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Purpose: To characterize factors associated with response to immune checkpoint inhibitors (ICI) in advanced sarcoma., Experimental Design: This is a retrospective study with a cohort of 216 patients with advanced sarcoma treated with ICI between 2016 and 2023 at Stanford Health Care. Overall survival, progression-free survival (PFS), objective response rates (ORR) per RECIST criteria, and reason for ICI discontinuation were analyzed across histologic subtypes, ICI regimens, tumor mutational burden, and PD-L1 expression., Results: The overall ORR in the cohort was 16.7%. The histologic subtypes with the highest ORR were Kaposi sarcoma (KS, 66.7%), alveolar soft part sarcoma (ASPS, 50%), angiosarcoma (33.3%), myxofibrosarcoma (MFS, 28.6%), and undifferentiated pleomorphic sarcoma (UPS, 27.8%). The subtypes with the lowest ORR were osteosarcoma (0%), synovial sarcoma (0%), and liposarcoma (3.7%). The subtypes with the highest median PFS were KS (median not reached), ASPS (median not reached), MFS (27.4 months), and UPS (11.3 months). The ORR for sarcomas with PD-L1 ≥ 1% was 27.8% (P = 0.02), whereas the ORR for sarcomas with tumor mutational burden ≥10 mutations per megabase of DNA was 28.6% (P = 0.20)., Conclusions: ORR and PFS were highly variable across sarcoma histologic subtypes. In this large analysis, KS, ASPS, angiosarcoma, MFS, and UPS demonstrated the highest ORR and longest PFS while osteosarcoma, synovial sarcoma, and liposarcoma had the lowest ORR and shortest PFS. PD-L1 expression was also associated with increased ORR. Our findings provide further insight into understanding the sarcoma histologic and immunologic factors that correspond with response to ICI., (©2024 American Association for Cancer Research.)

Background: Sarcomas do not belong to the Lynch Syndrome (LS)-tumour spectrum. A growing body literature has reported sarcomas in patients with LS. Clinical and tumour characteristics of these patients remain unknown., Patients and Methods: We set up the first national retrospective study, SarcLynch, describing the pathological and clinical characteristics of sarcomas developed in patients with LS. Patients were identified from two national networks and included from 23 centres in France., Results: Eighty-one patients participated in the SarcLynch study. Sixty-seven (83 %) tumours were soft-tissue sarcomas (STS) and 14 (17 %) bone sarcomas. Among STS, 59 (88 %) showed a pleomorphic component, with undifferentiated pleomorphic sarcoma (UPS) (36 %) and pleomorphic rhabdomyosarcoma (pRMS) (21 %) being the most represented subtypes. Sarcoma was the first neoplastic event in 32 patients (40 %). Thirty-two patients (40 %) were carriers of MSH2 germline pathogenic variants. Among patients who underwent an assessment of deficient mismatch repair (dMMR) by immunohistochemistry and/or molecular biology status, 75 % were dMMR by immunohistochemistry and 45 % were microsatellite instability high (MSI-H). Eight patients received immune checkpoint inhibitors and 4 (50 %) exhibited an objective response with 3 complete radiological response including 1 patient with pathological complete response. Duration of response ranged from 6 to 20 months., Conclusions: SarcLynch, the largest multicentric series describing sarcomas developed in patients with LS, revealed an enrichment in patients with pleomorphic sarcomas - especially UPS and pRMS. This finding strongly supports screening for MMR status evaluation in these rare histotypes both for oncogenetic screening and therapeutic interest. Considering an objective response rate of 50 %, access to immunotherapy should be considered in these tumours., Competing Interests: Declaration of Competing Interest 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., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Background: Kinase-rearranged spindle cell sarcomas are characterized by unique molecular features. The advent of next-generation sequencing (NGS) has enabled the detection of a multitude of kinase fusions, thereby contributing to the accurate categorization of these tumors., Case Presentation: A 37-year-old woman experienced the fourth recurrence of a cranial base tumor 25 years following the initial surgery and radiation therapy. Histological analysis disclosed spindle-shaped and oval tumor cells, along with a high number of mitotic figures. Immunohistochemistry showed a null immunophenotype, negative for pan-TRK, S-100, CD34, pan-CK, GFAP, and Olig2. Molecular analysis of the tumor tissue identified a novel ACVR2A::RAF1 fusion, comprising the first four exons of ACVR2A and the last nine exons of RAF1. The resulting fusion protein retains the extracellular and transmembrane domains of ACVR2A, while its kinase domain is replaced by the kinase domain of RAF1. This hybrid protein likely contributes to tumorigenesis by activating RAF1 signaling in response to ACVR2A ligands from the TGF-β superfamily., Treatment and Outcome: The patient was treated with the MEK1 inhibitor Trametinib, 2 mg per time and once a day. One month later, MRI showed significant tumor shrinkage and pain relief., Conclusion: The ACVR2A::RAF1 fusion represents a novel genomic profile in RAF1-rearranged spindle cell sarcoma, offering a rational basis for targeted therapy. This case highlights the importance of molecular diagnostics in identifying actionable targets and guiding treatment, potentially leading to significant clinical benefits., (© 2025 Wiley Periodicals LLC.)

Superficial anaplastic lymphoma kinase (ALK)-rearranged myxoid spindle cell neoplasms are a recently identified subtype of cutaneous soft tissue tumors, distinct for their co-expression of CD34 and S100 and characterized by ALK gene rearrangements. Although 72 cases have been reported primarily as isolated case reports, this tumor subtype has yet to be included in the WHO classification of soft tissue tumors, underscoring the need for further study. In this study, we diagnosed two additional cases, both arising in the dermis and subcutaneous tissue. These tumors exhibited characteristic pathological features, including linear or concentric whorl patterns, prominent myxoid and collagenized stroma, mild cellular atypia, and rare mitotic activity. The presence of infiltrative margins and the potential for recurrence after surgery suggest at least locally aggressive clinical behavior. Immunohistochemically, the tumors diffusely expressed S100 and CD34, with strong ALK-D5F3 positivity, confirmed by ALK gene rearrangement. These findings further expand the clinical and pathological spectrum of ALK-rearranged neoplasms and highlight the need for continued research on their biological behavior and classification., Competing Interests: Declaration of competing interest The authors have no conflicts of interest to declare., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Fibrosarcomas (FSA) are malignant mesenchymal tumors characterized by low chemo- and radiosensitivity. Development of novel treatment strategies for human adult FSA is hindered by the low incidence and the absence of suitable clinical models. Interestingly, aggressive FSA occur more frequently in domestic cats, hence potentially representing a clinically amenable model to assess novel therapies such as targeted imaging or theranostics. However, a lack of molecular characterization of FSA and adjacent normal tissue (NT) in both species hinders identification of tumor-specific targets and undermines the translational potential of feline FSA. Combining laser-capture microdissection, RNAsequencing and liquid chromatography-tandem mass spectrometry, we perform comprehensive profiling of 30 feline FSA and matched skeletal muscle, adipose and connective tissue. Clear inter-tissue differences allow identification of significantly upregulated and tumor-exclusive features that represent potential targets for diagnostic and therapeutic approaches. While feline FSA are characterized by hyperactive EIF2, TP53 and MYC signaling, immune-related and neuronal pathways emerge as modulators of tumor aggressiveness and immunosuppression. A high degree of molecular similarity with canine and adult FSA allows identification of tumor targets that are conserved across species. Significant enrichment in DNA repair pathways in feline FSA correlate with aggressive clinical behavior in human soft-tissue sarcoma. Finally, we leverage the molecular profiles to identify vulnerabilities, including sensitivity to ATR and PARP inhibition as potential treatment for feline FSA. In conclusion, this detailed landscape provides a rich resource to identify target candidates and therapeutic vulnerabilities within and across species and supports feline FSA as relevant models for the human disease., Competing Interests: Declaration of competing interest 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., (Copyright © 2024. Published by Elsevier Inc.)

Aims: CIC-rearranged sarcomas (CRS) are clinically aggressive undifferentiated round cell sarcomas (URCS), commonly driven by CIC::DUX4. Due to the repetitive nature of DUX4 and the variability of the fusion breakpoints, CIC::DUX4 fusion may be missed by molecular testing. Immunohistochemical (IHC) stains have been studied as surrogates for the CIC::DUX4 fusion. We aim to assess the performance of DUX4 IHC in the work-up of CRS and its expression in non-CRS round cell or epithelioid neoplasms., Methods and Results: Cases of molecularly confirmed CRS (n = 48) and non-CRS (n = 105) were included. CRS cases consisted of 35 females and 13 males, with ages ranging from less than 1 year to 67 years (median = 41 years). Among the molecularly confirmed non-CRS cases, C-terminal DUX4 expression was investigated in Ewing sarcomas (38 cases), alveolar rhabdomyosarcomas (18 cases), desmoplastic small round cell tumours (12 cases) and synovial sarcomas (n = five), as well as in non-mesenchymal neoplasms such as SMARCA4/SMARCB1-deficient tumours (n = five), carcinomas of unknown primary (n = three) and haematolymphoid neoplasms (four cases). DUX4 IHC was considered positive when strong nuclear expression was detected in more than 50% of neoplastic cells. When used as a surrogate for the diagnosis of CRS, the sensitivity and specificity of DUX4 IHC was 98 and 100%, respectively. Only one CRS case was negative for DUX4 IHC and harboured a CIC::FOXO4 fusion., Conclusions: DUX4 IHC is a highly sensitive and specific surrogate marker for the presence of CIC::DUX4 fusion, demonstrating its utility in establishing a diagnosis of CRS., (© 2024 The Author(s). Histopathology published by John Wiley & Sons Ltd.)

The clinicopathological spectrum of undifferentiated round cell sarcomas of bone and soft tissues is expanding after the 5th edition of the WHO classification. A 23-year-old male patient presented with a lump in his left thigh of 3 months' duration. Radiological examination revealed a well-defined, solid-cystic lobulated, soft tissue lesion in the proximal medial region of his left thigh, measuring 7.7 cm in the largest dimension. The referring diagnosis was an epithelioid sarcoma. Histopathological review of the tumor sections revealed a cellular tumor composed of malignant epithelioid to focally "rhabdoid-like" cells in a variable hyalinized and myxoid stroma with geographic areas of necrosis. In addition, there were areas reminiscent of hemangiopericytomatous vasculature. By immunohistochemistry, the tumor cells were diffusely positive for CD34, focally and distinctly for pan keratin (AE1/AE3). INI1/SMARCB1 and SMARCA4 (BRG1) were diffusely positive (normal). Next-generation sequencing with a wide sarcoma panel revealed EWSR1exon8::ZBT44exon4 fusion. The present example constitutes the first malignant epithelioid tumor with a hemangiopericytomatous growth pattern, exhibiting this rare fusion. The differential diagnoses of this tumor and their corresponding immunohistochemical profile are discussed. This example highlights the value of NGS in unraveling rare fusions and in differentiating these tumors from their several mimics., Competing Interests: Declaration of Conflicting InterestsThe authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Introduction: Liquid biopsy as a non-invasive method to investigate cancer biology and monitor residual disease has gained significance in clinical practice over the years. Whilst its applicability in carcinomas is well established, the low incidence and heterogeneity of bone and soft tissue sarcomas explains the less well-established knowledge considering liquid biopsy in these highly malignant mesenchymal neoplasms., Materials and Methods: A systematic literature review adhering to the PRISMA guidelines initially identified 920 studies, of whom 68 original articles could be finally included, all dealing with clinical applicability of liquid biopsy in sarcoma. Studies were discussed within two main chapters, i.e. translocation-associated and complex-karyotype sarcomas., Results: Overall, data on clinical applicability of liquid biopsy in 2636 patients with > 10 different entities of bone and soft tissue sarcomas could be summarised. The five most frequent tumour entities included osteosarcoma (n = 602), Ewing sarcoma (n = 384), gastrointestinal stromal tumour (GIST; n = 203), rhabdomyosarcoma (n = 193), and leiomyosarcoma (n = 145). Of 11 liquid biopsy analytes, largest evidence was present for ctDNA and cfDNA, investigated in 26 and 18 studies, respectively., Conclusions: This systematic literature review provides an extensive up-to-date overview about the current and potential future uses of different liquid biopsy modalities as diagnostic, prognostic, and disease monitoring markers in sarcoma., Competing Interests: Declarations. Conflict of interest: MAS has received travel grants by Alphamed Fischer, implantcast, ImplanTec, and PharmaMar, outside of the submitted work. AL has received institutional educational grants by DePuy Synthes, Alphamed, and ImplanTec, outside of the submitted work. PS has received educational grants, travel support, and honoraria for lectures by Medacta, DePuy Synthes, implantcast, outside of the submitted work. The remaining co-authors (MGS, KK, BAL, DAM) declare no conflicts of interest. Ethical approval: N/A. Informed consent: N/A., (© 2025. The Author(s).)

Background: N6-methyladenosine (m6A) methylation plays a key role in tumor progression. However, the significance of methyltransferase-like 3 (METTL3) in biological processes of soft tissue sarcoma (STS) patients, and the relationship between METTL3 and STS are unclear., Methods: The expression of METTL3 in STS and its relationship with patient prognosis were determined from database analyses. Immunohistochemical staining and 18 F-FDG radioautography were performed on tumor tissues from 39 patients with STS undergoing 18 F-FDG PET before treatment. METTL3 expression in tumor and peritumoral tissues was evaluated with the Wilcoxon test. The Mann-Whitney U test and Spearman's correlation analysis were used to explore correlations of METTL3 expression with both clinicopathological characteristics and 18 F-FDG uptake. One-way analysis of variance and ROC analysis were used to evaluate the efficacy of 18 F-FDG PET metabolic parameters in predicting METTL3 expression., Results: METTL3 expression was significantly higher in STS tumor tissues than normal tissues (all p values<0.01), and correlated with poor patient prognosis (p < 0.05). METTL3 expression was associated with histological differentiation (Z=-2.026, p = 0.043), but no significant difference was observed according to age, sex, tumor size, tumor location, or metastasis (all p values > 0.05). METTL3 expression positively correlated with the expression of CD163 (r = 0.502, p = 0.011), CD68 (r = 0.381, p = 0.017), and CD8 (r = 0.319, p = 0.048), and exhibited a trend toward correlation with CD4 expression (r = 0.310, p = 0.055). Moreover, 18 F-FDG metabolism positively correlated with METTL3 expression in STS (r = 0.580 for PET and r = 0.434 for radioautography, all p values<0.01). The SUVmax of PET was significantly higher in tumors with high rather than low METTL3 expression (Z=-2.979, p = 0.003)., Conclusions: METTL3 was overexpressed in STS, which may be a meaningful target of action in STS patients. The 18 F-FDG uptake was significantly elevated in tumors with high METTL3 expression, SUVmax could provide a meaningful imaging biomarker for its expression., Competing Interests: Declarations. Ethics approval and consent to participate: This retrospective study was approved and the requirement for the informed consent was waived by the Institutional Review Board of the First Affiliated Hospital of Dalian Medical University, and was carried out in accordance with the Declaration of Helsinki. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Human cancer cell lines are the mainstay of cancer research. Recent reports showed that highly mutated adult carcinoma cell lines (mainly HeLa and MCF-7) present striking diversity across laboratories and that long-term continuous culturing results in genomic/transcriptomic heterogeneity with strong phenotypical implications. Here, we hypothesize that oligomutated pediatric sarcoma cell lines mainly driven by a fusion transcription factor, such as Ewing sarcoma (EwS), are genetically and phenotypically more stable than the previously investigated adult carcinoma cell lines. A comprehensive molecular and phenotypic characterization of multiple EwS cell line strains, together with a simultaneous analysis during 12 months of continuous cell culture show that fusion-driven pediatric sarcoma cell line strains are genomically more stable than adult carcinoma strains, display remarkably stable and homogenous transcriptomes, and exhibit uniform and stable drug response. Additionally, the analysis of multiple EwS cell lines subjected to long-term continuous culture reveals that variable degrees of genomic/transcriptomic/phenotypic changes among fusion-driven cell lines, further exemplifying that the potential for reproducibility of in vitro scientific results may be rather understood as a spectrum, even within the same tumor entity., Competing Interests: Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Background: Paediatric sarcomas, including rhabdomyosarcoma, Ewing sarcoma and osteosarcoma, represent a group of malignancies that significantly contribute to cancer-related morbidity and mortality in children and young adults. These cancers share common challenges, including high rates of metastasis, recurrence or treatment resistance, leading to a 5-year survival rate of approximately 20% for patients with advanced disease stages. Despite the critical need, therapeutic advancements have been limited over the past three decades. The advent of chimeric antigen receptor (CAR)-based immunotherapies offers a promising avenue for novel treatments. However, CAR-T cells have faced significant challenges and limited success in treating solid tumours due to issues such as poor tumour infiltration, immunosuppressive tumour microenvironments and off-target effects. In contrast, the adaptation of CAR technology for natural killer (NK) cells has demonstrated potential in both haematological and solid tumours, suggesting a new therapeutic strategy for paediatric sarcomas., Methods: This study developed and validated a novel CAR-NK cell therapy targeting the ephrin type-A receptor-2 (EphA2) antigen, which is highly expressed in various paediatric sarcomas., Results: CAR expression was successfully detected on the surface of NK cells post-electroporation, indicating successful transfection. Significantly, EphA2-specific CAR-NK cells demonstrated enhanced cytotoxic activity against several paediatric sarcoma cell lines in vitro, including those of rhabdomyosarcoma, Ewing sarcoma and osteosarcoma, compared to unmodified NK cells. Transient messenger RNA (mRNA) transfection of NK cells is a safe approach in genetic engineering, with further chemical modifications to mRNA enhancing stability of temporal EphA2-CAR expression in NK cells, thereby promoting prolonged protein expression. Additionally, in vivo EphA2-CAR-NK cells showed promising anti-cancer activity in rhabdomyosarcoma and osteosarcoma mouse models., Conclusions: The study provides a foundational basis for the clinical evaluation of EphA2-targeted CAR-NK cell therapy across a spectrum of paediatric sarcomas. The enhanced anti-tumour effects observed in vitro/vivo suggests potential for improved therapeutic outcomes in hard-to-cure paediatric sarcomas., Key Points: Addressing unmet clinical needs in paediatric Sarcomas. Paediatric sarcomas, including rhabdomyosarcoma, Ewing sarcoma, and osteosarcoma, exhibit poor survival rates in advanced disease stages. The lack of significant therapeutic progress over the past three decades necessitates innovative treatment approaches. Advancing immunotherapy with CAR-NK cells. Natural killer (NK) cells modified with chimeric antigen receptors (CARs) represent a promising strategy to overcome the limitations of CAR-T cells, particularly in solid tumours. CAR-NK cells are associated with enhanced tumour targeting, reduced off-target effects, and improved safety profiles. EphA2 as a therapeutic target. EphA2, a receptor overexpressed in multiple paediatric sarcomas, is identified as a viable target for CAR-based immunotherapy due to its critical role in tumour progression and angiogenesis. Innovations in mRNA-based engineering. This study demonstrates the feasibility of transient mRNA transfection to engineer NK cells for CAR expression, offering a non-integrative and safer alternative to viral transduction. Enhancements in mRNA stability through chemical modifications, can further optimise protein expression. Preclinical efficacy of EphA2-CAR NK cells. EphA2-specific CAR-NK cells exhibit superior cytotoxicity against sarcoma cell lines in vitro and demonstrate significant anti-tumour activity in in vivo mouse models of rhabdomyosarcoma and osteosarcoma. Clinical translation potential. The findings establish a strong preclinical rationale for the clinical evaluation of EphA2-targeted CAR-NK therapy as a novel immunotherapeutic option for paediatric sarcomas. Future research directions: Combining EphA2-CAR NK cells with immune checkpoint inhibitors or other immunomodulatory agents could further enhance therapeutic outcomes and durability. Advanced preclinical models mimicking human tumour microenvironments are needed to refine and optimise this therapeutic approach., (© 2025 The Author(s). Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.)

Context.—: Non-smooth muscle uterine sarcomas are mostly represented by low-grade endometrial stromal sarcoma. However, several other rare, distinct types of uterine sarcoma are recognized, including high-grade endometrial stromal sarcoma, tumors with kinase fusions, uterine tumors resembling ovarian sex cord tumors, soft tissue-type sarcoma, and emerging entities such as KAT6A/B-rearranged tumors. The landscape of uterine sarcomas has changed, mostly because of the increasing knowledge concerning their molecular aberrations., Objective.—: To offer a comprehensive review of the literature focusing on fusions occurring in tumors other than smooth muscle mesenchymal uterine tumors with respect to their type, frequency, and overlap between diagnostic categories and entities., Data Sources.—: The data were mined from the PubMed/MEDLINE database covering the time period from January 1988 to June 2023. In total, 156 studies focusing on the problematics of fusions occurring in non-smooth muscle mesenchymal uterine tumors were selected, and thus became the basis for this review., Conclusions.—: One hundred ten fusions were identified in 703 tumors. The diagnostic significance of the molecular aberrations occurring in these tumors can be unclear in some cases. This can be related to the rare aberrations with a limited number of reported cases. Additionally, even well-known aberrations considered as specific for a certain distinct entity can occur in other lesions, the biological behavior and clinical significance of which can differ substantially., Competing Interests: The authors have no relevant financial interest in the products or companies described in this article., (© 2025 College of American Pathologists.)

Gene fusions involving JAZF1 are a recurrent event in low grade endometrial stromal sarcoma, and have been more recently described in few instances of endometrial stromal sarcoma-like tumors in the genitourinary tract of men. In this article, we describe a previously unreported spindle cell sarcoma harboring an in-frame JAZF1::NUDT5 gene fusion, arising in the chest wall of a 51-year-old man. The tumor had unique morphologic features resembling both endometrial stromal sarcoma and endometrial stromal sarcoma-like tumors, consisting of a mixture of cytologically bland and pleomorphic spindle cells with brisk mitotic activity, within an alternating myxoid and fibrous stroma. It had diffuse immunohistochemical expression of CD10, CD34 and CD56, and variable expression of androgen receptor. To our knowledge, neoplasms with these clinico-pathologic characteristics and novel gene fusion have not been previously reported in the English language literature., (© 2025 Wiley Periodicals LLC.)

Background: A liquid biopsy is a test that evaluates the status of a disease by analyzing a sample of bodily fluid, most commonly blood. In recent years, there has been progress in the development and clinical application of liquid biopsy methods to identify blood-based, tumor-specific biomarkers for many cancer types. However, the implementation of these technologies to aid in the treatment of patients who have a sarcoma remains behind other fields of cancer medicine. For this study, we chose to evaluate a sarcoma liquid biopsy based on circulating tumor DNA (ctDNA). All human beings have normal cell-free DNA (cfDNA) circulating in the blood. In contrast with cfDNA, ctDNA is genetic material present in the blood stream that is derived from a tumor. ctDNA carries the unique genomic fingerprint of the tumor with changes that are not present in normal circulating cfDNA. A successful ctDNA liquid biopsy must be able to target these tumor-specific genetic alterations. For instance, epidermal growth factor receptor (EGFR) mutations are common in lung cancers, and ctDNA liquid biopsies are currently in clinical use to evaluate the status of disease in patients who have a lung cancer by detecting EGFR mutations in the blood. As opposed to many carcinomas, sarcomas do not have common recurrent mutations that could serve as the foundation to a ctDNA liquid biopsy. However, many sarcomas have structural changes to their chromosomes, including gains and losses of portions or entire chromosomes, known as copy number alterations (CNAs), that could serve as a target for a ctDNA liquid biopsy. Murine double minute 2 (MDM2) amplification in select lipomatous tumors or parosteal osteosarcoma is an example of a CNA due to the presence of extra copies of a segment of the long arm of chromosome 12. Since a majority of sarcomas demonstrate a complex karyotype with numerous CNAs, a blood-based liquid biopsy strategy that searches for these CNAs may be able to detect the presence of sarcoma ctDNA. Whole-genome sequencing (WGS) is a next-generation sequencing technique that evaluates the entire genome. The depth of coverage of WGS refers to how detailed the sequencing is, like higher versus lower power on a microscope. WGS can be performed with high-depth sequencing (that is, > 60×), which can detect individual point mutations, or low-depth sequencing (that is, 0.1× to 5×), referred to as low-passage whole-genome sequencing (LP-WGS), which may not detect individual mutations but can detect structural chromosomal changes including gains and losses (that is, CNAs). While similar strategies have shown favorable early results for specific sarcoma subtypes, LP-WGS has not been evaluated for applicability to the broader population of patients who have a sarcoma., Questions/purposes: Does an LP-WGS liquid biopsy evaluating for CNAs detect ctDNA in plasma samples from patients who have sarcomas representing a variety of histologic subtypes?, Methods: This was a retrospective study conducted at a community-based, tertiary referral center. Nine paired (plasma and formalin-fixed paraffin-embedded [FFPE] tissue) and four unpaired (plasma) specimens from patients who had a sarcoma were obtained from a commercial biospecimen bank. Three control specimens from individuals who did not have cancer were also obtained. The paired and unpaired specimens from patients who had a sarcoma represented a variety of sarcoma histologic subtypes. cfDNA was extracted, amplified, and quantified. Libraries were prepared, and LP-WGS was performed using a NextSeq 500 next-generation sequencing machine at a low depth of sequencing coverage (∼1×). The ichorCNA bioinformatics algorithm, which was designed to detect CNAs from low-depth genomic sequencing data, was used to analyze the data. In contrast with the gold standard for diagnosis in the form of histopathologic analysis of a tissue sample, this test does not discriminate between sarcoma subtypes but detects the presence of tumor-derived CNAs within the ctDNA in the blood that should not be present in a patient who does not have cancer. The liquid biopsy was positive for the detection of cancer if the ichorCNA algorithm detected the presence of ctDNA. The algorithm was also used to quantitatively estimate the percent ctDNA within the cfDNA. The concentration of ctDNA was then calculated from the percent ctDNA relative to the total concentration of cfDNA. The CNAs of the paired FFPE tissue and plasma samples were graphically visualized using aCNViewer software., Results: This LP-WGS liquid biopsy detected ctDNA in 9 of 13 of the plasma specimens from patients with a sarcoma. The other four samples from patients with a sarcoma and all serum specimens from patients without cancer had no detectable ctDNA. Of those 9 patients with positive liquid biopsy results, the percent ctDNA ranged from 6% to 11%, and calculated ctDNA quantities were 0.04 to 5.6 ng/mL, which are levels to be expected when ctDNA is detectable., Conclusion: In this small pilot study, we were able to detect sarcoma ctDNA with an LP-WGS liquid biopsy searching for CNAs in the plasma of most patients who had a sarcoma representing a variety of histologic subtypes., Clinical Relevance: These results suggest that an LP-WGS liquid biopsy evaluating for CNAs to identify ctDNA may be more broadly applicable to the population of patients who have a sarcoma than previously reported in studies focusing on specific subtypes. Large prospective clinical trials that gather samples at multiple time points during the process of diagnosis, treatment, and surveillance will be needed to further assess whether this technique can be clinically useful. At our institution, we are in the process of developing a large prospective clinical trial for this purpose., Competing Interests: Each author certifies that there are no funding or commercial associations (consultancies, stock ownership, equity interest, patent/licensing arrangements, etc.) that might pose a conflict of interest in connection with the submitted article related to the author or any immediate family members. All ICMJE Conflict of Interest Forms for authors and Clinical Orthopaedics and Related Research® editors and board members are on file with the publication and can be viewed on request., (Copyright © 2024 by the Association of Bone and Joint Surgeons.)

Epithelioid sarcoma (ES) is a rare tumor hallmarked by the loss of INI1/SMARCB1 expression. Apart from this alteration, little is known about the biology of ES. Despite recent advances in treatment, the prognosis of ES remains unsatisfactory. To elucidate the molecular underpinnings of ES, and to identify diagnostic biomarkers and potential therapeutic vulnerabilities, we performed an integrated omics profiling (RNA sequencing and methylation array) of 24 primary, untreated ESs. Transcriptome and methylome analysis identified 2 distinct molecular clusters that essentially corresponded to the morphologic variants of ES, classic ES (C-ES) and the more aggressive proximal ES (P-ES). The P-ES group was characterized by hyperactivation of GATA3 and MYC pathways, with extensive epigenetic rewiring associated with EZH2 overexpression. Both DNA methylation and gene expression analysis indicated a striking similarity with the "MYC subgroup" of atypical teratoid/rhabdoid tumor, another SMARCB1-deficient tumor, implying a shared molecular background and potential therapeutic vulnerabilities. Conversely, the C-ES group exhibited an endothelial-like molecular profile, with expression of vascular genes and elevated proangiogenic SOX17 signaling. Immunohistochemistry validated the overexpression of the chromatin regulators GATA3 (9/12 vs 0/16) and EZH2 (7/7 vs 2/6) in P-ESs, and of the vascular factors SOX17 (8/8 vs 1/10) and N-cadherin (5/9 vs 0/10) in C-ESs. Therefore, these molecules emerge as potential diagnostic tools to fill the gap represented by the lack of ES subtype-specific biomarkers. In summary, our study shows that P-ES and C-ES represent distinct molecular entities defined by MYC/GATA3 and SOX17/endothelial molecular traits, respectively. Besides providing insights into the biology of ES, our study pinpoints subtype-specific biomarkers and potential therapeutic vulnerabilities., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

An amplicon-based targeted next-generation sequencing (NGS) assay for the detection of gene fusions in sarcomas was developed, validated, and implemented. This assay can detect fusions in targeted regions of 138 genes and BCOR internal tandem duplications. This study reviews our experience with testing on the first 652 patients analyzed. Gene fusions were detected in 238 (36.5%) of 652 cases, including 83 distinct fusions in the 238 fusion-positive cases, 10 of which had not been previously described. Among the 238 fusion-positive cases, the results assisted in establishing a diagnosis for 137 (58%) cases, confirmed a suspected diagnosis in 66 (28%) cases, changed a suspected diagnosis in 25 (10%) cases, and were novel fusions with unknown clinical significance in 10 (4%) cases. Twenty-six cases had gene fusions (ALK, ROS1, NTRK1, NTRK3, and COL1A1::PDGFB) for which there are targetable therapies. BCOR internal tandem duplications were identified in 6 (1.2%) of 485 patients. Among the 138 genes in the panel, 66 were involved in one or more fusions, and 72 were not involved in any fusions. There was little overlap between the genes involved as 5'-partners (31 different genes) and 3'-partners (37 different genes). This study shows the clinical utility of a next-generation sequencing gene fusion detection assay for the diagnosis and treatment of sarcomas., Competing Interests: Disclosure Statement None declared., (Copyright © 2025 Association for Molecular Pathology and American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Soft tissue sarcomas (STS) are heterogeneous and aggressive tumors, originating in connective tissues embryologically derived from the mesenchyme. Due to their rarity, crucial information about their biology is still lacking. In recent years, single-cell and spatial analyses have opened up new horizons in oncology, leading to the possibility of characterizing the internal architecture of the tumor at the single-cell and spatial levels. This review summarizes the first results acquired through these revolutionary methods for different types of STS. We discuss tumor cell populations and their evolution, interactions between tumor cells and the microenvironment, new prognostic markers, and clinically important targets. Finally, we examine the challenges presented by the single-cell and spatial omics of STS and the future perspectives in this field., Competing Interests: Competing interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Laser capture microdissection (LCM) of formalin-fixed, paraffin-embedded sections is a way to analyze gene expression of morphologically distinct areas of tissue, as microscopically visualized with stained tissue sections. Herein, I describe a method for laser dissecting lymphoid aggregates in canine cutaneous and subcutaneous sarcomas and their adjacent sarcoma tissue to determine the differential expression of RNA as determined by NanoString ® nCounter technology. Canine soft tissue sarcomas (STS) are diversely derived mesenchymal neoplasms that, regardless of exact histogenesis, behave similarly and thus have been grouped together as a diagnostic entity. The risk of recurrence and/or metastasis depends on the extent of surgical excision and histologic grade. Lymphoid aggregates are described in these tumors but have not been characterized. In humans, lymphoid aggregates characterized as tertiary lymphoid structures (TLS) improve the prognosis of several tumors, including sarcomas. We sought to determine if RNA expressed by lymphoid aggregates in canine sarcomas was compatible with TLS RNA expression. This chapter describes tissue preparation, staining, laser capture microdissection, and RNA isolation in preparation for digital RNA counting., (© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Background/aim: Soft tissue sarcoma (STS) is a mesenchymal tumor affecting multiple organs in dogs. Previous studies identified activation of the phosphatidylinositol-3 kinase (PI3K)/protein kinase B (PKB, AKT) pathway in canine STS cell lines and clinical samples, but the underlying mechanism remains unclear. This study investigated PTEN loss, PIK3CA mutation, and EGFR over-expression as potential drivers of PI3K/AKT pathway activation in STS., Materials and Methods: We analyzed 36 canine STS samples. PTEN and EGFR expression were evaluated using immunohistochemistry, while PIK3CA and EGFR mutations were assessed through DNA sequencing., Results: PTEN was expressed in all analyzed samples, with no evidence of loss. Weak PTEN expression was observed in 12 (33.3%) samples, while 24 (66.7%) showed normal expression. DNA sequencing of PIK3CA revealed a single point mutation (c.554 A>C, H554P) in one case, but no hotspot mutations were identified. High EGFR expression was significantly correlated with elevated phospho-AKT levels (p<0.0001). Immunolabelling indicated that 30 samples (83.3%) were EGFR-positive, and 27 of these also showed positive phospho-AKT labeling. Accordingly, one missense point mutation in exon 21 of EGFR (E868K) was identified in one of 12 samples., Conclusion: EGFR over-expression, rather than PTEN loss or PIK3CA mutations, may contribute to PI3K/AKT pathway dysregulation in canine STS. Further studies with larger sample sizes and additional validation techniques are necessary to confirm these findings., (Copyright © 2025, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Purpose: The genomic characteristics of uterine sarcomas have not been fully elucidated. This study aimed to explore the genomic landscape of the uterine sarcomas (USs)., Materials and Methods: Comprehensive genomic analysis through RNA-sequencing was conducted. Gene fusion, differentially expressed genes (DEGs), signaling pathway enrichment, immune cell infiltration, and prognosis were analyzed. A deep learning model was constructed to predict the survival of US patients., Results: A total of 71 US samples were examined, including 47 endometrial stromal sarcomas (ESS), 18 uterine leiomyosarcomas (uLMS), three adenosarcomas, two carcinosarcomas, and one uterine tumor resembling an ovarian sex-cord tumor. ESS (including high-grade ESS [HGESS] and low-grade ESS [LGESS]) and uLMS showed distinct gene fusion signatures; a novel gene fusion site, MRPS18A-PDC-AS1 could be a potential diagnostic marker for the pathology differential diagnosis of uLMS and ESS; 797 and 477 uterine sarcoma DEGs (uDEGs) were identified in the ESS vs. uLMS and HGESS vs. LGESS groups, respectively. The uDEGs were enriched in multiple pathways. Fifteen genes including LAMB4 were confirmed with prognostic value in USs; immune infiltration analysis revealed the prognositic value of myeloid dendritic cells, plasmacytoid dendritic cells, natural killer cells, macrophage M1, monocytes and hematopoietic stem cells in USs; the deep learning model named Max-Mean Non-Local multi-instance learning (MMN-MIL) showed satisfactory performance in predicting the survival of US patients, with the area under the receiver operating curve curve reached 0.909 and accuracy achieved 0.804., Conclusion: USs harbored distinct gene fusion characteristics and gene expression features between HGESS, LGESS, and uLMS. The MMN-MIL model could effectively predict the survival of US patients.

Several distinctive round cell sarcomas have emerged by leveraging new testing modalities to include immunohistochemistry, next-generation sequencing, methylation array, and others. While Ewing sarcoma has led the way as the prototypic round cell sarcoma, more recently described round cell sarcomas of bone and soft tissue are now recognized which have unique clinical, morphologic, immunophenotypic, and genetic signatures. While each of these entities is less common than Ewing sarcoma, it is important to distinguish these tumors for correct diagnosis, prognostication, and potential treatment management. The focus of this review will cover CIC-rearranged sarcoma, BCOR-altered sarcomas, and EWSR1-non-ETS sarcomas to include recent developments in desmoplastic small round cell tumor as well as sarcomas with EWSR1/FUS::NFATc2 and EWSR1::PATZ1 gene fusions, highlighting the clinical, morphologic, and immunophenotypic clues to the diagnosis with recognition of each molecular diagnostic hallmark., Competing Interests: Declarations. Compliance with ethical standards: Both authors complied with ethical standards. Conflict of interest: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Purpose: Myxoinflammatory fibroblastic sarcoma (MIFS) is a rare, low-grade sarcoma affecting with predilection the acral soft tissues of middle-aged adults. Clinically, MIFS is associated with a high rate of local recurrence but infrequent distant metastases. The diagnosis remains challenging due to their wide histologic spectrum and overlap with reactive, benign, and low-grade malignant lesions. Moreover, a significant limitation is that molecular confirmation is achieved in only a subset of cases, due to its broad range of genetic alterations which requires a multiplatform approach. Thus, a definitive diagnosis, especially at nonacral sites and in molecularly negative cases, remains uncertain. Our goal was to perform a detailed clinicopathologic and molecular reappraisal of MIFS managed at a single tertiary cancer center with dedicated orthopedic oncology expertise. Additionally, we examined potential outcomes correlating with specific genetic alterations., Patients and Methods: A cohort of 33 patients (12 males, 21 females, median age 52 years) was selected. Tumors were tested by FISH, Archer, and/or targeted NGS., Results: VGLL3 amplification was detected in 84%, BRAF fusions in 33% and combined TGFBR3/MGEA5 rearrangements in 32% of cases. Two novel fusions were detected, RRAGB::CCNB3 and FGFR1::ZBTB47. Other events included a YAP1::MAML2 fusion in two cases, one co-existing with a BRAF fusion. Overall, 8 (24%) patients recurred, 4 more than once, while 4 (12%) patients developed metastasis (3 locoregional, 1 pulmonary), all associated with VGLL3 gene amplification., Conclusion: Positive margin status was associated with increased recurrence and reduced disease-free survival (DFS, p = 0.02). Moreover, it emphasizes the impact of multiplatform molecular testing in confirming the diagnosis. The lack of both local recurrence and metastatic potential outside VGLL3 amplifications requires further investigation., (© 2025 Wiley Periodicals LLC.)

Background: Intimal sarcoma is a rare and aggressive soft-tissue sarcoma with limited treatment options. We explored genomic profiles of intimal sarcoma to uncover therapeutic implications., Materials and Methods: We analyzed tumor tissues from patients with intimal sarcoma who visited the Seoul National University Hospital (SNUH) using whole-exome, whole-transcriptome, and clinical next-generation sequencing (NGS), integrated with intimal sarcoma NGS data from two public cohorts. We examined expression characteristics and tumor-infiltrating lymphocytes (TILs) according to molecular subtypes., Results: Our study included 42 samples in total. Thirty-three patients showing copy number variation (CNV) enrichment with frequent CDK4/MDM2 amplifications were classified as the CNV-high (CNV-H) subtype. Five patients showing predominant MLH1 mutations or homozygous deletions were classified as the microsatellite instability-high-like (MSI-H-like) subtype. Hallmark pathways up-regulated in the CNV-H subtype included Wnt β-catenin and Hedgehog signaling. In the MSI-H-like subtype, interferon-γ response, tumor necrosis factor-α signaling via nuclear factor-κB, interferon-α response, inflammatory response, and interleukin-6-Jak-Stat3 signaling were up-regulated. CNV-H subtype samples predominantly showed an immune-desert phenotype, whereas MSI-H-like subtype samples predominantly showed an immune-inflamed phenotype. Two MSI-H-like subtype patients received pembrolizumab and experienced tumor shrinkage., Conclusions: We identified two intimal sarcoma molecular subtypes. Compared with CNV-H, MSI-H-like is enriched in pathways associated with tumor immune responses and TILs. Further efforts and clinical trials to better define these molecular subtypes are warranted to open new avenues for personalized treatment approaches and improve patient outcomes., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

BACKGROUND Limb-girdle muscular dystrophy recessive 1 (LGMDR1) is an autosomal recessive degenerative muscle disorder characterized by progressive muscular weakness caused by pathogenic variants in the CAPN3 gene. Desmoplastic small round cell tumors (DSRCT) are ultra-rare and aggressive soft tissue sarcomas usually in the abdominal cavity, molecularly characterized by the presence of a EWSR1::WT1 fusion transcript. Mouse models of muscular dystrophy, including LGMDR1, present an increased risk of soft tissue sarcomas. However, the DSRCT risk and general cancer risk in patients with LGMD is unknown. Here, we delineate the clinical, molecular, and genetic findings of a patient with LGMDR1 who developed a DSRCT. CASE REPORT The patient was a boy who was diagnosed at the age of 9 years with LGMDR1, caused by the biallelic pathogenic variants NP&#95;000061.1:p.(Arg448Cys) and NP&#95;000061.1:p.(Thr184ArgfsTer36) in CAPN3. At 17 years of age, a pathologic soft tissue mass was found in the right pelvis. Immunostaining was positive for Desmin and negative for Myogenin and MyoD1, and RNA sequencing showed a EWSR1::WT1 fusion transcript, confirming the diagnosis of DSRCT. The patient relapsed after 1 year and, following a second relapse, he was started on palliative treatment. No germline variants in childhood cancer predisposition genes were detected by whole genome sequencing. CONCLUSIONS We describe a patient with LGMDR1 who developed a DSRCT. Since associations between LGMD and pediatric cancer are hitherto unknown, further studies are warranted, as little information is currently published about the pediatric cancer risk in this patient group.

The mesenchymal transformations of infiltrating gliomas are uncommon events. This is particularly true of IDH-mutant astrocytomas and oligodendrogliomas, in which mesenchymal transformation is exceedingly rare. oligosarcoma is a newly recognized methylation class (MC) that represents transformed 1p/19q co-deleted oligodendrogliomas, but recent studies indicate it may be non-specific. Herein we report the diffuse sarcomatous transformation of a multifocal recurrent astrocytoma from a precursor IDH-mutant astrocytoma, CNS WHO grade 3, in a young patient following embolization therapy and matching to MC oligosarcoma. The sarcomatous recurrence and original tumor showed identical 17q breakpoints with loss of heterozygosity of TP53. Both lack the defining 1p/19q co-deletion or copy-neutral heterozygosity of an oligodendroglioma and oligosarcoma. The findings in this case report both contribute to the apparent heterogeneity of the novel MC oligosarcoma and describe a second reported mesenchymal transformation of an IDH-mutant astrocytoma., Competing Interests: Declarations. Ethics approval and consent to participate: Not applicable. Consent for publication: Consent was obtained from the patient described within this case report. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Neurotrophic Tyrosine Receptor Kinase (NTRK)-rearranged spindle cell tumors are a category of soft tissue tumors characterized by rearrangements of the NTRK gene family (NTRK1, NTRK2, and NTRK3), which leads to distinct molecular genetics, morphological, and immunophenotypic characteristics. The central feature of these tumors is the fusion of NTRK genes with other genes, resulting in abnormal expression and activation of tropomyosin receptor kinase proteins.In this report, we present the first documented case of an NTRK spindle cell tumor with SPECC1L-NTRK3 fusion located in the thoracic spine. This case underscores the diagnostic and therapeutic importance of next-generation sequencing in identifying tumor-specific genetic alterations and selecting targeted therapies. Following 1 month of entrectinib treatment, the patient experienced considerable tumor shrinkage and symptomatic improvement. For bone-derived NTRK-rearranged spindle cell sarcomas, entrectinib shows promising therapeutic efficacy and should be considered a preferred treatment option., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Objective: To investigate the clinicopathological features, immunophenotype, molecular characteristics, and differential diagnosis of primary intracranial DICER1-mutant sarcoma in order to better understand this tumor type. Methods: A retrospective analysis was conducted on 7 cases of primary intracranial DICER1-mutant sarcoma diagnosed in the Department of Pathology, Sun Yat-sen University Cancer Center, Guangzhou, China between 2021 and 2023 using next-generation sequencing. At the same time, 10 gliosarcomas, 4 intracranial FET::CREB fusion-positive mesenchymal tumors, 4 malignant meningiomas, 3 malignant solitary fibrous tumors, 3 malignant peripheral nerve sheath tumors, 3 synovial sarcomas and 3 rhabdomyosarcomas (total 30 cases) were selected as control. Results: Among the 7 patients with primary intracranial DICER1-mutant sarcoma, 6 were male and 1 was female, aged 10-32 years (median, 23 years). The tissue morphology was predominantly spindle or pleomorphic sarcoma-like, with 6 cases exhibiting eosinophilic globules, and 3 cases showing rhabdomyoblastic or rhabdomyosarcoma-like cell differentiation. Immunohistochemistry revealed focal desmin expression in 3 cases (3/7), ATRX loss in 3 cases (3/7), and p53 mutant pattern in 4 cases (4/7). Additionally, 4 cases (4/7) showed focal or diffuse SALL4 expression, whereas the control cases (30 cases) did not exhibit SALL4 protein expression, suggesting that SALL4 may possess certain auxiliary diagnostic value. Next-generation sequencing confirmed that all 7 cases of primary intracranial DICER1-mutant sarcoma harbored mutations in the DICER1 gene, with 5 cases having the mutation site at p.E1813D. Until May 2024, all 7 patients were alive. Conclusions: Primary intracranial DICER1-mutant sarcoma is a rare tumor. Understanding its morphological characteristics, immunohistochemical and molecular markers and differential diagnosis is crucial to avoid misdiagnosis and to improve diagnostic accuracy of this tumor.

Specific patterns of mitochondrial dynamics have been repeatedly reported to promote drug resistance in cancer. However, whether targeting mitochondrial fission- and fusion-related proteins could be leveraged to combat multidrug-resistant pediatric sarcomas is poorly understood. Here, we demonstrated that the expression and activation of the mitochondrial fission mediator DRP1 are affected by chemotherapy exposure in common pediatric sarcomas, namely, rhabdomyosarcoma and osteosarcoma. Unexpectedly, decreasing DRP1 activity through stable DRP1 knockdown neither attenuated sarcoma drug resistance nor affected growth rate or mitochondrial network morphology. The minimal impact on sarcoma cell physiology, along with the up-regulation of fission adaptor proteins (MFF and FIS1) detected in rhabdomyosarcoma cells, suggests an alternative DRP1-independent mitochondrial fission mechanism that may efficiently compensate for the lack of DRP1 activity. By exploring the upstream mitophagy and mitochondrial fission regulator, AMPKα1, we found that markedly reduced AMPKα1 levels are sufficient to maintain AMPK signaling capacity without affecting chemosensitivity. Collectively, our findings challenge the direct involvement of DRP1 in pediatric sarcoma drug resistance and highlight the complexity of yet-to-be-characterized noncanonical regulators of mitochondrial dynamics., (© 2024 Borankova et al.)

Significance: We show in mammalian settings that the capicua C1 functional domain is a supercharger for CIC::DUX4, a poorly studied fusion oncoprotein which drives a rare sarcoma with dismal outcomes., (©2024 The Authors; Published by the American Association for Cancer Research.)

In this report, a tumor exhibited EWSR1::RORß gene fusion, to our knowledge, is the first such reported case. The Ewing sarcoma breakpoint region 1 gene (EWSR1) is known to be associated with several soft tissue tumors although its specific role remains unclear. Its fusion with a member of the ETS family, including FLI1 and ERG, results in Ewing sarcoma, and its fusion with other genes unrelated to the ETS family, including NFATC2 and PATZ1, results in round cell sarcoma with EWSR1-non-ETS fusions, previously referred to as Ewing-like sarcoma. RORß encodes retinoic acid-related orphan receptor ß, a nuclear receptor (NR), and is involved in circadian rhythm modulation and cancer regulation. The specific role of RORß in tumorigenesis remains unclear; however, this case report suggests that it may form part of a new tumorigenic entity., Competing Interests: Declaration of interests None., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Purpose: Pediatric sarcomas are bone and soft tissue tumors that often exhibit high metastatic potential and refractory stem-like phenotypes, resulting in poor outcomes. Aggressive sarcomas frequently harbor a disrupted p53 pathway. However, whether pediatric sarcoma stemness is associated with abrogated p53 function and might be attenuated via p53 reactivation remains unclear., Methods: We utilized a unique panel of pediatric sarcoma models and tumor tissue cohorts to investigate the correlation between the expression of stemness-related transcription factors, p53 pathway dysregulations, tumorigenicity in vivo, and clinicopathological features. TP53 mutation status was assessed by next-generation sequencing. Major findings were validated via shRNA-mediated silencing and functional assays. The p53 pathway-targeting drugs were used to explore the effects and selectivity of p53 reactivation against sarcoma cells with stem-like traits., Results: We found that highly tumorigenic stem-like sarcoma cells exhibit dysregulated p53, making them vulnerable to drugs that restore wild-type p53 activity. Immunohistochemistry of mouse xenografts and human tumor tissues revealed that p53 dysregulations, together with enhanced expression of the stemness-related transcription factors SOX2 or KLF4, are crucial features in pediatric osteosarcoma, rhabdomyosarcoma, and Ewing's sarcoma development. p53 dysregulation appears to be an important step for sarcoma cells to acquire a fully stem-like phenotype, and p53-positive pediatric sarcomas exhibit a high frequency of early metastasis. Importantly, reactivating p53 signaling via MDM2/MDMX inhibition selectively induces apoptosis in aggressive, stem-like Ewing's sarcoma cells while sparing healthy fibroblasts., Conclusions: Our results indicate that restoring canonical p53 activity provides a promising strategy for developing improved therapies for pediatric sarcomas with unfavorable stem-like traits., Competing Interests: Declarations. Ethics approval and consent to participate: This study was conducted in accordance with the Declaration of Helsinki and approved by the Research Ethics Committee of Masaryk University (approval no. EKV-2019-051; granted on April 14, 2020). All human tumor tissues and noncommercial patient-derived sarcoma cell lines were obtained with written informed consent from patients or their legal guardians. The animal studies were approved by the Institutional Animal Care and Use Committee of Masaryk University and were registered by the Ministry of Education, Youth and Sports of the Czech Republic (ref. no. MSMT-4408/2016-6) as required by national legislation. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2024. Springer Nature Switzerland AG.)

Abstract: The standard treatment of choice for pleomorphic dermal sarcoma (PDS), a relatively uncommon soft tissue sarcoma and 1 morphologically similar to atypical fibroxanthoma, is wide local excision with close clinical follow-up. Studies regarding management of advanced/metastatic PDS with immune checkpoint inhibitors are limited as most STSs have historically been viewed as being immunologically inert. Contradicting this belief, in this report, we describe 2 cases of PDS with a robust host response. Histopathology of both cases revealed a dermal neoplasm comprising mitotically active, pleomorphic, spindled-to-ovoid cells, which were immunohistochemically negative for keratinocytic, melanocytic, and smooth muscle markers. An unusual feature in both cases was the presence of a brisk host response. Additional workup of the infiltrating lymphocyte population revealed an abnormal CD4:CD8 ratio in both cases, with the proportion of CD8 + lymphocytes surpassing (case 1) and equaling (case 2) that of the CD4 + T-lymphocyte population. The increased proportion of CD8 + lymphocytes prompted the additional workup of TCR gene rearrangement, which revealed a clonal population of T lymphocytes in both cases. The robust and clonal T-lymphocyte host response in both of our cases suggests that PDS appears to fit the classic model of an inflammatory-type tumor and may be a candidate for checkpoint inhibition. Future work includes additional reports of cases of PDS with an infiltrating clonal T-lymphocyte population and detailing the function and specificity of the infiltrating T lymphocytes to ascertain whether they have the potential to recognize and lyse the tumors they colonize., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

The spindle cell neoplasms include sinonasal tract angiofibroma (STA), glomangiopericytoma (GPC), and biphenotypic sinonasal sarcoma (BSNS). These entities share some clinical, histomorphologic, immunohistochemical, and even molecular characteristics. Nevertheless, there are features that are unique to each tumor type. STA shows heavily collagenized and vascular stroma, with apparent relationship to underlying hormonal changes. GPC consistently exhibits monomorphic, ovoid cytologic appearance with the constant feature of Beta-catenin nuclear accumulation by IHC. Lastly, BSNS is a deceptively bland and neural-like neoplasm featuring consistent co-expression of S100 and smooth muscle markers with specific genetic rearrangements serving to uniquely characterize this entity., Competing Interests: Disclosures The authors have nothing to disclose., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Sarcomas, rare malignant tumors of mesenchymal origin, are often underdiagnosed and have face diagnostic ambiguities and limited treatment options. The main objective of this study was to define the nanomechanical and biophysical properties of sarcoma cells, particularly examining how the cytoskeleton's remodeling and related cellular processes such as cell migration and invasion in response to environmental stimuli due to collagen content. Utilizing one murine fibrosarcoma and one osteosarcoma cell line we employed atomic force microscopy, immunostaining, advanced image processing, in vitro cellular assays, and molecular techniques to investigate cells' cytoskeleton remodeling in response to varying collagen concentration. Our study focused on how alterations in collagen content affects the cytoskeletal dynamics and correlate with changes in gene expression profiles relevant to metastasis and an aggressive cancer phenotypes. Our findings indicate that despite their shared classification, fibrosarcoma and osteosarcoma cells display distinct biophysical properties and respond differently to mechanical forces. Notably, this difference in cellular behavior renders mechanical properties a potent novel biomarkers. Furthermore, the metastasis-related identified genes related to metastatic capability, could be potential therapeutic targets. This study highlights the significance of understanding the unique traits of sarcoma cells to improve diagnostic precision and expand therapeutic strategies, for this rare type of cancer., Competing Interests: Declaration of competing interest 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., (Copyright © 2024 Elsevier B.V. All rights reserved.)

With the wide use of RNA sequencing technologies, the family of FET::CREB fusion mesenchymal neoplasms has expanded rapidly to include potentially aggressive neoplasms, not fitting any well established WHO entity. Recently, a group of intra-abdominal FET(EWSR1/FUS)::CREB(CREM/ATF1) fused unclassified neoplasms has been reported followed by recent recognition of an analogous extra-abdominal category of unclassified neoplasms carrying EWSR1::ATF1 fusions. We describe 9 additional tumors (5 extra-abdominal and 4 abdominal) carrying an EWSR1::CREM (n = 8) and FUS::CREM (n = 1) fusion. Patients were 7 females and 2 males aged 10 to 75 years (median, 34). Extra-abdominal tumors originated in the head and neck (2 sinonasal, 1 orbital) and soft tissues (1 gluteal, 1 inguinal). Abdominal tumors involved stomach (2), mesentery (1), and kidney (1). Tumor size ranged from 3.5 to 11 cm (median, 6). Treatment was radical surgery with (5) or without (2) neo/adjuvant radio/chemotherapy. Extended follow-up of 5 patients (21-52 months; median, 24) showed an aggressive course in two (40%); one died of disseminated metastases 52 months after several intensified chemotherapy regimens, and one was alive with progressive abdominal disease at 21 months. The immunophenotype of the two subcohorts was significantly overlapping with variable expression of EMA (7 of 8), keratin AE1/AE3 (5 of 9), CD99 (4 of 7), MUC4 (2 of 8), ALK (3 of 8), synaptophysin (3 of 9), chromogranin (1 of 8), CD34 (3 of 6), CD30 (1 of 6), PAX8 (1 of 7), and inhibin (1 of 7), but no reactivity with desmin (0 of 8), S100 (0 of 8), and SOX10 (0 of 8). This series further solidifies the notion that FET::CREB fusions are not limited to the triad of angiomatoid fibrous histiocytoma, clear cell sarcoma, and malignant gastrointestinal neuroectodermal tumor, but characterize an emerging family of potentially aggressive neoplasms occurring at both intra- and extra-abdominal sites. These tumors underscore the promiscuity of the FET::CREB fusions and highlight the pivotal role of phenotype-oriented classification of these neoplasms that share the same genotype, still featuring significant biological and behavioral distinctness., Competing Interests: Declarations. Ethics approval: Samples were used in accordance with ethical guidelines for the use of retrospective tissue samples provided by the local ethics committee of the Friedrich-Alexander University Erlangen-Nuremberg (ethics committee statements 24.01.2005 and 18.01.2012). Conflict of interest: AA is the Editor-in-Chief of Virchows Archiv. The authors have no financial or non-financial conflicts of interest to disclose., (© 2024. The Author(s).)

Pituitary neuroendocrine tumors (PitNET) that metastasize comprise ~ 0.2% of adenohypophyseal tumors are aggressive and are challenging to treat. However, many non-metastatic tumors are also aggressive. Herein, we review 21 specimens from 13 patients at UCSF with metastatic PitNETs (CSF or systemic, N = 7 patients), high-grade pituitary neuroendocrine neoplasms (HG-PitNEN, N = 4 patients), and/or PitNETs with sarcomatous transformation (PitNET-ST, N = 5 patients). We subtyped cases using the World Health Organization (WHO) and International Agency for Research on Cancer (IARC) criteria for neuroendocrine neoplasms (NENs). Lineage subtypes included acidophil stem cell, null cell, thyrotroph, corticotroph, lactotroph, and gonadotroph tumors. The median Ki-67 labeling index was 25% (range 5-70%). Lack of p16 was seen in 3 cases, with overexpression in 2. Strong diffuse p53 immunopositivity was present in 3 specimens from 2 patients. Loss of Rb expression was seen in 2 cases, with ATRX loss in one. Molecular analysis in 4 tumors variably revealed TERT alterations, homozygous CDKN2A deletion, aneuploidy, and mutations in PTEN, TP53, PDGFRB, and/or PIK3CA. Eight patients (62%) died of disease, 4 were alive at the last follow-up, and 1 was lost to the follow-up. All primary tumors had worrisome features, including aggressive lineage subtype, high mitotic count, and/or high Ki-67 indices. Additional evidence of high-grade progression included immunohistochemical loss of neuroendocrine, transcription factor, and/or hormone markers. We conclude that metastatic PitNET is not the only high-grade form of pituitary NEN. If further confirmed, these histopathologic and/or molecular features could provide advanced warning of biological aggressiveness and be applied towards a future grading scheme., Competing Interests: Declarations. Competing Interests: The authors declare no competing interests., (© 2024. The Author(s).)

Myxoid spindle cell sarcoma is a rare sarcoma with a demanding histopathologic diagnosis due to the absence of pathognomic immunohistochemistry markers. Genetics include complex karyotypic alterations without characteristic molecular abnormalities for this entity. NTRK alterations are rare findings with great clinical importance since they can be therapeutically targeted with two NTRK inhibitors. Herein we present a case of an adult unclassified myxoid spindle cell sarcoma with ETV6/NTRK3 fusion gene, which is a molecular finding characteristic for infantile fibrosarcoma. [ABSTRACT FROM AUTHOR]

DICER1-associated sarcoma is an emerging entity, defined by either somatic or germline dicer 1, ribonuclease III (DICER1) mutations and sharing characteristic morphologic features irrespective of the site of origin. In addition to the DICER1 driver mutation, concurrent genomic alterations, including tumor protein 53 (TP53) inactivation and RAS pathway activation, are frequently detected. Tumors that morphologically resemble malignant peripheral nerve sheath tumor (MPNST) have rarely been reported among DICER1 sarcomas and often pose diagnostic challenges. This study was prompted by a case involving morphologic features of MPNST, which harbored co-existing DICER1 and hotspot KRAS mutations. Hence, we investigated the incidence of these alterations in PNST from our molecular database compared to the genomic and morphologic spectrum of DICER1-mutant sarcomas. In total, we identified three cases diagnosed as MPNST with co-existing DICER1, ATRX chromatin remodeler (ATRX), and KRAS G12V/A alterations occurring in brain, cerebellopontine angle, and intra-abdominal sites. Two additional cases each of MPNSTs and neurofibromas were identified with hotspot KRAS mutations. All five MPNSTs lacked canonical neurofibromin 1 (NF1)/neurofibromin 2 (NF2) alterations, displaying a classic morphologic appearance with fascicular monomorphic spindle cells and followed a diverse clinical behavior. Among the 38 DICER1-associated sarcomas in our database, eight (21%) had secondary KRAS hotspot mutations, all composed of monomorphic spindle and/or round cells, including three with an MPNST-like histology. In contrast, all 10 (26%) DICER1-mutant sarcomas with TP53 mutations showed a pleomorphic phenotype. The DNA-based methylation profile of our index case clustered within the group of sarcomas with DICER1 alterations. Our results highlight a small subset of MPNST associated with DICER1 and/or KRAS mutations. However, their relationship with conventional MPNST remains to be determined in larger studies. © 2025 The Pathological Society of Great Britain and Ireland., (© 2025 The Pathological Society of Great Britain and Ireland.)

Capicua transcriptional repressor (CIC)-rearranged sarcoma (CRS) is a rare and recently described tumor that most commonly affects patients between 15 and 30 years of age. It is an undifferentiated round cell malignancy, with a disease defining CIC fusion, with double homeobox 4 (DUX4) being the most common partner. Here, we report a 77-year-old woman who presented with a cutaneous thigh mass with a clinical morphology suggesting Merkel cell carcinoma. Immunohistochemically, there was positivity for INSM1 (extensive) and synaptophysin (patchy) and granular expression of neurofilament (extensive), CAM5.2, and CK20 (focal, nonspecific). The majority of the tumor showed histopathologic features within the range of what can be seen in Merkel cell carcinoma, but there were divergent features, including a myxoid zone with corded and stranded tumor cells and a Ewing-sarcoma-like zone with confluent concentric membranous CD99 expression. WT-1 was strongly expressed, prompting RNA-based next generation sequencing for gene fusions, which identified the CIC:DUX4 [t(19;4)(19q13.2;4q35.2)]. A novel IRAK3:HMGA2 fusion was also identified. This example of CRS simulated MCC clinically, histopathologically, and immunohistochemically and represents a likely underrecognized diagnostic pitfall., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

NTRK (neurotropic tropomyosin receptor kinase)-rearranged spindle cell tumours represent a rare group of molecularly defined soft tissue neoplasms. These tumours, excluding infantile fibrosarcomas, are characterised by NTRK gene rearrangements and exhibit a range of histomorphologies, including spindle, epithelioid or rhabdoid cells with invasive growth. Their prognosis correlates with histological grade, and surgical resection is the primary treatment. The abnormally expressed oncogenic receptor tyrosine kinases TRK-A, TRK-B and TRK-C in this tumour have been shown to be therapeutical targetable, which may improve patient prognosis. We report a 17-year-old male patient presenting with a left axillary mass. Both fine-needle aspiration cytology (FNAC) and surgical resection specimens were submitted for examination. Comprehensive analysis of cytomorphology, immunohistochemical staining results and next-generation sequencing (NGS) data led to a final diagnosis of NTRK-rearranged spindle cell tumour. By comparing cytological and histological morphologies, we identified diagnostic clues in cytological specimens. NTRK-rearranged spindle cell tumours' definitive diagnosis enables targeted therapy. Fine-needle aspiration cytology, being minimally invasive, offers the potential for earlier and more definitive diagnoses, thereby improving patient treatment options., (© 2024 John Wiley & Sons Ltd.)

Canine cutaneous/subcutaneous soft-tissue sarcomas (STS) are diversely derived mesenchymal neoplasms with a risk of recurrence and/or metastasis depending on the extent of surgical excision and histologic grade. Lymphoid aggregates (LAs) are often described in these tumours but not characterised. In humans, LA characterised as tertiary lymphoid structures (TLSs) improve the prognosis of many tumours, including sarcomas. We sought to determine if LA meeting a size criterion (> 700 cells) in canine sarcomas met the criteria of TLS and the overall prevalence of LA of any size. RNA expression in large LAs versus aggregate-adjacent sarcoma tissue (AAS) was measured in laser capture microdissected tissue and compared to curl-derived RNA from aggregate-free sarcomas and lymph nodes. CD3, CD20, MUM-1 and PNAd expressions were measured using immunohistochemistry. CD20 and CD3 mRNA were more highly expressed in LA versus AAS (13.8 fold, p = 0.0003 and 2.3 fold, p = 0.043). This was supported by the IHC findings. The large LAs were also enriched in chemokine RNA expression characteristic of TLS (CXCR5 5.8 fold, p < 00001, CCL19 3.68 fold, p = 0.0209, CCL21 6.87 fold, p = 0.0209 and CXCL13 2.68 fold, p = 0.0924). Plasma cells and high endothelial venules were identified in LA containing tumours but not in control tissue. Large LAs were present in 12% of tumours, and LA of any size in 30%. We conclude that large LAs in canine STS are consistent with TLS., (© 2024 The Author(s). Veterinary and Comparative Oncology published by John Wiley & Sons Ltd.)

Background: Peritumoral edema on staging magnetic resonance imaging (MRI) is associated with higher local recurrence in soft tissue sarcoma (STS). This may relate to the presence of satellite malignant cells that are difficult to distinguish from benign atypia, leading to over- or undertreatment. This study evaluated a novel targeted molecular approach to identify malignancy in STS peritumoral planes as a means to improve personalized care., Methods: In the targeted molecular approach, whole-exome sequencing was employed to identify tumor-specific variants (TSVs), and peritumoral planes were assayed for malignancy, defined as two or more TSVs/plane, using droplet digital polymerase chain reaction (PCR). Feasibility was evaluated using a retrospective cohort (n = 8) in which planes with cellular atypia were tested. A prospective cohort (n = 8) then assayed all peritumoral planes with radiologic edema., Results: The targeted molecular approach identified malignancy in three of eight cases with cellular atypia of unknown significance (37.5%) and five of eight cases with peritumoral edema on staging MRI (62.5%). Peritumoral regions were heterogeneous; in none of the malignant cases did all sampled planes have evidence of tumor. Malignancy was also identified in regions without cellular atypia. Both cases with a local recurrence had molecular evidence of malignancy outside the main mass despite R0 margins., Conclusion: This study describes a novel personalized approach to detect malignancy in peritumoral regions in STS and is the first to identify molecular evidence of tumor outside the main mass. While development of a clinical tool is underway, these findings support the current approach of treating all peritumoral edema as malignant., Competing Interests: Disclosure: Miguel Alfonso V. Principe, Nalan Gokgoz, Patrick Prochazka, Victoria S. Coward, Sidharth Saini, Sonya MacParland, Rebecca Gladdy, Peter Ferguson, Jay S. Wunder, Irene L. Andrulis, Peter Chung, Anthony M. Griffin, Lawrence M. White, Brendan C. Dickson, and Kim M. Tsoi have no potential conflicts of interest to declare that may be relevant to the contents of this study., (© 2024. Society of Surgical Oncology.)

Multiple endocrine neoplasia type 1 (MEN1) is a hereditary tumor syndrome characterized by endocrine tumors, typically from parathyroid, pancreatic, or anterior pituitary origin. In addition, benign cutaneous soft tissue tumors are prevalent in MEN1 patients. Although sarcomas have been reported in MEN1 patients it is unclear if these tumors should be considered as part of the MEN1 syndrome. Here, five patients with a MEN1 syndrome and a sarcoma are described. In all five sarcomas loss of heterozygosity of the MEN1 gene and loss of expression of menin are shown, suggesting that sarcomas may be a phenotypic expression of MEN1 syndrome., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Pediatric sarcomas present heterogeneous morphology, genetics and clinical behavior posing a challenge for an accurate diagnosis. DNA methylation is an epigenetic modification that coordinates chromatin structure and regulates gene expression, determining cell type and function. DNA methylation-based tumor profiling classifier for sarcomas (known as sarcoma classifier) from the German Cancer Research Center (Deutsches Krebsforschungszentrum) was applied to 122 pediatric sarcomas referred to a reference pediatric oncology hospital. The classifiers reported 88.5% of agreement between histopathological and molecular classification confirming the initial diagnosis of all osteosarcomas and Ewing sarcomas. The Ewing-like sarcomas were reclassified into sarcomas with BCOR or CIC alterations, later confirmed by orthogonal diagnostic techniques. Regarding the CNAs profile, osteosarcomas had several chromosomal gains and losses as well as chromothripsis, whereas Ewing sarcomas had few large events, such as amplifications of chromosomes 8 and 12. The molecular classification together with clinical and histopathological assessment could improve the diagnosis of pediatric sarcomas although there are limitations to deal with more rare classes. This study provides an increase in the number of sarcomas evaluated for DNA methylation profiling in the pediatric population., Competing Interests: Declarations. Ethics approval and consent to participate: The study was conducted in accordance with the Declaration of Helsinki, and approved by the Institutional Ethics Committee of Boldrini Children’s Hospital (CAAE: 28386820.7.0000.5376, CAAE: 22737219.1.0000.5376, CAAE: 44219021.6.0000.5376). Informed consent was obtained from all subjects involved in the study. Consent for publication: Not Applicable. shared information does not compromise individuals' anonymity. Competing interests: The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results., (© 2024. The Author(s).)

Purpose: The IMMUNOSARC trial combined an antiangiogenic agent (sunitinib) with a PD1 inhibitor (nivolumab) in advanced sarcomas. Here, we present the first correlative studies of the soft-tissue sarcoma cohort enrolled in this trial., Experimental Design: Formalin-fixed paraffin-embedded and peripheral blood samples were collected at baseline and week 13. Formalin-fixed paraffin-embedded samples were used for transcriptomics and multiplex immunofluorescence, whereas peripheral blood samples were used for multiplexed immunoassays. Flow cytometry and Luminex assays were performed to validate translational findings in tumor-isolated cells and peripheral blood mononuclear cells derived from patients., Results: The density of intratumoral CD8+ T cells, measured by multiplexed immunophenotyping, was significantly increased after treatment. This augment was accompanied by the dynamic significant increase in the gene expressions of CD86, CHI3L1, CXCL10, CXCL9, LAG3, and VCAM1 and the decrease in the expression levels of NR4A1. In peripheral blood, 12 proteins were significantly modulated by treatment at week 13. A score integrating the dynamic expression of the 7 genes and the 12 soluble factors separated 2 groups with distinct progression-free survival (PFS): 4.1 months [95% confidence interval, 3.5-not reached (NR)] versus 17 months (95% confidence interval, 12.0-NR), P = 0.014. This molecular score was predictive of PFS when applied to the normalized data determined in the baseline samples., Conclusions: Treatment with sunitinib and nivolumab inflamed the sarcoma microenvironment, increasing CD8+ T-cell density and the expression of several genes/proteins with relevance in the response to PD1 inhibitors. A molecular signature identified two groups of patients with distinct PFS for the combination of antiangiogenics plus PD1 inhibitor therapy., (©2024 American Association for Cancer Research.)

MicroRNAs (miRNAs) are pivotal regulators of gene expression, influencing key cellular processes such as proliferation, differentiation, apoptosis, and metastasis. In the realm of sarcomas-a diverse group of malignant tumors affecting soft tissues and bone sarcomas-miRNAs have emerged as crucial players in tumorigenesis and tumor progression. This review delves into the intricate roles of miRNAs across various soft tissue sarcoma subtypes, including rhabdomyosarcoma, liposarcoma, leiomyosarcoma, synovial sarcoma, fibrosarcoma, angiosarcoma, undifferentiated pleomorphic sarcoma (UPS), and malignant peripheral nerve sheath tumor (MPNST). We explore how dysregulated miRNAs function as oncogenes or tumor suppressors, modulating critical pathways that define the aggressive nature of these cancers. Furthermore, we discuss the diagnostic and prognostic potential of specific miRNAs and highlight their promise as therapeutic targets. By understanding the miRNA-mediated regulatory networks, this review aims to provide a comprehensive overview of current research while pointing towards future directions for miRNA-based therapies. Our findings underscore the potential of miRNAs to transform the landscape of sarcoma treatment, offering hope for more precise, personalized, and effective therapeutic strategies.

Myxofibrosarcoma (MFS) and undifferentiated pleomorphic sarcoma (UPS) are two common and aggressive subtypes of soft tissue sarcoma. The aim of this study was to assess potential transcriptomic differences between MFS and UPS tumours and to evaluate the extent to which differences in gene expression profiles were associated with genomic and clinical features. The study included 162 patients with tumours diagnosed as MFS (N = 62) or UPS (N = 100). The patients had been diagnosed and treated at two Swedish sarcoma centres during a 30-year period. For gene expression profiling and gene fusion detection all tumours were analysed using RNA-sequencing and could be compared with data on clinical outcome (N = 155), global copy number profiles (N = 145), and gene mutations (N = 128). Gene expression profiling revealed three transcriptomic clusters (TCs) without any clear separation of MFS and UPS. One TC was associated with longer metastasis-free survival. These tumours had lower tumour mutation burden (TMB), were enriched for a copy number signature representative of focal LOH and chromosomal instability on a diploid background, and were relatively immune-depleted. MFS and UPS showed extensive genomic overlap, with whole genome doubling occurring more frequently among the latter. The results support the idea that MFS and UPS tumours have largely overlapping genomic and transcriptomic features, with UPS tumours showing more aggressive behaviour and more complex genomes. Independently of the tumour type, clinically relevant subgroups were revealed by gene expression analysis, and the finding of multiple genomic subgroups strongly suggest the existence of subgroups of relevance to treatment stratification. © 2024 The Author(s). The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland., (© 2024 The Author(s). The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.)