lines while exhibiting low toxicity to normal cells. Mechanistically, acetylshikonin induced the production of ROS, disrupted the mitochondrial membrane potential and promoted lipid peroxidation, ultimately leading to ferroptosis. Additionally, treatment with acetylshikonin led to decreased levels of GPX4 and increased intracellular ferrous ion (Fe²+) concentrations, further supporting its role in the induction of ferroptosis. In conclusion, the current study emphasized the potential of acetylshikonin as an effective agent in inducing ferroptosis in osteosarcoma cells. Acetylshikonin reduced osteosarcoma cell viability and selectively promoted ferroptosis by increasing ROS production, disrupting mitochondrial function and enhancing lipid peroxidation. Furthermore, its ability to downregulate GPX4 and increase intracellular Fe2+ levels indicated its role in triggering ferroptosis. These findings suggest that acetylshikonin may be a valuable therapeutic candidate for the treatment of osteosarcoma, potentially improving outcomes and addressing the limitations of current therapies.
Assuntos
Antraquinonas , Neoplasias Ósseas , Ferroptose , Peroxidação de Lipídeos , Osteossarcoma , Ferroptose/efeitos dos fármacos , Peroxidação de Lipídeos/efeitos dos fármacos , Humanos , Osteossarcoma/metabolismo , Osteossarcoma/patologia , Osteossarcoma/tratamento farmacológico , Linhagem Celular Tumoral , Espécies Reativas de Oxigênio/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Fosfolipídeo Hidroperóxido Glutationa Peroxidase/metabolismo , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/patologia , Neoplasias Ósseas/tratamento farmacológico , Antraquinonas/farmacologia , Apoptose/efeitos dos fármacosRESUMO
Osteosarcoma (OS) is a rare malignant tumor and has the second-highest mortality rate of malignant tumors in children. Due to its unclear pathogenesis and therapeutic targets, there has been no significant progress in the targeted therapy of OS in the past 50 years. Therefore, it is critically important to develop new drug targets for OS. In this study, a covalent molecule library consisting of 61 small molecules was constructed based on our previous research. Phenotypic screening revealed that small molecule 8e effectively inhibited the proliferation of OS 143B cells, with an IC50 value of 0.73 µM. Compound 8e also showed good antitumor effects and low toxicity in a xenograft model (30.1 % inhibition of OS growth in BALB/c nude mice). Using compound 9a as an efficient activity-based protein profiling (ABPP) probe, eukaryotic protein elongation factor 1 alpha 2 (EEF1A2) was then enriched and conveniently identified as a potential target. The potential target was validated by pull-down assay, cellular thermal shift assay (CETSA), mass spectrometry analysis, molecular docking, and in vitro and in vivo functional studies. Mechanistic studies suggest that compound 8e-induced 143B cell apoptotic is mediated by EEF1A2 inhibition of the AKT signaling pathway and EEF1A2 serves as a potential candidate for targeted OS therapy.
Assuntos
Antineoplásicos , Neoplasias Ósseas , Osteossarcoma , Fator 1 de Elongação de Peptídeos , Bibliotecas de Moléculas Pequenas , Humanos , Osteossarcoma/tratamento farmacológico , Osteossarcoma/patologia , Osteossarcoma/metabolismo , Antineoplásicos/farmacologia , Antineoplásicos/química , Antineoplásicos/síntese química , Fator 1 de Elongação de Peptídeos/metabolismo , Fator 1 de Elongação de Peptídeos/antagonistas & inibidores , Proliferação de Células/efeitos dos fármacos , Animais , Camundongos , Camundongos Endogâmicos BALB C , Ensaios de Seleção de Medicamentos Antitumorais , Camundongos Nus , Relação Estrutura-Atividade , Relação Dose-Resposta a Droga , Estrutura Molecular , Simulação de Acoplamento Molecular , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/patologia , Neoplasias Ósseas/metabolismo , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/patologia , Neoplasias Experimentais/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/síntese químicaRESUMO
Osteosarcoma is the most common primary malignant bone tumor, predominantly affecting young individuals. Despite standard chemotherapy and surgical resection, the overall survival rate has reached a plateau, emphasizing the need for more effective treatments. Flavonoids are antioxidant molecules with recognized anti-inflammatory and anticancer properties. In this study, we aimed to investigate the therapeutic potential of five flavonoids against four different osteosarcoma cell lines (MG-63, Saos-2 HOS, and 143B). Among the five structurally different flavonoids, robinetin exhibited the highest toxicity against osteosarcoma cells while sparing healthy human lung fibroblasts (MRC-5). Robinetin synergized with doxorubicin, reducing 143B cell viability, delaying migration, and downregulating metastasis-related transcription factors c-Jun, Snail, Slug, and Twist2. In vivo, robinetin inhibited the growth of osteosarcoma tumor xenografts in a chick chorioallantoic membrane model. Our study highlights and reports for the first time the therapeutic value of robinetin and demonstrates the potential of robinetin in osteosarcoma treatment.
Assuntos
Antineoplásicos , Neoplasias Ósseas , Movimento Celular , Flavonoides , Proteínas Nucleares , Osteossarcoma , Transdução de Sinais , Fatores de Transcrição da Família Snail , Proteína 1 Relacionada a Twist , Osteossarcoma/patologia , Osteossarcoma/tratamento farmacológico , Osteossarcoma/metabolismo , Humanos , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Fatores de Transcrição da Família Snail/metabolismo , Linhagem Celular Tumoral , Animais , Transdução de Sinais/efeitos dos fármacos , Proteína 1 Relacionada a Twist/metabolismo , Neoplasias Ósseas/patologia , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/metabolismo , Flavonoides/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto , Sobrevivência Celular/efeitos dos fármacos , Embrião de Galinha , Proteínas Nucleares/metabolismo , Antineoplásicos/farmacologiaRESUMO
BACKGROUND: Osteosarcoma is a malignant bone tumor primarily composed of interstitial cells; there is an urgent need to develop effective treatments to improve patient prognosis. Traditional Chinese medicine offers a promising direction for research. This study explores the inhibitory effects and mechanisms of eupatilin on osteosarcoma, as well as the feasibility of using exosomes loaded with eupatilin in the treatment of osteosarcoma. METHODS: The cell counting kit-8 (CCK-8) assay was utilized to determine the optimal experimental concentration of eupatilin and assess its effect on cell proliferation. Cell apoptosis, migration, and invasion were evaluated through flow cytometry, wound healing assay, transwell assay, and colony formation assay. The expression of neighbor of BRCA1 gene 2 (NBR2), microRNA-129-5p (miR-129-5p), and FKBP prolyl isomerase 11 (FKBP11) were assessed using real-time quantitative polymerase chain reaction and Western blot. Extracellular exosomes from bone marrow mesenchymal stem cells were extracted via ultracentrifugation. Exosomes overexpressing miR-129-5p were obtained by transfecting the stem cells, and exosomes loaded with eupatilin were prepared through co-incubation. The inhibitory effects of different exosome treatments were observed. RESULTS: Cytological experiments demonstrated that eupatilin significantly enhances the apoptosis rate of osteosarcoma cells, suppresses cell viability, and markedly diminishes the capacities for colony formation, migration, and invasion. PCR and WB analyses revealed that the expression levels of NBR2, FKBP11 gene, and protein were notably reduced, whereas the expression level of miR-129-5p was significantly elevated. Exosome-based therapy exhibited a pronounced inhibitory effect on osteosarcoma cells. CONCLUSION: Eupatilin exerts a reliable inhibitory effect on osteosarcoma cells through the NBR2/miR-129-5p/FKBP11 regulatory axis. Exosomes can effectively carry both eupatilin and miR-129-5p, enhancing their therapeutic efficacy.
Assuntos
Neoplasias Ósseas , Flavonoides , MicroRNAs , Osteossarcoma , Proteínas de Ligação a Tacrolimo , Humanos , Osteossarcoma/tratamento farmacológico , Osteossarcoma/metabolismo , Osteossarcoma/patologia , Osteossarcoma/genética , MicroRNAs/genética , MicroRNAs/metabolismo , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/patologia , Neoplasias Ósseas/genética , Movimento Celular/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Flavonoides/farmacologia , Proteínas de Ligação a Tacrolimo/metabolismo , Proteínas de Ligação a Tacrolimo/genética , Exossomos/metabolismo , Células Tumorais CultivadasRESUMO
Osteosarcoma is a highly aggressive type of bone cancer with a high rate of metastasis. The molecular mechanisms underlying osteosarcoma metastasis are not yet completely understood, representing an ongoing challenge for therapy. A possible therapeutic target is the hypoxia-inducible factor HIF-1α which is upregulated in metastatic osteosarcoma. Indeed, HIF-1α promotes proliferation, resistance to apoptosis and metabolic reprogramming towards glycolysis, whereas its downregulation increases apoptosis. The molecular mechanism mediated by the mitochondrial E3 ubiquitin ligase MUL1 could be exploited to target HIF-1α since low MUL1 protein levels result in HIF-1α accumulation and activity even under normoxic conditions, while high levels of MUL1 promote HIF-1α degradation. Here, we show that MUL1 protein levels inversely correlate with the aggressiveness of osteosarcoma cell lines. Induction of MUL1 in aggressive cells reduces HIF-1α levels, paired with a decrease in proliferation, migration and glycolysis and increase in apoptosis, whereas MUL1 inactivation in low-aggressive cells has opposite results. Therefore, the modulation of MUL1 protein levels affects cell proliferation, migration, apoptosis, and metabolism. This is the first report that reveals a tumor suppressor role for MUL1 in osteosarcoma, and suggests MUL1 induction as a potential therapeutic strategy to reduce HIF-1α activity in the metastatic progression of this cancer.
Assuntos
Neoplasias Ósseas , Mitocôndrias , Osteossarcoma , Ubiquitina-Proteína Ligases , Humanos , Osteossarcoma/patologia , Osteossarcoma/genética , Osteossarcoma/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Proliferação de Células/genética , Linhagem Celular Tumoral , Apoptose/genética , Mitocôndrias/metabolismo , Mitocôndrias/genética , Mitocôndrias/enzimologia , Neoplasias Ósseas/patologia , Neoplasias Ósseas/genética , Neoplasias Ósseas/metabolismo , Glicólise/genética , Movimento Celular/genética , Regulação Neoplásica da Expressão GênicaRESUMO
Chemoresistance is a major factor contributing to the poor prognosis of osteosarcoma. Increasing evidence underscores the pivotal role of enhanced tumor stemness in driving drug resistance. In this study we investigated the molecular mechanisms underlying the chemoresistance and stemness in osteosarcoma. Two cisplatin-resistant osteosarcoma cell line models (U2OS-DDPr and 143B-DDPr) were established by culturing parental U2OS and 143B cells with escalating cisplatin concentrations (250 ng/mL to 2.5 µg/mL) over a 6-month period. We found that the expression levels of suppressor of cytokine signaling 1 (SOCS1), an E3 ubiquitin ligase, were markedly downregulated in both chemo-resistant osteosarcoma cells and osteosarcoma tumor specimens, and the reduced expression in tumor specimens was correlated to poor prognosis in osteosarcoma patients. Silencing SOCS1 significantly reduced cisplatin sensitivity, enhanced spheroid formation capacity, and upregulated the expression of stem cell markers including SOX2, OCT4, and CD44. Conversely, restoring SOCS1 expression reversed these effects both in vitro and in vivo. Immunoprecipitation-mass spectrometry analysis revealed that SOCS1 bound to ACTN4 and suppressed its protein expression by promoting K63-linked ubiquitination, ultimately leading to proteasomal degradation. Specifically, the SH2 domain of SOCS1 interacted with the N-terminal region of ACTN4, with Lys66 of ACTN4 playing a critical role in facilitating this interaction and subsequent ubiquitination. In addition, the expression of ACTN4 was highly enriched in chemo-resistant tissues, and its overexpression was positively associated with advanced tumor staging. Importantly, ACTN4 functioned as an oncogene to promote cisplatin resistance and stemness in osteosarcoma. Furthermore, we found that wortmannin, an inhibitor of ACTN4, could markedly block the effect of SOCS1 silencing on osteosarcoma aggressiveness. In conclusion, SOCS1 deletion promotes stemness and chemoresistance in osteosarcoma by inhibiting ACTN4 ubiquitination and degradation, which offers promising therapeutic targets for potentiating chemosensitivity in osteosarcoma.
Assuntos
Actinina , Neoplasias Ósseas , Resistencia a Medicamentos Antineoplásicos , Células-Tronco Neoplásicas , Osteossarcoma , Proteína 1 Supressora da Sinalização de Citocina , Humanos , Osteossarcoma/tratamento farmacológico , Osteossarcoma/metabolismo , Osteossarcoma/patologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Proteína 1 Supressora da Sinalização de Citocina/metabolismo , Proteína 1 Supressora da Sinalização de Citocina/genética , Cisplatino/farmacologia , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/efeitos dos fármacos , Actinina/metabolismo , Linhagem Celular Tumoral , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/patologia , Animais , Antineoplásicos/farmacologia , Camundongos Nus , Proteólise , Camundongos , Ubiquitinação , Camundongos Endogâmicos BALB CRESUMO
BACKGROUND AND OBJECTIVE: Osteosarcoma is a highly malignant bone tumor that occurs primarily in children and adolescents. Ki67 protein expression level (detected through immunohistochemistry) is an important indicator for assessing tumor proliferative activity. This study aims to develop an efficient and low-cost artificial intelligence model to predict Ki67 expression levels from pathological images. METHODS: 73 hematoxylin and eosin-stained (H&E) whole slide images (WSIs) of osteosarcoma specimens were analyzed. Tumor regions were segmented using weakly supervised learning, followed by extraction of 215 nuclear features including shape, texture, spatial and topological features through the Hover-Net network. Feature selection was performed using five methods: least absolute shrinkage and selection operator (LASSO), mutual information (MI), recursive feature elimination (RFE), Wilcoxon rank sum test (WRST), and extreme gradient boosting (XGBoost), with the top 5 features selected from each method. These features were subsequently integrated with 8 machine learning classifiers: adaptive boosting (AdaBoost), balanced random forest (BalancedRF), k-nearest neighbors (KNN), light gradient boosting machine (LightGBM), multilayer perceptron (MLP), quadratic discriminant analysis (QDA), random forest (RF), and support vector machine (SVM) to determine the optimal hybrid model. RESULTS: By combining 5 key features with 8 machine learning classifiers, we selected the optimal hybrid model (XGBoost+SVM). This model demonstrated the best performance in accuracy (0.767 ± 0.018), recall (0.872 ± 0.036), F1-score (0.800 ± 0.012), and receiver operating characteristic-area under curve (ROC-AUC) (0.884 ± 0.045). The model showed both high accuracy and high sensitivity in Ki67 detection. CONCLUSION: Our model provides an automated and reliable solution for osteosarcoma Ki67 assessment, reducing dependence on traditional immunohistochemistry. Its excellent performance indicates strong potential for clinical translation.
Assuntos
Neoplasias Ósseas , Antígeno Ki-67 , Osteossarcoma , Humanos , Osteossarcoma/metabolismo , Osteossarcoma/patologia , Osteossarcoma/diagnóstico por imagem , Antígeno Ki-67/metabolismo , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/patologia , Criança , Adolescente , Aprendizado de Máquina , Algoritmos , Imuno-Histoquímica , Feminino , Curva ROC , Processamento de Imagem Assistido por Computador , MasculinoRESUMO
Cuproptosis and ferroptosis are difficult to synergize in tumor suppression due to their spatiotemporal asynchrony and mechanistic dispersion. Therefore, we proposed a new paradigm of Ferroptosis-Cuproptosis self-recycling. An ironsulfur cluster (Fe-S) metabolic collapse was constructed by hyaluronic acid (HA)-functionalized bimetallic nano-enzymes (HA@CuCo-NC) to initiate this tumoricidal mechanism. HA@CuCo-NC specifically accumulates in osteosarcoma cells through HA-mediated targeting. The multi-enzyme activity triggers a surge of reactive oxygen species (ROS) in the tumor microenvironment, which, along with Co2+, induces ferroptosis. This leads to severe mitochondrial damage and glutathione (GSH) depletion, which in turn activates the cuproptosis cascade. Cuproptosis causes Fe-S protein degradation and Fe2+ release into the labile iron pool (LIP) by inhibiting Fe-S cluster formation and dihydrolipoamide acetyltransferase (DLAT) oligomerization. The released Fe2+ enhances oxidative stress and promotes membrane disassembly via the Fenton reaction. The two act synergistically to form a dynamic self-reinforcing cycle: Ferroptosis-related mitochondrial damage exacerbates the toxicity of Cu2+. Cuproptosis-released Fe2+ further activates ferroptosis. In vivo experiments demonstrated that HA@CuCo-NC significantly inhibited the growth of osteosarcoma in situ without causing systemic toxicity. This research introduces a novel approach to treating solid tumors by reconstructing the metabolic interaction network of copper and iron.
Assuntos
Antineoplásicos , Neoplasias Ósseas , Cobre , Ferroptose , Proteínas Ferro-Enxofre , Nanoconjugados , Osteossarcoma , Osteossarcoma/tratamento farmacológico , Osteossarcoma/patologia , Osteossarcoma/metabolismo , Ferroptose/efeitos dos fármacos , Humanos , Antineoplásicos/farmacologia , Antineoplásicos/química , Animais , Camundongos , Cobre/química , Cobre/farmacologia , Nanoconjugados/química , Espécies Reativas de Oxigênio/metabolismo , Ensaios de Seleção de Medicamentos Antitumorais , Proliferação de Células/efeitos dos fármacos , Linhagem Celular Tumoral , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/patologia , Neoplasias Ósseas/metabolismo , Ferro/metabolismo , Ferro/química , Tamanho da Partícula , Propriedades de Superfície , Ácido Hialurônico/química , Proteínas Ferro-Enxofre/metabolismoRESUMO
Osteosarcoma, a prevalent age-related condition, often goes undiagnosed due to expensive and invasive detection methods. This study presents a novel, cost-effective, non-invasive electrochemical sensor for Osteosarcoma detection, leveraging peptide probes to selectively recognize key biomarkers like iron ions and osteocalcin. Using a phospholipid monolayer and a conductive substrate, the sensor utilizes peptide probes containing a tripeptide iron-binding sequence and an osteocalcin sequence to detect ferroptosis-induced iron ions and elevated osteocalcin levels, both indicative of early-stage Osteosarcoma. Electrochemical modulation facilitates the covalent assembly of osteocalcin into nanoscale aggregates, significantly amplifying the sensor's signal. This design avoids the need for complex antibodies or nanomaterials, enhancing affordability and simplicity. By integrating everyday components like toothpaste to form a low baseline signal and utilizing saliva as the sample, the sensor offers high sensitivity and a low-cost alternative to traditional diagnostic methods. This innovative approach combines bioinspired materials and electrochemical techniques to provide a promising solution for early Osteosarcoma detection, addressing the pressing need for accessible diagnostics in aging populations.
Assuntos
Técnicas Biossensoriais , Neoplasias Ósseas , Durapatita , Capacitância Elétrica , Técnicas Eletroquímicas , Ferroptose , Osteossarcoma , Peptídeos , Osteossarcoma/diagnóstico , Osteossarcoma/metabolismo , Humanos , Técnicas Eletroquímicas/métodos , Durapatita/química , Durapatita/metabolismo , Peptídeos/química , Técnicas Biossensoriais/métodos , Neoplasias Ósseas/diagnóstico , Ferro/análise , Osteocalcina/análise , Osteocalcina/químicaRESUMO
BACKGROUND/AIM: Recently, there have been numerous publications on the induction of ferroptosis by cysteine restriction in cancer cells. The present report aimed to determine whether cysteine restriction (CR) is a cancer-specific vulnerability in comparison with methionine restriction (MR), which is a known cancer-specific vulnerability. MATERIALS AND METHODS: Human cancer cell lines (HCT116 colon cancer, 143B osteosarcoma or HT1080 fibrosarcoma) and normal human fibroblasts (Hs27) were cultured in Dulbecco's modified Eagle's medium (DMEM) with dialyzed fetal bovine serum from which methionine or cysteine or both or neither had been depleted. Cancer and normal cells were co-cultured in 12-well plates under the above conditions. HCT116 cells expressing green fluorescent protein, and 143B and HT1080 cells expressing red fluorescent protein, were visualized by fluorescence microscopy. Normal fibroblasts and cancer cells were visualized by phase-contrast microscopy as well. RESULTS: In co-culture, of either 143B, HCT116 or HT1080 with Hs27 human fibrosarcoma, CR was toxic to Hs27 normal fibroblasts as well as to all three cancer cell lines. In contrast, MR was toxic only to the cancer cells but not normal fibroblasts. Dual CR and MR was toxic to normal and cancer cells. CONCLUSION: For all three cancer cell lines, HCT116 colon cancer, HT1080 fibrosarcoma and 143B osteosarcoma, both MR and CR were highly inhibitory in the co-cultures with Hs27 normal fibroblasts. In all cases MR had only a slight effect on normal fibroblasts, but CR was highly toxic to normal fibroblasts. Thus, MR is a cancer-specific vulnerability in contrast to CR which is toxic to both normal and cancer cells and is not a cancer-specificity vulnerability. Therefore, attempting to induce ferroptosis of cancer cells by CR does not appear to have potential as an effective cancer therapy.
Assuntos
Cisteína , Metionina , Neoplasias , Humanos , Cisteína/metabolismo , Cisteína/deficiência , Metionina/deficiência , Metionina/metabolismo , Fibroblastos/metabolismo , Linhagem Celular Tumoral , Células HCT116 , Técnicas de Cocultura , Neoplasias/metabolismo , Neoplasias/patologia , Osteossarcoma/metabolismo , Osteossarcoma/patologia , FerroptoseRESUMO
Canine appendicular osteosarcoma (OSA) is a highly metastatic tumor in dogs. Mortality due to metastatic disease is common and frequently occurs within 1 year of diagnosis despite standard-of-care treatment. Treatment includes amputation for palliation and chemotherapy for metastatic disease. Current histologic grading schemes and biomarkers are poor at predicting clinical outcome. Novel prognostic and therapeutic markers are required to improve patient care. MicroRNAs (miRNAs) are small molecules expressed by all cells and released into bodily fluids. Studies in human and canine OSA cell lines, tissues from the primary site, and blood have demonstrated the role of miRNAs in metastatic progression of OSA and its prognostication. We sought to investigate the miRNA profile of primary OSA tissue and compare it to pulmonary metastases and normal lung tissue by real-time quantitative polymerase chain reaction (PCR). Multiple miRNA and multiple variable models were investigated in primary OSA tissue to predict clinical outcome. Thirteen miRNAs had similar expression between primary and metastatic OSA but were different from normal lung tissue. MiR-9-5p, miR-196a-5p, and miR-196b were expressed in metastatic OSA but lacked expression in almost all normal lung samples. In multiple variable models for overall survival and disease-free interval, only miRNAs were selected as significant variables. This study found miRNAs that are nearly exclusively expressed in metastatic pulmonary OSA and could serve as novel therapeutic targets. MiRNAs were also found to be important prognostic biomarkers in tissue and improved prognostic ability as miRNA signatures.
Assuntos
Neoplasias Ósseas , Doenças do Cão , Neoplasias Pulmonares , MicroRNAs , Osteossarcoma , Animais , Cães , Osteossarcoma/veterinária , Osteossarcoma/patologia , Osteossarcoma/genética , Osteossarcoma/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Doenças do Cão/patologia , Doenças do Cão/genética , Doenças do Cão/metabolismo , Neoplasias Pulmonares/veterinária , Neoplasias Pulmonares/secundário , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Ósseas/veterinária , Neoplasias Ósseas/patologia , Neoplasias Ósseas/genética , Neoplasias Ósseas/metabolismo , Regulação Neoplásica da Expressão Gênica , Feminino , Masculino , Biomarcadores Tumorais/genética , Reação em Cadeia da Polimerase em Tempo Real/veterinária , PrognósticoRESUMO
Blue light (400-500 nm) photobiomodulation (PBM), particularly utilizing light-emitting diodes (LEDs), has garnered extensive attention for its therapeutic efficacy across various conditions, including oncology. Nevertheless, its impact on osteosarcoma cells remain inadequately characterized. This study sought to investigate the tumor-suppressive properties and underlying molecular mechanisms of LED blue light PBM in human osteosarcoma cell lines HOS and MG63. The results indicated that blue light PBM effectively suppressed cell proliferation and triggered G2/M phase cell cycle arrest. Moreover, blue light PBM activated apoptosis and ferroptosis, characterized by significant changes in ferroptosis-associated proteins (HO-1, PTGS2, GPX4). Concurrently, ROS accumulation triggered oxidative stress, as demonstrated by increased MDA and LPO levels, well-established markers of ferroptosis. These findings underscore the promising potential of LED blue light PBM as an innovative, non-pharmacological therapeutic strategy for osteosarcoma, warranting further investigation for future clinical applications.
Assuntos
Apoptose , Ferroptose , Luz , Osteossarcoma , Estresse Oxidativo , Espécies Reativas de Oxigênio , Humanos , Osteossarcoma/patologia , Osteossarcoma/metabolismo , Apoptose/efeitos da radiação , Linhagem Celular Tumoral , Espécies Reativas de Oxigênio/metabolismo , Ferroptose/efeitos da radiação , Estresse Oxidativo/efeitos da radiação , Proliferação de Células/efeitos da radiação , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos da radiação , Neoplasias Ósseas/patologia , Neoplasias Ósseas/metabolismo , Heme Oxigenase-1/metabolismo , Fosfolipídeo Hidroperóxido Glutationa Peroxidase/metabolismo , Ciclo-Oxigenase 2/metabolismo , Luz AzulRESUMO
PURPOSE: Development of Ce/Cu-modified black ceramics and investigation of the mechanism of anti-osteosarcoma activity. METHODS: Black ceramics doped with 0.3-0.7 wt% CeO2 and CuO were prepared and analyzed using energy-dispersive X-ray spectroscopy (EDS), compression stress-strain tests, weight change, and antibacterial assays. MNNG/HOS and MG63 cells were seeded on ceramic substrates, and their adhesion and morphology were analyzed using scanning electron microscopy (SEM) and phalloidin staining. In vitro, we first assessed the anti-tumor effects of ceramic materials, then validated their impact on cuproptosis by measuring Cu2+ levels and the expression of related proteins, including SLC31A1, HSP70, and ATP7A. Mitochondrial dysfunction and TCA abnormalities were assessed by measuring mitochondrial membrane potential, reactive oxygen species, mitochondrial structure, TCA cycle metabolites (citric acid and lactic acid), ATP levels, and DLAT and LIAS protein levels. The cuproptosis inhibitor TTM, PI3K activator 740YP, and AKT activator SC79 were used to explore the relationship between cuproptosis and the PI3K/AKT pathway in osteosarcoma cells treated with Ce/Cu-doped black ceramics. RESULTS: Our research confirmed the superior anti-tumor properties of the Ce/Cu-modified black ceramic and its potential mechanisms against osteosarcoma. The Ce/Cu-doped black ceramic caused mitochondrial dysfunction and disrupted TCA cycle metabolism, resulting in cuproptosis in MNNG/HOS and MG63 cells. The PI3K/AKT pathway was involved in cuproptosis in osteosarcoma cells treated with Ce/Cu-modified ceramics. CONCLUSION: Our research confirmed that Ce/Cu-modified black ceramics have superior anti-tumor effects and potential mechanisms, confirming the clinical potential of Ce/Cu-modified black ceramics in cancer treatment and providing theoretical basis for the clinical translation.
Assuntos
Antineoplásicos , Neoplasias Ósseas , Cerâmica , Cério , Ciclo do Ácido Cítrico , Cobre , Mitocôndrias , Osteossarcoma , Osteossarcoma/metabolismo , Osteossarcoma/patologia , Osteossarcoma/tratamento farmacológico , Humanos , Cerâmica/química , Cerâmica/farmacologia , Cobre/química , Cobre/farmacologia , Linhagem Celular Tumoral , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Cério/química , Cério/farmacologia , Ciclo do Ácido Cítrico/efeitos dos fármacos , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/patologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Antineoplásicos/farmacologia , Antineoplásicos/química , Espécies Reativas de Oxigênio/metabolismoRESUMO
Osteosarcoma, the most common primary malignant bone tumor, poses significant clinical challenges due to its aggressive nature, high metastatic potential, and resistance to conventional therapies. Despite improvements in surgical and chemotherapeutic approaches, survival rates for relapsed or metastatic disease remain poor. Recent advances in understanding the tumor immune microenvironment (TIME) and exosome biology have uncovered critical mechanisms driving osteosarcoma progression, immune evasion, and therapeutic resistance. Tumor-associated macrophages (TAMs), particularly the M2 phenotype, dominate the osteosarcoma immune landscape and contribute to immunosuppression through cytokine secretion and modulation of T cell function. Exosomes, as intercellular messengers, further exacerbate tumor progression by transporting oncogenic proteins, immunosuppressive factors (TGF-ß), miRNAs, and drug-resistance molecules. These vesicles also influence critical signaling cascades including Wnt/ß-catenin and TGF-ß pathways, shaping both local and systemic tumor responses. This review delineates the roles of immune cells and tumor-derived exosomes in osteosarcoma biology and evaluates emerging immunotherapeutic strategies, including immune checkpoint inhibitors, CAR-T cells, tumor vaccines, cytokine-targeted agents, and combination therapies. We highlight ongoing clinical trials, numerical efficacy metrics, and the translational promise of exosome-based diagnostics and therapeutics. Ultimately, integrated approaches targeting both the TIME and exosome-mediated mechanisms may yield more effective and durable treatments for osteosarcoma patients.
Assuntos
Neoplasias Ósseas , Exossomos , Osteossarcoma , Humanos , Osteossarcoma/imunologia , Osteossarcoma/terapia , Osteossarcoma/patologia , Osteossarcoma/metabolismo , Exossomos/imunologia , Exossomos/metabolismo , Microambiente Tumoral/imunologia , Neoplasias Ósseas/imunologia , Neoplasias Ósseas/terapia , Neoplasias Ósseas/patologia , Neoplasias Ósseas/metabolismo , Animais , Progressão da Doença , Imunoterapia/métodos , Macrófagos Associados a Tumor/imunologia , Macrófagos Associados a Tumor/metabolismo , Terapia de Alvo MolecularRESUMO
Osteosarcoma is a primary bone malignancy in which outcomes for patients with metastatic or relapsed disease remain unsatisfactory despite optimized surgery-chemotherapy backbones. Recently, advances in cancer neuroscience have highlighted neurotrophins-nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF)-and their Trk/p75NTR receptors as modulators of tumor behavior and immune tone, offering a new strategy to recondition the osteosarcoma microenvironment. Evidence has been accumulated with the real-world application of TRK inhibitors in fusion-positive cancers and anti-NGF biologics in bone pain, together with osteosarcoma specimens and functional models showing that NGF/BDNF signaling promotes invasion, angiogenesis, and immunosuppressive niches, while neuromodulatory agents may counter these programs. Notably, deployment of these agents as immunity enablers in an immunologically "cold" sarcoma remains controversial, with uncertainties around biomarker selection, pharmacodynamic monitoring, and rational combinations with checkpoint or cellular therapies. In this review, we summarized NGF/BDNF expression and receptor activity in osteosarcoma and conducted a translational, efficacy- and safety-oriented appraisal of tissue/biofluid biomarker readouts and drugging opportunities targeting neurotrophin and co-regulatory neural circuits. In addition, we further discussed the potential mechanisms by which neurotrophin and adrenergic pathways regulate the balance between microenvironmental immunosuppression and lasting anti-tumor immunity. The purpose of this article is to define NGF/BDNF as shapers of the osteosarcoma immune microenvironment and to delineate biomarker-guided therapeutic opportunities for clinical testing.