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Background Despite extensive immunohistochemical (IHC) and molecular studies combined with morphologic findings, a group of round/ovoid cell tumors histologically similar to Ewing sarcomas (ES) but lacking EWSR1 -rearrangements may remain unclassifiable. Design We retrospectively analyzed 41 Ewing-like tumors (formalin-fixed, paraffin-embedded) previously determined as negative or non-informative for EWSR1 -rearrangements by FISH and/or RT-PCR. A new histopathology revision and additional IHC and molecular analyses were carried out in order to investigate whether additional IHC and/or molecular testing in combination with the morphological findings may help in reaching a definitive diagnosis. Results Almost all the tumors ( n = 40) involved soft tissue and/or bone and half the patients died of disease. In the archival cases all diagnoses were Ewing sarcoma (ES), Ewing-like sarcoma (ELS), myoepithelial tumor and undifferentiated sarcoma (US). In the new review all the tumors were re-classified as, ES ( n = 16), Ewing-like tumor with EWSR1 rearrangement and amplification and possible EWSR1-NFATC2 gene fusion (n = 1), CIC -rearranged sarcomas or undifferentiated sarcoma, most consistent with CIC -rearranged sarcoma ( n = 7), sarcoma with BCOR-alteration or undifferentiated sarcoma, consistent with BCOR-associated sarcoma ( n = 3), neuroblastoma ( n = 2), unclassifiable neoplasm with neuroblastic differentiation ( n = 1), malignant rhabdoid tumor (n = 2), lymphoblastic lymphoma ( n = 1), clear cell sarcoma of the gastrointestinal tract ( n = 1), small cell carcinoma (n = 1), sclerosing rhabdomyosarcoma ( n = 1), desmoplastic small round cell tumor (n = 1), malignant peripheral sheath nerve tumor ( n = 1), poorly-differentiated synovial sarcoma ( n = 1), Possible gastrointestinal stromal tumor/GIST with predominant round cells (n = 1) and possible SMARCA4-deficient-sarcoma (n = 1). NKX2.2, ETV4 and BCOR immunoreactivity was observed in all ES, CIC -rearranged sarcomas and sarcomas with BCOR alteration, respectively. CIC -rearrangement by FISH was observed in many of the CIC -rearranged sarcomas. Conclusion Our analysis of 41 Ewing-like tumors confirms that there may be a significant pathological and IHC overlap among Ewing-like tumors, with prognostic and therapeutic impacts. Additional IHC (NKX2.2, ETV4 and BCOR) and molecular studies including FUS , CIC or BCOR analysis may support the final diagnosis when FISH or RT-PCR fail to detect EWSR1 -rearrangements. Any molecular findings should always be interpreted in relation to the specific clinical and pathological context. [ABSTRACT FROM AUTHOR]

Permanent synovial sarcoma cell lines are invaluable tools for understanding of the biology of this tumor. The present study reports the establishment of a new human cell line, PDSS-26, derived from a surgical specimen of a poorly differentiated synovial sarcoma. PDSS-26 has a doubling time of a 72 hours and grows as a monolayer of spindle cells that retain immunoreactivity for bcl-2 and vimentin. Karyotypic analysis revealed a rearrangement involving chromosomes 17 and 18, at the breakpoints q11.2 and q11.2, respectively, as the only structural aberrations. Analysis by reverse transcriptase polymerase chain reaction showed the presence of the SYT-SSX1 fusion transcript in both the primary tumor and the cell line. Cytoplasmic PTEN staining was detected by immunohistochemistry in both the PDSS-26 cell line and in original tumor, whereas no mutation was identified by automatic sequencing. Thus, PDSS-26 cells could be useful for future functional studies. [Copyright &y& Elsevier]

Objective: CD14+ monocyte cell lines can differentiate into an osteoclast (OC)-like lineage. However, the identification of human cell lines with stem cell characteristics, capable of differentiating into OCs, would provide a tool for the study of the molecular mechanisms regulating their commitment, differentiation, and function. Since the human acute myeloid leukemia cell line MUTZ-3 contains both CD34+ stem cell and CD14+ cell populations, we investigated the capacity of the stem/progenitor CD34+ population to differentiate into functional OCs. Materials and Methods: Sorted MUTZ-3-CD34+ and MUTZ-3-CD14+ cells were cultured in presence of M-CSF, RANK-L, and TNF-α to generate OCs. Differentiation was evaluated by TRAP staining and RT-PCR, which assessed the expression of c-fms, RANK, MMP-9, CATK, TRAP, and CTR in -CD34+OC and -CD14+OC cells. Resorption pit formation was also evaluated. CD34, CD14, M-CSF-R, RANK, and CTR expression was assessed by FACS analysis. Results: MUTZ-3-CD34+ differentiated into OCs, displaying the full range of differentiation markers; MMP-9, CATK, TRAP, and RANK mRNA were detected from day 3 of culture, whereas CTR from day 12. Stimulated MUTZ-3-CD34+ generated functional osteoclasts that formed extensive resorption lacunae on both mineralized surface and bone slices. Surprisingly, in both sorted populations we identified a population M-CSF-R+/RANK+ that at the same time co-expressed CD14 and CD34. Conclusions: These findings demonstrate that MUTZ-3 cells constitute an invaluable model to study the expression pattern in different developmental stages of commitment and differentiation. Importantly, the data indicate that the CD14+CD34+M-CSF-R+RANK+ population represents an intermediate stage of differentiation from CD34 precursors and monocytes to osteoclast. [Copyright &y& Elsevier]

Primary myelofibrosis (PMF) is a myeloproliferative neoplasm characterised by the clonal proliferation of the haematopoietic precursors together with the progressive development of bone marrow fibrosis. This stromal alteration is an important clinical issue and specific prognostic markers are not currently available. In bone marrow biopsies from 58 PMF patients, we explored the methylation pattern of genes encoding cytokines involved in the stromal reaction, namely platelet-derived growth factor-beta ( PDGFB ), transforming growth factor-beta ( TGFB ) and basic fibroblast growth factor ( FGF2 ). We also evaluated the methylation profile of the Long Interspersed Nucleotide Element 1 (LINE-1). PDGFB , FGF2 and LINE-1, but not TGFB , were significantly differently methylated in PMF compared to controls. Significantly, PDGFB hypomethylation (<16%) was correlated with a favourable PMF prognosis (grade of marrow fibrosis, p = 0.03; International Prognostic Scoring Systems p = 0.01 and Dynamic International Prognostic Scoring Systems, p = 0.02). Although the basis of the association of PDGFB hypomethylation with favourable prognosis remains to be clarified, we speculate that hypomethylation in PMF could represent the effect of acquired somatic mutations in genes involved in epigenetic regulation of the genome. [ABSTRACT FROM AUTHOR]

Background Tenosynovial giant cell tumour/pigmented villonodular synovitis (TGCT/PVNS) is a benign neoplasm of synovium and tendon sheath. We conducted a retrospective pooled analysis in three major referral centers. Methods Patients treated between 1998 and 2008 were examined. Only patients presenting with primary disease or first relapse were included. 5-year local failure free survival (5-year-LFFS) was analysed. Results 294 patients were included: 254 with new diagnosis and 40 in 1st local recurrence (171 F/123 M; median age: 36 years; tumour size ⩽2 cm in 27% of patients, >2 to ⩽5 cm in 41%, and >5 cm in 32%). A diffuse pattern was reported in 69%, localised in 31%. No metastases were documented. Local failure (LF) was reported in 28% of patients: 36% in diffuse pattern, 14% in localised ( p = 0.002); median time to LF: 16 months. With a median follow-up of 4.4 years, 5-year-LFFS was 66%, with multiple (up to five) local recurrences in 40% of relapsed patients. Size <2 cm, macroscopically complete resection, female gender and new diagnosis were associated with a better local control. After multivariate analysis, a previous relapse was independently associated with local failure. Conclusions This study underlines the propensity of TGCT/PVNS to multiple local recurrences. In absence of clinical factors, biological studies are needed to identify prognostic factors of local failure. After a first local recurrence, surgery does not seem to have a curative potential. In these high risk patients, studies addressing the role of target therapies are needed. [ABSTRACT FROM AUTHOR]

Tumor-induced osteomalacia (TIO) is a rare paraneoplastic syndrome due to a phosphaturic tumor, which overproduces Fibroblast Growth Factor 23 (FGF-23), causing hyperphosphaturia, hypophosphatemia, low 1,25(OH) 2 D and osteomalacia. Tumor localization is critical, diagnostic delay ranges from 2.5 to 28 years and to date surgical removal is considered effective treatment. We retrospectively evaluated patients with definite diagnosis of TIO referred to a tertiary Rheumatology Center between September 2000 and May 2020, investigating clinical management and disease outcome. We included 17 patients: 10 (58.8%) were females, mean age at diagnosis was 55.3 ± 13.9 years (mean ± standard deviation), with a diagnostic delay from symptoms onset to tumor detection of 6.6 ± 6.25 years. Biochemical data were: serum phosphorus 1.3 ± 0.4 mg/dL (Reference Range: 2.5–4.6), serum 1,25(OH) 2 D 31.8 ± 22.9 ng/mL (RR: 25–86), intact FGF-23, 358.9 ± 677 pg/mL (RR: 25–45); 24 h-Urine Phosphorus was increased in only 2 patients, while tubular reabsorption of phosphate (TRP) was decreased in all patients confirming a renal phosphate wasting. In 2013 68Ga- DOTA-based PET/CT was introduced in routinely practice and diagnostic delay was consistently reduced (from 8.6 ± 7.9 to 4.3 ± 2.4 years). Thirteen patients underwent surgery, one patient underwent radiofrequency ablation; 3 patients, not eligible for surgery, were treated only with supplements of phosphorus and calcitriol. One was started on Burosumab after several unsuccessful surgical attempts. After surgery or ablation, 8 patients had complete remission, 3 TIO persistence, and 3 had overtime relapse. Relapses were observed only in patients who previously underwent closed biopsy. To our knowledge, this is the widest European cohort of TIO patients in the last two decades. We confirm a usual diagnostic delay and recommend a stepwise diagnostic approach. Tumor biopsy is not recommended due to the potential cell spilling. Surgery is generally considered a definitive treatment, even though other approaches have been successful in curing TIO. Active surveillance on possible recurrence is always needed. Burosumab appears a promising therapy. • Diagnosis of TIO is often burdened with a long delay. • Tumor biopsy can cause cell spilling leading to a disease relapse. • 68Ga-DOTA-based PET/CT greatly reduces the diagnostic delay. • Burosumab appears a promising therapy for recurrent or persistent diseases. • Active surveillance is always needed to early detection of relapses. [ABSTRACT FROM AUTHOR]

Based on the French Federation Nationale des Centers de Lutte Contre le Cancer (FNCLCC) grading system, this study assesses the accuracy of conventional and modified core biopsy (CB) systems in predicting the final grade (low vs high) assigned to the resected specimen. Substituting Ki-67 immunoexpression for mitotic count, and radiological for histological assessment of necrosis, we used two modified FNCLCC CB grading systems: (1) Ki-67 immunoexpression alone, and (2) Ki-67 plus radiological assessment of necrosis. We graded 199 soft tissue sarcomas (STS) from nine centers, and compared the results for the conventional (obtained from local histopathology reports) and modified CB systems with the final FNCLCC grading of the corresponding resected specimens. Due to insufficient sample quality or lack of available radiologic data, five cases were not evaluated for Ki67 or radiological assessment of necrosis. The conventional FNCLCC CB grading system accurately identified 109 of the 130 high-grade cases (83.8%). The CB grading matched the final FNCLCC grading (low vs high) in 175 (87.9%) of the 199 resected tumors; overestimating the final grade in three cases and underestimating in 21 cases. Modified system 1 (Ki-67) accurately identified 117 of the 130 high-grade cases (90.0%). The CB grading matched the final FNCLCC grading (low vs high) in 175 (89.7%) of the 195 evaluated cases; overestimating seven and underestimating 13 cases. Modified system 2 (Ki-67 plus radiological necrosis) accurately identified 120 of the 130 high-grade cases (92.3%). This last matched the final FNCLCC grading (low vs high) in 177 (91.2%) of the 194 evaluated cases; overestimating seven and underestimating 10 cases. Modified system 2 obtained highest area under ROC curves, although not statistically significant. Underestimated CB grades did not correlate with histological subtypes, although many of the discrepant cases were myxoid tumors (myxofibrosarcomas or myxoid liposarcomas), leiomyosarcomas or undifferentiated pleomorphic/spindle cell sarcomas. Using modified FNCLCC CB grading systems to replace conventional mitotic count and histologic assessment of necrosis may improve the distinction between low and high-grade STS on CB. Our study confirms that classifying grade 1 as low grade and grades 2 and 3 as high grade improves correlation between CB and final grade by up to 21%, irrespective of CB system used. A higher than expected Ki-67 score in a low-grade sarcoma diagnosed on CB should raise concern that a higher-grade component may not have been sampled. Furthermore, correlation of all clinicopathological and radiological findings at multidisciplinary meetings is essential to assess the histological grade on CB as accurately as possible. [ABSTRACT FROM AUTHOR]

Purpose: Spinal lesion differential diagnosis remains challenging even in MRI. Radiomics and machine learning (ML) have proven useful even in absence of a standardized data mining pipeline. We aimed to assess ML diagnostic performance in spinal lesion differential diagnosis, employing radiomic data extracted by different software.Methods: Patients undergoing MRI for a vertebral lesion were retrospectively analyzed (n = 146, 67 males, 79 females; mean age 63 ± 16 years, range 8-89 years) and constituted the train (n = 100) and internal test cohorts (n = 46). Part of the latter had additional prior exams which constituted a multi-scanner, external test cohort (n = 35). Lesions were labeled as benign or malignant (2-label classification), and benign, primary malignant or metastases (3-label classification) for classification analyses. Features extracted via 3D Slicer heterogeneityCAD module (hCAD) and PyRadiomics were independently used to compare different combinations of feature selection methods and ML classifiers (n = 19).Results: In total, 90 and 1548 features were extracted by hCAD and PyRadiomics, respectively. The best feature selection method-ML algorithm combination was selected by 10 iterations of 10-fold cross-validation in the training data. For the 2-label classification ML obtained 94% accuracy in the internal test cohort, using hCAD data, and 86% in the external one. For the 3-label classification, PyRadiomics data allowed for 80% and 69% accuracy in the internal and external test sets, respectively.Conclusions: MRI radiomics combined with ML may be useful in spinal lesion assessment. More robust pre-processing led to better consistency despite scanner and protocol heterogeneity. [ABSTRACT FROM AUTHOR]

Purpose: To evaluate the diagnostic performance of machine learning for discrimination between low-grade and high-grade cartilaginous bone tumors based on radiomic parameters extracted from unenhanced magnetic resonance imaging (MRI).Methods: We retrospectively enrolled 58 patients with histologically-proven low-grade/atypical cartilaginous tumor of the appendicular skeleton (n = 26) or higher-grade chondrosarcoma (n = 32, including 16 appendicular and 16 axial lesions). They were randomly divided into training (n = 42) and test (n = 16) groups for model tuning and testing, respectively. All tumors were manually segmented on T1-weighted and T2-weighted images by drawing bidimensional regions of interest, which were used for first order and texture feature extraction. A Random Forest wrapper was employed for feature selection. The resulting dataset was used to train a locally weighted ensemble classifier (AdaboostM1). Its performance was assessed via 10-fold cross-validation on the training data and then on the previously unseen test set. Thereafter, an experienced musculoskeletal radiologist blinded to histological and radiomic data qualitatively evaluated the cartilaginous tumors in the test group.Results: After feature selection, the dataset was reduced to 4 features extracted from T1-weighted images. AdaboostM1 correctly classified 85.7 % and 75 % of the lesions in the training and test groups, respectively. The corresponding areas under the receiver operating characteristic curve were 0.85 and 0.78. The radiologist correctly graded 81.3 % of the lesions. There was no significant difference in performance between the radiologist and machine learning classifier (P = 0.453).Conclusions: Our machine learning approach showed good diagnostic performance for classification of low-to-high grade cartilaginous bone tumors and could prove a valuable aid in preoperative tumor characterization. [ABSTRACT FROM AUTHOR]

Aim: To evaluate the accuracy of magnetic resonance imaging (MRI) and computed tomography (CT) in assessing the resection margins of primary malignant bone tumours.Materials and Methods: Resected primary malignant bone tumour specimens removed from 46 patients (27 male; mean age: 48±22 years) were imaged using MRI (fat-saturated proton density-weighted and three-dimensional fat-suppressed T1-weighted gradient-recalled-echo) and CT immediately after surgery. A radiologist and an orthopaedist evaluated bone and soft-tissue margins of the specimens on both examinations. Histological evaluation was performed by a senior orthopaedic oncology pathologist. Margins were classified as R0 (safe margins), R1 (residuals between 0 and 1 mm), and R2 (macroscopic residuals). Cohen's k, chi-square, and McNemar's statistics were used.Results: Having histology as the reference standard, reproducibility of the radiologist ranged from moderate (k=0.544) to substantial (k=0.741) for bone and soft-tissue margins on CT, respectively, while that of the orthopaedist ranged from fair (k=0.316) to moderate (k=0.548). When comparing R2 and R0+R1 scores, reproducibility of readers' evaluation of bone margins increased ranging from substantial (k=0.655) to perfect (k=1.000). Inter-reader agreement ranged from fair (k=0.308) to substantial (k=0.633). Accuracy of the radiologist and orthopaedist ranged from 76% to 83% and from 68% to 72%, respectively. When comparing R2 and R0+R1 scores, the accuracy of both readers ranged from 83% to 100%. There was no association between local recurrence and margin scores of histology, MRI, and CT (p≥0.058).Conclusions: MRI and CT may be useful for extemporaneous analysis of resection margins of primary malignant bone tumours, although wide accuracy variability between the different imaging methods was observed. [ABSTRACT FROM AUTHOR]