Osteosarcoma, the most common primary bone malignancy, occurs most frequently in adolescents with a peak of incidence at 11–15 years. Melatonin, an indole amine hormone, shows a wide range of anticancer activities. The decrease in melatonin levels simultaneously concurs with the increase in bone growth and the peak age distribution of osteosarcoma during puberty, so melatonin has been utilized as an adjunct to chemotherapy to improve the quality of life and clinical outcomes. While a large amount of research has been conducted to understand the complex pleiotropic functions and the molecular and cellular actions elicited by melatonin in various types of cancers, a few review reports have focused on osteosarcoma. Herein, we summarized the anti-osteosarcoma effects of melatonin and its underlying molecular mechanisms to illustrate the known significance of melatonin in osteosarcoma and to address cellular signaling pathways of melatonin in vitro and in animal models. Even in the same kind of osteosarcoma, melatonin has been sparingly investigated to counteract tumor growth, apoptosis, and metastasis through different mechanisms, depending on different cell lines. We highlighted the underlying mechanism of anti-osteosarcoma properties evoked by melatonin, including antioxidant activity, anti-proliferation, induction of apoptosis, and the inhibition of invasion and metastasis. Moreover, we discussed the drug synergy effects of the role of melatonin involved and the method to fortify the anti-cancer effects on osteosarcoma. As a potential therapeutic agent, melatonin is safe for children and adolescents and is a promising candidate for an adjuvant by reinforcing the therapeutic effects and abolishing the unwanted consequences of chemotherapies.

Osteosarcoma, the most prevalent malignant bone tumor in the pediatric age group, is responsible for the great majority of cancer-associated deaths owing to its highly metastatic potential. The anti-metastatic effects of the new curcumin analogue L48H37 in human osteosarcoma are still unknown; hence, we investigated whether L48H37 represses human osteosarcoma cells’ biological behavior of migratory potential and invasive activities and attempted to delve into its underlying mechanisms. L48H37 up to 5 μM inhibited, without cytotoxicity, the motility, migration, and invasion of human osteosarcoma U2OS and MG-63 cells. In U2OS cells, the human protease array revealed an obvious decrease in urokinase plasminogen activator (uPA) expression after L48H37 treatment, and L48H37 actually reduced the level, protein and mRNA expression, and promoter activity of uPA dose-dependently. L48H37 decreased the phosphorylation of STAT3, JAK1, JAK2, and JAK3 in U2OS cells, but did not affect the phosphorylation of ERK, JNK, p38, and Akt. Using colivelin, an activator of STAT3, the L48H37-induced decrease in uPA and migratory potential could be countered as expected. Collectively, L48H37 represses the invasion and migration capabilities of U2OS and MG-63 cells by the suppression of uPA expression and the inhibition of JAK/STAT signaling. These results suggest that L48H37 may be a potential candidate for anti-metastatic treatment of human osteosarcoma.

Osteosarcoma is a highly common malignant bone tumor. Its highly metastatic properties are the leading cause of mortality for cancer. Niclosamide, a salicylanilide derivative, is an oral antihelminthic drug of known anticancer potential. However, the effect of niclosamide on osteosarcoma cell migration, invasion and the mechanisms underlying have not been fully clarified. Therefore, this study investigated niclosamide’s underlying pathways and antimetastatic effects on osteosarcoma. In this study, U2OS and HOS osteosarcoma cell lines were treated with niclosamide and then subjected to assays for determining cell migration ability. The results indicated that niclosamide, at concentrations of up to 200 nM, inhibited the migration and invasion of human osteosarcoma U2OS and HOS cells and repressed the transforming growth factor beta-induced protein (TGFBI) expression of U2OS cells, without cytotoxicity. After TGFBI knockdown occurred, cellular migration and invasion behaviors of U2OS cells were significantly reduced. Moreover, niclosamide significantly decreased the phosphorylation of ERK1/2 in U2OS cells and the combination treatment of the MEK inhibitor (U0126) and niclosamide resulted in the intensive inhibition of the TGFBI expression and the migratory ability in U2OS cells. Therefore, TGFBI derived from osteosarcoma cells via the ERK pathway contributed to cellular migration and invasion and niclosamide inhibited these processes. These findings indicate that niclosamide may be a powerful preventive agent against the development and metastasis of osteosarcoma.

Higher neutrophil-derived cytokine lipocalin-2 (LCN2) expression possesses a versatile role in a myriad of cancers, but little is known about the role of LCN2 on osteosarcoma metastasis. In this study, we demonstrated that higher LCN2 inhibited cellular motility, migration, and invasion of osteosarcoma cells. Moreover, using RNA sequencing technology, we found that LCN2 repressed MET gene expression in U2OS cells. Manipulation of LCN2 levels influenced the migratory potential of osteosarcoma cells as cellular migration was enhanced by transfecting with vectors containing a constitutively active LCN2 cDNA and recombinant human LCN2. Moreover, the phosphorylation of mitogen-activated protein kinases/extracellular signal-regulated kinase (ERK) kinase (MEK) 1/2 and ERK 1/2 was decreased by LCN2 knockdown. Furthermore, the use of ERK inhibitor (U0126) and activator (tBHQ) confirmed that the pharmaceutic inhibition of MEK–ERK augmented the LCN2-mediated MET suppression and migration of U2OS and HOS cells. Conclusively, LCN2 inhibits osteosarcoma cell metastasis by suppressing MET via the MEK–ERK pathway.

Osteosarcoma is the most common primary bone malignancy in teenagers and continues to confer a generally poor prognosis due to its highly metastatic potential. Poor solubility in water and instability of curcumin limits its bioavailability for use in the adjuvant situation to improve the prognosis and the long-term survival of patients with osteosarcoma. To further obtain information regarding the apoptosis induced by a new curcumin analog, GO-Y078, in human osteosarcoma cells, flow cytometric analysis, annexin V-FITC/PI apoptosis staining assay, human apoptosis array, and Western blotting were employed. GO-Y078 dose-dependently decreased viabilities of human osteosarcoma U2OS, MG-63, 143B, and Saos-2 cells and induced sub-G1 fraction arrest and apoptosis in U2OS and 143B cells. In addition to the effector caspase 3 and poly adenosine diphosphate-ribose polymerase, GO-Y078 significantly activated both initiators of extrinsic caspase 8 and intrinsic caspase 9, whereas cellular inhibitors of apoptosis 1 (cIAP-1) and X-chromosome-linked IAP (XIAP) in U2OS and 143B cells were significantly repressed. Moreover, GO-Y078 increased phosphorylation of extracellular signal-regulated protein kinases (ERK)1/2, c-Jun N-terminal kinases (JNK)1/2, and p38 in U2OS and 143B cells. Using inhibitors of JNK (JNK-in-8) and p38 (SB203580), GO-Y078′s increases in cleaved caspases 8, 9, and 3 could be expectedly suppressed, but they could not be affected by co-treatment with the ERK inhibitor (U0126). Altogether, GO-Y078 simultaneously induces both apoptotic pathways and cell arrest in U2OS and 143B cells through activating JNK and p38 signaling and repressing IAPs. These findings contribute to a better understanding of the mechanisms responsible for GO-Y078′s apoptotic effects on human osteosarcoma cells.

Metastatic osteosarcoma often results in poor prognosis despite the application of surgical en bloc excision along with chemotherapy. HO-3867 is a curcumin analog that induces cell apoptosis in several cancers, but the apoptotic effect and its mechanisms on osteosarcoma cells are still unknown. After observing the decrease in cellular viability of three human osteosarcoma U2OS, HOS, and MG-63 cell lines, and the induction of cellular apoptosis and arrest in sub-G1 phase in U2OS and HOS cells by HO-3867, the human apoptosis array showed that heme oxygenase (HO)-1 and cleaved caspase-3 expressions had significant increases after HO-3867 treatment in U2OS cells and vice versa for cellular inhibitors of apoptosis (cIAP)1 and X-chromosome-linked IAP (XIAP). Western blot analysis verified the results and showed that HO-3867 activated the initiators of both extrinsic caspase 8 and intrinsic caspase 9, and significantly increased cleaved PARP expression in U2OS and HOS cells. Moreover, with the addition of HO-3867, ERK1/2, and JNK1/2 phosphorylation were increased in U2OS and HOS cells. Using the inhibitor of JNK (JNK in 8), HO-3867’s increases in cleaved caspases 3, 8, and 9 could be expectedly suppressed, indicating that JNK signaling is responsible for both apoptotic pathways, including extrinsic and intrinsic, in U2OS and HOS cells caused by HO-3867. Through JNK signaling, HO-3867 has proven to be effective in causing both extrinsic and intrinsic apoptotic pathways of human osteosarcoma cells.