Biology and pathogenesis of human osteosarcoma

Abstract

Osteosarcoma (OS) is a bone tumor of mesenchymal origin, most frequently occurring during the rapid growth phase of long bones, and usually located in the epiphyseal growth plates of the femur or the tibia. Its most common feature is genome disorganization, aneuploidy with chromosomal alterations, deregulation of tumor suppressor genes and of the cell cycle, and an absence of DNA repair. This suggests the involvement of surveillance failures, DNA repair or apoptosis control during osteogenesis, allowing the survival of cells which have undergone alterations during differentiation. Epigenetic events, including DNA methylation, histone modifications, nucleosome remodeling and expression of non-coding RNAs have been identified as possible risk factors for the tumor. It has been reported that p53 target genes or those genes that have their activity modulated by p53, in addition to other tumor suppressor genes, are silenced in OS-derived cell lines by hypermethylation of their promoters. In osteogenesis, osteoblasts are formed from pluripotent mesenchymal cells, with potential for self-renewal, proliferation and differentiation into various cell types. This involves complex signaling pathways and multiple factors. Any disturbance in this process can cause deregulation of the differentiation and proliferation of these cells, leading to the malignant phenotype. Therefore, the origin of OS seems to be multifactorial, involving the deregulation of differentiation of mesenchymal cells and tumor suppressor genes, activation of oncogenes, epigenetic events and the production of cytokines.

Keywords: bone tumor; osteosarcoma; osteosarcoma biology; osteosarcoma pathogenesis.

Figure 1.

Epigenetic events which may contribute to the initiation and progression of OS. Target genes or those which have p53-modulated activities, including GDKN1A, HIC, GADD45, RASSF1A, TIMP3 and DAPK1, are silenced when hypermethylated. An inverse situation occurs with SEMA4D, RAF1 and PAK1 genes when they are hypomethylated which results in overexpression. However, both conditions may equally favor the development of OS. Silencing by hypermethylation of the RASSF1A gene leads to overexpression of the MDM2 gene, whose product promotes p53 degradation, which in turn results in uncontrolled cell cycle and absence of DNA repair and apoptosis inhibition. The hypomethylation of the IRX1 gene promotes activation of the CXCL14/NFkB signaling pathway, whereas the activation of the cMyc and PI3/Akt signaling pathways results in suppression of the GADD45 gene encoding the 5-hmC production, impeding the demethylation of other genes, which also favors tumor development. Mutation or methylation of the CDKN2A gene reduces the production of p16INK4a, which leads to overexpression of CDK4 and inactivation of pRB. It can also reduce the production of p14INK4a, which in turn suppresses p14ARF, resulting in high Mdm2 levels which degrades p53. Both mechanisms result in an uncontrolled cell cycle, lack of DNA repair and apoptosis inhibition, favoring tumor initiation and progression. OS, osteosarcoma.

Figure 2.

Participation of miRs in OS development. The suppression or activation of miR expression is an epigenetic event which may be involved in OS development. Suppression of miR-200b results in overexpression of the ZEB1 gene, whose product suppresses IL-2 gene expression, whereas the suppression of miR-101 results in overexpression of the ROCK1 gene which actuates in the PI3K/AKT and JAK/STAT signaling pathways. Both cases may increase the risk of initiation and progression of OS, since they cause impairment of the immune response. Suppression of miR-3928 increases the expression of the ERBB3, IL-6R and CDK6 genes, which may favor tumor development, promoting cell proliferation, uncontrolled cell cycle and apoptosis inhibition. The suppression of miR-143 results in the expression of versican and proteoglycan of extracellular matrix, favoring tumor progression. However, the expression of miR-17-92 increases the expression of miR-20a, which in turn reduces Fas expression, a cell death receptor, and inhibits apoptosis, whereas miR-574-3p expression suppresses expression of the SMAD4 tumor suppressor gene, which leads to increased cell proliferation and apoptosis inhibition. Therefore, both mechanisms may increase the risk of OS. miR, microRNA; OS, osteosarcoma.

Figure 3.

Role of lncRNAs and circRNAs in osteosarcoma. In general, lncRNAs and circRNAs act as sponges for miRNAs, causing its inactivation and favoring tumor development. Therefore, MAULTI acts by suppressing miR-129-5p which leads to activation of the RET-AKT signaling pathway; HOXD-AS1 suppresses the tumor suppressing action of p57 protein; TAG1 suppresses miR-212-3p, whereas SNHG1 suppresses miR-101-3p, which results in activation of ROCK1 gene expression. All these events favor the development of osteosarcoma. However, lncRNA SRA1 acts by suppressing miR-208a, inhibiting its tumorigenic action. Additionally, has-circRNA-101801 circRNA acts by suppressing miRNAs: has-miR-338-3p, has-miR-370-3p and has-miR-877-3p resulting in increased expression of HIF-1 and VEGF, and PI3K-Akt signaling pathway activation, thus favoring angiogenesis. Furthermore, circRNA TAD2A suppresses miR-203a-3p, which leads to increased CREB expression. This favors tumor development in both cases. circRNA, circular RNA; lncRNA, long non-coding RNA.

Figure 4.

Role of cytokines in human OS. Certain cytokines may contribute to OS development by activating cell signaling pathways, or by interfering in the differentiation process of MSCs. IL-6 activates the expression of JAK2 signal transducer, which in turn activates JAK-STAT signaling pathway that induces the differentiation of MSCs into osteoclasts. This leads to the production of VEGF and of proteins which act on bone resorption and induces angiogenesis, contributing to tumor development. TGFβ activates the MAPK pathway, which in turn activates the TGF-β/SMAD-2/-3 pathway, resulting in miR-143 suppression, with a consequent increase in versican expression of the extracellular matrix, thus contributing to tumor progression. TGF-β also acts on MSCs, inducing the production of IL-6, VEGF and additional TGF-β. IL-6 together with TNFα induces inflammation and the production of MMPs which (in synergistic action) inhibits MSC differentiation, increasing the risk of initiating OS. Furthermore, TNFα also increases the expression of CXC4 chemokine and its CXCR4 receptor, a condition which also favors tumor development. TNFα together with IL1β activates the production of IL-34 which binds to the M-CSF receptor, promoting growth and survival of myeloid cells and macrophage polarization for the M2 profile with recruitment of these cells into the tumor environment, therefore contributing to its progression. The STST3 transcription factor, upon being phosphorylated, activates cytokine IL-17 production, which (if connected to its IL-17RA receptor) stimulates VEGF, MM9 and CXCR4 production and promotes angiogenesis, thus contributing to tumor progression and formation of metastases. MSCs, mesenchymal stem cells; OS, osteosarcoma.