Shui-er Zheng

Analysis of prognostic factors in 333 Chinese patients with high-grade osteosarcoma treated by multidisciplinary combined therapy

To investigate prognostic factors for long‐term outcomes in Chinese patients with high‐grade osteosarcoma of the extremities or trunk treated by multidisciplinary combined therapy.

Clinical analysis of Chinese limb osteosarcoma patients treated by two combinations of methotrexate, cisplatin, doxorubicin and ifosfamide.

Aims:  The objective of this study was to investigate the efficacy and toxic side effects of two combinations of methotrexate, cisplatin, doxorubicin and ifosfamide on treating Chinese osteosarcoma patients.

Pirarubicin inhibits multidrug-resistant osteosarcoma cell proliferation through induction of G2/M phase cell cycle arrest.

Aim:Pirarubicin (THP) is recently found to be effective in treating patients with advanced, relapsed or recurrent high-grade osteosarcoma. In this study, the effects of THP on the multidrug-resistant (MDR) osteosarcoma cells were assessed, and the underlying mechanisms for the disruption of cell cycle kinetics by THP were explored.Methods:Human osteosarcoma cell line MG63 and human MDR osteosarcoma cell line MG63/DOX were tested. The cytotoxicity of drugs was examined using a cell proliferation assay with the Cell Counting Kit-8 (CCK-8). The distribution of cells across the cell cycle was determined with flow cytometry. The expression of cell cycle-regulated genes cyclin B1 and Cdc2 (CDK1), and the phosphorylated Cdc2 and Cdc25C was examined using Western blot analyses.Results:MG63/DOX cells were highly resistant to doxorubicin (ADM) and gemcitabine (GEM), but were sensitive or lowly resistant to THP, methotrexate (MTX) and cisplatin (DDP). Treatment of MG63/DOX cells with THP (200–1000 ng/mL) inhibited the cell proliferation in time- and concentration-dependent manners. THP (50–500 ng/mL) induced MG63/DOX cell cycle arrest at the G2/M phase in time- and concentration-dependent manners. Furthermore, the treatment of MG63/DOX cells with THP (200–1000 ng/mL) downregulated cyclin B1 expression, and decreased the phosphorylated Cdc2 at Thr161. Conversely, the treatment increased the phosphorylated Cdc2 at Thr14/Tyr15 and Cdc25C at Ser216, which led to a decrease in Cdc2-cyclin B1 activity.Conclusion:The cytotoxicity of THP to MG63/DOX cells may be in part due to its ability to arrest cell cycle progression at the G2/M phase, which supports the use of THP for managing patients with MDR osteosarcoma.

Heterogeneous expression and biological function of ubiquitin carboxy-terminal hydrolase-L1 in osteosarcoma

Ubiquitin carboxyl terminal hydrolase 1 (UCHL1), a member of the UCH class of DUBs, has been reported as either an oncogene or a tumor suppressor. However, the molecular mechanism underlying the biological function of UCHL1 in osteosarcoma is still unclear. This study was aimed at elucidating the roles of UCHL1 in regulating the biological behavior of osteosarcoma cells. In this study, we found that UCHL1 was elevated in osteosarcoma compared with normal bone tissue. Moreover, UCHL1 expression level was correlated with tumor maximum diameter, high rate of lung metastases and short survival time. Then, we found that knockdown of UCHL1 in osteosarcoma cell MG63 inhibited cell proliferation and significantly increased cell population in the G1 phase. Several cyclins promoting G1/S phase transition were reduced after UCHL1 knockdown, including cell cycle regulator cyclin D1, cyclin E1 and CDK6. Moreover, inhibition of UCHL1 in MG63 cells dramatically induced cell apoptosis. We also found that down-regulation of UCHL1 in MG63 significantly inhibited cell invasion. Then, we found that there was a positive correlation between UCHL1 expression level and the Akt and ERK phosphorylation status. Finally, in vivo data showed that knockdown of UCHL1 inhibited osteosarcoma growth in nude mice. These results indicate that UCHL1 could work as an oncogene and may serve as a promising therapeutic strategy for osteosarcoma.

Mediator of RNA polymerase II transcription subunit 19 promotes osteosarcoma growth and metastasis and associates with prognosis

Osteosarcoma (OS) is the most common primary malignant tumour of bone. Nearly 30-40% of OS patients have a poor prognosis despite multimodal treatments. Because the carcinogenesis of OS remains unclear, the identification of new oncogenes that control the tumourigenesis and progression of OS is crucial for developing new therapies. Here, we found that the expression of Mediator of RNA polymerase II transcription subunit 19 (Med19) was increased in OS samples from patients compared to normal bone tissues. Cyclin D1 and cyclin B1 are upregulated in Med19 positive OS tissues. Importantly, among 97 OS patients of Enneking stage IIB or IIIB, Med19 expression was correlated with metastasis (P<0.05) and poor prognosis (P<0.01). Med19 knockdown significantly induced growth inhibition, reduced colony-forming ability and suppressed migration in the OS cell lines Saos-2 and U2OS, along with the downregulated expression of cyclin D1 and cyclin B1. Med19 knockdown also induced apoptosis in Saos-2 cells via induction of caspase-3 and poly ADP-ribose polymerase (PARP). In addition, Med19 knockdown significantly suppressed tumour growth in an OS xenograft nude mouse model via suppression of cyclin D1 and cyclin B1. Simultaneously, Med19 downregulation decreased the expression of Ki67 and proliferating cell nuclear antigen (PCNA) in tumour samples from OS xenograft nude mice. Med19 depletion remarkably reduced tumour metastasis in a model of OS metastatic spreading. Taken together, our data suggest that Med19 acts as an oncogene in OS via a possible cyclin D1/cyclin B1 modulation pathway.

The overexpression of MRP4 is related to multidrug resistance in osteosarcoma cells.

Doxorubicin (Adriamycin, ADM) is an antimitotic drug used in the treatment of a wide range of malignant tumors, including acute leukemia, lymphoma, osteosarcoma, breast cancer, and lung cancer. Multidrug resistance-associated proteins (MRPs) are members of a superfamily of ATP-binding cassette (ABC) transporters, which can transport various molecules across extra- and intra-cellular membranes. The aim of this study was to investigate whether there was a correlation between MRP4 and primary ADM resistance in osteosarcoma cells. In this paper, we chose the human osteosarcoma cell line MG63, ADM resistant cell line MG63/DOX, and the patients primary cell GSF-0686. We checked the ADM sensitivity and cytotoxicity of all the three cells by cell proliferation assay. The intracellular drug concentrations were measured by using LC-MS/MS. We also examined MRP4 gene expression by RT-PCR and Western Blot. We found that the intracellular ADM concentration of the parent osteosarcoma cell line MG63 was higher than the ADM resistant osteosarcoma MG63/DOX cell line or the GSF-0686 cell after ADM treatment (P < 0.05). In addition, MRP4 mRNA and protein levels in ADM resistant osteosarcoma cells were higher than in MG63 cell (P < 0.05). Taking together, this work suggests that overexpression of MRP4 may confer ADM resistance in osteosarcoma cells.

Relationship of serum methotrexate concentration in high-dose methotrexate chemotherapy to prognosis and tolerability: A prospective cohort study in chinese adults with osteosarcoma

BACKGROUND Cancer that originates in the bone, termed primary bone cancer, is rare. Osteosarcoma (OS) occurs primarily in growing bone tissue and is more prevalent in children and adolescents. OS in adults is rare, with 3 to 5 cases per million population per year worldwide. There are limited data on treatment-related prognosis and adverse reactions in adults reported in the literature. OBJECTIVES The aims of this study were to investigate factors that influence serum methotrexate (MTX) concentrations used in chemotherapy in Chinese adult patients with OS, and to determine the correlations (based on age, sex, and dosage), if any, between MTX and prognosis, in terms of disease-free survival (DFS) and overall survival (OAS), and tolerability. METHODS Adult patients aged ≥30 years with OS received ≥3 courses (2 courses before surgery and 3-4 courses postsurgery) of high-dose MTX (6 or 8 g/m(2)) combined chemotherapy. The regimen consisted of day 1: MTX + folinic acid (herein referred to as citrovorum factor rescue); day 8: cisplatin; days 21 to 25: ifosfamide + mesna; and day 21: doxorubicin. Serum MTX concentrations were assessed immediately after the end of infusion (baseline) and at 24 and 48 hours using high-performance liquid chromatography. Changes in serum MTX concentrations, factors that influence serum MTX concentrations, and the relationship between serum MTX concentrations and prognosis and tolerability (determined by adverse reactions) were analyzed. Patients received a second course of treatment after a 3-week period. RESULTS Ninety patients (58 men, 32 women; age range, 30-67 years) with OS were included in the study. A total of 532 courses of combined chemotherapy were administered. The serum MTX concentrations ranged widely at baseline (244.31-929.68 mol/L, Cmin and Cmax, respectively) and at 24 hours (0.73-28.24 mol/L, respectively), suggesting that the serum MTX concentrations varied significantly between different individuals and within the same individual at different time points. The serum MTX concentrations in ~23% of cases (122/532) determined at 24 and/or 48 hours were numerically higher than the safety values (according to Nirenbergs reference: irreversible damage if MTX concentration was >10 umol/L and > 1 umol/L at 24 and 48 hours, respectively). No correlation was found between high serum MTX concentration at baseline and high serum MTX concentration at 24 hours (r = 0.401). The prevalences of the 3 most common adverse reactions in these patients were depressed white blood cell count (44.03%), dental ulcer (23.0%), and rash (18.0%). However, in the remaining 410 courses in which serum MTX concentrations were lower than the safety values, these prevalences were 14.6%, 3-9%, and 2.4%, respectively. Neither age nor sex was significantly associated with MTX Cmax, but dosage was (P < 0.05). Patients with a serum MTX Cmax concentration >500 μmol/L at baseline had a significantly longer DFS rate than those with ≤500 umol/L (P = 0.040). There were no significant between-group differences in the OAS rates. conclusions: In these Chinese patients with OS, serum MTX concentrations measured at different time points were varied. The findings suggest that adverse reactions occurred in patients whose serum MTX concentrations at 24 and/or 48 hours were higher than the safety values. The dosage appeared to have influenced MTX Cmax, while sex and age did not, and the Cmax was significantly related to DFS but not OAS.

Cyclophosphamide-hydroxycamptothecin as second-line chemotherapy for advanced Ewing's sarcoma: Experience of a single institution

To investigate the feasibility and efficacy of cyclophosphamide (CTX)‐hydroxycamptothecin (HCPT) as second‐line chemotherapy on advanced Ewings sarcoma.