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Synovial sarcoma (SS) is an aggressive soft tissue sarcoma with poor prognosis due to late recurrence and metastasis. Metastasis is an important prognostic factor of SS. This study aimed to identify the core genes and mechanisms associated with SS metastasis. Microarray data for GSE40021 and GSE40018 were obtained from the Gene Expression Omnibus database. 186 differentially expressed genes (DEGs) were identified. The biological functions and signalling pathways closely associated with SS metastasis included extracellular matrix (ECM) organization and ECM-receptor interaction. Gene set enrichment analysis showed that the terms cell cycle, DNA replication, homologous recombination and mismatch repair were significantly enriched in the metastasis group. Weighted gene co-expression network analysis identified the most relevant module and 133 hub genes, and 31 crossover genes were identified by combining DEGs. Subsequently, four characteristic genes, EXO1, NCAPG, POLQ and UHRF1, were identified as potential biomarkers associated with SS metastasis using the least absolute shrinkage and selection operator algorithm and validation dataset verification analysis. Immunohistochemistry results from our cohort of 49 patients revealed visible differences in the expression of characteristic genes between the non-metastatic and metastatic groups. Survival analysis indicated that high expression of characteristic genes predicted poor prognosis. Our data revealed that primary SS samples from patients who developed metastasis showed activated homologous recombination and mismatch repair compared to samples from patients without metastasis. Furthermore, EXO1, NCAPG, POLQ and UHRF1 were identified as potential candidate metastasis-associated genes. This study provides further research insights and helps explore the mechanisms of SS metastasis.
Assuntos
Biomarcadores Tumorais , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Metástase Neoplásica , Sarcoma Sinovial , Sarcoma Sinovial/genética , Sarcoma Sinovial/patologia , Sarcoma Sinovial/metabolismo , Humanos , Prognóstico , Biomarcadores Tumorais/genética , Redes Reguladoras de Genes , Feminino , Masculino , Bases de Dados Genéticas , Biologia Computacional/métodos , Pessoa de Meia-IdadeRESUMO
PURPOSE: Pulmonary metastasis was a negative factor of osteosarcoma prognosis. However, there is no universal criteria to confirm pulmonary metastasis at pulmonary micro nodule (PMN, Dmax ≤ 5 mm) stage other than pathology. We aimed to identify prevalence of PMNs, determine prognosis of osteosarcoma with PMNs, and analyze risk factors related to PMN progression. METHODS: We retrospectively reviewed 425 consecutive osteosarcoma patients. According to dynamic change in size and number of PMNs, patients were divided into PMN progression and non-progression group. Demographic data, initial laboratory data, radiological features, and oncological evaluations were analyzed. Cox regression was used to identify risk factors for PMN progression. Overall survival rate was measured and analyzed with Kaplan-Meier method. Differences with p < 0.05 were considered significant. RESULTS: PMNs were found in 74% (315/425) osteosarcoma patients, half of whom (157/315) suffering PMN progression. Overall survival rate was 70.2%, while survival rates for PMN progression group and non-progression group were 53.40% and 87.40%, respectively. Clinical risk factors for PMN progression in certain patients included blood vessel invasion, extrapulmonary metastases, low tumour cell necrosis rate, and large tumour size. Radiologic risk factors included greatest diameter, distance to pleura, CT value, solid components, and smooth border. CONCLUSION: PMN is quite common in osteosarcoma patients. PMN progression is related to both certain clinical and radiological factors, which could assist surgeons to determine its possibility to progress at an early stage.
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Neoplasias Ósseas , Neoplasias Pulmonares , Osteossarcoma , Neoplasias Ósseas/patologia , Humanos , Neoplasias Pulmonares/epidemiologia , Neoplasias Pulmonares/secundário , Prevalência , Estudos RetrospectivosRESUMO
BACKGROUND: Recent studies indicated that exosomal programmed death-ligand 1 (PD-L1) derived from cancers could induce immunosuppression and tumor pathogenesis. However, it is unclear how exosomes influence osteosarcoma (OS) progression and whether PD-L1 also exists in serum exosomes (Sr-exosomes) of patients with osteosarcoma. We examined serum exosomes from 70 OS patients, 9 patients with benign tumors and 22 healthy donors. OS-derived exosomes were functionally evaluated in vivo and in vitro. RESULTS: The characteristics of exosomes derived from OS patient serum and OS cell lines were confirmed by several methods. We found OS patients had a higher level of exosomal PD-L1 compared to healthy donors. Meanwhile, OS patients with pulmonary metastasis also showed a relatively higher level of exosomal PD-L1 than patients without metastasis. Next, bioinformatic analysis demonstrated that Sr-exosomes isolated from OS patients may involve in the important process of immune function and cancer pathogenesis for OS patients. Co-expression network centered with PD-L1 among Sr-exosomal differently expressed mRNA demonstrated exosomal N-cadherin had a close relationship with exosomal PD-L1 expression. Then, we confirmed higher level of Sr-exosomal N-cadherin in OS patients with pulmonary metastasis compared to ones without metastasis. Furthermore, we elucidated osteosarcoma-derived exosomes and exosomal-PD-L1 promoted the pulmonary metastasis in metastatic models. ROC (Receiver Operating Characteristic Curve) analysis showed AUC (Area Under Curve) of 0.823 for exosomal PD-L1, 0.806 for exosomal N-cadherin and 0.817 for exosomal N-cadherin/E-cadherin to distinguish OS patients with pulmonary metastasis from ones without metastasis. CONCLUSIONS: Osteosarcoma stimulates pulmonary metastasis by releasing exosomes, that carry PD-L1 and N-cadherin. Detection of exosomal PD-L1 and N-cadherin from serum of OS patients may predict pulmonary metastasis progression for OS patients.
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Antígeno B7-H1/sangue , Biomarcadores Tumorais/sangue , Caderinas/sangue , Exossomos/química , Neoplasias Pulmonares/diagnóstico , Metástase Neoplásica/diagnóstico , Osteossarcoma/metabolismo , Animais , Antígeno B7-H1/genética , Caderinas/genética , Linhagem Celular Tumoral , Permeabilidade da Membrana Celular , Biologia Computacional , Progressão da Doença , Feminino , Técnicas de Silenciamento de Genes , Humanos , Neoplasias Pulmonares/secundário , Camundongos Endogâmicos BALB C , RNA Mensageiro/metabolismo , Distribuição Tecidual , CicatrizaçãoRESUMO
Osteosarcoma is an aggressive bone malignancy with a poor prognosis. One putative proto-oncogene in osteosarcoma is SKP2, encoding a substrate recognition factor of the SCF E3 ubiquitin ligase. We previously demonstrated that Skp2 knockout in murine osteosarcoma improved survival and delayed tumorigenesis. Here, we performed RNA sequencing (RNA-seq) on tumors from a transgenic osteosarcoma mouse model with conditional Trp53 and Rb1 knockouts in the osteoblast lineage ("DKO": Osx1-Cre;Rb1lox/lox;p53lox/lox) and a triple-knockout model with additional Skp2 germline knockout ("TKO": Osx1-Cre;Rb1lox/lox;p53lox/lox;Skp2-/-), followed by qPCR and immunohistochemistry validation. To investigate the clinical implications of our results, we analyzed a human osteosarcoma patient cohort ("NCI-TARGET OS") with RNA-seq and clinical data. We found large differences in gene expression after SKP2 knockout. Surprisingly, we observed increased expression of genes related to immune microenvironment infiltration in TKO tumors, especially the signature genes for macrophages and to a lesser extent, T cells, B cells, and vascular cells. We also uncovered a set of relevant transcription factors that may mediate these changes. In osteosarcoma patient cohorts, high expression of genes upregulated in TKO was correlated with favorable overall survival, which was largely explained by the macrophage gene signatures. This relationship was further supported by our finding that SKP2 expression was negatively correlated with macrophage infiltration in the NCI-TARGET osteosarcoma and the TCGA Sarcoma cohorts. Overall, our findings indicate that SKP2 may mediate immune exclusion from the osteosarcoma tumor microenvironment, suggesting that SKP2 modulation in osteosarcoma may induce antitumor immune activation.
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Neoplasias Ósseas , Osteossarcoma , Animais , Humanos , Camundongos , Neoplasias Ósseas/genética , Modelos Animais de Doenças , Camundongos Knockout , Camundongos Transgênicos , Osteossarcoma/genética , Osteossarcoma/patologia , Prognóstico , Proteínas Quinases Associadas a Fase S/genética , Proteínas Quinases Associadas a Fase S/metabolismo , Microambiente Tumoral/genética , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismoRESUMO
Osteosarcoma (OS) is the most common primary pediatric bone malignancy. One promising new therapeutic target is SKP2, encoding a substrate recognition factor of the SCF E3 ubiquitin ligase responsible for ubiquitination and proteasome degradation of substrate p27, thus driving cellular proliferation. We have shown previously that knockout of Skp2 in an immunocompetent transgenic mouse model of OS improved survival, drove apoptosis, and induced tumor inflammation. Here, we applied single-cell RNA-sequencing (scRNA-seq) to study primary OS tumors derived from Osx-Cre driven conditional knockout of Rb1 and Trp53. We showed that murine OS models recapitulate the tumor heterogeneity and microenvironment complexity observed in patient tumors. We further compared this model with OS models with functional disruption of Skp2: one with Skp2 knockout and the other with the Skp2-p27 interaction disrupted (resulting in p27 overexpression). We found reduction of T cell exhaustion and upregulation of interferon activation, along with evidence of replicative and endoplasmic reticulum-related stress in the Skp2 disruption models, and showed that interferon induction was correlated with improved survival in OS patients. Additionally, our scRNA-seq analysis uncovered decreased activities of metastasis-related gene signatures in the Skp2-disrupted OS, which we validated by observation of a strong reduction in lung metastasis in the Skp2 knockout mice. Finally, we report several potential mechanisms of escape from targeting Skp2 in OS, including upregulation of Myc targets, DNA copy number amplification and overexpression of alternative E3 ligase genes, and potential alternative lineage activation. These mechanistic insights into OS tumor biology and Skp2 function suggest novel targets for new, synergistic therapies, while the data and our comprehensive analysis may serve as a public resource for further big data-driven OS research.
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Background: The percutaneous screw reconstruction technique, known as the "Tripod Technique," has demonstrated favorable clinical outcomes in the management of metastatic periacetabular lesions, as evidenced by our prior investigations and corroborated by independent studies. Nevertheless, there is a steep learning curve in handling this technique, with possible complications such as intraarticular screw placement. Methods: Preoperative pelvic CT scans were acquired before surgery and utilized for the guiding frame design. A convolutional neural network model was trained with annotated data to identify the starting point and trajectory of each potential screw. A model boundary intersection detection technology was used to determine the optimal diameter and length of each screw. A non-rigid registration technology was matched with a prefabricated model of the body surface to design personalized anchoring skin pads. Finally, a polylactic acid-based guiding frame for intraoperative was custom-made with a 3D printer. Results: 12 patients underwent a guiding frame-assisted Tripod procedure for treatment of periacetabular metastatic lesions. An intraoperative CT scan was performed in all cases to confirm screw trajectories. Among 36 screws that were implanted, 26 screws were implanted as designed. The remaining ten screws drifted, but all remained within the intra-osseous conduit without any complications. The mean surgical time was 1.22 h with the guiding frame compared with 2.3 h without the guiding frame. Following the surgical procedure, a noteworthy enhancement in pain management, as evidenced by a reduction in scores on the visual analog scale (p < 0.01), and an improvement in functional status, as assessed through the Eastern Cooperative Oncology Group score (p < 0.01), were observed when compared to the patient's pre-operative condition. Conclusion: This proof-of-concept investigation demonstrates that the amalgamation of AI-assisted surgical planning and additive manufacturing can improve surgical accuracy and shorten surgical duration. While access to this technology is currently constrained during its early stages of development, it is anticipated that these limitations will diminish as the potential of AI and additive manufacturing in facilitating complex orthopedic procedures becomes more evident, leading to a surge in interest and adoption of this approach.
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Osteosarcoma(OS) is a highly aggressive bone cancer for which treatment has remained essentially unchanged for decades. Although OS is characterized by extensive genomic heterogeneity and instability, RB1 and TP53 have been shown to be the most commonly inactivated tumor suppressors in OS. We previously generated a mouse model with a double knockout (DKO) of Rb1 and Trp53 within cells of the osteoblastic lineage, which largely recapitulates human OS with nearly complete penetrance. SKP2 is a repression target of pRb and serves as a substrate recruiting subunit of the SCFSKP2 complex. In addition, SKP2 plays a central role in regulating the cell cycle by ubiquitinating and promoting the degradation of p27. We previously reported the DKOAA transgenic model, which harbored a knock-in mutation in p27 that impaired its binding to SKP2. Here, we generated a novel p53-Rb1-SKP2 triple-knockout model (TKO) to examine SKP2 function and its potential as a therapeutic target in OS. First, we observed that OS tumorigenesis was significantly delayed in TKO mice and their overall survival was markedly improved. In addition, the loss of SKP2 also promoted an apoptotic microenvironment and reduced the stemness of DKO tumors. Furthermore, we found that small-molecule inhibitors of SKP2 exhibited anti-tumor activities in vivo and in OS organoids as well as synergistic effects when combined with a standard chemotherapeutic agent. Taken together, our results suggest that SKP2 inhibitors may reduce the stemness plasticity of OS and should be leveraged as next-generation adjuvants in this cancer.
Assuntos
Neoplasias Ósseas , Osteossarcoma , Animais , Humanos , Camundongos , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/genética , Carcinogênese , Inibidor de Quinase Dependente de Ciclina p27/genética , Camundongos Knockout , Osteossarcoma/tratamento farmacológico , Osteossarcoma/genética , Proteínas Quinases Associadas a Fase S/genética , Proteínas Quinases Associadas a Fase S/metabolismo , Microambiente TumoralRESUMO
Purpose: Osteosarcoma (OS) is an aggressive bone malignancy with a poor prognosis. One putative proto-oncogene in OS is SKP2, encoding a substrate recognition factor of the SCF E3 ubiquitin ligase. We previously demonstrated that SKP2 knockout in murine OS improved survival and delayed tumorigenesis. Here we aim to define the SKP2 drives transcriptional program and its clinical implication in OS. Experimental Design: We performed RNA-sequencing (RNA-seq) on tumors from a transgenic OS mouse model with conditional Trp53 and Rb1 knockouts in the osteoblast lineage ("DKO": Osx1-Cre;Rb1lox/lox;p53lox/lox) and a triple-knockout model with additional Skp2 germline knockout ("TKO": Osx1-Cre;Rb1lox/lox;p53lox/lox;SKP2-/-). We validated our RNA-seq findings using qPCR and immunohistochemistry. To investigate the clinical implications of our results, we analyzed a human OS patient cohort ("NCI-TARGET OS") with RNA-seq and clinical data. Results: We found large differences in gene expression after SKP2 knockout. Strikingly, we observed increased expression of genes related to immune microenvironment infiltration in TKO tumors. We observed significant increases in signature genes for macrophages and to a lesser extent, T cells, B cells and vascular cells. We also uncovered a set of relevant transcription factors that may mediate the changes. In OS patient cohorts, high expression of genes upregulated in TKO was correlated with favorable overall survival, which was largely explained by the macrophage gene signatures. This relationship was further supported by our finding that SKP2 expression was negatively correlated with macrophage infiltration in the NCI-TARGET OS and the TCGA Sarcoma cohort. Conclusion: Our findings indicate that SKP2 may mediate immune exclusion from the OS tumor microenvironment, suggesting that SKP2 modulation in OS may induce anti-tumor immune activation.