• Asian Pacific Journal of Cancer Prevention
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• 제14권4호
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• pp.2311-2315
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• 2013

Objective: To investigate the expression level of TUG1 and one of its transcript variants (n377360) in osteosarcoma cells and assess the role of TUG1 in proliferation and apoptosis in the U2OS cell line. Methods: TUG1 and n377360 expression levels in patients with osteosarcomas and the U2OS human osteosarcoma cell line were evaluated using real-time quantitative PCR. U2OS cells were transected with TUG1 and n377360 siRNA or non-targeting siRNA. MTS was performed to assess the cell proliferation and flow cytometry was applied to analyze apoptosis. Results: We found significantly higher TUG1 and n377360 expression levels in osteosarcoma tissues compared with matched non-tumorous tissues. In line with this, suppression of TUG1 and n377360 expression by siRNA significantly impaired the cell proliferation potential of osteosarcoma cells. Furthermore, inhibition of TUG1 expression significantly promoted osteosarcoma cell apoptosis. Conclusions: The overexpression of TUG1 and n377360 in osteosarcoma specimens and the functional role of TUG1 and n377360 regarding cell proliferation and apoptosis in an osteosarcoma cell line provided evidence that the use of TUG1 or n377360 may be a viable but an as yet unexplored therapeutic strategy in tumors that over express these factors.

• Asian Pacific Journal of Cancer Prevention
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• 제14권2호
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• pp.1101-1106
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• 2013

Background: Osteosarcoma is the most common primary bone tumor in childhood and adolescence. Carboplatin, a platinum-derived agent, is used as neoadjuvant chemotherapy for pediatric osteosarcoma because of its anti-tumor activity and had low toxicity as compared to cisplatin. Objective: To determine demographic data, prognostic factors and outcome of childhood osteosarcoma treated with a carboplatin-based chemotherapeutic protocol at Chiang Mai University. Method: A retrospective analysis was conducted on 34 osteosarcoma patients aged less than 18 years and treated between 2003 and 2011. Results: Overall limb-salvage and amputation rates were 23.5% and 70.6%, respectively. With the mean follow-up time of 29.5 months (1.5-108.9), the Kaplan-Meier analysis for 3-year disease-free survival (DFS) and 3-year overall survival (OS) were $20.2{\pm}7.7%$ and $47.1{\pm}9.5%$ respectively. Patients who had initial pulmonary metastasis were at significantly greater risk for developing recurrence (p=0.02, OR=7; 1.2-40.1) and had a tendency to have lower 3-year OS compared to those without initial pulmonary metastasis ($28.1{\pm}13%$, $63.1{\pm}12.3%$, respectively, p=0.202). On univariate analysis, age at diagnosis >14 years and patients who were declined surgery were significantly associated with lower 3-year OS (p=0.008 and <0.05, respectively). However, age at diagnosis, sex, tumor size and histological subtypes were not found to significantly affect recurrence or survival. Conclusions: In our study, the survival rate was far lower than those reported from developed countries. These might indicate the ineffectiveness of carboplatin in combination with doxorubicin as frontline treatment of pediatric osteosarcoma, especially in those with initial pulmonary metastasis. Refinement in risk and treatment stratification and dose intensification for pediatric osteosarcoma constitutes a future challenge to improve outcomes, especially in metastatic patients who may need a more intensive regimen.

• Asian Pacific Journal of Cancer Prevention
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• 제15권22호
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• pp.9823-9829
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• 2014

Background: High-dose methotrexate (HD-MTX) is recognized as an efficient component of therapy against pediatric osteosarcoma in combination with other drugs such as cisplatin (CDP), carboplatin (CBDCA), doxorubicin (ADM), etoposide (VP-16) and ifosfamide (IFO). Objectives: To demonstrate the feasibility and effectiveness of the HD-MTX/CDP/DOX/VP-16/IFO [MTX(+)] protocol comparable to CDP/ADM/CBDCA/IFO [MTX(-)] for treating childhood osteosarcoma at Ramathibodi Hospital (1999-2014). Materials and Methods: A retrospective analysis was conducted of osteosarcoma patients aged less than 18 years treated with two chemotherapeutic regimens between 1999 and 2014. A total of 45 patients received the MTX(-) and 21 the MTX(+) protocol. Results: Overall limb-salvage and amputation rate were 12.9% and 77.7%, respectively. Kaplan-Meier analysis results for 3-year disease free survival (DFS) and overall survival (OS) regardless of treatment regimens were $43.4{\pm}6.0%$ and $53.2{\pm}6.1%$ respectively. The 3-year DFS and OS were improved significantly with the MTX(+) protocol compared to MTX(-) protocol (p=0.010 and p=0.009, log rank test) [$69.8{\pm}10.5%$, $79.8{\pm}9.1%$ for MTX(+) and $31.1{\pm}6.9%$, $42.2{\pm}7.4%$ for MTX(-) protocol, respectively]. Patients with metastatic osteosarcoma treated with the MTX(+) protocol had statistically significant higher 3-year DFS and OS than those treated with the MTX(-) protocol ($66.7{\pm}13.6%$ and $15.0{\pm}8.0%$ for 3-year DFS, p=0.010, $73.3{\pm}13.2%$ and $20{\pm}8.9%$ for 3-year OS, p=0.006, respectively). The independent risk factors for having inferior 3-year DFS and OS were poor histological response (tumor necrosis <90%) and treatment with the MTX(-) protocol. The multivariate analysis identified only the treatment with the MTX(-) protocol as an independent predictor of inferior OS with a hazard ratio (HR) of 3.53 (95% confidence interval of 1.2-10.41, p=0.022). Conclusions: Our study demonstrated the tolerability, feasibility and efficacy of the HDMTX-based regimen improving the survival rate in pediatric osteosarcoma cases, in line with reports from developed countries.

• BMB Reports
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• 제46권2호
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• pp.113-118
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• 2013

TTRAP is a multi-functional protein that is involved in multiple aspects of cellular functions including cell proliferation, apoptosis and the repair of DNA damage. Here, we demonstrated that the lentivirus-mediated overexpression of TTRAP significantly inhibited cell growth and induced apoptosis in osteosarcoma cells. The ectopic TTRAP suppressed the growth and colony formation capacity of two osteosarcoma cell lines, U2OS and Saos-2. Cell apoptosis was induced in U2OS cells and the cell cycle was arrested at G2/M phase in Saos-2 cells. Exogenous expression of TTRAP in serum-starved U2OS and Saos-2 cells induced an increase in caspase-3/-7 activity and a decrease in cyclin B1 expression. In comparison with wild-type TTRAP, mutations in the 5'-tyrosyl-DNA phosphodiesterase activity of TTRAP, in particular $TTRAP^{E152A}$, showed decreased inhibitory activity on cell growth. These results may aid in clarifying the physiological functions of TTRAP, especially its roles in the regulation of cell growth and tumorigenesis.

• Asian Pacific Journal of Cancer Prevention
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• 제15권24호
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• pp.10797-10801
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• 2015

Background: To explore the molecular mechanisms of metastatic osteosarcoma (OS) by using the microarray expression profiles of metastatic and non-metastatic OS samples. Materials and Methods: The gene expression profile GSE37552 was downloaded from Gene Expression Omnibus database, including 2 human metastatic OS cell line models and 2 two non-metastatic OS cell line models. The differentially expressed genes (DEGs) were identified by Multtest package in R language. In addition, functional enrichment analysis of the DEGs was performed by WebGestalt, and the protein-protein interaction (PPI) networks were constructed by Hitpredict, then the signal pathways of the genes involved in the networks were performed by Kyoto Encyclopaedia of Genes and Genomes (KEGG) automatic annotation server (KAAS). Results: A total of 237 genes were classified as DEGs in metastatic OS. The most significant up- and down-regulated genes were A2M (alpha-2-macroglobulin) and BCAN (brevican). The DEGs were significantly related to the response to hormone stimulus, and the PPI network of A2M contained IL1B (interleukin), LRP1 (low-density lipoprotein receptor-related protein 1) and PDGF (platelet-derived growth factor). Furthermore, the MAPK signaling pathway and focal adhesion were significantly enriched. Conclusions: A2M and its interactive proteins, such as IL1B, LRP1 and PDGF may be candidate target molecules to monitor, diagnose and treat metastatic OS. The response to hormone stimulus, MAPK signaling pathway and focal adhesion may play important roles in metastatic OS.

• Clinical and Experimental Pediatrics
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• 제62권2호
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• pp.62-67
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• 2019

Purpose: We aimed to determine the prognostic significance of lymphocyte counts and the lymphocytemonocyte ratio (LMR) in pediatric patients with osteosarcoma. Methods: We retrospectively reviewed the medical records of 27 pediatric patients with localized extremity osteosarcoma, treated at the Korea Cancer Center Hospital between May 2002 and March 2016. Leukocyte counts and LMR before treatment and on day 14 (LMR14) of the first cisplatin-doxorubicin chemotherapy round were evaluated. Patients were dichotomized according to the median value of these parameters, and survival rates were compared. Results: The median age of the 27 patients was 9.9 years (range, 3.2-14.1 years) and tumor sites were: distal femur (n=14), proximal humerus (n=7), proximal tibia (n=2), proximal fibula (n=2), and elsewhere (n=2). Patients were followed up on for a median of 76.4 months (range, 4.5-174.7 months), and 5-year overall (OS) and event-free survival (EFS) rates were $66.0%{\pm}9.8%$ and $60.9%{\pm}9.7%$, respectively. Patients with a higher pretreatment lymphocyte count (${\geq}2,320/{\mu}L$) had better OS (90.9% vs. 46.2%, P=0.04) and EFS (83.9% vs. 38.5%, P=0.02). However, the day 14 lymphocyte count was not associated with survival. While no survival difference was observed between patients grouped according to pretreatment LMR (median value, 6.3), patients with a higher LMR14 (${\geq}5$) fared better than those with lower LMR14 (5-year OS: 83.3% vs. 46.3%, P=0.04). Conclusion: Pretreatment lymphocyte count and LMR during chemotherapy had prognostic significance in pediatric osteosarcoma patients. Further studies involving larger cohorts are necessary to validate our findings.

• Asian Pacific Journal of Cancer Prevention
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• 제15권4호
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• pp.1817-1822
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• 2014

Objective: The aim of this study was to screen for possible biomarkers of metastatic osteosarcoma (OS) using a DNA microarray. Methods: We downloaded the gene expression profile GSE49003 from Gene Expression Omnibus database, which included 6 gene chips from metastatic and 6 from non-metastatic OS patients. The R package was used to screen and identify differentially expressed genes (DEGs) between metastatic and non-metastatic OS patients. Then we compared the expression of DEGs in the two groups and sub-grouped into up-regulated and down-regulated, followed by functional enrichment analysis using the DAVID system. Subsequently, we constructed an miRNA-DEG regulatory network with the help of WebGestalt software. Results: A total of 323 DEGs, including 134 up-regulated and 189 down-regulated, were screened out. The up-regulated DEGs were enriched in 14 subcategories and most significantly in cytoskeleton organization, while the down-regulated DEGs were prevalent in 13 subcategories, especially wound healing. In addition, we identified two important miRNAs (miR-202 and miR-9) pivotal for OS metastasis, and their relevant genes, CALD1 and STX1A. Conclusions: MiR-202 and miR-9 are potential key factors affecting the metastasis of OS and CALD1 and STX1A may be possible targets beneficial for the treatment of metastatic OS. However, further experimental studies are needed to confirm our results.

• 대한골관절종양학회지
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• 제20권1호
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• pp.1-6
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• 2014

목적: 골육종에서 Connective Tissue Growth Factor (CTGF)의 발현 정도를 확인하고 발암기전에서의 역할을 살펴보고자 하였다. 대상 및 방법: 환자에서 수립한 23개의 골육종 세포주에서 CTGF의 발현 정도를 관찰하였으며, siRNA를 이용하여 CTGF의 발현을 억제한 후 세포침습과 세포 증식에서 CTGF의 역할을 분석하였다. 결과: 17명(74%)의 세포주에서 control 세포인 정상 골모세포보다 CTGF의 발현이 증가되어 있었다. CTGF의 발암기전에서의 역할을 관찰하기 위해 불멸화된 골육종 세포주 SaOS-2와 MG63에서 siRNA로 CTGF의 발현을 억제한 후 siRNA를 transfection한 세포에서 유의하게 세포침습이 억제되고 세포 증식이 억제되는 것을 관찰하였다. 결론: 골육종 세포주에서 CTGF의 발현이 높았고 세포침습, 세포 성장에 관여하는 바 CTGF가 골육종의 발암기전에서 중요한 역할을 하는 것으로 판단된다.

• Asian Pacific Journal of Cancer Prevention
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• 제14권2호
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• pp.795-799
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• 2013

Ku70/80 heterodimer is a central element in the nonhomologous end joining (NHEJ) DNA repair pathway, Ku80 playing a key role in regulating the multiple functions of Ku proteins. It has been found that the Ku80 protein located at telomeres is a major contributor to radiosensitivity in some telomerase positive human cancer cells. However, in ALT human osteosarcoma cells, the precise function in radiosensitivity and telomere maintenance is still unknown. The aim of this study was to investigate the effects of Ku80 depletion in the U2OS ALT cell line cell line. Suppression of Ku80 expression was performed using a vector-based shRNA and stable Ku80 knockdown in cells was verified by Western blotting. U2OS cells treated with shRNA-Ku80 showed lower radiobiological parameters (D0, Dq and SF2) in clonogenic assays. Furthermore, shRNA-Ku80 vector transfected cells displayed shortening of the telomere length and showed less expression of TRF2 protein. These results demonstrated that down-regulation of Ku80 can sensitize ALT cells U2OS to radiation, and this radiosensitization is related to telomere length shortening.

• 대한골관절종양학회지
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• 제13권2호
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• pp.88-95
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• 2007

목적: wild-type p53 단백질을 가지고 있는 U2OS 인간 골육종 세포주의 세포자멸사에서 MMP억제제와 doxorubicin의 작용을 비교하고자 하였다. 대상 및 방법: 배양한U2OS 세포주에 Doxorubicin, MMP억제제(MMPI III)를 따로 투여한 것과 두 약물을 동시에 투여한 것을 비교하였다. 또한 Doxorubicin의 작용이 Fas/FasL 경로를 통해 유발되는지 알아보기 위해 Fas 중화항체를 doxorubicin과 함께 투여하였다. 약물을 처리하여 배양한 세포에서 발생한 세포자멸사와 세포괴사를 확인하기 위해서 유세포 검사를 시행하였다. 결과: Doxorubicin을 처리한 세포가 처리하지 않은 세포보다 약물 농도에서 더 많은 세포괴사(p=0.000)를 보였다. 이에 반해 MMPI III를 처리한 세포는 약물 농도와 반응시간에 따른 세포자멸사 및 세포괴사에서 차이를 보이지 않았다. 두 약물을 동시에 투여한 군은 그렇지 않은 군에 비해 세포 자멸사 및 세포괴사에서 차이가 없었다. Doxorubicin에 Fas 중화항체를 추가한 군과 doxorubicin 단독 투여 군 사이에 세포자멸사 및 세포괴사에서 차이가 없었다. 결론: Doxorubicin과 같이 MMP 억제제를 골육종 치료에 사용하려면 wild-type p53 뿐만 아니라 wild-type p14를 가지고 있는 골육종 세포에 대한 검사가 필요할 것으로 사료된다.