What Is the Prevalence of Clinically Important Findings Among Incidentally Found Osseous Lesions?

BACKGROUND: Patients with incidentally found musculoskeletal lesions are regularly referred to orthopaedic oncology. Most orthopaedic oncologists understand that many incidental findings are nonaggressive and can be managed nonoperatively. However, the prevalence of clinically important lesions (defined as those indicated for biopsy or treatment, and those found to be malignant) remains unknown. Missing clinically important lesions can result in harm to patients, but needless surveillance may exacerbate patient anxiety about their diagnosis and accrue low-value costs to the payor. QUESTIONS/PURPOSES: (1) What percentage of patients with incidentally discovered osseous lesions referred to orthopaedic oncology had lesions that were clinically important, defined as those receiving biopsy or treatment or those found to be malignant? (2) Using standardized Medicare reimbursements as a surrogate for payor expense, what is the value of reimbursements accruing to the hospital system for the imaging of incidentally found osseous lesions performed during the initial workup period and during the surveillance period, if indicated? METHODS: This was a retrospective study of patients referred to orthopaedic oncology for incidentally found osseous lesions at two large academic hospital systems. Medical records were queried for the word "incidental," and matches were confirmed by manual review. Patients evaluated at Indiana University Health between January 1, 2014, and December 31, 2020, and those evaluated at University Hospitals between January 1, 2017, and December 31, 2020, were included. All patients were evaluated and treated by the two senior authors of this study and no others were included. Our search identified 625 patients. Sixteen percent (97 of 625) of patients were excluded because their lesions were not incidentally found, and 12% (78 of 625) were excluded because the incidental findings were not bone lesions. Another 4% (24 of 625) were excluded because they had received workup or treatment by an outside orthopaedic oncologist, and 2% (10 of 625) were excluded for missing information. A total of 416 patients were available for preliminary analysis. Among these patients, 33% (136 of 416) were indicated for surveillance. The primary indication for surveillance included lesions with a benign appearance on imaging and low clinical suspicion of malignancy or fracture. A total of 33% (45 of 136) of these patients had less than 12 months of follow-up and were excluded from further analysis. No minimum follow-up criteria were applied to patients not indicated for surveillance because this would artificially inflate our estimated rate of clinically important findings. A total of 371 patients were included in the final study group. Notes from all clinical encounters with orthopaedic and nonorthopaedic providers were screened for our endpoints (biopsy, treatment, or malignancy). Indications for biopsy included lesions with aggressive features, lesions with nonspecific imaging characteristics and a clinical picture concerning for malignancy, and lesion changes seen on imaging during the surveillance period. Indications for treatment included lesions with increased risk of fracture or deformity, certain malignancies, and pathologic fracture. Diagnoses were determined using biopsy results if available or the documented opinion of the consulting orthopaedic oncologist. Imaging reimbursements were obtained from the Medicare Physician Fee Schedule for 2022. Because imaging charges vary across institutions and reimbursements vary across payors, this method was chosen to enhance the comparability of our findings across multiple health systems and studies. RESULTS: Seven percent (26 of 371) of incidental findings were determined to be clinically important, as previously defined. Five percent (20 of 371) of lesions underwent tissue biopsy, and 2% (eight of 371) received surgical intervention. Fewer than 2% (six of 371) of lesions were malignant. Serial imaging changed the treatment of 1% (two of 136) of the patients, corresponding to a rate of one in 47 person-years. Median reimbursements to work up the incidental findings analyzed was USD 219 (interquartile range USD 0 to 404), with a range of USD 0 to 890. Among patients indicated for surveillance, the median annual reimbursement was USD 78 (IQR USD 0 to 389), with a range of USD 0 to 2706. CONCLUSION: The prevalence of clinically important findings among patients referred to orthopaedic oncology for incidentally found osseous lesions is modest. The likelihood of surveillance resulting in a change of management was low, but the median reimbursements associated with following these lesions was also low. We conclude that after appropriate risk stratification by orthopaedic oncology, incidental lesions are rarely clinically important, and judicious follow-up with serial imaging can be performed without incurring high costs. LEVEL OF EVIDENCE: Level III, therapeutic study.

Ischial osteochondroma as an unusual source of pregnancy-related sciatic pain: a case report.

BACKGROUND: While most cases of sciatica result from degenerative conditions of the low back, some cases result from conditions of the hip and pelvic region. Sciatica developing in relation to pregnancy or labor also presents unique considerations. CASE PRESENTATION: A 37-year-old African American woman with a history of hypertension and polycystic ovary syndrome presented to a chiropractor at a hospital-based outpatient clinic with a seven-week history of low back pain with radiation into the right lower extremity which began during labor. The chiropractor performed a brief trial of care, yet when the patient's symptoms worsened, ordered lumbar spine radiographs, followed by lumbar magnetic resonance imaging (MRI), which were both normal. The chiropractor then ordered hip radiographs, which were suggestive of ischial osteochondroma, and referred the patient to an orthopedic oncologist. MRI findings were compatible with an osteochondroma with associated adventitial bursitis and mass effect on the sciatic nerve. The patient initially chose conservative management with bursa aspiration and therapeutic injection. Despite initial relief, there was eventual return of symptoms. The patient elected to undergo surgical removal, with a positive outcome. CONCLUSION: The key distinguishing features that led to a diagnosis of osteochondroma in this case included attention to the patient-reported symptoms and history, worsening of symptoms despite conservative care, and lack of explanatory findings on lumbar imaging. This case highlights the benefit of evaluating the hip and pelvis when the clinical features of sciatica cannot be ascribed to a lumbar etiology. This case also illustrates the role of a chiropractor working in an integrative health system to facilitate timely imaging and referrals to resolve a challenging diagnosis.

The Natural History of Benign Bone Tumors of the Extremities in Asymptomatic Children: A Longitudinal Radiographic Study.

BACKGROUND: Benign bone tumors are common incidental findings in the pediatric population during radiographic evaluation. Counseling these patients requires reassurance and raises questions about the natural history of these tumors over time. The purpose of this study was to estimate the prevalence and observe the behavior of benign childhood bone tumors in an asymptomatic population. METHODS: A historical, longitudinal radiographic collection of healthy children was reviewed, which included comprehensive left-sided radiographs of the extremities at yearly intervals. In this study, 262 subjects with 25,555 radiographs were screened for benign bone tumors at a median age of 8 years (range, 0 to 18 years). All potential tumors were reviewed by a multidisciplinary panel, which confirmed the radiographic diagnosis of each lesion, the age at which the lesion first appeared, and the age at which it had resolved. Prevalence rates were calculated using the number of distinct subjects available for each radiographic location and age. RESULTS: Thirty-five tumors were identified in 33 subjects, including 19 nonossifying fibromas, 8 enostoses, 6 osteochondromas, and 2 enchondromas. The prevalence rate for all tumors combined increased with age and was 18.9% overall. The overall prevalence rates for specific tumor types were 7.5% for nonossifying fibromas, 5.2% for enostoses, 4.5% for osteochondromas, and 1.8% for enchondromas. Nonossifying fibromas demonstrated a bimodal distribution of prevalence, with a peak at 5 years (10.8%) and another after skeletal maturity (13.3%). The median age at the first appearance for all tumors combined was 9 years (range, 2 to 15 years), but varied by tumor type. Nonossifying fibromas often resolved (7 [37%] of 19), with further resolution possible beyond the last available radiograph. Enostoses, osteochondromas, and enchondromas persisted until the last available radiographs in all subjects. CONCLUSIONS: The prevalence of benign childhood bone tumors of the extremities was 18.9% in a historical asymptomatic population. Longitudinal radiographs allowed observation of the timing of the first appearance and the potential for resolution for each tumor type. These findings provide unique evidence to answer many commonly encountered questions when counseling patients and their families on benign bone tumors. LEVEL OF EVIDENCE: Prognostic Level IV. See Instructions for Authors for a complete description of levels of evidence.

Comparison of Publication Rates for Musculoskeletal Oncology Abstracts Presented at National Meetings.

BACKGROUND: Scientific meetings provide a forum to disseminate new research and advance patient care. The American Academy of Orthopaedic Surgeons (AAOS), Connective Tissue Oncology Society (CTOS), and Musculoskeletal Tumor Society (MSTS) annual meetings are examples of such gatherings in the field of musculoskeletal oncology. After a review of select MSTS abstracts from 1991 to 1999 revealed a 41% publication rate in scientific journals, previous authors cautioned meeting attendees that the majority of abstracts may not survive rigorous peer review and may not be scientifically valid. Since two decades have passed, this study reexamined publication rates and characteristics in a contemporary and expanded cohort of oncology abstracts presented at the AAOS, CTOS, and MSTS annual meetings. METHODS: 1408 podium and poster abstracts from the AAOS (oncology-focused from 2013 to 2015), CTOS (2012 to 2014), and MSTS (2012 to 2014) annual meetings were reviewed to allow for a four-year publication window. Searches were performed with PubMed and Google Scholar databases to identify full-text publications using abstract keywords. Characteristics of each abstract and resulting publication were collected. Statistical analysis was performed using the chi-square and Kruskal-Wallis tests for time-independent comparisons, and the log-rank test after reverse Kaplan-Meier analysis for time-dependent comparisons. RESULTS: Abstract publication rates overall were higher for podium presentations (67%, 280 of 415) compared to poster presentations (53%, 530 of 993; p < 0.001). When both abstract types were combined, differences between meetings did not meet statistical significance (AAOS: 65%, 106 of 162; CTOS: 57%, 521 of 909; MSTS: 54%, 183 of 337, p=0.06). Abstracts from AAOS meetings were more often published prior to the first day of the meeting (AAOS: 24%, 25 of 106; CTOS: 10%, 52 of 521; MSTS: 14%, 25 of 183; p < 0.01). After excluding previously published abstracts, AAOS abstracts had the shortest time to publication (median: 10.8 months, interquartile range (IQR): 4.4 to 18.8 months), compared to those from CTOS (16.0 months, 8.4 to 25.9 months, p < 0.01) and MSTS (15 months, 7.9 to 25.0 months, p < 0.01) meetings. CTOS abstracts were published in higher impact journals (median: 3.7, IQR: 2.9 to 5.9), compared to those from AAOS (2.9, 1.9 to 3.2, p < 0.01) and MSTS (3.1, 2.3 to 3.1, p < 0.01) meetings. Finally, 7.7% (62 of 810) of published abstracts were presented at more than one meeting. CONCLUSIONS: Publication rates in this study were higher than previous reports in musculoskeletal oncology and comparable or better than recent reports for other orthopedic meetings. Comparisons across the AAOS, CTOS, and MSTS annual meetings highlight notable differences but suggest similarity overall in the quality of evidence presented with little overlap between meetings. Taken together, this study points to progress in the review processes used by the program committees, reaffirms the importance of critical appraisal when considering abstract findings, and supports the continued organization of multiple scientific meetings in musculoskeletal oncology.

The Interval Between Preoperative Radiation and Surgery Is Not Associated with Overall Survival for Soft-tissue Sarcomas: An Analysis of the National Cancer Database.

BACKGROUND: Most cancer centers prefer preoperative radiation therapy (preRT) over postoperative therapy to treat soft-tissue sarcoma (STS) to limit long-term fibrosis, joint stiffness, and edema. Surgery is often delayed after preRT to allow for tissue recovery and to reduce wound complications. However, the association between the time interval between preRT and surgery and survival is unknown. QUESTIONS/PURPOSES: (1) What factors are associated with the preRT-surgery interval in patients with STS? (2) Is the preRT-surgery interval associated with overall survival? METHODS: The National Cancer Database, a nationwide registry that includes 70% of all new cancers in the United States with 90% follow-up, was reviewed to identify 6378 patients who underwent preRT and surgical resection for a localized extremity or pelvic STS from 2004 to 2014. Patients were excluded if they had lymphatic or metastatic disease at diagnosis (23%; n = 1438), underwent neoadjuvant chemotherapy (24%; 1531), were missing vital status (8%; 487), had chemosensitive histologies (9%; 603), underwent radiation other than external beam (1%; 92), were missing preRT-surgery interval (1%; 45), or had a preRT-surgery interval greater than 120 days (< 1%; 6). A total of 2176 patients were included for analysis, with a mean preRT-surgery interval of 35 ± 16 days. A multiple linear regression model was generated to assess demographic, clinicopathologic, and treatment characteristics associated with the preRT-surgery interval. A Kaplan-Meier survival analysis was then conducted, stratified by the preRT-surgery interval, to assess survival over 10 years. Finally, a multivariate Cox regression analysis model was constructed to further evaluate the association between the preRT-surgery interval and overall survival, adjusted for demographic, clinicopathologic, and treatment characteristics. RESULTS: A longer preRT-surgery interval was associated with higher age (ß = 0.002 per year [95% CI 0.0 to 0.004]; p = 0.026), tumor location in the pelvis (compared with the lower extremity; ß = 0.15 [95% CI 0.082 to 0.22]; p < 0.001), and malignant peripheral nerve sheath tumor subtype (compared with undifferentiated pleomorphic sarcoma; ß = 0.17 [95% CI 0.044 to 0.29]; p = 0.008). A shorter preRT-surgery interval was associated with higher facility volume (ß = -0.002 per case [95% CI -0.003 to -0.002]; p = 0.026) and higher tumor stage (compared with Stage I; ß = -0.066 [95% CI -0.13 to -0.006]; p = 0.03 for Stage II; ß = -0.12 [95% CI -0.17 to -0.065]; p < 0.001 for Stage III). The 5-year overall survival rates were similar across all preRT-surgery interval groups: less than 3 weeks (66% [95% CI 60 to 72]), 3 to 4 weeks (65% [95% CI 60 to 71]), 4 to 5 weeks (65% [95% CI 60 to 71]), 5 to 6 weeks (66% [95% CI 60 to 72]), 6 to 7 weeks (63% [95% CI 54 to 72]), 7 to 9 weeks (66% [95% CI 58 to 74]), and more than 9 weeks (59% [95% CI 48 to 69]). Over 10 years, no difference in overall survival was observed when stratified by the preRT-surgery interval (p = 0.74). After controlling for potentially confounding variables, including age, sex, Charlson/Deyo comorbidity score, histology, tumor size, stage and surgery type, the preRT-surgery interval was not associated with survival (hazard ratio = 1 per day [95% CI 1 to 1]; p = 0.88). CONCLUSION: With the numbers available, this study demonstrates that a delay in surgery up to 120 days after radiation is not associated with poorer survival. Therefore, clinicians may be able to delay surgery to minimize the risks of wound complications and modifiable comorbidities without affecting overall survival.Level of Evidence Level III, therapeutic study.

Targeting the Cancer Epigenome with Histone Deacetylase Inhibitors in Osteosarcoma.

Epigenetic deregulation is an emerging hallmark of cancer that enables tumor cells to escape surveillance by tumor suppressors and ultimately progress. The structure of the epigenome consists of covalent modifications of chromatin components, including acetylation by histone acetyltransferases (HATs) and deacetylation by histone deacetylases (HDACs). Targeting these enzymes with inhibitors to restore epigenetic homeostasis has been explored for many cancers. Osteosarcoma, an aggressive bone malignancy that primarily affects children and young adults, is notable for widespread genetic and epigenetic instability. This may explain why therapy directed at unique molecular pathways has failed to substantially improve outcomes in osteosarcoma over the past four decades. In this review, we discuss the potential of targeting the cancer epigenome, with a focus on histone deacetylase inhibitors (HDACi) for osteosarcoma. We additionally highlight the safety and tolerance of HDACi, combination chemotherapy with HDACi, and the ongoing challenges in the development of these agents.

Receptor Tyrosine Kinases in Osteosarcoma: 2019 Update.

The primary conclusions of our 2014 contribution to this series were as follows: Multiple receptor tyrosine kinases (RTKs) likely contribute to aggressive phenotypes in osteosarcoma and, therefore, inhibition of multiple RTKs is likely necessary for successful clinical outcomes. Inhibition of multiple RTKs may also be useful to overcome resistance to inhibitors of individual RTKs as well as resistance to conventional chemotherapies. Different combinations of RTKs are likely important in individual patients. AXL, EPHB2, FGFR2, IGF1R, and RET were identified as promising therapeutic targets by our in vitro phosphoproteomic/siRNA screen of 42 RTKs in the highly metastatic LM7 and 143B human osteosarcoma cell lines. This chapter is intended to provide an update on these topics as well as the large number of osteosarcoma clinical studies of inhibitors of multiple tyrosine kinases (multi-TKIs) that were recently published.

Six-Week Interval Between Preoperative Radiation and Surgery Is Associated With Fewer Major Wound Complications in Soft Tissue Sarcoma.

OBJECTIVES: A total of 30% to 40% of soft tissue sarcoma (STS) patients develop major wound complications (MWCs) after preoperative radiation (preRT). The optimal preRT-surgery interval and its association with MWCs is unknown. This study investigated whether a longer preRT-surgery interval is associated with fewer MWCs compared with historical controls. METHODS: All patients treated by a single surgeon after preRT with limb-sparing wide resection for extremity and trunk STS were retrospectively reviewed from 2004 to 2014. The primary outcome was MWCs defined as a secondary operation, invasive procedure, wound packing, or readmission for wound care. Secondary outcomes of local recurrence and survival were followed and independent variables were analyzed for an association with MWCs. RESULTS: Fifty-four patients were included with a median follow-up of 32 months and age of 61 years. The majority of tumors were deep (91%), large (median size of 11 cm), high grade (78%) and within the lower extremity (78%). The median preRT-surgery interval was 43 days and 80% of patients received surgery 35 to 49 days after radiation. MWCs were observed in 15% of patients and 88% occurred within 40 days. Predictors for MWCs on multivariate analysis were peripheral vascular disease (P=0.03), location in the medial compartment of the thigh (P=0.03), and neurovascular involvement (P=0.03). CONCLUSIONS: This study presents a cohort of STS patients with an extended preRT-surgery interval of ~6 weeks. MWCs in this population occurred at a lower rate than historical controls. Overall these findings support the exploration of a longer interval to reduce MWCs and their associated morbidity.

Metachronous, Polyostotic Aneurysmal Bone Cysts in an Adolescent Female: A Case Report.

UPDATE: This article was updated on December 17, 2019, because a new reference was added post-publication. This new reference (Niemeier TE, Leddy LR, Chapin RW, Smith TM. Metachronous Aneurysmal Bone Cysts in a Fourteen-Year-Old Girl: A Case Report and Review of the Literature. JBJS Case Connect. 2013 Jun 12;3[2 Suppl 8]:1-7) has been inserted as reference 26, and the original references 26 and 29 through 35 have been renumbered accordingly. Additionally, Table I and several passages in the text have been updated to reflect the addition of the new reference. Specifically, in Table I, the study by Niemeier et al. has been inserted as the fifth row between the "Amer et al." and "Current case" rows. On page 3, the text that had read "To our knowledge, there are only 4 published cases of metachronous, polyostotic ABCs in 4 male patients-. The longest documented interval between clinical presentation of the first and second lesions is 15 years, whereas the shortest interval is 15 months." now reads "To our knowledge, there are only 5 published cases of metachronous, polyostotic ABCs in 4 male patients and 1 female patient-. The longest documented interval between clinical presentation of the first and second lesions is 15 years, whereas the shortest interval is 3 months." On page 7, the passage that had read "However, to our knowledge, there are only 4 published cases of metachronous, polyostotic ABCs, and all patients were males-. The current case demonstrates that females may also develop polyostotic disease. None of the 4 previously reported cases of metachronous, polyostotic ABCs demonstrated a previous history of malignancy-." now reads "However, to our knowledge, there are only 5 published cases of metachronous, polyostotic ABCs with only 1 case describing lesions in a female patient-. This is the second case demonstrating that females may also develop polyostotic disease. None of the 5 previously reported cases of metachronous, polyostotic ABCs demonstrated a previous history of malignancy-." Also on page 7, the sentence that had read "Metachronous ABCs are rare and have been reported to present anywhere from 15 months to 15 years after diagnosis of the initial lesion,." now reads "Metachronous ABCs are rare and have been reported to present anywhere from 3 months to 15 years after diagnosis of the initial lesion-."An erratum has been published: JBJS Case Connect. 2019 Dec 26;9(4):e0263ER. CASE: We present a case involving an adolescent female who developed metachronous, polyostotic aneurysmal bone cysts (ABCs) of the left hemipelvis and left proximal tibia within a 16-month interval. At age 12 years, the left periacetabular ABC was initially treated with selective arterial embolization and percutaneous sclerotherapy, followed by intralesional curettage and bone grafting. At age 14 years, the left proximal tibia ABC was treated with intralesional curettage, bone grafting, and prophylactic internal fixation. She showed no evidence of recurrence of either lesion after 32 and 12 months, respectively. CONCLUSIONS: Metachronous, polyostotic ABCs may occur in females. Metachronous lesions may present years after the initial ABC; therefore, additional imaging to rule out polyostotic disease is not indicated in the routine management of a solitary ABC. Patients with multiple ABCs should be managed by following the standard approach for treatment of each lesion.

Are Limb-sparing Surgical Resections Comparable to Amputation for Patients With Pelvic Chondrosarcoma? A Case-control, Propensity Score-matched Analysis of the National Cancer Database.

BACKGROUND: Chondrosarcoma is the second most frequent primary sarcoma of bone and frequently occurs in the pelvis. Surgical resection is the primary treatment with the two main operative modalities being limb-sparing resection and amputation. Contemporary management has trended toward limb-sparing procedures; however, whether this approach has an adverse effect on long-term survival is unclear. QUESTIONS/PURPOSES: (1) What are the 5- and 10-year survival rates after limb-sparing surgery and amputation? (2) What factors are associated with survival after contemporary surgical management of pelvic chondrosarcoma? METHODS: The 2004-2014 National Cancer Database, a nationwide registry that includes approximately 70% of all new cancers in the United States with requirement for 90% followup, was reviewed for patients diagnosed with pelvic chondrosarcoma who had undergone limb-sparing surgery or amputation. To compare survival, patient demographics, tumor attributes, and treatment characteristics were used to generate one-to-one propensity score-matched cohorts. Other factors associated with survival were determined through multivariable Cox regression. Three hundred eighty-five patients (75%) underwent limb-sparing surgery and 131 (25%) underwent amputation. Propensity score matching resulted in two balanced cohorts of 131 patients. RESULTS: With the numbers available, we could not demonstrate a difference in overall survival between limb-sparing procedures and amputation. The 5-year survivorship was 70% (95% confidence interval [CI], 62%-79%) for limb-sparing surgery versus 70% (95% CI, 62%-79%) for amputation. The 10-year survivorship was 60% (95% CI, 48%-75%) for limb-sparing surgery versus 59% (95% CI, 48%-72%) for amputation. Kaplan-Meier survival analysis did not demonstrate a difference in survival (p = 0.9). Older age (hazard ratio [HR] = 1.029, p < 0.001), higher Charlson/Deyo comorbidity score (HR = 3.039, p = 0.004), higher grade (HR = 2.547, p = 0.005 for Grade 2; HR = 7.269, p < 0.001 for Grade 3; HR = 10.36, p < 0.001 for Grade 4), and positive surgical margins (HR = 1.61, p = 0.039) were associated with decreased survival. CONCLUSIONS: Our findings support the trend toward increased use of limb-sparing surgery for patients with pelvic chondrosarcoma. Orthopaedic oncologists may use our results when counseling their patients regarding treatment options. However, the choice of limb-sparing surgery versus amputation for these challenging patients should still be a highly individualized decision with careful discussion between the patient and the surgeon. LEVEL OF EVIDENCE: Level III, therapeutic study.

Micrometastatic Drug Screening Platform Shows Heterogeneous Response to MAP Chemotherapy in Osteosarcoma Cell Lines.

BACKGROUND: Approximately 80% of patients with osteosarcoma harbor subclinical pulmonary micrometastases at diagnosis. Conventional chemotherapy includes methotrexate, doxorubicin, and cisplatin (MAP); however, this regimen and thus overall survival (60%-70%) have remained largely unchanged for 30 years. It therefore is necessary to identify novel therapeutics targeting the metastatic progression of osteosarcoma. QUESTIONS/PURPOSES: This laboratory study explored application of osteosarcoma spheroids (sarcospheres) for drug screening with the following purposes: (1) to characterize sarcosphere size; (2) to establish accurate measurement of sarcosphere growth; (3) to confirm sarcosphere uniformity; and (4) to apply the platform to evaluate MAP chemotherapy. METHODS: Sarcospheres were first characterized to establish accurate measurement of sarcosphere growth and uniform production. The refined platform then was applied to evaluate MAP chemotherapy to validate its use in drug screening. Sarcospheres were generated from highly metastatic human cell lines (143B, MG-63.3, and LM7) by centrifugation to form three-dimensional aggregates modeling micrometastases. Sarcospheres were matured for 24 hours and then incubated with or without drug from Days 0 to 2. Size was assessed by diameter and volume using brightfield microscopy. Growth was measured by volume and resazurin reduction in viable cells. Sarcosphere uniformity was assessed by diameter and resazurin reduction at Day 0 and the Z' factor, a measure of assay suitability for high-throughput screening, was calculated at Day 2. Sarcospheres were treated with individual MAP agents (0 to 1000 µmol/L) to determine concentrations at which 50% of growth from Days 0 to 2 was inhibited (GIC50). Cell lines resistant to MAP in sarcospheres were treated in monolayer for comparison. RESULTS: Sarcosphere diameter and growth from Days 0 to 2 were quantitatively dependent on the number of cells seeded and the cell line used. Accurate measurement of growth occurred after resazurin incubation for 6 hours, without EDTA-mediated permeabilization, and was correlated with the number of cells seeded and sarcosphere volume for 143B (Spearman's r: 0.98; p < 0.001), MG-63.3 (0.99; p < 0.001), and LM7 (0.98; p < 0.001). Sarcospheres met established criteria for screening applications as mean Z' factors were greater than 0.5 for all cell lines. Response to MAP therapy was cell line-dependent, because MG-63.3 and LM7 sarcospheres exhibited greater than 2000-fold resistance to methotrexate (GIC50 = 88 ± 36 µmol/L and 174 ± 16 µmol/L, respectively) compared with the 143B cell line (GIC50 = 0.04 ± 0.01 µmol/L; p < 0.001 for MG-63.3 and LM7). MG-63.3 monolayers were more sensitive to methotrexate (GIC50 = 0.01 ± 0.01 µmol/L; p < 0.001) than MG-63.3 sarcospheres, whereas LM7 monolayers remained chemoresistent (GIC50 not reached). CONCLUSIONS: This study developed and validated a drug screening platform for progression of osteosarcoma micrometastases. It also highlights heterogeneity among osteosarcoma cell lines. These findings appear to reflect known patient-to-patient heterogeneity and underscore the importance of evaluating multiple tumor models when testing drugs for the treatment of osteosarcoma. CLINICAL RELEVANCE: The described approach is a promising starting point for drug screening in osteosarcoma because it is tailored to evaluate micrometastatic disease. A reliable and rapid method to identify novel therapeutics is critical to improve stagnant outcomes for patients with osteosarcoma.

An Osteological Study on the Prevalence of Osteochondromas.

BACKGROUND: Osteochondromas are frequently reported to be the most common benign tumor of bone. However, there is not definitive data on their prevalence rate as they are often clinically silent, with previous studies estimating 1-3%. METHODS: We examined a large osteologic collection of 2954 skeletons to identify likely osteochondromas. One author examined all bones excluding the skull in each skeleton for atypical cortical projections. These candidates were then evaluated by an orthopedic resident and then a fellowship trained pediatric orthopedic surgeon for final inclusion using strict criteria. RESULTS: 13 cases of osteochondroma were identified from 2954 skeletons to yield a prevalence of 0.44% (95% confidence interval 0.20% - 0.68%) in the study population. All were located on long bones: three in the humerus, six in the femur and four in the tibia. CONCLUSIONS: We found a prevalence of 0.44% for osteochondromas, which is approximately half the lowest value reported in previous literature.

Receptor tyrosine kinases in osteosarcoma: not just the usual suspects.

Despite aggressive surgical and chemotherapy protocols, survival rates for osteosarcoma patients have not improved over the last 30 years. Therefore, novel therapeutic agents are needed. Receptor tyrosine kinases have emerged as targets for the development of new cancer therapies since their activation leads to enhanced proliferation, survival, and metastasis. In fact, aberrant expression and activation of RTKs have been associated with the progression of many cancers. Studies from our lab using phosphoproteomic screening identified RTKs that are activated and thus may contribute to the signaling within metastatic human osteosarcoma cells. Functional genomic screening using siRNA was performed to distinguish which of the activated RTKs contribute to in vitro phenotypes associated with metastatic potential (motility, invasion, colony formation, and cell growth). The resulting RTK hits were then validated using independent validation experiments. From these results, we identified four RTKs (Axl, EphB2, FGFR2, and Ret) that have not been previously studied in osteosarcoma and provide targets for the development of novel therapeutics.

Osteosarcoma Phenotype Is Inhibited by 3,4-Methylenedioxy-ß-nitrostyrene.

ß-nitrostyrene compounds, such as 3,4-methylenedioxy-ß-nitrostyrene (MNS), inhibit growth and induce apoptosis in tumor cells, but no reports have investigated their role in osteosarcoma. In this study, human osteosarcoma cell families with cell lines of varying tumorigenic and metastatic potential were utilized. Scrape motility assays, colony formation assays, and colony survival assays were performed with osteosarcoma cell lines, both in the presence and absence of MNS. Effects of MNS on human osteoblasts and airway epithelial cells were assessed in monolayer cultures. MNS decreased metastatic cell line motility by 72-76% and colony formation by 95-100%. MNS consistently disrupted preformed colonies in a time-dependent and dose-dependent manner. MNS had similar effects on human osteoblasts but little effect on airway epithelial cells. An inactive analog of MNS had no detectable effects, demonstrating specificity. MNS decreases motility and colony formation of osteosarcoma cells and disrupts preformed cell colonies, while producing little effect on pulmonary epithelial cells.

Osteosarcoma.

Osteosarcoma, the most common bone sarcoma, affects approximately 560 children and adolescents annually in the United States. The incidence of new diagnoses peaks in the second decade of life. Twenty percent of patients present with clinically detectable metastases, with micrometastases presumed to be present in many of the remaining patients. Treatment typically includes preoperative chemotherapy, surgical resection, and postoperative chemotherapy. Limb-salvage procedures with wide surgical margins are the mainstay of surgical intervention. Advances in chemotherapy protocols have led to a 5-year survival rate of 60% to 78%. Among the goals of future treatment regimens are improved chemotherapeutic agents with higher specificity and lower toxicity.

Specific tyrosine kinase inhibitors regulate human osteosarcoma cells in vitro.

Inhibitors of specific tyrosine kinases are attractive lead compounds for development of targeted chemotherapies for many tumors, including osteosarcoma. We asked whether inhibition of specific tyrosine kinases would decrease the motility, colony formation, and/or invasiveness by human osteosarcoma cell lines (TE85, MNNG, 143B, SAOS-2, LM-7). An EGF-R inhibitor reduced motility of all five cell lines by 50% to 80%. In contrast, an IGF-1R inhibitor preferentially reduced motility by 42% in LM-7 cells and a met inhibitor preferentially reduced motility by 80% in MNNG cells. The inhibitors of EGF-R, IGF-1R, and met reduced colony formation by more than 80% in all tested cell lines (TE85, MNNG, 143B). The EGF-R inhibitor reduced invasiveness by 62% in 143B cells. The JAK inhibitor increased motility of SAOS-2 and LM7 cells without affecting colony formation or invasiveness. Inhibitors of HER-2, NGF-R, and PDGF-Rs did not affect motility, invasiveness, or colony formation. These results support the hypothesis that specific tyrosine kinases regulate tumorigenesis and/or metastasis in osteosarcoma.