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Background: Mesocosm experiments have become increasingly popular in climate change research as they bridge the gap between small-scale, less realistic, microcosm experiments, and large-scale, more complex, natural systems. Characteristics of aquatic mesocosm designs (e.g., mesocosm volume, study duration, and replication) vary widely, potentially affecting the magnitude and direction of effect sizes measured in experiments. In this global systematic review we aim to identify the type, direction and strength of climate warming effects on aquatic species, communities and ecosystems in mesocosm experiments. Furthermore, we will investigate the context-dependency of the observed effects on several a priori determined effect moderators (ecological and methodological). Our conclusions will provide recommendations for aquatic scientists designing mesocosm experiments, as well as guidelines for interpretation of experimental results by scientists, policy-makers and the general public. Methods: We will conduct a systematic search using multiple online databases to gather evidence from the scientific literature on the effects of warming experimentally tested in aquatic mesocosms. Data from relevant studies will be extracted and used in a random effects meta-analysis to estimate the overall effect sizes of warming experiments on species performance, biodiversity and ecosystem functions. Experimental characteristics (e.g., mesocosm size and shape, replication-level, experimental duration and design, biogeographic region, community type, crossed manipulation) will be further analysed using subgroup analyses.

BackgroundFour international study groups undertook a large study in resectable osteosarcoma, which included two randomised controlled trials, to determine the effect on survival of changing post-operative chemotherapy based on histological response.Patients and methodsPatients with resectable osteosarcoma aged ≤40 years were treated with the MAP regimen, comprising pre-operatively of two 5-week cycles of cisplatin 120 mg/m(2), doxorubicin 75 mg/m(2), methotrexate 12 g/m(2) × 2 (MAP) and post-operatively two further cycles of MAP and two cycles of just MA. Patients were randomised after surgery. Those with ≥10% viable tumour in the resected specimen received MAP or MAP with ifosfamide and etoposide. Those with

To determine if tumor necrosis factor alpha (TNF-alpha) and interleukin-6 (IL-6). IL-6 gene expression is influenced by amount and source of dietary fat, 30 weanling female rats were randomly assigned to a moderate-fat soybean oil (MFS; 22% of total energy fed as fat), high-fat (HF) soybean oil (HFS; 39% of total energy fed as fat), or HF tallow (HFT; 39% of total energy fed as fat) diet treatments. Oral glucose tolerance tests (OGTT) were conducted serially over 10 weeks of treatment. HFT and HFS rats gained more weight and had greater body fat than the MFS rats fed similar amounts of energy. Both groups of HF-fed rats had greater (P<.05) insulin resistance (homeostasis model assessment) than MFS-fed rats. TNF-alpha mRNA abundance quantified by real-time reverse transcriptase-polymerase chain reaction was greater (P<.05) in liver and lower (P<.05) in adipose tissue in HFT compared to HFS and MFS rats. There were positive correlations (P<.05) between hepatic TNF-alpha mRNA and insulin resistance, and negative correlations between insulin sensitivity and hepatic TNF-alpha mRNA and hepatic IL-6 mRNA. During Week 3 and Week 6 OGTTs, hyperinsulinemic responses were observed in the HFT group, after which, on Week 9, insulin secretion was diminished in response to the OGTT, suggesting impaired pancreatic insulin secretion. HFS rats exhibited insulin resistance on Week 9 OGTT. In summary, an HFT diet fed to growing female rats caused insulin resistance associated with increased hepatic TNF-alpha mRNA leading to pancreatic insufficiency. Early-onset insulin resistance related to the inflammatory process in obesity is influenced by the amount and type of fat in the diet. [ABSTRACT FROM AUTHOR]

Background: Nonbacterial osteitis (NBO), a term referring to sterile bone lesions with nonspecific histopathological features of inflammation, may be either unifocal or multifocal, acute (< or =6 months) or chronic, and recurrent. Only when the condition is chronic, recurrent, and multifocal is it appropriate to use the term chronic recurrent multifocal osteomyelitis (CRMO). We present our clinical experience as the largest reported series of children with NBO to date. Methods: We report a retrospective clinical, histopathological, and radiological study of 41 children with nonbacterial osteitis. Results: Of 41 children (2-16 years of age) diagnosed with NBO in our institution over the last 6 years, 21 (51%) had recurrent disease and 18 (44%) had multifocal disease. The most common bones affected were the clavicle, femur, and tibia (in order of decreasing prevalence) accounting for 44 (63%) of a total of 70 lesions. Only one individual had SAPHO syndrome (synovitis, acne, pustulosis, hyperostosis, osteitis) and no other patients had evidence of bowel or skin disease. In the absence of evidence for an infective etiology, we recommend nonsteroidal anti-inflammatory agents as the firstline therapy and bisphosphonates only in cases of resistant disease. Conclusions: On the basis of our findings, we propose using a patient questionnaire and protocol for investigating and managing patients who present with NBO to orthopedic surgeons. We predict that this will benefit patients with this disorder by improving our knowledge of the presenting signs and symptoms and related disorders, rationalizing the therapeutic approach, and allowing us to learn about the natural history of the disease. [ABSTRACT FROM AUTHOR]

Conservation breeding programs for endangered species face challenges, notably in the development of husbandry techniques, complicated by the impracticality of conducting controlled experiments. To reduce uncertainty regarding what works in conservation breeding programs, it is essential to capture data. In avian breeding programs, the construction of quality nests and appropriate incubation and handling of eggs by the parents are essential prerequisites to the successful production of offspring. Here, we study factors influencing nest-building and parental incubation outcomes in 'alalā (Corvus hawaiiensis), which is extinct in the wild and numbers fewer than 130 individuals in human care. Using parent-incubated egg data from 2018 to 2021 (171 clutches, 55 pairs), we evaluated the role of husbandry factors in determining the quality of nests constructed by 'alalā and the likelihood of nest quality and other social and environmental factors to predict egg loss (breaking, cannibalization, or ejection of an egg from the nest). More than half of all eggs laid failed to reach the hatch date. Nest quality was the most influential predictor of egg loss, with eggs in higher quality nests more likely to reach the hatch date. Male age also influenced egg loss, with very young (3 years old) and older males (⪞12 years old) experiencing more egg loss. Furthermore, the two facilities and two aviary types also differed in the quality of nests 'alalā constructed, indicating that nest-building behavior can be influenced by husbandry practices. These findings reduce uncertainty and provide insights for recommended management interventions to facilitate successful reproduction in human care., (© 2024 Wiley Periodicals LLC.)

Osteosarcoma and Ewing sarcoma are bone tumors mostly diagnosed in children, adolescents, and young adults. Despite multimodal therapy, morbidity is high and survival rates remain low, especially in the metastatic disease setting. Trials investigating targeted therapies and immunotherapies have not been groundbreaking. Better understanding of biological subgroups, the role of the tumor immune microenvironment, factors that promote metastasis, and clinical biomarkers of prognosis and drug response are required to make progress. A prerequisite to achieve desired success is a thorough, systematic, and clinically linked biological analysis of patient samples, but disease rarity and tissue processing challenges such as logistics and infrastructure have contributed to a lack of relevant samples for clinical care and research. There is a need for a Europe-wide framework to be implemented for the adequate and minimal sampling, processing, storage, and analysis of patient samples. Two international panels of scientists, clinicians, and patient and parent advocates have formed the Fight Osteosarcoma Through European Research consortium and the Euro Ewing Consortium. The consortia shared their expertise and institutional practices to formulate new guidelines. We report new reference standards for adequate and minimally required sampling (time points, diagnostic samples, and liquid biopsy tubes), handling, and biobanking to enable advanced biological studies in bone sarcoma. We describe standards for analysis and annotation to drive collaboration and data harmonization with practical, legal, and ethical considerations. This position paper provides comprehensive guidelines that should become the new standards of care that will accelerate scientific progress, promote collaboration, and improve outcomes., (©2024 The Authors; Published by the American Association for Cancer Research.)

Tumors frequently display high chromosomal instability and contain multiple copies of genomic regions. Here, we describe GRITIC, a generic method for timing genomic gains leading to complex copy number states, using single-sample bulk whole-genome sequencing data. By applying GRITIC to 6,091 tumors, we found that non-parsimonious evolution is frequent in the formation of complex copy number states in genome-doubled tumors. We measured chromosomal instability before and after genome duplication in human tumors and found that late genome doubling was followed by an increase in the rate of copy number gain. Copy number gains often accumulate as punctuated bursts, commonly after genome doubling. We infer that genome duplications typically affect the landscape of copy number losses, while only minimally impacting copy number gains. In summary, GRITIC is a novel copy number gain timing framework that permits the analysis of copy number evolution in chromosomally unstable tumors.

Recently, DNA methylation clocks have been proven to be precise age predictors, and the application of these clocks in cancer tissue has revealed a global age acceleration in a majority of cancer subtypes when compared to normal tissue from the same individual. The polycomb repressor complex 2 plays a pivotal role in the aging process, and its targets have been shown to be enriched in CpG sites that gain methylation with age. This complex is further regulated by the chromatin remodeling complex SWItch/Sucrose Non-Fermentable and its core subunit, notably the tumor suppressor gene SMARCB1, which under physiological conditions inhibits the activity of the polycomb repressor complex 2. Hence, the loss of function of core members of the SWItch/sucrose non-fermentable complex, such as the tumor suppressor gene SMARCB1, results in increased activity of polycomb repressor complex 2 and interferes with the aging process. SMARCB1-deficient neoplasms represent a family of rare tumors, including amongst others malignant rhabdoid tumors, atypical teratoid and rhabdoid tumors, and epithelioid sarcomas. As aging pathways have recently been proposed as therapeutic targets for various cancer types, these tumors represent candidates for testing such treatments. Here, by deriving epigenetic age scores from more than 1000 tumor samples, we identified epigenetic age acceleration as a hallmark feature of epithelioid sarcoma. This observation highlights the potential of targeting aging pathways as an innovative treatment approach for patients with epithelioid sarcoma., (© 2024. The Author(s).)

There has been little change to the standard treatment for osteosarcoma (OS) over the last 25 years and there is an unmet need to identify new biomarkers and novel therapeutic approaches if outcomes are to improve. Furthermore, there is limited evidence on the impact of OS treatment on patient-reported outcomes (PROs). ICONIC (Improving Outcomes through Collaboration in Osteosarcoma; NCT04132895) is a prospective observational cohort study recruiting newly diagnosed OS patients across the United Kingdom (UK) with matched longitudinal collection of clinical, biological, and PRO data. During Stage 1, which assessed the feasibility of recruitment and data collection, 102 patients were recruited at 22 sites with representation from patient groups frequently excluded in OS studies, including patients over 50 years and those with less common primary sites. The feasibility of collecting clinical and biological samples, in addition to PRO data, has been established and there is ongoing analysis of these data as part of Stage 2. ICONIC will provide a unique, prospective cohort of newly diagnosed OS patients representative of the UK patient population, with fully annotated clinical outcomes linked to molecularly characterised biospecimens, allowing for comprehensive analyses to better understand biology and develop new biomarkers and novel therapeutic approaches.

Paget disease of bone is a metabolic disorder with a strong genetic component, characterised by pronounced disorganised bone remodelling. Complications of this disease include an increased risk of developing bone neoplasms. Here, we describe the case of a 60-year-old Italian patient with Paget disease of bone, presenting with an osteoclast-rich tumour. Our analysis of this entity, based on the clinical, morphological and genetic data (whole exome sequencing), suggests that osteoclast-rich lesions in Paget disease of bone are genetically distinct from classical giant cell tumour of bone. We discuss the importance of differentiating these osteoclast-rich lesions., (© 2023. The Author(s).)

Background: Over the last two decades, many elective procedures have transitioned to day-case surgery thanks to the introduction of 'enhanced recovery' protocols. Only recently has total hip arthroplasty been considered a candidate for day-case surgery, as it was once associated with significant pain, mobility impairment and prolonged postoperative recovery. The National Orthopaedic Hospital Cappagh became the first public hospital in Ireland to set up a day-case total hip arthroplasty service in June 2018, and since then has performed over 109 such cases., Aims: We outline our day-case total hip arthroplasty pathway, with specific focus on anaesthetic considerations. We report rates of failed discharge and readmission., Results: We achieved successful same-day discharge in 90.8% of our first 109 cases. Readmission rate was 4.6%., Conclusion: Our experience of implementing a day-case total hip arthroplasty pathway was highly positive and congruent with expectations from the literature. With appropriate patient selection and education, day-case total hip arthroplasty is not just safe, but of benefit to both patients and healthcare systems., Competing Interests: Declaration of conflicting interestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

For some critically endangered species, conservation breeding is a vital steppingstone toward re-establishing wild populations. The 'Alalā (Hawaiian crow, Corvus hawaiiensis), currently extinct in the wild, exists today only in a conservation breeding program, which, for many years, utilized successful hands-on husbandry approaches such as separating and resocializing pairs, providing partially manmade nests, artificially incubating eggs, and puppet rearing nestlings. Yet, a top priority of any conservation breeding program is to retain natural behaviors essential to postrelease survival and reproduction, to achieve successful reintroduction and restoration to the wild. We describe how we are adapting 'Alalā husbandry techniques to strengthen pair bonds through full-time socialization, enable pairs to build robust nests, encourage females to incubate eggs to hatch, and provide pairs and their offspring with vital parental rearing experiences. We discuss the use of standardized, data-driven methods to objectively track our progress towards successful parental breeding and to select release candidates based on their likelihood to survive and breed in the wild. The information shared in this report can be applied to other conservation breeding programs, particularly those implementing or transitioning to husbandry techniques geared towards preparing species to thrive in the wild., (© 2023 The Authors. Zoo Biology published by Wiley Periodicals LLC.)

Identification of mutations in histones in a number of human neoplasms and developmental syndromes represents the most compelling evidence to date for a causal role of epigenetic perturbations in human disease. In most cases, these mutations have gain of function properties that cause deviation from normal developmental processes leading to embryo defects and/or neoplastic transformation. These exciting discoveries represent a step-change in our understanding of the role of chromatin (dys)regulation in development and disease. However, the mechanisms of action of oncogenic histone mutations (oncohistones) remain only partially understood. Here, we critically assess existing literature on oncohistones focussing mainly on bone neoplasms. We show how it is possible to draw parallels with some of the cell-autonomous mechanisms of action described in paediatric brain cancer, although the functions of oncohistones in bone tumours remain under-investigated. In this respect, it is becoming clear that histone mutations targeting the same residues display, at least in part, tissue-specific oncogenic mechanisms. Furthermore, it is emerging that cancer cells carrying oncohistones can modify the surrounding microenvironment to support growth and/or alter differentiation trajectories. A better understanding of oncohistone function in different neoplasms provide potential for identification of signalling that could be targeted therapeutically. Finally, we discuss some of the main concepts and future directions in this research area, while also drawing possible connections and parallels with other cancer epigenetic mechanisms., (© 2023. The Author(s).)

Introduction: Several studies have linked increased risk of osteosarcoma with tall stature, high birthweight, and early puberty, although evidence is inconsistent. We used genetic risk scores (GRS) based on established genetic loci for these traits and evaluated associations between genetically inferred birthweight, height, and puberty timing with osteosarcoma., Methods: Using genotype data from two genome-wide association studies, totaling 1039 cases and 2923 controls of European ancestry, association analyses were conducted using logistic regression for each study and meta-analyzed to estimate pooled odds ratios (ORs) and 95% confidence intervals (CIs). Subgroup analyses were conducted by case diagnosis age, metastasis status, tumor location, tumor histology, and presence of a known pathogenic variant in a cancer susceptibility gene., Results: Genetically inferred higher birthweight was associated with an increased risk of osteosarcoma (OR =1.59, 95% CI 1.07-2.38, P = 0.02). This association was strongest in cases without metastatic disease (OR =2.46, 95% CI 1.44-4.19, P = 9.5 ×10 -04 ). Although there was no overall association between osteosarcoma and genetically inferred taller stature (OR=1.06, 95% CI 0.96-1.17, P = 0.28), the GRS for taller stature was associated with an increased risk of osteosarcoma in 154 cases with a known pathogenic cancer susceptibility gene variant (OR=1.29, 95% CI 1.03-1.63, P = 0.03). There were no significant associations between the GRS for puberty timing and osteosarcoma., Conclusion: A genetic propensity to higher birthweight was associated with increased osteosarcoma risk, suggesting that shared genetic factors or biological pathways that affect birthweight may contribute to osteosarcoma pathogenesis., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2023. Published by Elsevier Ltd.)

Epithelioid sarcoma is a rare and aggressive mesenchymal tumour, the genetic hallmark of which is the loss of expression of SMARCB1, a key member of the SWItch/Sucrose Non-Fermentable (SWI/SNF) chromatin remodelling complex. Hampered by its rarity, epithelioid sarcoma has received little research attention and therapeutic options for this disease remain limited. SMARCB1-deficient tumours also include malignant rhabdoid tumour, atypical teratoid and rhabdoid tumour, epithelioid malignant peripheral nerve sheath tumour, and poorly differentiated chordoma. Histologically, it can be challenging to distinguish epithelioid sarcoma from malignant rhabdoid tumour and other SMARCB1-deficient tumours, whereas methylation profiling shows that they represent distinct entities and facilitates their classification. Methylation studies on SMARCB1-deficient tumours, although not including epithelioid sarcomas, reported methylation subgroups which resulted in new clinical stratification and therapeutic approaches. In addition, emerging evidence indicates that immunotherapy, including immune checkpoint inhibitors, represents a promising therapeutic strategy for SMARCB1-deficient tumours. Here, we show that some epithelioid sarcomas share methylation patterns of malignant rhabdoid tumours indicating that this could help to distinguish these entities and guide treatment. Using gene expression data, we also showed that the immune environment of epithelioid sarcoma is characterised by a predominance of CD8 + lymphocytes and M2 macrophages. These findings have potential implications for the management of patients with epithelioid sarcoma. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland., (© 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.)

Rhabdomyosarcoma accounts for roughly 1% of adult sarcomas, with pleomorphic rhabdomyosarcoma (PRMS) as the most common subtype. Survival outcomes remain poor for patients with PRMS, and little is known about the molecular drivers of this disease. To better characterize PRMS, we performed a broad array of genomic and immunostaining analyses on 25 patient samples. In terms of gene expression and methylation, PRMS clustered more closely with other complex karyotype sarcomas than with pediatric alveolar and embryonal rhabdomyosarcoma. Immune infiltrate levels in PRMS were among the highest observed in multiple sarcoma types and contrasted with low levels in other rhabdomyosarcoma subtypes. Lower immune infiltrate was associated with complete loss of both TP53 and RB1 . This comprehensive characterization of the genetic, epigenetic, and immune landscape of PRMS provides a roadmap for improved prognostications and therapeutic exploration., Competing Interests: The authors declare no competing interests., (© 2023 The Authors.)

Malignant peripheral nerve sheath tumor (MPNST), an aggressive soft-tissue sarcoma, occurs in people with neurofibromatosis type 1 (NF1) and sporadically. Whole-genome and multiregional exome sequencing, transcriptomic, and methylation profiling of 95 tumor samples revealed the order of genomic events in tumor evolution. Following biallelic inactivation of NF1, loss of CDKN2A or TP53 with or without inactivation of polycomb repressive complex 2 (PRC2) leads to extensive somatic copy-number aberrations (SCNA). Distinct pathways of tumor evolution are associated with inactivation of PRC2 genes and H3K27 trimethylation (H3K27me3) status. Tumors with H3K27me3 loss evolve through extensive chromosomal losses followed by whole-genome doubling and chromosome 8 amplification, and show lower levels of immune cell infiltration. Retention of H3K27me3 leads to extensive genomic instability, but an immune cell-rich phenotype. Specific SCNAs detected in both tumor samples and cell-free DNA (cfDNA) act as a surrogate for H3K27me3 loss and immune infiltration, and predict prognosis., Significance: MPNST is the most common cause of death and morbidity for individuals with NF1, a relatively common tumor predisposition syndrome. Our results suggest that somatic copy-number and methylation profiling of tumor or cfDNA could serve as a biomarker for early diagnosis and to stratify patients into prognostic and treatment-related subgroups. This article is highlighted in the In This Issue feature, p. 517., (©2023 The Authors; Published by the American Association for Cancer Research.)

The causes of pediatric cancers' distinctiveness compared to adult-onset tumors of the same type are not completely clear and not fully explained by their genomes. In this study, we used an optimized multilevel RNA clustering approach to derive molecular definitions for most childhood cancers. Applying this method to 13,313 transcriptomes, we constructed a pediatric cancer atlas to explore age-associated changes. Tumor entities were sometimes unexpectedly grouped due to common lineages, drivers or stemness profiles. Some established entities were divided into subgroups that predicted outcome better than current diagnostic approaches. These definitions account for inter-tumoral and intra-tumoral heterogeneity and have the potential of enabling reproducible, quantifiable diagnostics. As a whole, childhood tumors had more transcriptional diversity than adult tumors, maintaining greater expression flexibility. To apply these insights, we designed an ensemble convolutional neural network classifier. We show that this tool was able to match or clarify the diagnosis for 85% of childhood tumors in a prospective cohort. If further validated, this framework could be extended to derive molecular definitions for all cancer types., (© 2023. The Author(s).)

The FOS gene family has been implicated in tumourigenesis across several tumour types, particularly mesenchymal tumours. The rare fibrous tumour desmoplastic fibroblastoma is characterised by overexpression of FOSL1. However, previous studies using cytogenetic and molecular techniques did not identify an underlying somatic change involving the FOSL1 gene to explain this finding. Prompted by an unusual index case, we report the discovery of a novel FOSL1 rearrangement in desmoplastic fibroblastoma using whole-genome and targeted RNA sequencing. We investigated 15 desmoplastic fibroblastomas and 15 fibromas of tendon sheath using immunohistochemistry, in situ hybridisation and targeted RNA sequencing. Rearrangements in FOSL1 and FOS were identified in 10/15 and 2/15 desmoplastic fibroblastomas respectively, which mirrors the pattern of FOS rearrangements observed in benign bone and vascular tumours. Fibroma of tendon sheath, which shares histological features with desmoplastic fibroblastoma, harboured USP6 rearrangements in 9/15 cases and did not demonstrate rearrangements in any of the four FOS genes. The overall concordance between FOSL1 immunohistochemistry and RNA sequencing results was 90%. These findings illustrate that FOSL1 and FOS rearrangements are a recurrent event in desmoplastic fibroblastoma, establishing this finding as a useful diagnostic adjunct and expanding the spectrum of tumours driven by FOS gene family alterations. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland., (© 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.)

Osteosarcoma is the most common primary malignant tumour of the bone. Osteosarcoma incidence is bimodal, peaking at 18 and 60 years of age, and is slightly more common in males. The key pathophysiological mechanism involves several possible genetic drivers of disease linked to bone formation, causing malignant progression and metastasis. While there have been significant improvements in the outcome of patients with localized disease, with event-free survival outcomes exceeding 60%, in patients with metastatic disease, event-free survival outcomes remain poor at less than 30%. The suspicion of osteosarcoma based on radiographs still requires pathological evaluation of a bone biopsy specimen for definitive diagnosis and CT imaging of the chest should be performed to identify lung nodules. So far, population-based screening and surveillance strategies have not been implemented due to the rarity of osteosarcoma and the lack of reliable markers. Current screening focuses only on groups at high risk such as patients with genetic cancer predisposition syndromes. Management of osteosarcoma requires a multidisciplinary team of paediatric and medical oncologists, orthopaedic and general surgeons, pathologists, radiologists and specialist nurses. Survivors of osteosarcoma require specialized medical follow-up, as curative treatment consisting of chemotherapy and surgery has long-term adverse effects, which also affect the quality of life of patients. The development of osteosarcoma model systems and related research as well as the evaluation of new treatment approaches are ongoing to improve disease outcomes, especially for patients with metastases., (© 2022. Springer Nature Limited.)

Oncohistones represent compelling evidence for a causative role of epigenetic perturbations in cancer. Giant cell tumours of bone (GCTs) are characterised by a mutated histone H3.3 as the sole genetic driver present in bone-forming osteoprogenitor cells but absent from abnormally large bone-resorbing osteoclasts which represent the hallmark of these neoplasms. While these striking features imply a pathogenic interaction between mesenchymal and myelomonocytic lineages during GCT development, the underlying mechanisms remain unknown. We show that the changes in the transcriptome and epigenome in the mesenchymal cells caused by the H3.3-G34W mutation contribute to increase osteoclast recruitment in part via reduced expression of the TGFβ-like soluble factor, SCUBE3. Transcriptional changes in SCUBE3 are associated with altered histone marks and H3.3 G34W enrichment at its enhancer regions. In turn, osteoclasts secrete unregulated amounts of SEMA4D which enhances proliferation of mutated osteoprogenitors arresting their maturation. These findings provide a mechanism by which GCTs undergo differentiation in response to denosumab, a drug that depletes the tumour of osteoclasts. In contrast, hTERT alterations, commonly found in malignant GCT, result in the histone-mutated neoplastic cells being independent of osteoclasts for their proliferation, predicting unresponsiveness to denosumab. We provide a mechanism for the initiation of GCT, the basis of which is dysfunctional cross-talk between bone-forming and bone-resorbing cells. The findings highlight the role of tumour/microenvironment bidirectional interactions in tumorigenesis and how this is exploited in the treatment of GCT., (© 2022. The Author(s).)

Brown tumors are rare and generally self-limiting mass lesions of bone occurring in the context of hyperparathyroidism. Although commonly regarded as endocrine-driven tumor-like lesions, we detected pathogenic hotspot KRAS mutations in 10/16 brown tumors (62%) with similar frequencies found in cases affecting the peripheral and axial skeleton. Pathogenic mutations in other driver genes of the RAS-MAPK pathway were not identified. Our findings suggest brown tumors to represent true neoplasms driven by the activation of the RAS-MAPK signaling pathway. The frequent regression of brown tumors after normalization of hyperparathyroidism points to a second hit mediated by endocrine stimulation to be required for tumor development. Our findings underline the pathogenic relation of brown tumors to nonossifying fibroma and giant cell granuloma of the jaws which both appear histologically similar to brown tumors and are also driven by RAS-MAPK signaling pathway activation., Competing Interests: Conflicts of Interest and Source of Funding: D.T., S.H., B.A., and D.B. were supported by the Swiss National Science Foundation, the Foundation of the Bone Tumor Reference Centre, the Gertrude von Meissner Stiftung and the Stiftung für krebskranke Kinder, Regio Basiliensis. A.M.F. is supported by the National Institute for Health Research, the Cancer Research UK University College London Experimental Cancer Medicine Centre, the RNOH Research and Development Department and the RNOH Biobank Team and the Tom Prince Trust. The authors have disclosed that they have no significant relationships with, or financial interest in, any commercial companies pertaining to this article., (Copyright © 2022 The Author(s). Published by Wolters Kluwer Health, Inc.)

Background: Central conventional chondrosarcoma (CS) is the most common subtype of primary malignant bone tumour in adults. Treatment options are usually limited to surgery, and prognosis is challenging. These tumours are characterised by the presence and absence of IDH1 and IDH2 mutations, and recently, TERT promoter alterations have been reported in around 20% of cases. The effect of these mutations on clinical outcome remains unclear. The purpose of this study was to determine if prognostic accuracy can be improved by the addition of genomic data, and specifically by examination of IDH1, IDH2, and TERT mutations., Methods: In this study, we combined both archival samples and data sourced from the Genomics England 100,000 Genomes Project (n = 356). Mutations in IDH1, IDH2, and TERT were profiled using digital droplet PCR (n = 346), whole genome sequencing (n=68), or both (n = 64). Complex events and other genetic features were also examined, along with methylation array data (n = 84). We correlated clinical features and patient outcomes with our genetic findings., Results: IDH2-mutant tumours occur in older patients and commonly present with high-grade or dedifferentiated disease. Notably, TERT mutations occur most frequently in IDH2-mutant tumours, although have no effect on survival in this group. In contrast, TERT mutations are rarer in IDH1-mutant tumours, yet they are associated with a less favourable outcome in this group. We also found that methylation profiles distinguish IDH1- from IDH2-mutant tumours. IDH wild-type tumours rarely exhibit TERT mutations and tend to be diagnosed in a younger population than those with tumours harbouring IDH1 and IDH2 mutations. A major genetic feature of this group is haploidisation and subsequent genome doubling. These tumours evolve less frequently to dedifferentiated disease and therefore constitute a lower risk group., Conclusions: Tumours with IDH1 or IDH2 mutations or those that are IDHwt have significantly different genetic pathways and outcomes in relation to TERT mutation. Diagnostic testing for IDH1, IDH2, and TERT mutations could therefore help to guide clinical monitoring and prognostication., (© 2022. The Author(s).)

CRISPR/Cas9, base editors and prime editors comprise the contemporary genome editing toolbox. Many studies have optimized the use of CRISPR/Cas9, as the original CRISPR genome editing system, in substituting single nucleotides by homology directed repair (HDR), although this remains challenging. Studies describing modifications that improve editing efficiency fall short of isolating clonal cell lines or have not been validated for challenging loci or cell models. We present data from 95 transfections using a colony forming and an immortalized cell line comparing the effect on editing efficiency of donor template modifications, concentration of components, HDR enhancing agents and cold shock. We found that in silico predictions of guide RNA efficiency correlated poorly withactivity in cells. Using NGS and ddPCR we detected editing efficiencies of 5-12% in the transfected populations which fell to 1% on clonal cell line isolation. Our data demonstrate the variability of CRISPR efficiency by cell model, target locus and other factors. Successful genome editing requires a comparison of systems and modifications to develop the optimal protocol for the cell model and locus. We describe the steps in this process in a flowchart for those embarking on genome editing using any system and incorporate validated HDR-boosting modifications for those using CRISPR/Cas9., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Usher, Ligammari, Ahrabi, Hepburn, Connolly, Bond, Flanagan and Cottone.)

Gains and losses of DNA are prevalent in cancer and emerge as a consequence of inter-related processes of replication stress, mitotic errors, spindle multipolarity and breakage-fusion-bridge cycles, among others, which may lead to chromosomal instability and aneuploidy 1,2 . These copy number alterations contribute to cancer initiation, progression and therapeutic resistance 3-5 . Here we present a conceptual framework to examine the patterns of copy number alterations in human cancer that is widely applicable to diverse data types, including whole-genome sequencing, whole-exome sequencing, reduced representation bisulfite sequencing, single-cell DNA sequencing and SNP6 microarray data. Deploying this framework to 9,873 cancers representing 33 human cancer types from The Cancer Genome Atlas 6 revealed a set of 21 copy number signatures that explain the copy number patterns of 97% of samples. Seventeen copy number signatures were attributed to biological phenomena of whole-genome doubling, aneuploidy, loss of heterozygosity, homologous recombination deficiency, chromothripsis and haploidization. The aetiologies of four copy number signatures remain unexplained. Some cancer types harbour amplicon signatures associated with extrachromosomal DNA, disease-specific survival and proto-oncogene gains such as MDM2. In contrast to base-scale mutational signatures, no copy number signature was associated with many known exogenous cancer risk factors. Our results synthesize the global landscape of copy number alterations in human cancer by revealing a diversity of mutational processes that give rise to these alterations., (© 2022. The Author(s).)

Superficial CD34-positive fibroblastic tumor (SCD34FT) is a recently recognized soft tissue tumor that is considered to be of borderline malignancy. The pathogenesis of this tumor remains incompletely understood, but it has been suggested that SCD34FT overlaps with tumors showing fusions involving the PRDM10 gene. Previous analyses of PRDM10-rearranged tumors have demonstrated that they have a distinct gene expression profile, resulting in high expression of CADM3 (also known as SynCam3), which can be detected immunohistochemically. Here, we investigated a series (n = 43) of SCD34FT or PRDM10-rearranged tumors and potential mimics (n = 226) with regard to morphological, genetic, and immunohistochemical features. The results show that SCD34FT and PRDM10-rearranged tumor are morphologically indistinguishable; 41 of 43 tumors of both entities are CADM3-positive. Hence, we suggest that they constitute a single entity, preferably referred to as SCD34FT. Expression of CADM3 was only rarely seen in other soft tissue tumors, except in tumors with Schwann cell differentiation. Thus, IHC for CADM3, in combination with the characteristic morphological features, is a valuable adjunct in the diagnosis of SCD34FT., (© 2021. The Author(s), under exclusive licence to United States & Canadian Academy of Pathology.)

Aim: Osteosarcoma (OS) is the most common primary bone tumour in children and adolescents. Circulating free (cfDNA) and circulating tumour DNA (ctDNA) are promising biomarkers for disease surveillance and prognostication in several cancer types; however, few such studies are reported for OS. The purpose of this study was to discover and validate methylation-based biomarkers to detect plasma ctDNA in patients with OS and explore their utility as prognostic markers., Methods: Candidate CpG markers were selected through analysis of methylation array data for OS, non-OS tumours and germline samples. Candidates were validated in two independent OS datasets (n = 162, n = 107) and the four top-performing markers were selected. Methylation-specific digital droplet PCR (ddPCR) assays were designed and experimentally validated in OS tumour samples (n = 20) and control plasma samples. Finally, ddPCR assays were applied to pre-operative plasma and where available post-operative plasma from 72 patients with OS, and findings correlated with outcome., Results: Custom ddPCR assays detected ctDNA in 69% and 40% of pre-operative plasma samples (n = 72), based on thresholds of one or two positive markers respectively. ctDNA was detected in 5/17 (29%) post-operative plasma samples from patients, which in four cases were associated with or preceded disease relapse. Both pre-operative cfDNA levels and ctDNA detection independently correlated with overall survival (p = 0.0015 and p = 0.0096, respectively)., Conclusion: Our findings illustrate the potential of mutation-independent methylation-based ctDNA assays for OS. This study lays the foundation for multi-institutional collaborative studies to explore the utility of plasma-derived biomarkers in the management of OS., Competing Interests: Conflict of interest statement The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Crown Copyright © 2022. Published by Elsevier Ltd. All rights reserved.)

Benign bone-forming tumors comprise osteomas, osteoid osteomas, and osteoblastomas. Osteomas affect a wide age range and are usually discovered incidentally. They occur predominantly in the craniofacial skeleton and are classically composed of compact bone. Osteoid osteomas and osteoblastomas are painful lesions occurring in young patients. They are morphologically similar and characterized by FOS gene rearrangement and c-FOS expression at a protein level. Osteoid osteomas are usually smaller than 2 cm in maximum dimension with limited growth potential; osteoblastomas are larger than 2 cm and may be locally aggressive. Histologically both are composed of anastomosing trabeculae of woven bone., Competing Interests: Disclosure The authors have nothing to disclose., (Copyright © 2021 Elsevier Inc. All rights reserved.)

This review provides an overview of the spectrum of tumors showing notochordal differentiation. This spectrum encompasses benign entities that are mostly discovered incidentally on imaging, reported as benign notochordal cell tumor, usually not requiring surgical intervention; slowly growing and histologically low-grade tumors referred to as conventional chordoma but associated with a significant metastatic potential and mortality; and more aggressive disease represented by histologically higher-grade tumors including dedifferentiated chordoma, a high-grade biphasic tumor characterized by a conventional chordoma juxtaposed to a high-grade sarcoma, usually with a spindle or pleomorphic cell morphology, and associated with a poor prognosis and poorly differentiated chordoma., (Copyright © 2021 Elsevier Inc. All rights reserved.)