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BackgroundAlthough 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 resultsWe 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.ConclusionsAfter 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.

Background Beti-cel ex vivo gene therapy integrates a modified HBB gene into hematopoietic stem cells of patients with TDT, aiming to enable lifelong, stable production of functional adult hemoglobin (Hb). The efficacy and safety of the treatment have been demonstrated in a total of 63 patients treated across 4 clinical trials (HGB-204,-HGB-205, HGB-207, and HGB-212). Here, we present the first patient who received beti-cel outside of the clinical trial setting, a 14-year-old male with a β 0/β + (IVS-1-6) genotype. Methods Following hematopoietic stem cell collection via granulocyte-colony stimulating factor plus plerixafor mobilization and apheresis, CD34+ cells were transduced with the BB305 lentiviral vector encoding HbA T87Q. The patient received hypertransfusion before mobilization and conditioning, maintaining a pre-transfusion Hb level of >11 g/dL. Six days prior to beti-cel infusion, single-agent busulfan myeloablation was initiated (16 single doses at 0.8 mg/kg body weight; 3.2 mg/kg/24 h) with concomitant clonazepam (see Table for treatment timeline). Ursodeoxycholic acid therapy was continued as hepatic veno-occlusive disease (VOD) prophylaxis through inpatient treatment. Results The patient was diagnosed with TDT at the age of 2 years in his home country and has been treated in Germany since the age of 9. Regular transfusion therapy was initiated soon after diagnosis (Table). Aged 9, the patient was started on desferasirox for iron elimination therapy. His annualized red blood cell (RBC) transfusion volume was 174 ml/kg in 2018 and 185 ml/kg in 2019, maintaining his pre-transfusion Hb at or above 9 g/dl. No HLA-related donor was available for allogeneic transplant. At informed consent, the patient was 13 years old and met the eligibility criteria for beti-cel treatment as outlined in the summary of product characteristics (SmPC). The patient was physically fit, with a 90% Lansky score and regular participation in school sports, but reported physical limitations when running extensively. The patient underwent a thorough assessment before admission (Table), which did not reveal any remarkable abnormalities except TDT-related splenomegaly and signs of slight iron overload (liver iron content, 2.0 mg/g dry weight [normal range, 0.17-1.8]). On 11/Feb/2021, the patient was infused with 5.1 × 10 6 CD34+ cells/kg. The patient received 4 RBC and 8 platelet transfusions following infusion until Day 13 and 27, respectively (Table). Neutrophil and platelet engraftment occurred on day 27 post beti-cel infusion. The patient was discharged from inpatient treatment the same day, in excellent general condition, with 90% Lansky score, Hb of 8.2 g/dl, a reticulocyte count of 9.3%, a total white cell count of 1.55/nl, a neutrophil count of 0.75/nl, and a platelet count of 24/nl. At last follow-up (+100 days), the patient felt well and exhibited normal exercise tolerance. He has received neither red blood cell nor platelet transfusions or chelation therapy since discharge. Total Hb was 11.8 g/dl (Table). Granulocytes and lymphocytes had recovered to normal levels. The patient showed continued, albeit slowly improving, thrombocytopenia (platelet count, 31/nl [29/nl at +60 days]), consistent with previous observations after beti-cel therapy. Myeloablation and beti-cel infusion were tolerated well. Adverse events post infusion were febrile neutropenia, elevated C-reactive protein levels, pruritus, gingivitis, mild mucositis, and vertigo, consistent with the SmPC. At +23 and +26 days, the patient experienced transient subjective hearing loss (quickly resolved). No VOD events occurred. Conclusions This is the first real-world patient with TDT treated with beti-cel therapy. The treatment regimen had a tolerability profile consistent with that of mobilization, apheresis, and busulfan myeloablation, matching clinical trial observations. Following treatment, this 14-year-old patient reached a total Hb of 11.8 g/dL at +100 days without requirement of red cell transfusions and continues to exhibit prolonged but slowly improving and asymptomatic thrombocytopenia. Figure 1 Figure 1. Disclosures Schmitt: TolerogenixX Ltd: Current Employment; Therakos/Mallinckrodt: Research Funding; Hexal: Other: Travel grant; Jazz Pharmaceuticals: Other: Travel grant. Schmitt: Bluebird Bio: Other: Travel grants; Novartis: Other: Travel grants, Research Funding; TolerogenixX: Current holder of individual stocks in a privately-held company; Apogenix: Research Funding; MSD: Membership on an entity's Board of Directors or advisory committees; Hexal: Other: Travel grants, Research Funding; Kite Gilead: Other: Travel grants. Kulozik: Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Sanofi: Consultancy, Honoraria; BioMedX: Consultancy, Honoraria; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; bluebird bio, Inc.: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau.

Progress in the systemic control of osteosarcoma has been limited over the past decades thus indicating the urgent clinical need for the development of novel treatment strategies. Therefore, we have recently developed new preclinical models to study promising novel agents for the treatment of pediatric osteosarcoma. The checkpoint kinase (chk) inhibitor prexasertib (LY2606368) and its salt form (LSN2940930) have recently been shown to be active in adult and pediatric malignancies, including sarcoma. We have now tested the potency of prexasertib in clonogenic survival assays in two new lines of primary patient‐derived osteosarcoma cells and in two established osteosarcoma cell lines as a single agent and in combination with cisplatin and the poly ADP‐ribose polymerase (PARP) inhibitor talazoparib. Prexasertib alone results in strongly reduced clonogenic survival at low nanomolar concentrations and acts by affecting cell cycle progression, induction of apoptosis and induction of double‐stranded DNA breakage at concentrations that are well below clinically tolerable and safe plasma concentrations. In combination with cisplatin and talazoparib, prexasertib acts in a synergistic fashion. Chk1 inhibition by prexasertib and its combination with the DNA damaging agent cisplatin and the PARP‐inhibitor talazoparib thus emerges as a potential new treatment option for pediatric osteosarcoma which will now have to be tested in preclinical primary patient derived in vivo models and clinical studies., What's new? Treatment options and outcomes for osteosarcoma have changed very little in decades. New treatment strategies are thus urgently needed. In this study, the authors found that the checkpoint inhibitor ‘prexasertib’ increased apoptosis and DNA damage in patient‐derived osteosarcoma cells, at very low concentrations. In addition, when combined with cisplatin and talazoparib, prexasertib acted in a synergistic fashion. This drug thus represents a promising therapeutic candidate for further preclinical and clinical development in osteosarcoma.

BACKGROUND: The identification of strategies to improve mutant CFTR function remains a key priority in the development of new treatments for cystic fibrosis (CF). Previous studies demonstrated that the K⁺ channel opener 1-ethyl-2-benzimidazolone (1-EBIO) potentiates CFTR-mediated Cl⁻ secretion in cultured cells and mouse colon. However, the effects of 1-EBIO on wild-type and mutant CFTR function in native human colonic tissues remain unknown. METHODS: We studied the effects of 1-EBIO on CFTR-mediated Cl⁻ secretion in rectal biopsies from 47 CF patients carrying a wide spectrum of CFTR mutations and 57 age-matched controls. Rectal tissues were mounted in perfused micro-Ussing chambers and the effects of 1-EBIO were compared in control tissues, CF tissues expressing residual CFTR function and CF tissues with no detectable Cl⁻ secretion. RESULTS: Studies in control tissues demonstrate that 1-EBIO activated CFTR-mediated Cl⁻ secretion in the absence of cAMP-mediated stimulation and potentiated cAMP-induced Cl⁻ secretion by 39.2±6.7% (P