ABSTRACT
Osteosarcoma is the most common malignant tumor in adolescent bone malignancies. It has the characteristics of a high metastasis rate, high mortality and poor prognosis. As a subclass of endogenous noncoding RNAs, circRNAs have been identified to be related to the occurrence, development and prognosis of different kinds of cancers, but the mechanism of their effect on osteosarcoma is not clear. In the present study, we identified a novel circRNA, hsa_circ_0087302, by RNA-seq, and we found that it was expressed at low levels in osteosarcoma. Using RT-PCR, we confirmed that the expression of hsa_circ_0087302 in osteosarcoma cells was lower than that in osteoblasts. Functional validation experiments revealed that hsa_circ_0087302 overexpression inhibited proliferation, cell cycle, migration, and invasion in osteosarcoma cells. Furthermore, Western blotting experiments demonstrated that hsa_circ_0087302 affected the expression of cell cycle- and Wnt/ß-catenin signaling pathway-related proteins. For the first time, we identified that hsa_circ_0087302 may affect the malignant biological behavior of osteosarcoma cells through the Wnt/ß-catenin signaling pathway. In summary, hsa_circ_0087302 may provide a new direction for the diagnosis and treatment of osteosarcoma.
Subject(s)
Bone Neoplasms , MicroRNAs , Osteosarcoma , Adolescent , Cell Line, Tumor , Cell Movement , Cell Proliferation , Gene Expression Regulation, Neoplastic , Humans , RNA, Circular , Wnt Signaling PathwayABSTRACT
Background: Osteosarcoma (OS) is the most common primary malignant bone tumour in children and adolescents, with rapid growth, frequent metastasis, and a poor prognosis, but its pathogenesis has not been fully elucidated. Exploring the pathogenesis of OS is of great significance for improving diagnoses and finding new therapeutic targets. Methods: Differentially expressed circRNAs (DECs), miRNAs (DEMs), methylated DNA sites (DMSs), and mRNAs (DEGs) were identified between OS and control cell lines. GSEA of DEGs and functional enrichment analysis of methylated DEGs were carried out to further identify potential biological processes. Online tools were used to predict the miRNA binding sites of DECs and the mRNA binding sites of DEMs, and then construct a circRNA-miRNA-mRNA network. Next, an analysis of the interaction between methylated DEGs was performed with a protein-protein interaction (PPI) network, and hub gene identification and survival analysis were carried out. The expression pattern of circRNA-miRNA-mRNA was validated by real-time PCR. Results: GSEA and functional enrichment analysis indicated that DEGs and methylated DEGs are involved in important biological processes in cancer. Hsa_circ_0001753/has_miR_760/CD74 network was constructed and validated in cell lines. Low expression levels of CD74 are associated with poor overall survival times and show good diagnostic ability. Conclusion: Methylated DEGs may be involved in the development of OS, and the hsa_circ_0001753/has_miR_760/CD74 network may serve as a target for the early diagnosis of and targeted therapy for OS.
ABSTRACT
BACKGROUND: Breast cancer (BRCA) is the most common cancer in women, while the bones are one of the most common sites of metastasis. Although new diagnostic methods or radiation or chemotherapies and targeted therapies have made huge advances, the occurrence of bone metastasis is also linked with poorer survival. Enhancer RNAs (eRNAs) have been demonstrated to participate in the progression of tumorigenesis and metastasis. However, the role of eRNAs in BRCA bone metastasis remains largely unclear. METHOD: Gene expression profiling of 1,211 primary BRCA and 17 bone metastases samples were retrieved from The Cancer Genome Atlas (TCGA) database, and the significant prognostic eRNAs were identified by Cox regression and least absolute shrinkage and selection operator (LASSO) regression. The acceptable accuracy and discrimination of the nomogram were indicated by the receiver operating characteristic (ROC) and the calibration curves. Then target genes of eRNA, immune cell percentage by CIBERSORT analysis, immune genes by single-sample gene set enrichment analysis (ssGSEA), hallmark of cancer signaling pathway by gene set variation analysis (GSVA), and reverse phase protein array (RPPA) protein chip were used to build a co-expression regulation network and identified the key eRNAs in bone metastasis of BRCA. Finally, Cell Counting Kit-8 (CCK8) assay, cell cycle assay, and transwell assay were used to study changes in cell proliferation, migration, and invasiveness. Immunoprecipitation assay and Western blotting were used to test the interaction and the regulation signaling pathways. RESULTS: The 27 hub eRNAs were selected, and a survival-related linear risk assessment model with a relatively high accuracy (area under curve (AUC): 0.726) was constructed. In addition, seven immune-related eRNAs (SLIT2, CLEC3B, LBPL1, FRY, RASGEF1B, DST, and ITIH5) as prognostic signatures for bone metastasis of BRCA were further confirmed by LASSO and multivariate Cox regression and CIBERSORT analysis. Finally, in vitro assay demonstrated that overexpression of SLIT2 reduced proliferation and metastasis in BRCA cells. Using high-throughput co-expression regulation network, we identified that SLIT2 may regulating P38 MAPK/c-Fos signaling pathway to promote the effects of metastasis. CONCLUSION: Based on the co-expression network for bone metastasis of BRCA, we screened key eRNAs to explore a prognostic model in predicting the bone metastasis by bioinformatics analysis. Besides, we identified the potential regulatory signaling pathway of SLIT2 in BRCA bone metastasis, which provides a promising therapeutic strategy for metastasis of BRCA.
ABSTRACT
Osteosarcoma (OS) is the most common type of malignant bone tumor that affects children and adolescents. Still, the cellular and molecular mechanisms driving the development of this disease remain poorly understood. In this study, numerous dysregulated lncRNAs were identified by RNA-seq. As a result, we were able to find a novel lncRNA Lnc-MAP6-1:3 which is highly expressed in osteosarcoma. Using a set of approaches including gene knockdown, RT-PCR, oncogenic function assay and western blotting, we observed that knockdown of Lnc-MAP6-1:3 expression suppressed cell proliferation and colony formation, and promoted apoptosis in vitro. For the first time, we have identified that Lnc-MAP6-1:3 potentially influence the malignant behavior of osteosarcoma via Bax/Bcl-2 and Wnt/ß-catenin signaling pathways. Henceforth, Lnc-MAP6-1:3 may provide a new molecular route of research and therapeutic applications for the diagnosis and treatment of osteosarcoma.
