Transcriptional regulation and therapeutic potential of cyclin-dependent kinase 9 (CDK9) in sarcoma.

Sarcomas include various subtypes comprising two significant groups - soft tissue and bone sarcomas. Although the survival rate for some sarcoma subtypes has improved over time, the current methods of treatment remain efficaciously limited, as recurrent, and metastatic diseases remain a major obstacle. There is a need for better options and therapeutic strategies in treating sarcoma. Cyclin dependent kinase 9 (CDK9) is a transcriptional kinase and has emerged as a promising target for treating various cancers. The aberrant expression and activation of CDK9 have been observed in several sarcoma subtypes, including rhabdomyosarcoma, synovial sarcoma, osteosarcoma, Ewing sarcoma, and chordoma. Enhanced CDK9 expression has also been correlated with poorer prognosis in sarcoma patients. As a master regulator of transcription, CDK9 promotes transcription elongation by phosphorylation and releasing RNA polymerase II (RNAPII) from its promoter proximal pause. Release of RNAPII from this pause induces transcription of critical genes in the tumor cell. Overexpression and activation of CDK9 have been observed to lead to the expression of oncogenes, including MYC and MCL-1, that aid sarcoma development and progression. Inhibition of CDK9 in sarcoma has been proven to reduce these oncogenes' expression and decrease proliferation and growth in different sarcoma cells. Currently, there are several CDK9 inhibitors in preclinical and clinical investigations. This review aims to highlight the recent discovery and results on the transcriptional role and therapeutic potential of CDK9 in sarcoma.

Pedicled omental flaps for complex wound reconstruction following surgery for primary spine tumors of the mobile spine and sacrum.

OBJECTIVE: Surgery for primary tumors of the mobile spine and sacrum often requires complex reconstruction techniques to cover soft-tissue defects and to treat wound and CSF-related complications. The anatomical, vascular, and immunoregulatory characteristics of the omentum make it an excellent local substrate for the management of radiation soft-tissue injury, infection, and extensive wound defects. This study describes the authors' experience in complex wound reconstruction using pedicled omental flaps to cover defects in surgery for mobile spine and sacral primary tumors. METHODS: A retrospective cohort analysis was conducted on 34 patients who underwent pedicled omental flap reconstruction after en bloc resection of primary sacral and mobile spine tumors between 2010 and 2020. The study focused on assessing the indications for omental flap usage, including soft-tissue coverage, protection against postoperative radiation therapy, infection management, vascular supply for bone grafts, and dural defect and CSF leak repair. Patient demographic characteristics, tumor characteristics, surgical outcomes, and follow-up data were analyzed to determine the procedure's efficacy and complication rates. RESULTS: From 2010 to 2020, 34 patients underwent pedicled omental flap reconstruction after en bloc resection of sacral (24 of 34 [71%]) and mobile spine (10 of 34 [29%]) primary tumors, mostly chordomas. The patient cohort included 21 men and 13 women with a median (range) age of 60 (32-89) years. The most common indication for omental flap was soft-tissue coverage (20 of 34 [59%]). Other indications included protecting abdominopelvic organs for postoperative radiation therapy (6 of 34 [18%]), treating infections (5 of 34 [15%]), providing vascular supply for free fibular bone graft (1 of 34 [3%]), and repairing large dural defects and CSF leak (2 of 34 [6%]). The median (range) follow-up was 24 (0-132) months, during which 71% (24 of 34) of patients did not require additional surgery for wound-related complications. At last follow-up, 59% (20 of 34) had stable disease and 32% (11 of 34) had recurrence, had progression of disease, or had been discharged to hospice after treatment. CONCLUSIONS: The pedicled omentum is an effective local tissue graft that can be used for complex wound reconstruction and management of high-risk closures in primary spine tumors. This technique may have a lower rate of complications than other approaches and may influence surgical planning and flap selection in challenging cases.

Erratum: ATR inhibition sensitizes liposarcoma to doxorubicin by increasing DNA damage.

[This corrects the article on p. 1577 in vol. 12, PMID: 35530299.].

Pilot Study: Short Term Impact of Radiation Therapy on Bone Mineral Density and Bone Metabolism.

Despite the risk of complications, high dose radiation therapy is increasingly utilized in the management of selected bone malignancies. In this study, we investigate the impact of moderate to high dose radiation (over 50 Gy) on bone metabolism and structure. Between 2015 and 2018, patients with a primary malignant bone tumor of the sacrum that were either treated with high dose definitive radiation only or a combination of moderate to high dose radiation and surgery were prospectively enrolled at a single institution. Quantitative CTs were performed before and after radiation to determine changes in volumetric bone mineral density (BMD) of the irradiated and non-irradiated spine. Bone histomorphometry was performed on biopsies of the irradiated sacrum and the non-irradiated iliac crest of surgical patients using a quadruple tetracycline labeling protocol. In total, 9 patients were enrolled. Two patients received radiation only (median dose 78.3 Gy) and 7 patients received a combination of preoperative radiation (median dose 50.4 Gy), followed by surgery. Volumetric BMD of the non-irradiated lumbar spine did not change significantly after radiation, while the BMD of the irradiated sacrum did (pre-radiation median: 108.0 mg/cm3 (IQR 91.8-167.1); post-radiation median: 75.3 mg/cm3 (IQR 57.1-110.2); p = 0.010). The cancellous bone of the non-irradiated iliac crest had a stable bone formation rate, while the irradiated sacrum showed a significant decrease in bone formation rate [pre-radiation median: 0.005 mm3/mm2/year (IQR 0.003-0.009), post-radiation median: 0.001 mm3/mm2/year (IQR 0.001-0.001); p = 0.043]. Similar effects were seen in the cancellous and endocortical envelopes. This pilot study shows a decrease of volumetric BMD and bone formation rate after high-dose radiation therapy. Further studies with larger cohorts and other endpoints are needed to get more insight into the effect of radiation on bone. Level of evidence: IV.

Mesenchymal Stem/Stromal Cells: Immunomodulatory and Bone Regeneration Potential after Tumor Excision in Osteosarcoma Patients.

Osteosarcoma (OS) is a type of bone cancer that is derived from primitive mesenchymal cells typically affecting children and young adults. The current standard of treatment is a combination of neoadjuvant chemotherapy and surgical resection of the cancerous bone. Post-resection challenges in bone regeneration arise. To determine the appropriate amount of bone to be removed, preoperative imaging techniques such as bone and CT scans are employed. To prevent local recurrence, the current standard of care suggests maintaining bony and soft tissue margins from 3 to 7 cm beyond the tumor. The amount of bone removed in an OS patient leaves too large of a deficit for bone to form on its own and requires reconstruction with metal implants or allografts. Both methods require the bone to heal, either to the implant or across the allograft junction, often in the setting of marrow-killing chemotherapy. Therefore, the issue of bone regeneration within the surgically resected margins remains an important challenge for the patient, family, and treating providers. Mesenchymal stem/stromal cells (MSCs) are potential agents for enhancing bone regeneration post tumor resection. MSCs, used with scaffolds and growth factors, show promise in fostering bone regeneration in OS cases. We spotlight two MSC types-bone marrow-derived (BM-MSCs) and adipose tissue-derived (ASCs)-highlighting their bone regrowth facilitation and immunomodulatory effects on immune cells like macrophages and T cells, enhancing therapeutic outcomes. The objective of this review is two-fold: review work demonstrating any ability of MSCs to target the deranged immune system in the OS microenvironment, and synthesize the available literature on the use of MSCs as a therapeutic option for stimulating bone regrowth in OS patients post bone resection. When it comes to repairing bone defects, both MB-MSCs and ASCs hold great potential for stimulating bone regeneration. Research has showcased their effectiveness in reconstructing bone defects while maintaining a non-tumorigenic role following wide resection of bone tumors, underscoring their capability to enhance bone healing and regeneration following tumor excisions.

Discoidin Domain Receptor Tyrosine Kinase 1 (DDR1) Is a Novel Therapeutic Target in Liposarcoma: A Tissue Microarray Study.

BACKGROUND: Liposarcoma is the most commonly diagnosed subtype of soft tissue sarcoma. As these tumors often arise near vital organs and neurovascular structures, complete resection can be challenging; consequently, recurrence rates are high. Additionally, available chemotherapeutic agents have shown limited benefit and substantial toxicities. There is, therefore, a clear and unmet need for novel therapeutics for liposarcoma. Discoidin domain receptor tyrosine kinase 1 (DDR1) is involved in adhesion, proliferation, differentiation, migration, and metastasis in several cancers. However, the expression and clinical importance of DDR1 in liposarcoma are unknown. QUESTIONS/PURPOSES: The purposes of this study were to assess (1) the expression, (2) the association between DDR1 and survival, and (3) the functional roles of DDR1 in liposarcoma. METHODS: The correlation between DDR1 expression in tumor tissues and clinicopathological features and survival was assessed via immunohistochemical staining of a liposarcoma tissue microarray. It contained 53 samples from 42 patients with liposarcoma and 11 patients with lipoma. The association between DDR1 and survival in liposarcoma was analyzed by Kaplan-Meier plots and log-rank tests. The DDR1 knockout liposarcoma cell lines were generated by CRISPR-Cas9 technology. The DDR1-specific and highly selective DDR1 inhibitor 7RH was applied to determine the impact of DDR1 expression on liposarcoma cell growth and proliferation. In addition, the effect of DDR1 inhibition on liposarcoma growth was further accessed in a three-dimensional cell culture model to mimic DDR1 effects in vivo. RESULTS: The results demonstrate elevated expression of DDR1 in all liposarcoma subtypes relative to benign lipomas. Specifically, high DDR1 expression was seen in 55% (23 of 42) of liposarcomas and no benign lipomas. However, DDR1 expression was not found to be associated with poor survival in patients with liposarcoma. DDR1 knockout or treatment of 7RH showed decreased liposarcoma cell growth and proliferation. CONCLUSION: DDR1 is aberrantly expressed in liposarcoma, and it contributes to several markers of oncogenesis in these tumors. CLINICAL RELEVANCE: This work supports DDR1 as a promising therapeutic target in liposarcoma.

Transcriptional Profiling Supports the Notochordal Origin of Chordoma and Its Dependence on a TGFB1-TBXT Network.

Chordoma is a rare malignant tumor demonstrating notochordal differentiation. It is dependent on brachyury (TBXT), a hallmark notochordal gene and transcription factor, and shares histologic features and the same anatomic location as the notochord. This study involved a molecular comparison of chordoma and notochord to identify dysregulated cellular pathways. The lack of a molecular reference from appropriate control tissue limits our understanding of chordoma and its relationship to notochord. Therefore, an unbiased comparison of chordoma, human notochord, and an atlas of normal and cancerous tissue was conducted using gene expression profiling to clarify the chordoma/notochord relationship and potentially identify novel drug targets. The study found striking consistency in gene expression profiles between chordoma and notochord, supporting the hypothesis that chordoma develops from notochordal remnants. A 12-gene diagnostic chordoma signature was identified and the TBXT/transforming growth factor beta (TGF-ß)/SOX6/SOX9 pathway was hyperactivated in the tumor, suggesting that pathways associated with chondrogenesis were a central driver of chordoma development. Experimental validation in chordoma cells confirmed these findings and emphasized the dependence of chordoma proliferation and survival on TGF-ß. The computational and experimental evidence provided the first molecular connection between notochord and chordoma and identified core members of a chordoma regulatory pathway involving TBXT. This pathway provides new therapeutic targets for this unique malignant neoplasm and highlights TGF-ß as a prime druggable candidate.

Analysis of Questions in Sections of the Orthopaedic In-Training Examination: A Scoping Review.

PURPOSE: The purpose of this review was to assess all available studies that analyzed the types of questions in individual sections of the Orthopaedic In-Training Examination, which may be used as a reference for residents studying for their examination. METHODS: Following the Providing Innovative Service Models and Assessment extension for Scoping Reviews guidelines, a systematic review was conducted on studies that report on sections or question categories of the Orthopaedic In-Training Examination using PubMed, MEDLINE, and Web of Science databases. Two reviewers and an arbitrator reviewed and extracted relevant data from 20 included studies which made up the systematic review. RESULTS: All 20 studies in the review reported the mean number of questions per section, with the highest coming from musculoskeletal trauma (18.9% to 19.0%). 18 studies reported the Buckwalter taxonomic classification; 42.0% of questions were T1, 18.2% were T2, and 39.5% were T3 with a wide range from section to section. Primary sources were nearly three times more likely to be cited when compared with textbook sources. There were 12 journals that were commonly cited with the most being the Journal of Bone and Joint Surgery: American Volume (17/18). DISCUSSION: This study accurately portrays the characteristics of each section of the Orthopaedic In-Training Examination over the past 10 years. These data suggest that orthopaedic residents may be inclined to focus on musculoskeletal trauma, topics related to clinical management, and primary journal sources for studying. In addition, residency programs may choose to focus on higher yield sources or material to prepare their residents for the examination.

Advances in the development of chordoma models for drug discovery and precision medicine.

Chordomas are malignant bone tumors that arise from remnants of the notochord. These tumors are generally slow-growing, locally aggressive, and invasive. Chordomas are typically resistant to conventional chemo- and radiotherapy. The clinical management of this disease is very challenging, usually, treatment is surgical resection, which may be combined with radiotherapy. Although chordomas have undergone histologic and genetic analysis, the molecular mechanisms that drive their pathogenesis and resistance are still largely unknown. For many years this could be attributed to the lack of accurate and reliable in vitro and in vivo tumor models. Yet, over the past decade, many efforts have been made to prioritize the generation of useful chordoma cell lines, and tumor models that have shed more light on this malignancy and have made efficacious drug discovery a greater possibility. This review summarizes and discusses recent enhancements and improvements made to generate useful chordoma models and their applications in drug discovery and precision medicine.

In situ hybridization to detect DNA amplification in extracellular vesicles.

EVs have emerged as an important component in tumour initiation, progression and metastasis. Although notable progresses have been made, the detection of EV cargoes remain significantly challenging for researchers to practically use; faster and more convenient methods are required to validate the EV cargoes, especially as biomarkers. Here we show, the possibility of examining embedded EVs as substrates to be used for detecting DNA amplification through ultrasensitive in situ hybridization (ISH). This methodology allows the visualization of DNA targets in a more direct manner, without time consuming optimization steps or particular expertise. Additionally, formalin-fixed paraffin-embedded (FFPE) blocks of EVs allows long-term preservation of samples, permitting future studies. We report here: (i) the successful isolation of EVs from liposarcoma tissues; (ii) the EV embedding in FFPE blocks (iii) the successful selective, specific ultrasensitive ISH examination of EVs derived from tissues, cell line, and sera; (iv) and the detection of MDM2 DNA amplification in EVs from liposarcoma tissues, cell lines and sera. Ultrasensitive ISH on EVs would enable cargo study while the application of ISH to serum EVs, could represent a possible novel methodology for diagnostic confirmation. Modification of probes may enable researchers to detect targets and specific DNA alterations directly in tumour EVs, thereby facilitating detection, diagnosis, and improved understanding of tumour biology relevant to many cancer types.

Pedicled vastus lateralis myocutaneous flap for sacropelvic defects after wide oncologic resection: Wound complications and outcomes.

BACKGROUND AND OBJECTIVES: Adequate coverage of the soft tissue defects from wide resection of sacropelvic malignancies remains challenging. The vastus lateralis flap has been described for coverage in the setting of trauma and infection. This flap has not been described for coverage of sacropelvic tumor defects. METHODS: This is a retrospective cohort study of adult patients who underwent wide resection of a primary sacropelvic malignancy with reconstruction employing a pedicled vastus lateralis flap at two tertiary care centers. Patient demographics, tumor staging, and rate of complications were assessed. RESULTS: Twenty-eight patients were included, with a median age of 51 years. The most common primary tumor was chondrosarcoma followed by chondroblastic osteosarcoma. The median follow-up was 1.1 years. There were 10 cases of wound infection requiring re-operation and three cases of flap failure. CONCLUSIONS: We describe a pedicled vastus lateralis flap for coverage of defects after wide resection of sacropelvic malignancies. A large proportion of our cohort had independent risk factors for wound complications. Even with a cohort with high baseline risk for wound complications, we show that the use of a pedicled vastus lateralis flap is a safe reconstructive option with a wound complication rate in line with the literature.

ATR inhibition sensitizes liposarcoma to doxorubicin by increasing DNA damage.

Liposarcomas account for approximately 20% of all adult sarcomas and have limited therapeutic options outside of surgery. Inhibition of ataxia-telangiectasia and Rad3 related protein kinase (ATR) has emerged as a promising chemotherapeutic strategy in various cancers. However, its activation, expression, and function in liposarcoma remain unkown. In this study, we investigated the expression, function, and potential of ATR as a therapeutic target in liposarcoma. Activation and expression of ATR in liposarcoma was analyzed by immunohistochemistry, which was further explored for correlation with patient clinical characteristics. ATR-specific siRNA and the ATR inhibitor VE-822 were applied to determine the effect of ATR inhibition on liposarcoma cell proliferation and anti-apoptotic activity. Migration activity and clonogenicity were examined using wound healing and clonogenic assays. ATR (p-ATR) was overexpressed in 88.1% of the liposarcoma specimens and correlated with shorter overall survival in patients. Knockdown of ATR via specific siRNA or inhibition with VE-822 suppressed liposarcoma cell growth, proliferation, migration, colony-forming ability, and spheroid growth. Importantly, ATR inhibition significantly and synergistically enhanced liposarcoma cell line chemosensitivity to doxorubicin. Our findings support ATR as critical to liposarcoma proliferation and doxorubicin resistance. Therefore, the addition of ATR inhibition to a standard doxorubicin regimen is a potential treatment strategy for liposarcoma.

Expression and Clinical Significance of High-Mobility Group AT-hook 2 (HMGA2) in Osteosarcoma.

OBJECTIVE: Although high-mobility group AT-hook 2 (HMGA2) has been shown to have crucial roles in the pathogenesis and metastasis of various malignancies, its expression and significance in osteosarcoma remain unknown. Here we evaluate the expression, clinical prognostic value, and overall function of HMGA2 in osteosarcoma. METHODS: Sixty-nine osteosarcoma patient specimens within a tissue microarray (TMA) were analyzed by immunohistochemistry for HMGA2 expression. Demographics and clinicopathological information including age, gender, tumor location, metastasis, recurrence, chemotherapy response, follow-up time, and disease status were also collected. After validation of expression, we determined whether there was a correlation between HMGA2 expression and patient clinicopathology. HMGA2 expression was also evaluated in osteosarcoma cell lines and patient tissues by Western blot, we analyzed the expression of HMGA2 in the human osteosarcoma cell lines MG63, 143B, U2OS, Saos-2, MNNG/HOS, and KHOS. HMGA2-specific siRNA and clonogenic assays were then used to determine the effect of HMGA2 inhibition on osteosarcoma cell proliferation, growth, and chemosensitivity. RESULTS: HMGA2 expression was elevated in the osteosarcoma patient specimens and human osteosarcoma cell lines. HMGA2 was differentially expressed in human osteosarcoma cell lines. Specifically, a relatively high expression of HMGA2 was present in KHOS, MNNG/HOS, 143B and a relatively low expression was in MG63, U2OS as well as Saos-2. HMGA2 expression is correlated with metastasis and shorter overall survival. High HMGA2 expression is an independent predictor of poor osteosarcoma prognosis. There was no significant correlation between HMGA2 expression and the age, gender, or tumor site of the patient. HMGA2 expression is predominantly within the nucleus. The expression of HMGA2 also directly correlated to neoadjuvant chemoresistance. There was a significant reduction of HMGA2 expression in the siRNA transfection group. After the use of siRNA, the proliferation of osteosarcoma cells is decreased and the chemosensitivity of osteosarcoma cells is significantly increased. CONCLUSION: Our study supports HMGA2 as a potential prognostic biomarker and therapeutic target in osteosarcoma.

Inhibition of CDK7-dependent transcriptional addiction is a potential therapeutic target in synovial sarcoma.

Synovial sarcoma is typical aggressive malignant without satisfactory treatment outcome in adult series. Cyclin-dependent kinases (CDKs) in transcription have been considered promising molecular targets in cancer. Among these, CDK7 has been shown to play important roles in the pathogenesis of malignancies. However, the modulation mechanism of CDK7-regulated transcription in synovial sarcoma is unknown. In the present study, we aim to determine the expression and function of CDK7 in the transcription cycle of RNA polymerase II (RNAP II), and evaluate its prognostic and therapeutic significance in synovial sarcoma. Results showed that overexpression of CDK7 correlates with higher clinical stage and grade, and worse outcomes in clinic. High CDK7 expression was confirmed in all tested human synovial sarcoma cell lines and CDK7 was largely localized to the cell nucleus. Downregulation through siRNA or inhibition with the CDK7-targeting agent BS-181 exhibited dose-dependent cytotoxicity and prevented cell colony formation. Western blots demonstrated that inhibition of CDK7 paused transcription by a reduction of RNAP II phosphorylation. Blocking CDK7-dependent transcriptional addiction was accompanied by promotion of apoptosis. Furthermore, the CDK7-specific inhibitor reduced 3D spheroid formation and migration of synovial sarcoma. Collectively, our findings highlight the role of CDK7-dependent transcriptional addiction in human synovial sarcoma. CDK7-specific cytotoxic agents are therefore promising novel treatment options for synovial sarcoma.

Definitive high-dose, proton-based radiation for unresected mobile spine and sacral chordomas.

BACKGROUND/PURPOSE: Treatment of spine and sacral chordoma generally involves surgical resection, usually in conjunction with radiation therapy.In certain locations, resection may result in significant neurological dysfunction, so definitive radiation has been used as an alternative to surgery. The purpose of this study is to report the results of high-dose, proton-based definitive radiotherapy for unresected spinal and sacral chordomas. MATERIALS/METHODS: Retrospective review of 67 patients with newly diagnosed, unresected spinal chordomas treated with high-dose definitive, proton-based radiotherapy at our center from 1975 to 2019. RESULTS: Reasons for radiotherapy alone included medical inoperability and/or concern for neurological dysfunction based on spine level or patient choice. Tumor locations included cervical (n = 10), thoracic (n = 1), lumbar (n = 4) spine, and sacrum (n = 52). Median maximal tumor diameter was 7.4 cm (range 1.8-25 cm). Median total dose was 77.4 Gy (RBE) (range 73.8-85.9 Gy RBE). Analysis with median follow-up of 56.2 months (range, 4-171.7 months) showed overall survival of 83.5 % (95%CI: 69.4-91.5%) and 65.9% (95%CI: 47.3-79.3%), disease-free survival of 64% (95%CI: 49.3-75.4) and 44.1% (95%CI: 27.8-59.2%), local control of 81.8% (95%CI: 67.6-90.2%) and 63.6% (95%CI: 44.7-77.5%), and distant control of 77.4% (95%CI: 63.6-86.5%) and 72.5% (95%CI: 55.7-83.8%) at 5 and 8 years respectively. The most common late side effect was insufficiency fracture. CONCLUSION: These results continue to support the use of high-dose definitive radiotherapy for patients with medically inoperable or otherwise unresected mobile spine or sacrococcygeal chordomas. There is a trend towards better disease-free survival with doses > 78 Gy (RBE).

SMARCB1 expression is a novel diagnostic and prognostic biomarker for osteosarcoma.

BACKGROUND: Although weak SWI/SNF related matrix-associated actin-dependent regulator of chromatin subfamily B member 1 (SMARCB1) expression is a known diagnostic and prognostic biomarker in several malignancies, its expression and clinical significance in osteosarcoma remain unknown. The aim of the present study was to investigate SMARCB1 expression in osteosarcoma and its clinical significance with respect to chemosensitivity and prognosis. METHODS: We obtained 114 specimens from 70 osteosarcoma patients to construct a tissue microarray (TMA) and assess SMARCB1 protein expression via immunohistochemistry (IHC). The mRNA expression of SMARCB1 was in-silico analyzed using open-access RNA sequencing (RNA-Seq) and clinicopathological data provided by the Therapeutically Applicable Research to Generate Effective Treatments on Osteosarcoma (TARGET-OS) project. The correlations between SMARCB1 expression and clinical features were statistically analyzed. RESULTS: Weak SMARCB1 expression occurred in 70% of the osteosarcoma patient specimens in the TMA, and significantly correlated with poor neoadjuvant response as well as shorter overall and progression-free survival (PFS). In addition, mRNA in-silico analysis confirmed that SMARCB1 expression correlates with chemotherapeutic response and prognosis in osteosarcoma patients. CONCLUSION: To our knowledge, the present study is the first to analyze SMARCB1 expression in osteosarcoma. SMARCB1 may serve as a novel diagnostic and prognostic biomarker in osteosarcoma.

Establishment and Characterization of a Novel Dedifferentiated Chondrosarcoma Cell Line DDCS2.

BACKGROUND: The dedifferentiated variant of chondrosarcoma is highly aggressive and carries an especially grim prognosis. While chemotherapeutics has failed to benefit patients with dedifferentiated chondrosarcoma significantly, preclinical chemosensitivity studies have been limited by a scarcity of available cell lines. There is, therefore, an urgent need to expand the pool of available cell lines. METHODS: We report the establishment of a novel dedifferentiated chondrosarcoma cell line DDCS2, which we isolated from the primary tumor specimen of a 60-year-old male patient. We characterized its short tandem repeat (STR) DNA profile, growth potential, antigenic markers, chemosensitivity, and oncogenic spheroid and colony-forming capacity. RESULTS: DDCS2 showed a spindle to polygonal shape and an approximate 60-hour doubling time. STR DNA profiling revealed a unique genomic identity not matching any existing cancer cell lines within the ATCC, JCRB, or DSMZ databases. There was no detectable contamination with another cell type. Western blot and immunofluorescence assays were consistent with a mesenchymal origin, and our MTT assay revealed relative resistance to conventional chemotherapeutics, which is typical of a dedifferentiated chondrosarcoma. Under ex vivo three-dimensional (3D) culture conditions, the DDCS2 cells produced spheroid patterns similar to the well-established CS-1 and SW1353 chondrosarcoma cell lines. CONCLUSION: Our findings confirm DDCS2 is a novel model for dedifferentiated chondrosarcoma and therefore adds to the limited pool of current cell lines urgently needed to investigate the chemoresistance within this deadly cancer.

Aggressive osteoblastoma with a secondary aneurysmal bone cyst treated with denosumab.

Osteoblastomas and aneurysmal bone cysts (ABC) are rare benign bone tumors that make up about 1%-2% of primary bone malignancies, typically occurring in young patients with a median age of 20 years, most commonly effecting the axial skeleton. ABCs may develop independently as primary lesions, or secondary to other bony lesions including osteoblastomas, chondroblastomas, and giant cell tumors. Treatment of unresectable or extensive osteoblastomas can be challenging. In 2013, the Food and Drug Administration (FDA) approved denosumab for the treatment of giant cell tumors of the bone due to its efficacy in these morbid bony lesions. Various case reports have shown that osteoblastomas can respond to denosumab. Furthermore, numerous ABC case reports have described the efficacy of denosumab in these situations. We herein describe a unique case of a young patient with an aggressive osteoblastoma and secondary ABCs who was successfully treated with denosumab.

Cyclin-dependent kinase 7 (CDK7) is an emerging prognostic biomarker and therapeutic target in osteosarcoma.

BACKGROUND: Overexpression of cyclin-dependent kinase 7 (CDK7) is a well-known pathogenic feature of various malignancies and a sign of a more dismal prognosis. As relatively little is known about CDK7 in osteosarcoma, we elected to evaluate its expression, prognostic value, and function. METHODS: We began by analyzing the publicly available data sets on CDK7 expression, including RNA sequencing data from the Therapeutically Applicable Research to Generate Effective Treatments on Osteosarcoma (TARGET-OS) and the Gene Expression database of Normal and Tumor tissues 2 (GENT2). The correlation between patient tissue CDK7 expression and their clinicopathological features and prognosis was assessed via immunohistochemical staining of a unique tissue microarray constructed from osteosarcoma specimens. Furthermore, we analyzed CDK7 expression in osteosarcoma cell lines and tissues by Western blot. CDK7-specific siRNA and a highly-selective CDK7 inhibitor, BS-181, were applied to determine the function of CDK7 on osteosarcoma cell growth and proliferation. In addition, the effect of CDK7 inhibition on clonogenicity was evaluated using a clonogenic assay, and a 3D cell culture model was used to mimic CDK7 effects in an in vivo environment. RESULTS: Our results demonstrate that higher CDK7 expression significantly correlates with recurrence, metastasis, and shorter overall survival in osteosarcoma patients. Therapeutically, we show that CDK7 knockdown with siRNA or selective inhibition with BS-181 decreases proliferation and induces apoptosis of osteosarcoma cells. CONCLUSION: This study supports CDK7 overexpression as an independent predictor of poor prognosis and promising therapeutic target for osteosarcoma.

T-LAK cell-originated protein kinase (TOPK): an emerging prognostic biomarker and therapeutic target in osteosarcoma.

T-lymphokine-activated killer (T-LAK) cell-originated protein kinase (TOPK) is an emerging target with critical roles in various cancers; however, its expression and function in osteosarcoma remain unexplored. We evaluated TOPK expression using RNA sequencing and gene expression data from public databases (TARGET-OS, CCLE, GTEx, and GENT2) and immunohistochemistry in an osteosarcoma tissue microarray (TMA). TOPK gene expression was significantly higher in osteosarcoma than normal tissues and directly correlated with shorter overall survival. TOPK was overexpressed in 83.3% of the osteosarcoma specimens within our TMA and all osteosarcoma cell lines, whereas normal osteoblast cells had no aberrant expression. High expression of TOPK associated with metastasis, disease status, and shorter overall survival. Silencing of TOPK with small interfering RNA (siRNA) decreased cell viability, and inhibition with the selective inhibitor OTS514 suppressed osteosarcoma cell proliferation, migration, colony-forming ability, and spheroid growth. Enhanced chemotherapeutic sensitivity and a synergistic effect were also observed with the combination of OTS514 and either doxorubicin or cisplatin in osteosarcoma cell lines. Taken together, our study demonstrated that TOPK is a potential prognostic biomarker and therapeutic target for osteosarcoma treatment.