Systematic analysis of RNA-binding proteins identifies targetable therapeutic vulnerabilities in osteosarcoma.

Osteosarcoma is the most common primary malignant bone tumor with a strong tendency to metastasize, limiting the prognosis of affected patients. Genomic, epigenomic and transcriptomic analyses have demonstrated the exquisite molecular complexity of this tumor, but have not sufficiently defined the underlying mechanisms or identified promising therapeutic targets. To systematically explore RNA-protein interactions relevant to OS, we define the RNA interactomes together with the full proteome and the transcriptome of cells from five malignant bone tumors (four osteosarcomata and one malignant giant cell tumor of the bone) and from normal mesenchymal stem cells and osteoblasts. These analyses uncover both systematic changes of the RNA-binding activities of defined RNA-binding proteins common to all osteosarcomata and individual alterations that are observed in only a subset of tumors. Functional analyses reveal a particular vulnerability of these tumors to translation inhibition and a positive feedback loop involving the RBP IGF2BP3 and the transcription factor Myc which affects cellular translation and OS cell viability. Our results thus provide insight into potentially clinically relevant RNA-binding protein-dependent mechanisms of osteosarcoma.

Primary Multi-Systemic Metastases in Osteosarcoma: Presentation, Treatment, and Survival of 83 Patients of the Cooperative Osteosarcoma Study Group.

BACKGROUND: To evaluate patient and tumour characteristics, treatment, and their impact on survival in patients with multi-systemic metastases at initial diagnosis of high-grade osteosarcoma. Precedure: Eighty-three consecutive patients who presented with multi-systemic metastases at initial diagnosis of high-grade osteosarcoma were retrospectively reviewed. In cases of curative intent, the Cooperative Osteosarcoma Study Group recommended surgical removal of all detectable metastases in addition to complete resection of the primary tumour and chemotherapy. RESULTS: Eighty-three eligible patients (1.8%) were identified among a total of 4605 individuals with high-grade osteosarcoma. Nine (10.8%) of these achieved complete surgical remission, of whom seven later had recurrences. The median follow-up time was 12 (range, 1-165) months for all patients. Actuarial event-free survival after 1, 2, and 5 years was 9.6 ± 3.2%, 1.4 ± 1.4%, and 1.4 ± 1.4%, and overall survival was 54.0 ± 5.6%, 23.2 ± 4.9%, and 8.7 ± 3.3%. In univariate analyses, elevated alkaline phosphatase before chemotherapy, pleural effusion, distant bones as metastatic sites, and more than one bone metastasis were negative prognostic factors. Among treatment-related factors, the microscopically complete resection of the primary tumour, a good response to first-line chemotherapy, the macroscopically complete resection of all affected tumour sites, and local treatment (surgery ± radiotherapy) of all bone metastases were associated with better outcomes. Tumour progression under first-line treatment significantly correlated with shorter survival times. CONCLUSION: The outlook for patients with multi-systemic primary metastases from osteosarcoma remains very poor. The utmost importance of surgical resection of all tumour sites was confirmed. For unresectable bone metastases, radiotherapy might be considered. In the patient group studied, standard chemotherapy was often insufficiently effective. In the case of such advanced disease, alternative treatment options are urgently required.

Osteosarcomas With Few Chromosomal Alterations or Adult Onset Are Genetically Heterogeneous.

Osteosarcoma is the most common primary bone malignancy, often detected in children and adolescents and commonly associated with TP53 alterations along with a high number of chromosomal rearrangements. However, osteosarcoma can affect patients of any age, and some tumors display less genetic complexity. Besides TP53 variants, data on key driving mutations are lacking for many osteosarcomas, particularly those affecting adults. To detect osteosarcoma-specific alterations, we screened transcriptomic and genomic sequencing and copy number data from 150 bone tumors originally diagnosed as osteosarcomas. To increase the precision in gene fusion detection, we developed a bioinformatic tool denoted as NAFuse, which extracts gene fusions that are verified at both the genomic and transcriptomic levels. Apart from the already reported genetic subgroups of osteosarcoma with TP53 structural variants, or MDM2 and/or CDK4 amplification, we did not identify any recurrent genetic driver that signifies the remaining cases. Among the plethora of mutations identified, we found genetic alterations characteristic of, or similar to, those of other bone and soft tissue tumors in 8 cases. These mutations were found in tumors with relatively few other genetic alterations or in adults. Due to the lack of clinical context and available tissue, we can question the diagnosis only on a genetic basis. However, our findings support the notion that osteosarcomas with few chromosomal alterations or adult onset seem genetically distinct from conventional osteosarcomas of children and adolescents.

Spontaneous remission and loss of monosomy 7: a window of opportunity for young children with SAMD9L syndrome.

Monosomy 7 is the most common cytogenetic abnormality in pediatric myelodysplastic syndrome (MDS) and associated with a high risk of disease progression. However, in young children, spontaneous loss of monosomy 7 with concomitant hematologic recovery has been described, especially in the presence of germline mutations in SAMD9 and SAMD9L genes. Here, we report on our experience of close surveillance instead of upfront hematopoietic stem cell transplantation (HSCT) in seven patients diagnosed with SAMD9L syndrome and monosomy 7 at a median age of 0.6 years (range, 0.4-2.9). Within 14 months from diagnosis, three children experienced spontaneous hematological remission accompanied by a decrease in monosomy 7 clone size. Subclones with somatic SAMD9L mutations in cis were identified in five patients, three of whom attained hematological remission. Two patients acquired RUNX1 and EZH2 mutations during the observation period, of whom one progressed to myelodysplastic syndrome with excess of blasts (MDS-EB). Four patients underwent allogeneic HSCT at a median time of 26 months (range, 14-40) from diagnosis for MDSEB, necrotizing granulomatous lymphadenitis, persistent monosomy 7, and severe neutropenia. At last follow-up, six patients were alive, while one passed away due to transplant-related causes. These data confirm previous observations that monosomy 7 can be transient in young children with SAMD9L syndrome. However, they also indicate that delaying HSCT poses a substantial risk of severe infection and disease progression. Finally, surveillance of patients with SAMD9L syndrome and monosomy 7 is critical to define the evolving genetic landscape and to determine the appropriate timing of HSCT (clinicaltrials gov. Identifier: NCT00662090).

Disruption of the TP53 locus in osteosarcoma leads to TP53 promoter gene fusions and restoration of parts of the TP53 signalling pathway.

TP53 is the most frequently mutated gene in human cancer. This gene shows not only loss-of-function mutations but also recurrent missense mutations with gain-of-function activity. We have studied the primary bone malignancy osteosarcoma, which harbours one of the most rearranged genomes of all cancers. This is odd since it primarily affects children and adolescents who have not lived the long life thought necessary to accumulate massive numbers of mutations. In osteosarcoma, TP53 is often disrupted by structural variants. Here, we show through combined whole-genome and transcriptome analyses of 148 osteosarcomas that TP53 structural variants commonly result in loss of coding parts of the gene while simultaneously preserving and relocating the promoter region. The transferred TP53 promoter region is fused to genes previously implicated in cancer development. Paradoxically, these erroneously upregulated genes are significantly associated with the TP53 signalling pathway itself. This suggests that while the classical tumour suppressor activities of TP53 are lost, certain parts of the TP53 signalling pathway that are necessary for cancer cell survival and proliferation are retained. In line with this, our data suggest that transposition of the TP53 promoter is an early event that allows for a new normal state of genome-wide rearrangements in osteosarcoma. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Survival in primary hemophagocytic lymphohistiocytosis, 2016 to 2021: etoposide is better than its reputation.

ABSTRACT: Primary hemophagocytic lymphohistiocytosis (pHLH) is a life-threatening hyperinflammatory syndrome that develops mainly in patients with genetic disorders of lymphocyte cytotoxicity and X-linked lymphoproliferative syndromes. Previous studies with etoposide-based treatment followed by hematopoetic stem cell transplantation (HSCT) resulted in 5-year survival of 50% to 59%. Contemporary data are lacking. We evaluated 88 patients with pHLH documented in the international HLH registry from 2016-2021. In 12 of 88 patients, diagnosis was made without HLH activity, based on siblings or albinism. Major HLH-directed drugs (etoposide, antithymocyte globulin, alemtuzumab, emapalumab, ruxolitinib) were administered to 66 of 76 patients who were symptomatic (86% first-line etoposide); 16 of 57 patients treated with etoposide and 3 of 9 with other first-line treatment received salvage therapy. HSCT was performed in 75 patients; 7 patients died before HSCT. Three-year probability of survival (pSU) was 82% (confidence interval [CI], 72%-88%) for the entire cohort and 77% (CI, 64%-86%) for patients receiving first-line etoposide. Compared with the HLH-2004 study, both pre-HSCT and post-HSCT survival of patients receiving first-line etoposide improved, 83% to 91% and 70% to 88%. Differences to HLH-2004 included preferential use of reduced-toxicity conditioning and reduced time from diagnosis to HSCT (from 148 to 88 days). Three-year pSU was lower with haploidentical (4 of 9 patients [44%]) than with other donors (62 of 66 [94%]; P < .001). Importantly, early HSCT for patients who were asymptomatic resulted in 100% survival, emphasizing the potential benefit of newborn screening. This contemporary standard-of-care study of patients with pHLH reveals that first-line etoposide-based therapy is better than previously reported, providing a benchmark for novel treatment regimes.

Telehealth Needs and Concerns of Stakeholders in Pediatric Palliative Home Care.

Pediatric palliative home care (PPHC) provides care for children, adolescents, and young adults with life-limiting illnesses in their own homes. Home care often requires long travel times for the PPHC team, which is available to the families 24/7 during crises. The complementary use of telehealth may improve the quality of care. In this pilot study we identify the needs and concerns of patients, teams, and other stakeholders regarding the introduction of telehealth. As a first step, focus groups were conducted in three teams. For the second step, semi-structured interviews were conducted with patients and their families (n = 15). Both steps were accompanied by quantitative surveys (mixed methods approach). The qualitative data were analyzed using content analysis. A total of 11 needs were identified, which were prioritized differently. Highest priority was given to: data transmission, video consultation, access to patient records, symptom questionnaires, and communication support. The concerns identified were related to the assumption of deterioration of the status quo. Potential causes of deterioration were thought to be the negative impact on patient care, inappropriate user behavior, or a high level of technical requirements. As a conclusion, we define six recommendations for telehealth in PPHC.

Solitary pulmonary metastases at first recurrence of osteosarcoma: Presentation, treatment, and survival of 219 patients of the Cooperative Osteosarcoma Study Group.

BACKGROUND: To evaluate patient and tumour characteristics, treatment and their impact on survival in patients with a solitary pulmonary metastasis at first relapse of high-grade osteosarcoma. PROCEDURE: Two-hundred and nineteen consecutive patients who had achieved a complete surgical remission and then developed a solitary pulmonary metastasis at first recurrence of high-grade osteosarcoma were retrospectively reviewed. RESULTS: Two hundred and three (94.9%) of 214 patients achieved a second complete remission. After a median time from initial diagnosis of osteosarcoma to first relapse of 2.3 years (range, 0.3-18.8 years), actuarial post-relapse overall survival after 2 and 5 years was 72.0% and 51.2%. Post-relapse event-free survival was 39.1% and 31.1%. Median follow-up time was 3.2 years (range, 0.1-29.4 years). A longer time until first relapse and diagnosis due to imaging were positive prognostic factors in uni- and multivariate analyses, as were a second complete surgical remission and, in regard to death, the absence of a subsequent relapse. The use of salvage chemotherapy and radiotherapy were not associated with patient outcomes, nor was the surgical approach (thoracoscopy vs. thoracotomy) nor the exploration (uni- vs. bilateral). CONCLUSION: Approximately half of the patients who experience a solitary pulmonary relapse at first recurrence of osteosarcoma remain alive 5 years after this first relapse. Only one third will remain disease-free. A complete surgical resection of the lesion is essential for long-term survival while relapse chemotherapy does not seem to improve survival. Innovative therapies are required to improve outcomes.

Patient-tailored adoptive immunotherapy with EBV-specific T cells from related and unrelated donors.

BACKGROUNDAdoptive transfer of EBV-specific T cells can restore specific immunity in immunocompromised patients with EBV-associated complications.METHODSWe provide results of a personalized T cell manufacturing program evaluating donor, patient, T cell product, and outcome data. Patient-tailored clinical-grade EBV-specific cytotoxic T lymphocyte (EBV-CTL) products from stem cell donors (SCDs), related third-party donors (TPDs), or unrelated TPDs from the allogeneic T cell donor registry (alloCELL) at Hannover Medical School were manufactured by immunomagnetic selection using a CliniMACS Plus or Prodigy device and the EBV PepTivators EBNA-1 and Select. Consecutive manufacturing processes were evaluated, and patient outcome and side effects were retrieved by retrospective chart analysis.RESULTSForty clinical-grade EBV-CTL products from SCDs, related TPDs, or unrelated TPDs were generated for 37 patients with refractory EBV infections or EBV-associated malignancies with and without a history of transplantation, within 5 days (median) after donor identification. Thirty-four patients received 1-14 EBV-CTL products (fresh and cryopreserved). EBV-CTL transfer led to a complete response in 20 of 29 patients who were evaluated for clinical response. No infusion-related toxicity was reported. EBV-specific T cells in patients' blood were detectable in 16 of 18 monitored patients (89%) after transfer, and their presence correlated with clinical response.CONCLUSIONPersonalized clinical-grade manufacture of EBV-CTL products via immunomagnetic selection from SCDs, related TPDs, or unrelated TPDs in a timely manner is feasible. Overall, EBV-CTLs were clinically effective and well tolerated. Our data suggest EBV-CTL transfer as a promising therapeutic approach for immunocompromised patients with refractory EBV-associated diseases beyond HSCT, as well as patients with preexisting organ dysfunction.TRIAL REGISTRATIONNot applicable.FUNDINGThis study was funded in part by the German Research Foundation (DFG, 158989968/SFB 900), the Deutsche Kinderkrebsstiftung (DKS 2013.09), Wilhelm-Sander-Stiftung (reference 2015.097.1), Ellen-Schmidt-Program of Hannover Medical School, and German Federal Ministry of Education and Research (reference 01EO0802).

Constitutional Microsatellite Instability, Genotype, and Phenotype Correlations in Constitutional Mismatch Repair Deficiency.

BACKGROUND & AIMS: Constitutional mismatch repair deficiency (CMMRD) is a rare recessive childhood cancer predisposition syndrome caused by germline mismatch repair variants. Constitutional microsatellite instability (cMSI) is a CMMRD diagnostic hallmark and may associate with cancer risk. We quantified cMSI in a large CMMRD patient cohort to explore genotype-phenotype correlations using novel MSI markers selected for instability in blood. METHODS: Three CMMRD, 1 Lynch syndrome, and 2 control blood samples were genome sequenced to >120× depth. A pilot cohort of 8 CMMRD and 38 control blood samples and a blinded cohort of 56 CMMRD, 8 suspected CMMRD, 40 Lynch syndrome, and 43 control blood samples were amplicon sequenced to 5000× depth. Sample cMSI score was calculated using a published method comparing microsatellite reference allele frequencies with 80 controls. RESULTS: Thirty-two mononucleotide repeats were selected from blood genome and pilot amplicon sequencing data. cMSI scoring using these MSI markers achieved 100% sensitivity (95% CI, 93.6%-100.0%) and specificity (95% CI 97.9%-100.0%), was reproducible, and was superior to an established tumor MSI marker panel. Lower cMSI scores were found in patients with CMMRD with MSH6 deficiency and patients with at least 1 mismatch repair missense variant, and patients with biallelic truncating/copy number variants had higher scores. cMSI score did not correlate with age at first tumor. CONCLUSIONS: We present an inexpensive and scalable cMSI assay that enhances CMMRD detection relative to existing methods. cMSI score is associated with mismatch repair genotype but not phenotype, suggesting it is not a useful predictor of cancer risk.

Drug sensitivity profiling of 3D tumor tissue cultures in the pediatric precision oncology program INFORM.

The international precision oncology program INFORM enrolls relapsed/refractory pediatric cancer patients for comprehensive molecular analysis. We report a two-year pilot study implementing ex vivo drug sensitivity profiling (DSP) using a library of 75-78 clinically relevant drugs. We included 132 viable tumor samples from 35 pediatric oncology centers in seven countries. DSP was conducted on multicellular fresh tumor tissue spheroid cultures in 384-well plates with an overall mean processing time of three weeks. In 89 cases (67%), sufficient viable tissue was received; 69 (78%) passed internal quality controls. The DSP results matched the identified molecular targets, including BRAF, ALK, MET, and TP53 status. Drug vulnerabilities were identified in 80% of cases lacking actionable (very) high-evidence molecular events, adding value to the molecular data. Striking parallels between clinical courses and the DSP results were observed in selected patients. Overall, DSP in clinical real-time is feasible in international multicenter precision oncology programs.

FOXR2 Is an Epigenetically Regulated Pan-Cancer Oncogene That Activates ETS Transcriptional Circuits.

Forkhead box R2 (FOXR2) is a forkhead transcription factor located on the X chromosome whose expression is normally restricted to the testis. In this study, we performed a pan-cancer analysis of FOXR2 activation across more than 10,000 adult and pediatric cancer samples and found FOXR2 to be aberrantly upregulated in 70% of all cancer types and 8% of all individual tumors. The majority of tumors (78%) aberrantly expressed FOXR2 through a previously undescribed epigenetic mechanism that involves hypomethylation of a novel promoter, which was functionally validated as necessary for FOXR2 expression and proliferation in FOXR2-expressing cancer cells. FOXR2 promoted tumor growth across multiple cancer lineages and co-opted ETS family transcription circuits across cancers. Taken together, this study identifies FOXR2 as a potent and ubiquitous oncogene that is epigenetically activated across the majority of human cancers. The identification of hijacking of ETS transcription circuits by FOXR2 extends the mechanisms known to active ETS transcription factors and highlights how transcription factor families cooperate to enhance tumorigenesis. SIGNIFICANCE: This work identifies a novel promoter that drives aberrant FOXR2 expression and delineates FOXR2 as a pan-cancer oncogene that specifically activates ETS transcriptional circuits across human cancers. See related commentary by Liu and Northcott, p. 2977.

Integrative medicine during the intensive phase of chemotherapy in pediatric oncology in Germany: a randomized controlled trial with 5-year follow up.

BACKGROUND: Integrative medicine is used frequently alongside chemotherapy treatment in pediatric oncology, but little is known about the influence on toxicity. This German, multi-center, open-label, randomized controlled trial assessed the effects of complementary treatments on toxicity related to intensive-phase chemotherapy treatment in children aged 1-18 with the primary outcome of the toxicity sum score. Secondary outcomes were chemotherapy-related toxicity, overall and event-free survival after 5 years in study patients. METHODS: Intervention and control were given standard chemotherapy according to malignancy & tumor type. The intervention arm was provided with anthroposophic supportive treatment (AST); given as anthroposophic base medication (AMP), as a base medication for all patients and additional on-demand treatment tailored to the intervention malignancy groups. The control was given no AMP. The toxicity sum score (TSS) was assessed using NCI-CTC scales. RESULTS: Data of 288 patients could be analyzed. Analysis did not reveal any statistically significant differences between the AST and the control group for the primary endpoint or the toxicity measures (secondary endpoints). Furthermore, groups did not differ significantly in the five-year overall and event-free survival follow up. DISCUSSION: In this trial findings showed that AST was able to be safely administered in a clinical setting, although no beneficial effects of AST between group toxicity scores, overall or event-free survival were shown.

Methylation and copy number profiling: emerging tools to differentiate osteoblastoma from malignant mimics?

Rearrangements of the transcription factors FOS and FOSB have recently been identified as the genetic driver event underlying osteoid osteoma and osteoblastoma. Nuclear overexpression of FOS and FOSB have since then emerged as a reliable surrogate marker despite limitations in specificity and sensitivity. Indeed, osteosarcoma can infrequently show nuclear FOS expression and a small fraction of osteoblastomas seem to arise independent of FOS/FOSB rearrangements. Acid decalcification and tissue preservation are additional factors that can negatively influence immunohistochemical testing and make diagnostic decision-making challenging in individual cases. Particularly aggressive appearing osteoblastomas, also referred to as epithelioid osteoblastomas, and osteoblastoma-like osteosarcoma can be difficult to distinguish, underlining the need for additional markers to support the diagnosis. Methylation and copy number profiling, a technique well established for the classification of brain tumors, might fill this gap. Here, we set out to comprehensively characterize a series of 77 osteoblastomas by immunohistochemistry, fluorescence in-situ hybridization as well as copy number and methylation profiling and compared our findings to histologic mimics. Our results show that osteoblastomas are uniformly characterized by flat copy number profiles that can add certainty in reaching the correct diagnosis. The methylation cluster formed by osteoblastomas, however, so far lacks specificity and can be misleading in individual cases.

DNA methylation-based classifier and gene expression signatures detect BRCAness in osteosarcoma.

Although osteosarcoma (OS) is a rare cancer, it is the most common primary malignant bone tumor in children and adolescents. BRCAness is a phenotypical trait in tumors with a defect in homologous recombination repair, resembling tumors with inactivation of BRCA1/2, rendering these tumors sensitive to poly (ADP)-ribose polymerase inhibitors (PARPi). Recently, OS was shown to exhibit molecular features of BRCAness. Our goal was to develop a method complementing existing genomic methods to aid clinical decision making on administering PARPi in OS patients. OS samples with DNA-methylation data were divided to BRCAness-positive and negative groups based on the degree of their genomic instability (n = 41). Methylation probes were ranked according to decreasing variance difference between two groups. The top 2000 probes were selected for training and cross-validation of the random forest algorithm. Two-thirds of available OS RNA-Seq samples (n = 17) from the top and bottom of the sample list ranked according to genome instability score were subjected to differential expression and, subsequently, to gene set enrichment analysis (GSEA). The combined accuracy of trained random forest was 85% and the average area under the ROC curve (AUC) was 0.95. There were 449 upregulated and 1,079 downregulated genes in the BRCAness-positive group (fdr < 0.05). GSEA of upregulated genes detected enrichment of DNA replication and mismatch repair and homologous recombination signatures (FWER < 0.05). Validation of the BRCAness classifier with an independent OS set (n = 20) collected later in the course of study showed AUC of 0.87 with an accuracy of 90%. GSEA signatures computed for this test set were matching the ones observed in the training set enrichment analysis. In conclusion, we developed a new classifier based on DNA-methylation patterns that detects BRCAness in OS samples with high accuracy. GSEA identified genome instability signatures. Machine-learning and gene expression approaches add new epigenomic and transcriptomic aspects to already established genomic methods for evaluation of BRCAness in osteosarcoma and can be extended to cancers characterized by genome instability.

The Pediatric Precision Oncology INFORM Registry: Clinical Outcome and Benefit for Patients with Very High-Evidence Targets.

INFORM is a prospective, multinational registry gathering clinical and molecular data of relapsed, progressive, or high-risk pediatric patients with cancer. This report describes long-term follow-up of 519 patients in whom molecular alterations were evaluated according to a predefined seven-scale target prioritization algorithm. Mean turnaround time from sample receipt to report was 25.4 days. The highest target priority level was observed in 42 patients (8.1%). Of these, 20 patients received matched targeted treatment with a median progression-free survival of 204 days [95% confidence interval (CI), 99-not applicable], compared with 117 days (95% CI, 106-143; P = 0.011) in all other patients. The respective molecular targets were shown to be predictive for matched treatment response and not prognostic surrogates for improved outcome. Hereditary cancer predisposition syndromes were identified in 7.5% of patients, half of which were newly identified through the study. Integrated molecular analyses resulted in a change or refinement of diagnoses in 8.2% of cases. SIGNIFICANCE: The pediatric precision oncology INFORM registry prospectively tested a target prioritization algorithm in a real-world, multinational setting and identified subgroups of patients benefiting from matched targeted treatment with improved progression-free survival, refinement of diagnosis, and identification of hereditary cancer predisposition syndromes.See related commentary by Eggermont et al., p. 2677.This article is highlighted in the In This Issue feature, p. 2659.

The early evolutionary landscape of osteosarcoma provides clues for targeted treatment strategies.

Osteosarcomas are aggressive primary tumors of bone that are typically detected in locally advanced stages; however, which genetic mutations drive the cancer before its clinical detection remain unknown. To identify these events, we performed longitudinal genome-sequencing analysis of 12 patients with metastatic or refractory osteosarcoma. Phylogenetic and molecular clock analyses were carried out next to identify actionable mutations, and these were validated by integrating data from additional 153 osteosarcomas and pre-existing functional evidence from mouse PDX models. We found that the earliest and thus clinically most promising mutations affect the cell cycle G1 transition, which is guarded by cyclins D3, E1, and cyclin-dependent kinases 2, 4, and 6. Cell cycle G1 alterations originate no more than a year before the primary tumor is clinically detected and occur in >90% and 50% of patients of the discovery and validation cohorts, respectively. In comparison, other cancer driver mutations could be acquired at any evolutionary stage and often do not become pervasive. Consequently, our data support that the repertoire of actionable mutations present in every osteosarcoma cell is largely limited to cell cycle G1 mutations. Since they occur in mutually exclusive combinations favoring either CDK2 or CDK4/6 pathway activation, we propose a new genomically-based algorithm to direct patients to correct clinical trial options. © 2021 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd. on behalf of The Pathological Society of Great Britain and Ireland.

Germline RET variants underlie a subset of paediatric osteosarcoma.

BACKGROUND: Although considerable effort has been put into decoding of the osteosarcoma genome, very little is known about germline mutations that underlie this primary malignant tumour of bone. METHODS AND RESULTS: We followed here a coincidental finding in a multiple endocrine neoplasia family in which a 32-year-old patient carrying a germline pathogenic RET mutation developed an osteosarcoma 2 years after the resection of a medullary thyroid carcinoma. Sequencing analysis of additional 336 patients with osteosarcoma led to the identification of germline activating mutations in the RET proto-oncogene in three cases and somatic amplifications of the gene locus in five matched tumours (4%, n=5/124 tumours). Functional analysis of the pathogenic variants together with an integrative analysis of osteosarcoma genomes confirmed that the mutant RET proteins couple functional kinase activity to dysfunctional ligand binding. RET mutations further co-operated with alterations in TP53 and RB1, suggesting that osteosarcoma pathogenesis bears reminiscence to the stepwise model of medullary thyroid carcinoma. CONCLUSIONS: After Li-Fraumeni-predisposing mutations in TP53, RET becomes the second most mutated cancer-predisposing gene in the germline of patients with osteosarcoma. Hence, early identification of RET mutation carriers can help to identify at-risk family members and carry out preventive measures.

Overactivation of the IGF signalling pathway in osteosarcoma: a potential therapeutic target?

Osteosarcoma is the most common primary malignant bone tumour in children and adolescents. More than a third of patients do not respond to standard therapy and urgently require alternative treatment options. Due to a high degree of inter- and intra-tumoural genomic heterogeneity and complexity, recurrent molecular alterations that could serve as prognostic predictors or therapeutic targets are still lacking in osteosarcoma. Copy number (CN) gains involving the IGF1R gene, however, have been suggested as a potential surrogate marker for treating a subset of patients with IGF1R inhibitors. In this study, we screened a large set of osteosarcomas and found specific CN gains of the IGF1R gene in 18 of 253 (7.1%) cases with corresponding IGF1R overexpression. Despite the discouraging results observed in clinical trials in other tumours so far, focusing only on selected patients with osteosarcoma that show evidence of IGF pathway activation might represent a promising new and innovative treatment approach.