Biomarker For Osteosarcoma Research Articles

N6-Methyladenosine-Related LncRNAs Are Potential Remodeling Indicators in the Tumor Microenvironment and Prognostic Markers in Osteosarcoma.

N6-Adenosine methylation, yielding N6-methyladenosine (m6A), is a reversible epigenetic modification found in messenger RNAs and long non-coding RNAs (lncRNAs), which affects the fate of modified RNA molecules and is essential for the development and differentiation of immune cells in the tumor microenvironment (TME). Osteosarcoma (OS) is the most common primary bone tumor in children and adolescents, and is characterized by high mortality. Currently, the possible role of m6A modifications in the prognosis of OS is unclear. In the present study, we investigated the correlation between m6A-related lncRNA expression and the clinical outcomes of OS patients via a comprehensive analysis. Clinical and workflow-type data were obtained from the Genotype-Tissue Expression Program and The Cancer Genome Atlas. We examined the relationship between m6A modifications and lncRNA expression, conducted Kyoto Encyclopedia of Genes analysis and also gene set enrichment analysis (GSEA), implemented survival analysis to investigate the association of clinical survival data with the expression of m6A-related lncRNAs, and utilized Lasso regression to model the prognosis of OS. Furthermore, we performed immune correlation analysis and TME differential analysis to investigate the infiltration levels of immune cells and their relationship with clinical prognosis. LncRNA expression and m6A levels were closely associated in co-expression analysis. The expression of m6A-related lncRNAs was quite low in tumor tissues; this appeared to be a predicting factor of OS in a prognostic model, independent of other clinical features. The NOD-like receptor signaling pathway was the most significantly enriched pathway in GSEA. In tumor tissues, SPAG4 was overexpressed while ZBTB32 and DEPTOR were downregulated. Tissues in cluster 2 were highly infiltrated by plasma cells. Cluster 2 presented higher ESTIMATE scores and stromal scores, showing a lower tumor cell purity in the TME. In conclusion, m6A-related lncRNA expression is strongly associated with the occurrence and development of OS, and can be used to as a prognostic factor of OS. Moreover, m6A-related lncRNAs and infiltrating immune cells in the TME could serve as new therapeutic targets and prognostic biomarkers for OS.

SMARCB1 expression is a novel diagnostic and prognostic biomarker for osteosarcoma.

Background: Although weak SWI/SNF related matrix-associated actin-dependent regulator of chromatin subfamily B member 1 (SMARCB1) expression is a known diagnostic and prognostic biomarker in several malignancies, its expression and clinical significance in osteosarcoma remain unknown. The aim of the present study was to investigate SMARCB1 expression in osteosarcoma and its clinical significance with respect to chemosensitivity and prognosis.Methods: We obtained 114 specimens from 70 osteosarcoma patients to construct a tissue microarray (TMA) and assess SMARCB1 protein expression via immunohistochemistry (IHC). The mRNA expression of SMARCB1 was in-silico analyzed using open-access RNA sequencing (RNA-Seq) and clinicopathological data provided by the Therapeutically Applicable Research to Generate Effective Treatments on Osteosarcoma (TARGET-OS) project. The correlations between SMARCB1 expression and clinical features were statistically analyzed.Results: Weak SMARCB1 expression occurred in 70% of the osteosarcoma patient specimens in the TMA, and significantly correlated with poor neoadjuvant response as well as shorter overall and progression-free survival (PFS). In addition, mRNA in-silico analysis confirmed that SMARCB1 expression correlates with chemotherapeutic response and prognosis in osteosarcoma patients.Conclusion: To our knowledge, the present study is the first to analyze SMARCB1 expression in osteosarcoma. SMARCB1 may serve as a novel diagnostic and prognostic biomarker in osteosarcoma.

N6-methyladenosine Modification-Related Long Non-Coding RNAs are Potential Biomarkers for Predicting the Prognosis of Patients With Osteosarcoma.

Background: The role of N6-methyladenosine (m6A)-related long non-coding RNAs (lncRNAs) in osteosarcoma (OS) has not been fully studied yet. We aimed to identify m6A-related lncRNAs that could act as prognostic biomarkers for OS. Methods: Pearson correlation was performed to identify m6A-related lncRNAs. Univariate and multivariate Cox regression analyses were performed to construct the risk model and assess whether the risk score was an independent prognostic factor for patients with OS. Gene Set Enrichment Analysis (GSEA) was performed to analyze the functions of genes in high-risk and low-risk groups. StarBase and Cytoscape were used to construct a competing endogenous RNA (ceRNA) network based on m6A-related prognostic lncRNA signature. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed to analyze the function of genes involved in the ceRNA network. Results: We extracted 122 common lncRNAs from TCGA and Gene Expression Omnibus (GEO) databases. Pearson correlation results revealed 59 significant m6A-related lncRNAs in The Cancer Genome Atlas (TCGA) database, from which 2 were screened to construct a risk signature in TCGA dataset, which was then validated in the GEO dataset. A corresponding risk score was calculated and shown to be an independent prognostic factor for patients with OS. Enrichment analysis indicated that cell proliferation-related biological processes were more common in the high-risk group, while immune-related biological processes were more common in the low-risk group. Moreover, we established a nomogram that had a good ability to predict the overall survival of patients with OS. Additionally, a ceRNA network based on small nucleolar RNA host gene 7 (SNHG7) and small nucleolar RNA host gene 12 (SNHG12) was constructed, with genes that were enriched in hepatocellular carcinoma, gastric cancer, and non-small-cell lung cancer pathways. Conclusion: Our study revealed the prognostic role of m6A-related lncRNAs in OS and identified SNHG7 and SNHG12 as potential biomarkers for predicting the prognosis of patients with OS. These findings have enriched our understanding of the role of m6A modification in the dysregulation of lncRNAs in OS.

MCAM is associated with metastasis and poor prognosis in osteosarcoma by modulating tumor cell migration.

BackgroundAlthough there are standard treatment options for osteosarcoma (OS), the prognoses of patients with OS remain varied. Therefore, it is important to profile OS patients at a high risk of mortality to develop focused interventions. Although tumor biomarkers are closely associated with clinical outcomes, data on prognostic biomarkers for OS remain scarce.MethodsWe collected RNA expression profiles and clinical data of 90 OS patients from the GEO database (dataset GSE21257 and GSE39055) and 96 patients in the TARGET program. The data were analyzed using univariate Kaplan‐Meier survival analysis to screen candidate gene sets that might be associated with OS survival.ResultsOur analysis demonstrated that melanoma cell adhesion molecule (MCAM) was associated with overall survival of patients with OS in the three cohorts. The data showed that MCAM was upregulated in OS patients who had metastases within 5 years compared to those without metastases. GO analysis revealed that genes correlated with MCAM were mainly involved in cell migration and wound healing processes. In addition, wound healing assays and gene set enrichment analysis results from RNA sequencing data of small interfering (si)‐MCAM‐transfected OS cells demonstrated that MCAM modulated tumor cell migration.ConclusionsOur data demonstrate that MCAM may be a novel prognostic biomarker for OS. MCAM is associated with increased cell migration ability and risk of metastasis, thus leading to poor prognoses in OS patients.

The Clinical Significance and Potential Molecular Mechanism of Upregulated CDC28 Protein Kinase Regulatory Subunit 1B in Osteosarcoma.

Background CDC28 Protein Kinase Regulatory Subunit 1B (CKS1B) is a member of cyclin-dependent kinase subfamily and the relationship between CKS1B and osteosarcoma (OS) remains to be explored. Methods 80 OS and 41 nontumor tissue samples were arranged to conduct immunohistochemistry (IHC) to evaluate CKS1B expression between OS and nontumor samples. The standard mean deviation (SMD) was calculated based on in-house IHC and tissue microarrays and exterior high-throughput datasets for further verification of CKS1B expression in OS. The effect of CKS1B expression on clinicopathological and overall survival of OS patients was measured through public high-throughput datasets, and analysis of immune infiltration and single-cell RNA-seq was applied to ascertain molecular mechanism of CKS1B in OS. Results A total of 197 OS samples and 83 nontumor samples (including tissue and cell line) were obtained from in-house IHC, microarrays, and exterior high-throughput datasets. The analysis of integrated expression status demonstrated upregulation of CKS1B in OS (SMD = 1.38, 95% CI [0.52–2.25]) and the significant power of CKS1B expression in distinguishing OS samples from nontumor samples (Area under the Curve (AUC) = 0.89, 95% CI [0.86–0.91]). Clinicopathological and prognosis analysis indicated no remarkable significance but inference of immune infiltration and single-cell RNA-seq prompted that OS patients with overexpressed CKS1B were more likely to suffer OS metastasis while MYC Protooncogene may be the upstream regulon of CKS1B in proliferating osteoblastic OS cells. Conclusions In this study, sufficient evidence was provided for upregulation of CKS1B in OS. The advanced effect of CKS1B on OS progression indicates a foreground of CKS1B as a biomarker for OS.

Long Non-Coding RNA ANRIL as a Potential Biomarker of Chemosensitivity and Clinical Outcomes in Osteosarcoma.

Simple SummaryOsteosarcoma is a bone cancer with a poor prognosis. This is, in part, due to resistance to current standard-of-care chemotherapeutic treatment. As personalized treatment plans become more widely utilized, the role of patient-specific genome markers may serve to identify individuals with chemo-resistant disease at the outset of diagnosis. ANRIL, a long non-coding RNA, has promise as a predictive biomarker. Utilizing osteosarcoma cell lines, we observed that altering the expression of ANRIL significantly alters the sensitivity to cisplatin and doxorubicin, two agents that are a standard-of-care for treatment. Analysis of clinical data from the TARGET dataset confirmed higher ANRIL expression portending poorer prognosis, as evidenced by association with death and metastases at diagnosis.Osteosarcoma has a poor prognosis due to chemo-resistance and/or metastases. Increasing evidence shows that long non-coding RNAs (lncRNAs) can play an important role in drug sensitivity and cancer metastasis. Using osteosarcoma cell lines, we identified a positive correlation between the expression of a lncRNA and ANRIL, and resistance to two of the three standard-of-care agents for treating osteosarcoma—cisplatin and doxorubicin. To confirm the potential role of ANRIL in chemosensitivity, we independently inhibited and over-expressed ANRIL in osteosarcoma cell lines followed by treatment with either cisplatin or doxorubicin. Knocking-down ANRIL in SAOS2 resulted in a significant increase in cellular sensitivity to both cisplatin and doxorubicin, while the over-expression of ANRIL in both HOS and U2OS cells led to an increased resistance to both agents. To investigate the clinical significance of ANRIL in osteosarcoma, we assessed ANRIL expression in relation to clinical phenotypes using the osteosarcoma data from the Therapeutically Applicable Research to Generate Effective Treatments (TARGET) dataset. Higher ANRIL expression was significantly associated with increased rates of metastases at diagnosis and death and was a significant predictor of reduced overall survival rate. Collectively, our results suggest that the lncRNA ANRIL can be a chemosensitivity and prognosis biomarker in osteosarcoma. Furthermore, reducing ANRIL expression may be a therapeutic strategy to overcome current standard-of-care treatment resistance.

Low GNG12 Expression Predicts Adverse Outcomes: A Potential Therapeutic Target for Osteosarcoma.

BackgroundG protein subunit gamma 12 (GNG12) is observed in some types of cancer, but its role in osteosarcoma is unknown. This study hypothesized that GNG12 may be a potential biomarker and therapeutic target. We aimed to identify an association between GNG12 and osteosarcoma based on the Gene Expression Omnibus and the Therapeutically Applicable Research to Generate Effective Treatments (TARGET) databases.MethodsOsteosarcoma samples in GSE42352 and TARGET database were selected as the test cohorts. As the external validation cohort, 78 osteosarcoma specimens from The Second Affiliated Hospital of Nanchang University were collected. Patients with osteosarcoma were divided into high and low GNG12 mRNA-expression groups; differentially expressed genes were identified as GNG12-related genes. The biological function of GNG12 was annotated using Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, gene set enrichment analysis, and immune infiltration analysis. Gene expression correlation analysis and competing endogenous RNA regulatory network construction were used to determine potential biological regulatory relationships of GNG12. Overall survival, Kaplan–Meier analysis, and log-rank tests were calculated to determine GNG12 reliability in predicting survival prognosis.ResultsGNG12 expression decreased in osteosarcoma samples. GNG12 was a highly effective biomarker for osteosarcoma [area under the receiver operating characteristic (ROC) curve (AUC) = 0.920], and the results of our Kaplan–Meier analysis indicated that overall survival and progression-free survival differed significantly between low and high GNG-expression group (p < 0.05). Functional analyses indicated that GNG12 may promote osteosarcoma through regulating the endoplasmic reticulum. Expression correlation analysis and competing endogenous RNA network construction showed that HOTTIP/miR-27a-3p may regulate GNG12 expression. Furthermore, the subunit suppresses adaptive immunity via inhibiting M1 and M2 macrophage infiltration. GNG12 was inhibited in metastatic osteosarcoma compared with non-metastatic osteosarcoma, and its expression predicted survival of patients (1, 3, and 5-year AUCs were 0.961, 0.826, and 0.808, respectively).ConclusionThis study identified GNG12 as a potential biomarker for osteosarcoma prognosis, highlighting its potential as an immunotherapy target.

Circulating MicroRNAs as Novel Potential Diagnostic Biomarkers for Osteosarcoma: A Systematic Review.

Osteosarcoma (OS) is a fast-progressing bone tumor with high incidence in children and adolescents. The main diagnostic methods for OS are imaging exams and biopsies. In spite of the several resources available for detecting the disease, establishing an early diagnosis is still difficult, resulting in worse prognosis and lower survival rates for patients with OS. The identification of novel biomarkers would be helpful, and recently, circulating microRNAs (miRNAs) have been pointed to as possible non-invasive biomarkers. In order to assess the effectiveness of miRNA research, we performed a systematic review to assess the potential role of circulating miRNAs as biomarkers for OS diagnosis. We performed a search in various databases—PubMed, LILACS (Literatura Latino-americana e do Caribe em Ciências da Saúde), VHL (Virtual Health Library), Elsevier, Web of Science, Gale Academic One File—using the terms: “Circulating microRNAs” OR “plasma microRNAs” OR “serum microRNAs” OR “blood microRNAs” OR “cell-free microRNAs” OR “exosome microRNAs” OR “extracellular vesicles microRNAs” OR “liquid biopsy” AND “osteosarcoma” AND “diagnostic”. We found 35 eligible studies that were independently identified and had had their quality assessed according to Quality Assessment of Diagnostic Accuracy Studies (QUADAS-2) guidelines. Despite the useful number of publications on this subject and the fact that several microRNAs showed excellent diagnostic performance for OS, the lack of consistency in results suggests that additional prospective studies are needed to confirm the role of circulating miRNAs as non-invasive biomarkers in OS.

Silica nanoparticle-modified microcomb electrode for voltammetry detection of osteopontin with high sensitivity.

Osteosarcoma is a commonly occurring bone malignancy, and it is the second most common cause of cancer deaths in adolescents and children. A sensitive silica nanoparticle (Si-NP) modified current-volt sensor was introduced to identify the osteopontin antigen, a well-known biomarker for osteosarcoma. Si-NP was extracted from the rice husk ash and utilized for the surface functionalization on the interdigitated microelectrode sensing surface. Extracted Si-NP has a spherical shape with uniform distribution, and it is confirmed by field emission scanning electron microscopy and field-emission transmission electron microscopy. Si-NP was layered on the electrode surface through a (3-aminopropyl)triethoxysilane amine linker, and the antibody was immobilized on Si-NP through a glutaraldehyde linker. Osteopontin was effectively detected on the antibody-attached surface, and the determination limit was 0.6 ng/mL. The regression was determined as y = 0.9366x - 1.1113 and the R2 value was 0.9331 and the detection limit of osteopontin was 0.6 ng/mL in the range between 0.3 and 5 ng/mL. In addition, control performance with nonimmune antibodies and albumin did not change the current volt, showing the specific osteopontin identification. This research work brings out the easy and cost-effective method to diagnose osteosarcoma and its etiology.

Identification of PSMD14 as a potential novel prognosis biomarker and therapeutic target for osteosarcoma.

BackgroundOsteosarcoma is the most common primary bone tumor. The survival rate of osteosarcoma patients has not significantly increased in the past decades. Uncovering the mechanisms of malignancy, progression, and metastasis will shed light on the development of new therapeutic targets and treatment for osteosarcoma.AimThe aim of this study is to identify potential osteosarcoma biomarker and/or therapeutic targets by using integrated bioinformatics analysis.Methods and resultsWe utilized existing gene expression datasets to identify differential expressed genes (DEGs) that could serve as osteosarcoma biomarkers or even as therapeutic targets. We found 48 DEGs were overlapped in three datasets. Among these 48 DEGs, PSMD14 was on the top of the up‐regulated gene list. We further found that higher PSMD14 expression was correlated with higher risk group (younger age group, ≤20.83 years of age), metastasis within 5 years and higher grade of tumor. Higher PSMD14 expression in osteosarcoma had positive correlation with higher infiltration of CD8+ T cells, neutrophils and myeloid dendritic cells. Kaplan‐Myer survival data further revealed that higher expression of PSMD14 predicted significantly worse prognosis (p = .013). Gene set enrichment analysis was further performed for the DEGs related to PSMD14 in osteosarcoma. We found that lower PSMD14 expression group had more immune responses such as interferon γ, α responses, inflammation response etc. However, the higher PSMD14 expression group had more cell proliferation‐related biological processes, such as G2M checkpoints and Myc targets. Through establishing protein–protein interaction networks using PSMD14 related DEGs, we identified 10 hub genes that were all ribosomal proteins. These hub genes may play roles in osteosarcoma tumorigenesis, progression and/or metastasis.ConclusionWe identified PSMD14 gene as a possible osteosarcoma biomarker, and/or a possible therapeutic target.

Identification of MAP3K15 as a potential prognostic biomarker and correlation with immune infiltrates in osteosarcoma

BackgroundOsteosarcoma (OS) is a type of primary malignant tumor, and increasing evidence shows the clinical benefits of immunotherapy in treating OS. However, the lack of comprehensive studies on the complex OS immune microenvironment hinders the application of immunotherapy. Thus, this study aimed to systematically explore the immune characteristics of OS and identify novel biomarkers for OS treatment.MethodsWe systematically studied the immune score and proportions of infiltrating immune cells in OS in the Therapeutically Applicable Research to Generate Effective Treatments (TARGET) and Gene Expression Omnibus (GEO) databases using the ESTIMATE and CIBERSORT algorithms. Differential expression and functional analyses were used to identify dysregulated genes and explore their functions. Survival and Cox regression analyses were applied to establish an immune-related prognostic signature. Additionally, qPCR and immunohistochemistry were performed to validate the results.ResultsA total of 103 differentially expressed immune genes (DEIGs) were found in the TARGET-OS and GSE39058 databases, and these DEIGs were mainly enriched in leukocyte proliferation, leukocyte differentiation, osteoclast differentiation, natural killer (NK) cell-mediated cytotoxicity, and the adaptive immune system. A predictive signature was constructed based on the survival analysis, with an area under the receiver operating characteristic (ROC) curve (AUC) of 0.65. Moreover, we found that mitogen-activated protein kinase kinase kinase 15 (MAP3K15) can predict the prognosis of patients with OS and is closely related to CD4+ T cells and macrophages. The OS patients with high MAP3K15 expression had a significantly poorer prognosis.ConclusionsOur study found that MAP3K15, whose expression level is closely related to immune activity in tumors, is a critical immune-related biomarker, and our findings may provide a basis for OS immunotherapy.

Diaphanous related formin 3 knockdown suppresses cell proliferation and metastasis of osteosarcoma cells

Osteosarcoma is a malignant osteoblastic tumor that can gravely endanger the lives and health of children and adolescents. Therefore, there is an urgent need to explore new biomarkers for osteosarcoma and determine new targeted therapies to improve the efficacy of osteosarcoma treatment. Diaphanous related formin 3 (DIAPH3) promotes tumorigenesis in hepatocellular carcinoma and lung adenocarcinoma, suggesting that DIAPH3 may be a target for tumor therapy. To date, there have been no reports on the function of DIAPH3 in osteosarcoma. DIAPH3 protein expression in osteosarcoma tissues and healthy bone tissues adjacent to cancer cells was examined by immunohistochemical staining. DIAPH3 mRNA expression correlates with overall survival and reduced disease-free survival. DIAPH3 protein is upregulated in osteosarcoma tissues, and its expression is significantly associated with tumor size, tumor stage, node metastasis, and distant metastasis. Functional in vitro experiments revealed that DIAPH3 knockdown suppressed cell proliferation and suppressed cell migration and invasion of osteosarcoma cell lines MG-63 and HOS. Functional experiments demonstrated that DIAPH3 knockdown inhibited subcutaneous tumor growth and lung metastasis in vivo. In conclusion, DIAPH3 expression can predict the clinical outcome of osteosarcoma. In addition, DIAPH3 is involved in the proliferation and metastasis of osteosarcoma, and as such, DIAPH3 may be a potential therapeutic target for osteosarcoma.

Abstract 672: Prognostic significance of urokinase plasminogen activator system in a naturally occurring model of osteosarcoma

Abstract Introduction Human pediatric and canine osteosarcoma (OSA) are both highly metastatic primary bone cancers that share molecular and clinical similarities. Urokinase plasminogen activator (uPA) and its receptor uPAR promote invasion, metastasis and angiogenesis in multiple cancer types and have been shown to play critical roles in malignant characteristics of OSA in vitro, yet their clinical significance has not been investigated in OSA patients. Methods Serum samples of dogs with naturally-occurring appendicular OSA (n=26) and clinically healthy dogs (n=6) were analyzed using commercially available canine-specific ELISA and uPA activity kits. Dogs with OSA that presented without metastatic disease at diagnosis and received treatment with amputation plus adjuvant carboplatin chemotherapy were included. Survival data and patient characteristics were obtained from medical records. Differences in serum uPA/uPAR results between the groups was tested by t test or Wilcoxan rank sum test. Kaplan Meier survival curve and log rank test were used for survival analysis in the OSA cohort after subgrouping by X-Tyle software. Well-established prognostic factors and serum uPA/uPAR results were tested for prognostic significance by cox regression analysis in both univariate and multivariate analyses. Results Dogs with OSA had significantly higher serum uPA (p=0.008; 1905.4 vs 1439.5 pg/mL) but not uPAR (p=0.866) levels compared to healthy dogs. OSA-bearing dogs with high serum uPA had significantly shorter progression-free (p=0.003; 94 vs 266 days) and overall survival (p=0.01; 123 vs 338 days) than dogs with low uPA level. Similarly, dogs with high serum uPA activity had significantly shorter progression-free survival (p=0.043; 227 vs 266 days). Serum uPAR was not prognostic in this cohort. Multivariate analysis revealed increased serum uPA level (p=0.024) as a poor prognostic indicator in addition to other established prognostic factors in dogs such as humeral location (p=0.022) and increased monocyte count (p=0.015). Conclusions Circulating uPA quantity and activity are increased in canine OSA patients with more aggressive disease, indicating importance of the uPA system in OSA disease progression. While an OSA-associated increase in circulating uPAR was not evident, this receptor likely contributes to OSA malignancy subsequent to the increased uPA ligand production. Serum uPA may be an attractive prognostic biomarker in OSA and the up-regulated uPA/uPAR axis should be further investigated as a potential therapeutic target for this disease. Citation Format: Arata Matsuyama, Anthony J. Mutsaers. Prognostic significance of urokinase plasminogen activator system in a naturally occurring model of osteosarcoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 672.

MiRNA identification by nuclease digestion in ELISA for diagnosis of osteosarcoma.

Osteosarcoma is a bone cancer formed by the cells of the bone. Children, young adults, and teens are highly affected by osteosarcoma. Early identification of osteosarcoma is mandatory to improve the treatment and increase the lifespan of the patients. MicroRNA-195 (miR-195) was shown to be a suitable biomarker for osteosarcoma, and the present study describes a sensitive method of miR-195 identification by nuclease digestion in ELISA to detect and quantify the level of miR-195. S1 nuclease catalyzed endo- and exonucleolytic digestion of single-stranded (ss) RNA and DNA on ELISA polystyrene substrate, which helped to identify duplexed miR-195. This method selectively and specifically identified miR-195 without any biofouling interactions and reached the limit of detection at 10fM within the range from 10fM to 10nM. Due to complete digestion of ssDNA, single- and triple-mismatched sequences failed to increase the ELISA signal, indicating specific miRNA detection. Furthermore, human serum spiked with miR-195 did not interfere with the detection, confirming selective identification. This method identified miR-195 at a lower level and will help to diagnose earlier stages of osteosarcoma.

M6A regulators are associated with osteosarcoma metastasis and have prognostic significance: A study based on public databases.

Background:Osteosarcoma represents the most common malignant bone tumor with high metastatic potential and inferior prognosis. RNA methylation (N6-methyladenosine [m6A]) is a prevalent RNA modification that epigenetically influences numerous biological processes including tumorigenesis. This study aims to determine that m6A regulators are significant biomarkers for osteosarcoma, and establish a prognostic model to predict the survival of patients.Methods:In this study, we comprehensively analyzed the underlying associations between m6A regulators’ mRNA expressions and metastasis as well as prognosis of osteosarcoma patients in the Cancer Genome Atlas. Multivariate Cox-regression analysis was used to screen regulators that were significantly associated with overall survival of osteosarcoma patients. Least absolute shrinkage and selection operator (LASSO) Cox-regression analysis was used for constructing m6A regulator-based osteosarcoma prognostic signature.Results:Some of the regulators exhibited aberrant mRNA levels between osteosarcoma samples with and without metastasis. Multivariate Cox-regression analysis identified several regulators with potential prognostic significance. A risk score formula consisted of methyltransferase-like 3, YTH domains of Homo sapiens, and fat mass and obesity-associated protein was obtained through which patients could be prognostically stratified independently of potential confounding factors. The signature was also significantly associated with the metastatic potential of osteosarcoma. All the analyses could be well reproduced in another independent osteosarcoma cohort from the Gene Expression Omnibus.Conclusions:In conclusion, this study first revealed potential roles of m6A regulators in osteosarcoma metastasis and prognosis, which should be helpful for its clinical decision-making.

TRIM68, PIKFYVE, and DYNLL2: The Possible Novel Autophagy- and Immunity-Associated Gene Biomarkers for Osteosarcoma Prognosis

IntroductionOsteosarcoma is among the most common orthopedic neoplasms, and currently, there are no adequate biomarkers to predict its prognosis. Therefore, the present study was aimed to identify the prognostic biomarkers for autophagy-and immune-related osteosarcoma using bioinformatics tools for guiding the clinical diagnosis and treatment of this disease.Materials and MethodsThe gene expression and clinical information data were downloaded from the Public database. The genes associated with autophagy were extracted, followed by the development of a logistic regression model for predicting the prognosis of osteosarcoma using univariate and multivariate COX regression analysis and LASSO regression analysis. The accuracy of the constructed model was verified through the ROC curves, calibration plots, and Nomogram plots. Next, immune cell typing was performed using CIBERSORT to analyze the expression of the immune cells in each sample. For the results obtained from the analysis, we used qRT-PCR validation in two strains of human osteosarcoma cells.ResultsThe screening process identified a total of three genes that fulfilled all the screening criteria. The survival curves of the constructed prognostic model revealed that patients with the high risk presented significantly lower survival than the patients with low risk. Finally, the immune cell component analysis revealed that all three genes were significantly associated with the immune cells. The expressions of TRIM68, PIKFYVE, and DYNLL2 were higher in the osteosarcoma cells compared to the control cells. Finally, we used human pathological tissue sections to validate the expression of the genes modeled in osteosarcoma and paracancerous tissue.ConclusionThe TRIM68, PIKFYVE, and DYNLL2 genes can be used as biomarkers for predicting the prognosis of osteosarcoma.

A panel of eight mRNA signatures improves prognosis prediction of osteosarcoma patients.

Genetic alterations are vital to the progression of osteosarcoma carcinoma. The present study investigated a panel of gene signatures that could evaluate prognosis in osteosarcoma based on data from the Therapeutically Applicable Research To Generate Effective Treatments initiative. Osteosarcoma messenger RNA (mRNA) profiles and clinical data were downloaded from the therapeutically applicable research to generate effective treatments database. Patients with osteosarcoma were divided into two groups based on findings at diagnosis: with and without metastasis. Differentially expressed mRNAs were compared and analyzed between groups. Univariate and multivariate Cox regression analyses identified a set of eight mRNAs with the ability to classify patients into high-risk and low-risk groups with significantly different overall survival times. Further analysis indicated that the eight-mRNA signature was an independent prognostic factor after adjusting for other clinical factors. Receiver operating characteristic curve analysis demonstrated a good performance of the eight-mRNA signature. Further, the biological processes and signaling pathways of the eight-mRNA signature were reviewed using Gene Ontology and Kyoto Encyclopedia of Genes and Genomes resources. Finally, the results of the TCGA analysis were verified by other cohorts from Gene Expression Omnibus database. The identification of an eight-mRNA signature not only provides a prognostic biomarker of osteosarcoma but also offers the potential of novel therapeutic targets for its treatment.

Increased expression of microRNA-26a-5p predicted a poor survival outcome in osteosarcoma patients: An observational study.

MicroRNA (miR)-26a-5p is an oncogene significantly associated with osteosarcoma. We try to evaluate expression of circulating miR-26a-5p in osteosarcoma patients and evaluate its significance.A total of 243 consecutive osteosarcoma patients and 96 healthy participates were enrolled. Circulating miR-26a-5p levels were evaluated by using real-time quantitative reverse transcription polymerase chain reactions (RT-PCR). The association between circulating miR-26a-5p level and survival outcomes was evaluated by univariate and multivariate analysis.Circulating miR-26a-5p levels in osteosarcoma patients was significantly higher than that of healthy volunteers (P < .05). Upregulated miR-26a-5p was significantly related to advanced cancer and metastasis (both P < .05). Moreover, patients with a high serum miR-26a-5p had a poorer overall survival than those with a low serum miR-26a-5p levels (P < .05). Circulating miR-26a-5p level also been showed as independent risk factor for osteosarcoma in multivariate analysis (hazard ratio [HR], 0.38; 95% confidence interval [CI]: 0.11–0.98; P < .01).Circulating miR-26a-5p was significantly upregulated in osteosarcoma patients and remarkably associated with poor prognosis, indicating that circulating miR-26a-5p might serve as a useful diagnostic and prognostic biomarker for osteosarcoma.

Inhibiting proliferation and metastasis of osteosarcoma cells by downregulation of long non-coding RNA colon cancer-associated transcript 2 targeting microRNA-143.

Osteosarcoma is a malignant bone tumor, which has a high incidence in children and adolescents. However, the pathogenesis of osteosarcoma remains unclear. Long noncoding RNA (lncRNA) is a new potential therapeutic target and diagnostic biomarker for osteosarcoma. Hence, the present study aimed to explore the effect of lncRNA colon cancer-associated transcript (CCAT2) on osteosarcoma and its potential underlying mechanisms. For this purpose, the proliferation of osteosarcoma cells was measured using the CCK-8 assay. The scratch-wound and cell invasion assays were used to determine the migration and invasion of osteosarcoma cells, respectively. LncRNA CCAT2 and microRNA (miR)-143 binding sites were identified by the dual-luciferase reporter assay. RNA and protein expression levels were detected by reverse-transcription quantitative PCR and western blotting, respectively. Downregulation of lncRNA CCAT2 inhibited the proliferation, migration, and invasion of osteosarcoma cells. The findings also revealed that miR-143 bound directly to lncRNA CCAT2. The expression of miR-143 was upregulated by the knockdown of lncRNA CCAT2. Downregulation of the FOS-like antigen 2 was also observed after knockdown of lncRNA CCAT2. The function of lncRNA CCAT2 in osteosarcoma cells was attenuated by co-transfection with anti-miR-143 oligodeoxyribonucleotide. In conclusion, downregulation of lncRNA CCAT2 inhibited the proliferation and metastasis of osteosarcoma cells by targeting miR-143. lncRNA CCAT2 was identified as a potential target for osteosarcoma treatment.

Long noncoding RNA CRNDE functions as a diagnostic and prognostic biomarker in osteosarcoma, as well aspromotes itsprogression via inhibition of miR-335-3p.

This study was performed to evaluate the diagnostic and prognostic value, as well as the role of long-chain noncoding RNA (lncRNA) colorectal neoplasia differentially expressed (CRNDE) in osteosarcoma (OS). A quantitative real-time polymerase chain reaction assay was to determine lncRNA CRNDE and microRNA-335-3p(miR-335-3p) expressions. The Kaplan-Meier analysis was to analyze the relationship between lncRNA CRNDE expression and survival in patients with OS. Receiver operating characteristic curves were to evaluate the diagnostic value of lncRNA CRNDE in OS. Bioinformatics analysis and luciferase reporter assays were used to predict and confirm the relationship between lncRNA CRNDE and miR-335-3p. Cell counting Kit-8 and transwell migration assays assessed the role of lncRNA CRNDE and miR-335-3p in OS cells. lncRNA CRNDE expression was upregulated and miR-355-3p expression was downregulated in OS. In patients with OS, low lncRNA CRNDE expression demonstrated higher overall survival, whereas high lncRNA CRNDE expression was an independent poor prognostic factor. Furthermore, increased lncRNA CRNDE expression was associated with distant metastasis and the tumor-node-metastasisstage in patients with OS, which can be considered as an independent diagnostic biomarker in OS. We revealed that miR-335-3p was the target of lncRNA CRNDE. It also demonstrated that knockdown of lncRNA CRNDE inhibited OS cell proliferation, migration, and invasion,and inhibition of miR-355-3p promoted this effect. Finally, miR-335-3p partially mediated the stimulatory effects of lncRNA CRNDE in OS. We demonstrated that lncRNA CRNDE is a potential diagnostic and prognostic biomarker for OS, and the lncRNA CRNDE/miR-335-3p axis participates in OS progression.