Metastatic Osteosarcoma Patients Research Articles

Circular RNA expression profile identifies circMGEA5 as a novel metastasis-promoting factor and potential biomarker in osteosarcoma.

Osteosarcoma (OS) is associated with a high incidence of lung metastasis, which leads to a high risk of cancer death. Circular RNA (circRNA), a novel class of noncoding RNA, is emerging as a key player in human cancer. Herein, we explored the role of circMGEA5 in OS metastasis by conducting circRNA expression microarray. CircMGEA5 was significantly upregulated in metastatic OS tissues compared to primary tissues. High circMGEA5 was positively related with shorter overall and disease-free survival time. Knockdown of circMGEA5 suppressed OS cell migration, invasion, and epithelial-mesenchymal transition (EMT). Mechanistically, circMGEA5 acted as a competing endogenous RNA (ceRNA) to directly sponge miR-153-3p and miR-8084, resulting in increasing ZEB1 and Snail expression, respectively, thereby inducing EMT and metastasis. In turn, ZEB1 and Snail were capable to bind to circMGEA5 promoter, activating circMGEA5 transcription, thus forming a positive feedback loop. Furthermore, we established the tail vein injection model and found that circMGEA5 depletion remarkably reduced lung metastasis nodules generated by OS cells. In sum, our findings, for the first time, reveal the metastasis-promoting role of circMGEA5 in OS. Targeting of this newly identified ceRNA axis may be crucial in the development of novel therapies for metastatic OS patients.

1507P Distinct enomic features between osteosarcomas initially metastasing to bone and to lung

Osteosarcoma (OS) initially metastasing to bone only shows distinct biological features compared to osteosarcoma that initially metastasizes to the lung, which suggests an underlying difference in genomic pathogenetic mechanism. However, related studies are lacking. We analyzed whole-exome sequencing (WES) data for 38 metastatic OS patients with different relapse patterns. Multiplex Immunohistochemistry was also applied to investigate the immune- microenvironment. We tried to redefine disease subclassifications for OS based on genetic alterations to guide treatment strategies, and we correlated these genetic profiles with clinical treatment courses and prognosis to elucidate potential evolving cladograms. We investigated whole-exome sequencing of 12/38 patients with high-grade OS (31.6%) with initial bone metastasis (group A) and 26/38 (68.4%) with initial pulmonary metastasis (group B), of whom 15/38 (39.5%) had paired samples of primary lesions and metastatic lesions for these two cohorts with different relapse patterns. We found that osteosarcoma in group A mainly carries single-nucleotide variations while those in group B mainly exhibits structural variants. Additionally, osteosarcoma in group A displayed better immunogenicity, including a higher tumor mutation burden and neoantigen load and more tertiary lymphoid structures in tumor microenvironment.Table: 1507PThe difference of osteosarcoma patients’ genomic alterations between group A and group BGroup AGroup BStatistical methodp valuen (%)12 (31.6)26 (68.4)Genomic type, n (%)Chi-square0.0180aSNV10 (26.3)11 (28.9)SV or mixed2 (5.3)15 (39.5)TMBWilcoxon0.0100aRange1.1-32.90-14.2Median (Q1, Q3)4.85 (2.75, 11.98)2.4 (1.38, 4.45)NeoantigenWilcoxon0.002∗Range57-20840-1539Median (Q1, Q3)743 (316.5, 1034.5)128.5 (49, 200.5)MSI, n (%)Chi-square0.3160MSS11 (29.0)26 (68.4)MSI-H1 (2.6)0 (0.0)HED-status, n (%)Chi-square1.0000Low10 (26.3)22 (57.9)High2 (5.3)4 (10.5) Open table in a new tab Osteosarcoma with different relapse patterns has distinct genomic landscapes. This study suggests that osteosarcoma with mainly single-nucleotide variations other than structural variants might exhibit biological behavior predisposing toward bone metastases; immunotherapy is a promising therapeutic option for this group of patients.

Galectin-3 inhibition boosts the therapeutic efficacy of Semliki Forest virus in pediatric osteosarcoma

The outcomes of metastatic and nonresponder pediatric osteosarcoma patients are very poor and have not improved in the last 30 years. These tumors harbor a highly immunosuppressive environment, making existing immunotherapies ineffective. Here, we evaluated the use of Semliki Forest virus (SFV) vectors expressing galectin-3 (Gal3) inhibitors as therapeutic tools, since both the inhibition of Gal3, which is involved in immunosuppression and metastasis, and virotherapy based on SFV have been demonstrated to reduce tumor progression in different tumor models. In vitro, inhibitors based on the Gal3 amino-terminal domain alone (Gal3-N) or fused to a Gal3 peptide inhibitor (Gal3-N-C12) were able to block the binding of Gal3 to the surface of activated T cells. In vivo, SFV expressing Gal3-N-C12 induced strong antitumor responses in orthotopic K7M2 and MOS-J osteosarcoma tumors, leading to complete regressions in 47% and 30% of mice, respectively. Pulmonary metastases were also reduced in K7M2 tumor-bearing mice after treatment with SFV-Gal3-N-C12. Both the antitumor and antimetastatic responses were dependent on modulation of the immune system, primarily including an increase in tumor-infiltrating lymphocytes and a reduction in the immunosuppressive environment inside tumors. Our results demonstrated that SFV-Gal3-N-C12 could constitute a potential therapeutic agent for osteosarcoma patients expressing Gal3.

REGOMAIN: A randomized, placebo-controlled, double-blinded, multicenter, comparative phase II study of the efficacy of regorafenib as maintenance treatment in patients (pts) with high-grade bone sarcomas (HGBS) at diagnosis or relapse and without complete remission after standard treatment.

TPS11585 Background: Primary metastatic osteosarcoma (OS) patients are treated with a curative intent following the same principles of non-metastatic OS, while the treatment of recurrent OS is primarily surgical in the case of isolated lung metastases. When complete removal of all metastases cannot be achieved, the prognosis remains poor, with a median Progression-Free Survival (PFS) between 3 to 8 months, and therefore there is a clinical need to reduce the risk of progression after the initial treatment sequence. The REGOBONE study reported a significant PFS benefit of regorafenib (REG) compared to placebo (in osteosarcomas: median PFS: 16.4 versus 4.1 weeks) and a manageable safety profile in patients with histologically confirmed HGBS (i.e., osteosarcoma or other bone sarcomas with the exception of Ewing sarcomas, chondrosarcoma and chordoma). Methods: This multicenter trial is ongoing to study the efficacy and safety of maintenance REG in pts > = 16 years old with HGBS, without complete remission but with no progressive disease after standard treatment, either at diagnosis or at relapse. Sixty pts will be randomly allocated in a 1:1 ratio to receive either oral REG at a daily dose of 120mg or its matching placebo, continuously for a maximum of 12 months. Randomization is stratified according to the setting of the disease: initial diagnosis versus relapse. The primary objective is to compare the efficacy (PFS) between the 2 arms. The expected 4-month PFS rates are 30% in the control arm and 60% in the REGO arm (HR = 0.42). Fifty-two events will provide 87% power to show significant improvement in PFS, using a 2-sided log-rank test at a 5% level. Secondary endpoints include Overall Response Rate (ORR), Disease Control Rate (DCR), Time to Treatment Failure (TTF), Overall Survival (OS), Quality of Life (QoL), and safety profile. Radiological endpoints will be evaluated using the RECIST 1.1 with tumor assessments every 2 months (first 6 months) and then every 3 months. Translational objectives will identify predictive biomarkers for efficacy of REG as maintenance therapy. Pts of the control arm who experience disease progression may switch to receive open label REG. As of Feb 1st, 2022, 3 patients have been randomized. 14 sites of the French Sarcoma Group will participate. An amendment is being implemented to lower the age limit (12 years old) and to expand tumor types to other HGBS (Ewing sarcomas, chondrosarcomas, Undifferentiated Pleomorphic Sarcomas, Leiomyosarcomas and angiosarcomas). Clinical trial information: NCT04698785.

Characterizing the critical role of metabolism in osteosarcoma based on establishing novel molecular subtypes.

Osteosarcoma is the third most frequently diagnosed cancer among adolescents. Immunotherapy is an effective curative treatment for metastatic osteosarcoma patients. This study aimed to further reveal the significance of metabolism in tumor progression, and to categorize molecular subtypes for guiding personalized therapy. Univariate Cox regression analysis was performed to screen metabolism-related genes associated with osteosarcoma prognosis. A molecular subtyping system was developed by unsupervised consensus clustering. Survival analysis and functional analysis were used to evaluate the performance of subtyping and characterize the TME of subtypes. Stepwise Akaike information criterion (stepAIC) was employed to optimize the prognostic model. C1 and C2 subtypes showed distinct prognosis, with more favorable survival in C2 subtype. C2 subtype presented a higher immune infiltration and active anti-tumor response. Notably, C2 subtype was predicted to have better immune response to immune checkpoint blockade. In addition, a 5-gene prognostic signature with robust ability to classify patients into high-risk and low-risk groups was developed. The study revealed the critical role of metabolism in tumorigenesis by comparing the features between the two subtypes. Oncogenic pathways including epithelial mesenchymal transition (EMT), glycolysis and hypoxia may be closely involved in the correlation with metabolism. Importantly, we developed a novel subtyping system and a 5-gene signature with high potential to be applied in clinical practice.

Recent and Ongoing Research into Metastatic Osteosarcoma Treatments.

The survival rate for metastatic osteosarcoma has not improved for several decades, since the introduction and refinement of chemotherapy as a treatment in addition to surgery. Over two thirds of metastatic osteosarcoma patients, many of whom are children or adolescents, fail to exhibit durable responses and succumb to their disease. Concerted efforts have been made to increase survival rates through identification of candidate therapies via animal studies and early phase trials of novel treatments, but unfortunately, this work has produced negligible improvements to the survival rate for metastatic osteosarcoma patients. This review summarizes data from clinical trials of metastatic osteosarcoma therapies as well as pre-clinical studies that report efficacy of novel drugs against metastatic osteosarcoma in vivo. Considerations regarding the design of animal studies and clinical trials to improve survival outcomes for metastatic osteosarcoma patients are also discussed.

Prognostic significance of the ratio of surgically resected to radiologically detected lung nodules in patients with metastatic osteosarcoma

The factors that affect the prognosis of patients’ metastatic osteosarcoma are still poorly understood. In this study, we investigated a new prognostic factor, the ratio of surgically resected to radiologically detected osteosarcoma lung nodules (SR/RD), which may have predictive value. Patients and methodsData from patients with metastatic osteosarcoma who underwent metastasectomy between January 2009 and December 2020, in a single center, were reviewed. The relationships between survival and the SR/RD ratio, timing of lung metastases, number of nodules, laterality, and presence of tumor necrosis at first metastasectomy were investigated. ResultsAmong the 125 metastatic osteosarcoma patients, 80 patients had an SR/RD ratio ≤1. The median duration of follow-up was 72 months, ranging from 6 to 118 months. The five-year overall survival (OS) and postmetastasectomy event-free survival (EFS) for all patients were 36.5% and 18.1%, respectively. The five-year OS of patients with a low SR/RD ratio was 49.6% and that of patients with a high SR/RD ratio was 11.8 (P = 0.001). The two-year postmetastasectomy EFS rates of the high and low ratio groups were 24.1% and 9.4%, respectively (P = 0.001). The SR/RD ratio, number of nodules, and tumor necrosis had significant effects on OS and postmetastasectomy EFS in univariate analysis. A Cox proportional hazard model demonstrated that tumor necrosis and an SR/RD ratio >1 were associated with OS (HR = 1.8 and 2.01) and postmetastasectomy EFS (HR = 1,69 and 1.97). ConclusionsA high SR/RD ratio of greater than 1 and poor tumor necrosis were significantly associated with poor survival among patients with metastatic osteosarcoma who had lung metastasectomy. The high SR/RD ratio may be a surrogate outcome for incomplete metastatic tumor resection.

Targeting the VEGF Pathway in Osteosarcoma.

Osteosarcoma is the most common primary tumor of the bones affecting mainly young adults. Despite the advances in the field of systemic anticancer therapy, the prognosis of relapsed of metastatic osteosarcoma patients remain dismal with very short survival. However, the better understanding of the pathophysiology of this subtype of sarcoma has led to the identification of new targeted agents with significant activity. In fact, increased angiogenesis plays a major role in the tumor growth and survival of osteosarcoma patients. Several targeted agents have demonstrated a significant anti-tumor activity including multi-kinase inhibitors. In this review, we will discuss the pathophysiology, rationale, and role of targeting angiogenesis via the VEGF pathway in patients with osteosarcoma with emphasis on the published clinical trials and future directions.

MYLK4 promotes tumor progression through the activation of epidermal growth factor receptor signaling in osteosarcoma

BackgroundOsteosarcoma (OS) is the most common primary bone cancer in adolescents and lung metastasis is the leading cause of death in patients with OS. However, the molecular mechanisms that promote OS growth and metastasis remain unknown.MethodsWe investigated the expression of myosin light chain kinase family members between metastasis and non-metastasis patients in the TARGET database and ensured that only myosin light chain kinase family member 4 (MYLK4) had higher expression in metastatic osteosarcoma patients. Then we confirmed the results by immunohistochemistry (IHC) and Western blotting (WB) of OS tissues. The effect of MYLK4 on the metastasis and proliferation of OS cells was investigated by wound healing, Transwell and colony-formation assays. Mass spectrum analysis was used to ensure the new binding protein of MYLK4. Tissue microarrays analysis was used to show the correlation between MYLK4 and pEGFR (Y1068). A series of in vivo experiments were conducted to reveal the mechanisms by which MYLK4 modulated the metastasis and proliferation of OS.ResultsMyosin Light Chain Kinase Family Member 4 (MYLK4) was significantly upregulated in metastatic human OS tissues. Growth and metastasis of OS could be accelerated by MYLK4 overexpression, whereas silencing MYLK4 expression resulted in decreased cell growth and metastasis. Mechanistically, mass spectrum analysis showed that MYLK4 interacted with the epidermal growth factor receptor (EGFR) in osteosarcoma cells and promoted growth and metastasis via the EGFR signaling pathway. Tissue microarrays analysis also showed that MYLK4 expression had a positive correlation with the expression of pEGFR (Y1068). Moreover, the EGFR inhibitor gefitinib could partially reverse the effect of cell proliferation and metastasis caused by MYLK4 overexpression. Importantly, the combination of MYLK4 and EGFR inhibitors had synergistic effects on growth and metastasis of OS in vitro and in vivo.ConclusionOur results indicate that MYLK4 promotes OS growth and metastasis by activating the EGFR signaling pathway and can be a novel therapeutic target for the treatment of OS patients.

11-O-Galloyl Bergenin from Corylopsis coreanas Leaves Induces Autophagy and Apoptosis in Human Osteosarcoma.

Osteosarcoma is the most common malignant bone-forming tumor, wherein most patients with high grade osteosarcomas are treated with chemotherapy. Despite this, survival for metastatic or relapsed osteosarcoma patients has remained at an overall 5-year survival rate of 20%. In particular, the extracts of Corylopsis coreana (Korean winter hazel), a cultivated woody plant in South Korea, have shown beneficial anti-inflammatory, anti-oxidative, anti-osteoclastic, and antihyperuricemic properties. Therefore, this study aimed to demonstrate the antitumor activities and underlying mechanism of 11-O-Galloyl bergenin (OGAL) isolated from Corylopsis coreanas leaves in human osteosarcoma cells. Herein, we found that OGAL inhibited MG63 cell proliferation and induced cellular apoptosis as evidenced by cleaved-PARP, cleaved-caspase 3, TUNEL-positive cells, and Annexin V-positive cells. Specifically, OGAL-induced apoptosis was accompanied by p53 and p21 upregulation, BAX expression, and decreased Bcl-2 and cdk2. Moreover, OGAL induced autophagy via AKT inactivation, LC3II upregulation, and MG63 cell autophagosome formation. OGAL-induced autophagy was also accompanied by increased p38 phosphorylation, whereas JNK and ERK1/2 activities were found to be unaffected upon examining the MAPK signaling pathway. Furthermore, wound healing and Boyden chamber assays showed that OGAL suppressed MG63 cell migration and invasion. Given these findings, this study provided evidence that OGAL has antitumor effects by apoptosis and autophagy enhancement through increased p53, AKT, and p38 signaling, suggesting that OGAL may be a potential therapeutic strategy for osteosarcoma treatment.

Adoptive transfer of TILs plus anti-PD1 therapy: An alternative combination therapy for treating metastatic osteosarcoma

AimWe sought to investigate the efficacy of adoptive transfer of TILs plus anti-PD1 therapy in metastatic osteosarcoma patients. Materials and methodsA total of 30 patients received anti-PD1 therapy (Group 1) while 30 patients were subjected to TILs plus anti-PD1 therapy (Group 2). Progression-free survival time (PFS) and overall survival time (OS) were analyzed using Kaplan-Meier analysis. Potential prognostic factors were analyzed using univariate and multivariate analyses. ResultsThe ORR in Group 2 is 33.3%, which is significantly higher than Group1 (6.67%). In addition, we found significantly prolonged mPFS (5.4 months) and mOS (15.2 months) in Group 2 compared to those in Group 1, which recorded mPFS and mOS of 3.8 and 6.6 months, respectively. Univariate and multivariate analyses indicate that patients with more infusions of TIL numbers and CD8+TILs or less infusions of CD8+ PD1+TILs and CD4+FoxP3+ TILs show increased PFS and OS. Moreover, PD1hi is another good prognostic factor that predict PFS and OS. ConclusionOverall, these findings indicated that TILs plus anti-PD1 therapy has significant clinical outcomes in metastatic osteosarcoma patients. However, further studies are essential to validate and characterize the therapeutic activity of TILs plus anti-PD1.

Retrospective Analysis of Adoptive TIL Therapy plus Anti-PD1 Therapy in Patients with Chemotherapy-Resistant Metastatic Osteosarcoma

Background The pathological subtype of osteosarcoma is one of the most common malignant bone tumors. Notably, chemotherapy-resistant metastatic osteosarcoma has been reported to cause significant mortality and shows poor prognosis with the currently available multidisciplinary treatments. This study investigated whether combined adoptive TIL and anti-PD1 therapy improves the prognosis of patients with chemotherapy-resistant metastatic osteosarcoma. Methods A total of 60 patients with chemotherapy-resistant metastatic osteosarcoma between June 2016 and March 2018 were enrolled. The primary endpoint was to evaluate the safety and adverse effects (AEs) of infusions of TIL and anti-PD1 therapy in the patients. Besides, secondary endpoints included assessing the objective response rate (ORR), progression-free survival time (PFS), and overall survival time (OS). Results We reported that combined TIL therapy and anti-PD1 therapy is safe and all treatment-related AEs were reversible or manageable. The ORR of all the patients is 36.67%, and patients with more infusions of TIL and CD8+TIL, less infusions of CD8+PD1+TIL, and less infusion of CD4+FoxP3+TIL exhibited increased PFS and OS. Conclusion This study determined that combined TIL and anti-PD1 therapy is safe and effective in metastatic osteosarcoma patients with chemotherapy resistance.

422P Response rate and time to progression after first line chemotherapy with cisplatin and adriamycin in patients with metastatic osteosarcoma at presentation

Metastatic osteosarcoma at presentation forms 10 to 20% of total osteosarcomas diagnosed. They have a very poor prognosis compared to non-metastatic subset. Patients with resectable metastasis have a better DFS and OS compared to unresectable metastatic disease at presentation. This study was done to find the response rate to first line cisplatin and adriamycin combination chemotherapy to upfront metastatic osteosarcoma patients and time to progression after first line chemotherapy. Secondary objective was to find the overall survival. Patients diagnosed with osteosarcoma at madras medical college were identified from the database for a period of 5.5 years from January 2013 to June 2018. From that patients who had metastasis at presentation were further sorted out and taken for analysis. 77 patients were diagnosed to have osteosarcoma during the 5.5year period. Among them 36 patients had metastatic disease at presentation. 28 patients were male (77.8%) and 8 patients were female (22.2%). Median age of the patients was 19.5 years. 23 patients had ECOG PS 2 (63.9%) and 12 had ECOG PS 1 (33.3%) and 1 had ECOG PS 3 (2.8%). Tibia and femur were the most common primary site (75%). 5 patients had resectable lung metastasis and underwent pulmonary metastasectomy and were excluded from analysis and 1 patient was on palliative care only. Among remaining 30 patients, 27 patients had multiple lung metastasis (90%) and 3 patients had bone metastasis (10%). 30 patients received 6 cycles of chemotherapy with adriamycin 25mg/m2 on d1-d3 and cisplatin 100mg/m2 on d1 repeated every 21 days except 3 who progressed during chemotherapy. 17 patients (56.7%) had grade 1-2 vomiting. No one developed any acute cardiotoxicity. No treatment related death occurred. At the end of 6 cycles, 22 patients (73.3%) had partial response and 5 patients (16.7%) had stable disease and 3 patients (10%) had progressive disease. Median time to progression was 11.5 months. 2year OS was 73.3%. This platinum doublet is a reasonable option for these patients as most of them are young.

Transient NK Cell Depletion Facilitates Pulmonary Osteosarcoma Metastases After Intravenous Inoculation in Athymic Mice.

Two thirds of metastatic osteosarcoma patients die within 5 years of diagnosis. Improved experimental models of osteosarcoma metastasis will facilitate the development of more effective therapies. Intravenous cancer cell injection can produce lung metastases in nude mice, but this "experimental metastasis" technique has been predominantly applied to a single osteosarcoma cell line (143B) and required injection of 1-2 million cells. Using two human osteosarcoma cell lines, we discovered that transient Natural Killer cell depletion dramatically enhanced the efficiency of experimental pulmonary osteosarcoma metastasis. This technique for modeling osteosarcoma metastasis may enable the identification of better treatments for this aggressive cancer.

The efficacy and safety of Apatinib mesylate in the treatment of metastatic osteosarcoma patients who progressed after standard therapy and the VEGFR2 gene polymorphism analysis.

The aim of this study was to investigate the efficacy and safety of Apatinib mesylate in the treatment of metastatic osteosarcoma patients who progressed after standard therapy and the VEGFR2 gene polymorphism analysis. Designed as a retrospective study, a total of 105 metastatic osteosarcoma patients who progressed after standard therapy were included in this study. The metastatic osteosarcoma patients received 500-750mg Apatinib mesylate according to body surface area until disease progression or unacceptable toxicity with 28days one cycle. Overall response was evaluated after two cycles Apatinib treatment, then progression-free survival (PFS) and overall survival (OS) were evaluated, and safety data were recorded. Additionally. peripheral blood and peripheral blood mononuclear cell (PBMC) specimens in the osteosarcoma patients were collected for the genotyping of VEGFR2 genetic variation and mRNA expression, respectively. Analysis on the association between genotype and baseline characteristics and VEGFR2 gene mRNA expression was analyzed. The univariate analysis of genotypes and prognosis was carried out by Kaplan-Meier survival analysis, and multivariate analysis was adjusted by Cox regression analysis. The objective response rate (ORR) of the 105 metastatic osteosarcoma patients was 37.14%, disease control rate (DCR) was 77.14%, median PFS was 4.1months, and median OS was 9.0months. Regarding the VEGFR2 gene polymorphisms analysis, only - 906T > C was of clinical significance. The prevalence of - 906T > C in VEGFR2 among the study population was as follows: TT genotype 62 cases (59.05%), TC genotype 36 cases (34.29%) and CC genotype 7 cases (6.66%), minor allele frequency of - 906T > C was 0.24. Compared with patients with TC/CC genotype, patients with TT genotype showed longer median PFS (5.0 versus 3.1months, P = 0.011) and median OS (9.8 versus 7.6months, P = 0.032). There was no correlation between the polymorphism and adverse reactions. Additionally, the mRNA expression in 69 randomly selected sample indicated that the mRNA expression of VEGFR2 of the patients with CC/TC genotypes were significantly higher than those of the TT genotype patients (P < 0.001). Apatinib was safe and effective in the treatment of metastatic osteosarcoma patients who progressed after standard therapy. The clinical outcomes of Apatinib may be influenced by the polymorphism - 906T > C of VEGFR2 through mediating the mRNA expression of VEGFR2.

Elevated miR-216b Enhances Cisplatin-Induced Apoptosis in Osteosarcoma by Downregulating JMJD2C and Regulating HIF-1α/HES1 Signaling Axis

Background: Cisplatin-based chemotherapy is the standard regimen for osteosarcoma (OS). However, chemoresistance to cisplatin commonly contributes to treatment failure and poor prognosis for metastatic OS patients. The study aims to identify whether miR-216b is dysregulated in OS and its relation to OS cell resistance to cisplatin. Methods: Tumor and paracancerous tissues were collected from OS patients to determine expression patterns of miR-216b and JMJD2C. Pearson correlation analysis was used to evaluate the expression between miR-216b and JMJD2C. CCK8 assay and flow cytometry were used to determine cell viability and apoptosis. Targeting relationship of miR-216b and JMJD2C was determined by dual luciferase reporter assay. Co-immunoprecipitation (IP) assay was used to determine binding between JMJD2C and HIF1α. Chromatin IP (ChIP)-qPCR was used to examine the level of HES1 promoter fragment enriched by H3K9me3 antibodies. Results: miR-216b expression was reduced in OS and positively correlated with patient survival. miR-216b overexpression further enhanced cisplatin-induced apoptosis in OS cells. miR-216b targeted and inhibited JMJD2C. JMJD2C bound and enhanced HIF1α to remove the H3K9 methylation modification at the HES1 promoter region, leading to upregulation of HES1. miR-216b overexpression further reduced cisplatin-induced decrease in tumor growth and weight in nude mice. HES1 overexpression weakened all effects of miR-216b in OS cells and in nude mice xenografts. Conclusion: miR-216b further enhances the effects of cisplatin through downregulating JMJD2C and regulating HIF1α/HES1 signaling axis in OS, suggesting that miR-216b may be an adjunct to cisplatin chemotherapy in the treatment of OS. Funding Statement: This study was supported by General Program of National Natural Science Foundation of China (No. 81572632 & No. 8187100734). Declaration of Interests: The author declares no competing interest exists. Ethics Approval Statement: Human tissue collection protocol was approved by Institutional ethics committee. All animal procedures were approved by Institutional Animal Care and Ethics Committee.

Novel classifiers with clinical laboratory parameters for early detection of osteosarcoma.

BackgroundOsteosarcoma (OS) is one of the most common malignant bone tumors. It is essential to explore early diagnostic indicators with high sensitivity and specificity due to the rapid progression and metastasis of OS and the poor survival of metastatic OS patients. However, a few indicators of diagnostic significance have been described.MethodsA total of 458 OS patients, 312 healthy individuals, and 228 patients with primary benign bone lesions were included. Logistic regression was performed on 46 clinical laboratory parameters to establish the diagnostic classifiers, which were evaluated by analysis of the receiver operating characteristic (ROC) curves.ResultsWe established three diagnostic classifiers, called C os for all ages, C los for low ages, and C hos for high ages, with clinical laboratory parameters to distinguish OS from healthy individuals. All classifiers showed better diagnostic performances than alkaline phosphatase (ALP) in the independent validation cohort. In addition, these classifiers had better ability than ALP to discriminate OS from primary benign bone lesions. Furthermore, C os, C los, and C hos had larger AUC than ALP to identify small‐size and early‐stage OS and could also detect ALP‐negative OS effectively.ConclusionOur study suggests the potential of C os, C los, and C hos as non‐invasive biomarkers for early OS.

Survival analysis of patients with metastatic osteosarcoma: a Surveillance, Epidemiology, and End Results population-based study.

The present study is aimed at investigating whether (1) primary tumour surgery confers an improved survival on patients with metastatic osteosarcoma and (2) primary tumour surgery influences survival of patients with metastatic osteosarcoma differently according to primary tumour site. We retrospectively identified 517 patients with high-grade, metastatic osteosarcoma in the Surveillance, Epidemiology, and End Results (SEER) database between 1994 and 2013. The effect of primary tumour surgery on survival was assessed using Kaplan-Meier analyses, log-rank tests, and multivariate Cox proportional hazard regression modeling. Of those 517 patients with metastatic osteosarcoma in the cohort, 351 patients (68%) underwent primary surgery, and 166 patients (32%) did not undergo surgery. Primary tumour surgery was associated with increased overall survival (hazard ratio (HR) = 0.457, 95% CI 0.354-0.590, p < 0.001) and cancer-specific survival (HR = 0.422, 95% CI 0.325-0.550, p < 0.001). When we focused on different primary tumour sites, receipt of primary tumour surgery significantly prolonged the survival of patients with extremity osteosarcoma (p < 0.05 for overall and cancer-specific survival). However, for patients with pelvis/spine osteosarcoma, both univariate and multivariate analyses indicated that primary tumour surgery might not be associated with improved survival (p > 0.05 for overall and cancer-specific survival). Our study is the first population-based analysis to provide evidence of a favourable prognostic impact of primary tumour surgery on metastatic extremity osteosarcoma patients but not metastatic axial (pelvis/spine) osteosarcoma patients. Moreover, we found that surgery type (resection of the primary tumor without amputation vs. amputation) did not influence survival in patients with metastatic osteosarcoma.

Prognostic and clinicopathological significance of circulating tumor cells in osteosarcoma

Osteosarcoma is the most common form of primary malignant bone tumor, with metastasis playing an essential role in determining a patient's prospects for survival. It is essential that new and better molecular targets that respond effectively to therapies and are predictive of the risk of tumor metastasis are identified. We have therefore undertaken the present prospective study to ascertain the clinical significance of circulating tumor cells (CTCs) in osteosarcoma patients. Peripheral blood was obtained from patients both pre- and post-surgery then processed using a CanPatrol™ system, an enrichment technique allowing isolation of CTCs by virtue of their size at baseline. Multiplex RNA in situ hybridization (RNA-ISH) was subsequently conducted to characterize the CTCs based on various molecular markers including MTA1, CD45, EpCAM, CK8, CK19, Vimentin and Twist. MTA1 expression was further validated by immunohistochemistry of the tumor tissue. Besides defining a diagnosis and prognosis for osteosarcoma patients, the correlation between CTC count and their molecular and clinicopathological characteristics was found to assist in the analysis of the response of patients to neoadjuvant chemotherapy. Our results revealed that the number of CTCs was significantly higher at baseline in metastatic patients than in those whose osteosarcomas were localized. The variation was attributed to the neoadjuvant chemotherapy treatment. A cut-off value of 7 CTCs/5 mL was found to effectively distinguish patients who had either a favorable or unfavorable prognosis. Notably, the ratio of mesenchymal CTCs at baseline was found to be higher in metastatic vs. localized osteosarcoma patients. In addition, the expression of MTA1 was higher in mesenchymal CTCs than the other CTC phenotypes. Furthermore, immunohistochemical analysis demonstrated a higher expression of MTA1 in tumor tissues from metastatic osteosarcoma patients. Taken together, our findings conclusively establish that the number and molecular phenotype of CTCs are predictive of tumor metastasis and the response of patients to neoadjuvant chemotherapy.

Pazopanib in metastatic osteosarcoma patients: Report of 9 cases treated off label at Rizzoli Orthopedic Institute.

e23501Background: Osteosarcoma is a rare disease recurring mainly to the lungs. In unoperable cases high dose ifosfamide, gemcitabine/docetaxel, /etoposide are employed. Very few studies are availa...