High-throughput sequencing has transformed clinical diagnostics of rare diseases (RD), cancer and infectious diseases by enabling the identification of disease-causing genetic alterations and facilitating individualised treatment and care. In response to these advances, Genomic Medicine Sweden (GMS) was established in 2017 as a national collaborative effort to accelerate implementation of genomics-based precision medicine within Sweden's regionally organized, publicly funded healthcare system. GMS brings together the seven university healthcare regions and their associated medical faculties, in collaboration with healthcare regions across Sweden, Science for Life Laboratory, patient organizations, industry and governmental agencies. Activities are coordinated through national disease-specific expert groups, supported by cross-cutting functions in bioinformatics, health economics, ethics, education and patient engagement. At the operational level, seven Genomic Medicine Centres, embedded at university hospitals, develop and deliver harmonised genomic diagnostics nationwide. The National Genomics Platform provides secure infrastructure for large-scale data storage, analysis, and national and international data sharing. Following initial project-based funding, GMS now receives long-term governmental support. This review describes the national implementation of genomic-based precision diagnostics, discusses challenges and lessons learnt, and highlights key milestones across disease areas, including whole-genome sequencing in RD and paediatric cancer, comprehensive genomic profiling of haematological malignancies and solid tumours, pathogen genomics in microbiology, pharmacogenomic testing and emerging applications of polygenic risk scores in complex diseases. Collectively, these efforts have contributed to more than 500,000 genomic tests being performed within Swedish healthcare between 2017 and 2025. Finally, we outline future diagnostic needs and priority areas to ensure sustainable, scalable and equitable access to precision medicine.

ABSTRACT Ovarian tissue cryopreservation enables fertility preservation in females undergoing gonadotoxic therapies, restoring fertility in adults. Although offered even before puberty, the childhood ovary and its vulnerability to therapy remain poorly characterized. Here, ovarian tissue from 16 patients undergoing fertility preservation (aged 1-16 years) and 11 adult controls (aged 22-32 years) was analyzed using single-cell RNA sequencing, spatial transcriptomics, and multiplex immunostaining. In chemotherapy-naïve samples, 13 somatic cell populations underwent extracellular matrix remodeling, vascular, neural, and stromal maturation during puberty, whereas changes in germline related to chromatin remodeling. Spatial transcriptomics resolved 23 clusters across, revealing distinct tissue organization and follicular niche composition between children and adults. Chemotherapy exposure depleted perifollicular and vascular cells, suppressed intercellular signaling, and dysregulated over half of puberty-associated genes, converging on stress responses and extracellular matrix remodeling, with SEPTIN7 as a potential biomarker. These findings uncover critical developmental vulnerabilities of the pediatric ovary relevant to fertility preservation.

Tyrosine kinase inhibitors (TKIs) have improved outcomes in Philadelphia chromosome-positive (Ph+) acute lymphoblastic leukemia (ALL) and are increasingly incorporated into treatment protocols of Philadelphia chromosome-like (Ph-like, ABL-class) ALL. However, central nervous system (CNS) relapse remains a significant challenge. Imatinib, a first-generation TKI, demonstrates limited CNS penetration in adults, yet data in children are sparse. This prospective, multicenter study investigated cerebrospinal fluid (CSF) and matched plasma concentrations of imatinib and its primary bioactive metabolite (N-desmethyl-imatinib) in children and young adults with Ph + or Ph-like (ABL-class) ALL. Plasma and CSF samples were analyzed with liquid chromatography tandem mass spectrometry (LC-MS/MS). Linear mixed-effects models were used to assess concentrations across compartments and over time. Between January 2023 and June 2025, 32 paired plasma and CSF samples were collected from ten patients (range 1–8 samples/patient; median: 3). In total, 78

Levels of oncogenic ALK mutations in tissue samples

ABSTRACT Background The European ALLTogether protocol for childhood acute lymphoblastic leukemia, initiated in Sweden 2019, introduced earlier asparaginase during induction, dexamethasone instead of prednisone for all patients, and omitted anthracyclines from low‐risk induction to reduce treatment‐related toxicity. Consolidation‐1 was based on the Induction 1B‐phase developed by the BFM‐group, replacing mercaptopurine, methotrexate, and asparaginase used in the previous NOPHO ALL2008 protocol, ALL2008. Following implementation, early treatment toxicity was considered unacceptably high, prompting a protocol amendment. We compared the prevalence of 14 predefined toxicities, the number of inpatient days, and weight changes during induction and consolidation‐1 between the two protocols. Methods We conducted a population‐based cohort study in Sweden, reviewing patient records from 117 children treated under ALLTogether protocol and 234 matched controls under ALL2008 protocol. Results The mean number of toxicities per patient was similar between the protocols (2.5 [290/117] vs. 2.3 [547/234]). ALLTogether cohort had significantly greater weight gain, with over 50% experiencing a > 10% increase ( p < 0.01). Hyperglycemia (OR 5.17, 95% CI 1.93–13.82) and osteonecrosis (3.4% vs. 0%, p = 0.012) were more common, while liver dysfunction (0.59, 0.38–0.93) was less frequent in the ALLTogether protocol. The number of inpatient days was similar across protocols, except for the initial hospitalization, which was longer in ALLTogether (median 11 vs. 7 days, p < 0.01). Conclusions The early introduction of asparaginase likely contributed to increased weight gain, hyperglycemia, and osteonecrosis. While overall toxicity burden remained similar between protocols, the shift in toxicity profile may explain the perception of increased early toxicity during treatment with the ALLTogether protocol. Trial Registration ClinicalTrials.gov identifier: NCT03911128; EudraCT numbers: 2018‐001795‐38 and 2008‐003235‐20

Coverage with the NB-ALK sequencing panel

INTRODUCTION:Infection remains the most common treatment-related toxicity of childhood ALL, emphasizing the need to identify patients at risk and to tailor treatment strategies accordingly. AIMS:The primary aim was to compare infectious toxicity during early treatment for childhood ALL following the ALLTogether and NOPHO ALL-2008 (ALL-2008) protocols, and second, to identify risk factors for infectious toxicity. METHODS:A national retrospective matched cohort study was conducted, including 345 patients aged 1-17 years diagnosed with ALL and treated in Sweden according to the ALLTogether or ALL-2008 protocols. Nonparametric tests were used to compare infectious outcomes between protocols, and regression modeling was used to identify risk factors of the infectious outcomes. RESULTS:Treatment following ALL-2008 showed higher infectious toxicity during induction, whereas treatment following ALLTogether showed increased infectious toxicity during consolidation 1. Overall, treatment according to ALL-2008 was associated with a higher incidence of infections. Anthracycline use and young age (1-9 years) were associated with both higher infectious incidence and more severe infectious complications. Dexamethasone was associated with both lower incidence and lower severity of infectious complications as compared to prednisone. CONCLUSIONS:A notable shift in the timing of infectious toxicity was observed between the two treatment protocols. Risk factors for infectious toxicity during early treatment include anthracycline use and young age. Dexamethasone as an induction steroid was associated with lower infectious burden, although its effect is difficult to isolate from the simultaneous anthracycline effect. The findings suggest that treatment composition plays a central role in determining both the extent and timing of infectious complications.

Sequencing results, ALK hotspot mutation sites

STUDY QUESTION:Which spermatogonial differentiation states are present in prepubertal testes under normal conditions and following chemotherapy-induced depletion of spermatogonia in paediatric patients with cancer? SUMMARY ANSWER:Single-cell transcriptomic analysis reveals that only undifferentiated spermatogonia are present in prepubertal boys, while differentiated states emerge during puberty, with reduced protein expression of advanced spermatogonial markers observed in younger patients, those treated with alkylating agents, or those with a diminished spermatogonial pool. WHAT IS KNOWN ALREADY:Paediatric oncology treatments often involve gonadotoxic therapies that can impair spermatogonial stem cells, increasing the risk of subfertility. While five distinct spermatogonial subpopulations have been identified in adult testes via single-cell RNA sequencing, their presence in prepubertal testes of childhood cancer patients remains to be confirmed through marker protein expression. STUDY DESIGN, SIZE, DURATION:Gene expression profiles of spermatogonial subpopulations were investigated using single-cell RNA sequencing data from six testicular samples of healthy boys aged 0-17 years. Protein expression patterns were examined via immunofluorescence staining in 14 biobank control samples (median age: 4.9 years; range: 0.6-13.1 years) and in 31 prepubertal testicular tissue samples of paediatric patients with cancer (median age: 6.8 years; range: 0.7-13.1 years). PARTICIPANTS/MATERIALS, SETTING, METHODS:Gene expression profiles of UTF1 (states 0-1), ID4 (states 0-1), PIWIL4 (states 0-1), FGFR3 (states 0-2), and KIT (state 4), were analysed in testicular cells of paediatric origin obtained from our previously published open-access data source (GSE134144 and GSE120508). The protein expression of these spermatogonial subpopulation markers was evaluated by counting immunofluorescence-positive cells per analysed area. Marker expression was correlated with prior chemotherapy exposure and spermatogonia numbers. Exposure to alkylating agents was quantified as the cumulative cyclophosphamide equivalent dose (CED), and anthracycline exposure as the cumulative doxorubicin isoequivalent dose equivalents (DIE). A depleted spermatogonia pool was defined as having S/T Z-scores lower than -7 SD. MAIN RESULTS AND THE ROLE OF CHANCE:Transcriptomic analysis confirmed that germ cells in the prepubertal testis consist solely of undifferentiated spermatogonia. The expression of KIT protein, defining differentiated spermatogonia, was positively correlated with age (P < 0.001). A reduction in the number of spermatogonia expressing ID4 protein was associated with higher CED (P = 0.001), and spermatogonia expressing KIT protein with higher CED and DIE exposure (P = 0.005, and P = 0.035, respectively). A depleted spermatogonia pool (S/T Z-score <-7 SD) correlated with fewer spermatogonia expressing ID4 (P = 0.033), FGFR3 (P = 0.050), and KIT (P = 0.051) proteins. These results indicate that distinct protein expression patterns were observed following chemotherapy-induced reduction of the spermatogonial pool, with reduced expression of ID4, FGFR3, and KIT proteins. Numbers of spermatogonia positive for markers indicating more naïve, undifferentiated states, such as UTF1 and PIWIL4, did not correlate with spermatogonial pool reduction. LIMITATIONS, REASONS FOR CAUTION:The study population was heterogeneous in terms of age and treatment exposure. Moreover, the impact of specific cancer treatments could not be individually assessed. Limited tissue availability reduced the statistical power of the study, and repeated double or triple immunofluorescence staining could not be performed. As a result, the correlations between the expression of different spermatogonial markers can only be considered indicative trends. Child testicular control tissue samples were considered normal for inclusion if no testicular pathology was reported. However, detailed information on prior medical treatments or testicular volumes for the patients in this biobank was unavailable. WIDER IMPLICATIONS OF THE FINDINGS:Our observations suggest that alkylating agents have dose-dependent effects on all spermatogonial subpopulations. However, spermatogonial subtypes expressing the protein markers UTF1 and PIWIL4 were more resistant to chemotherapy-induced depletion of the spermatogonial pool, potentially representing true reserve stem cells. The identification of reserve stem cells could provide a valuable method for evaluating the fertility potential of testicular tissue collected for fertility preservation in prepubertal and peripubertal boys. STUDY FUNDING/COMPETING INTEREST(S):This study was supported by grants from the Swedish Childhood Cancer Fund (PR2019-0123; PR2022-0115; TJ2020-0023) (J.-B.S.), Finnish Cancer Society (K.J.), Finnish Foundation for Paediatric Research (K.J.), Swedish Research Council (2018-03094; 2021-02107) (J.-B.S.), and Birgitta and Carl-Axel Rydbeck's Research Grant for Paediatric Research (2020-00348; 2020-00335; 2021-00073; 2022-00317, 2024-00255) (J.-B.S., K.J.). Y.C. and Y.Y. received a scholarship from the Chinese Scholarship Council. J.S. was supported by a grant from Mary Béves Foundation for Childhood Cancer Research. H.B.O. was supported by the Sultan Qaboos University in Oman. The authors declare no competing interests. TRIAL REGISTRATION NUMBER:N/A.

Patients with anaplastic lymphoma kinase (ALK)-driven neuroblastoma may respond to tyrosine kinase inhibitors, but resistance to treatment occurs and methods currently used for detection of residual disease have limited sensitivity. Here, we present a national unselected cohort of five patients with relapsed or refractory ALK-driven neuroblastoma treated with lorlatinib as monotherapy and test the potential of targeted circulating tumor DNA (ctDNA) analysis as a guide for treatment decisions in these patients. We developed a sequencing panel for ultrasensitive detection of ALK mutations associated with neuroblastoma or resistance to tyrosine kinase inhibitors and used it for ctDNA analysis in 83 plasma samples collected longitudinally from the four patients who harbored somatic ALK mutations. All four patients with ALK p.R1275Q experienced major responses and were alive 35 to 61 months after starting lorlatinib. A fifth patient with ALK p.F1174L initially had a partial response but relapsed after 10 months of treatment. In all cases, ctDNA was detected at the start of lorlatinib single-agent treatment and declined gradually, correlating with clinical responses. In the two patients exhibiting relapse, ctDNA increased 9 and 3 months, respectively, before clinical detection of disease progression. In one patient harboring HRAS p.Q61L in the relapsed tumor, retrospective ctDNA analysis showed that the mutation appeared de novo after 8 months of lorlatinib treatment. We conclude that some patients with relapsed or refractory high-risk neuroblastoma show durable responses to lorlatinib as monotherapy, and targeted ctDNA analysis is effective for evaluation of treatment and early detection of relapse in ALK-driven neuroblastoma. SIGNIFICANCE:We present five patients with ALK-driven relapsed or refractory neuroblastoma treated with lorlatinib as monotherapy. All patients responded to treatment, and four of them were alive after 3 to 5 years of follow-up. We performed longitudinal ctDNA analysis with ultra-deep sequencing of the ALK tyrosine kinase domain. We conclude that ctDNA analysis may guide treatment decisions in ALK-driven neuroblastoma, also when the disease is undetectable using standard clinical methods.

Introduction Allopurinol can be used in maintenance therapy (MT) for acute lymphoblastic leukemia (ALL) to mitigate hepatic toxicity in patients with skewed 6-mercaptopurine (6MP) metabolism. These patients have high erythrocyte levels of methylated mercaptopurine metabolites (e-MeMP) associated with liver toxicity, including transaminitis and hypoglycemia, and low erythrocyte levels of thioguanine nucleotides (e-TGN), the key intermediate metabolites mediating the antileukemic effect. Retrospective studies and case reports show that the unfavorable metabolite ratio (high e-MeMP/TGN) can be modified by adding allopurinol leading to lower MeMP and higher TGN. In a recent publication https://doi.org/10.3324/haematol.2023.284390 we showed that allopurinol leads to a similar metabolic shift also in unselected pediatric ALL-patients, without previous severe liver toxicity or other signs of skewed 6MP metabolism. We have now analysed the levels of DNA incorporated TGN (DNA-TG) in our cohort since more recent studies, e.g. https://doi.org/10.1016/s1470-2045(17)30154-7, suggests that relapse-free survival is better correlated to DNA-TG than to e-TGN. Methods Pediatric ALL patients on NOPHO ALL-2008 non-high risk protocols with thiopurine methyltransferase wild-type were studied in a prospective before-after trial, NCT03022747. 6MP metabolites were measured, in total 9 blood samples per patient, during 12 weeks of standard MT, followed by 12 weeks of MT with addition of allopurinol 50 mg/m2 and finally 4 weeks of MT without allopurinol. Mean DNA-TG for each patient was calculated for all study phases separately. DNA-TG/e-TGN ratio was calculated as there was a concern that a decrease in e-MeMP might reduce the DNA incorporation of TGN, since e-MeMP is known to inhibit de novo purine synthesis. The 6MP dose was reduced by 50% when allopurinol was initiated to prevent excessive myelosuppression. DNA-TG was quantified using the same method as in the study cited above, with 1−2 µg DNA purified from whole blood and thioguanine measured with ultra-performance liquid chromatography tandem mass spectrometry. Results 51 patients from Sweden and Finland, age 0-15 (median 4) years, were included, of whom 48 completed the study. In paired analysis DNA-TG was 393 fmol/mg DNA higher (1248 vs 855) (p<0.001), e-TGN 193 nmol/mmol Hb higher (457 vs 264) (p<0.001) and e-MeMP 6324 nmol/mmol Hb lower (2614 vs 8938) (p<0.001) when allopurinol was added, Mean DNA-TG/e-TGN ratio did not change significantly, 3.15 on allopurinol compared to 3.51 before, (p=0.10). Four weeks after allopurinol was discontinued, mean DNA-TG decreased 564 fmol/mg DNA (p<0.001), e-TGN decreased 271 nmol/mmol Hb (p<0.001) whereas e-MeMP showed a non-significant increase of 777 nmol/mmol Hb (p=0.17). Alanine aminotransferase decreased by 39% a few weeks after the e-MeMP-levels decreased. Mean absolute neutrophil count (ANC) was lower on allopurinol, 1.35 x 109/L, compared to 1.72 before (p<0.001). Allopurinol did not increase the number of severe adverse events (SAE) nor cause any life-threatening episodes. In accordance with study protocol the mean given dose 6MP was halved during the weeks on allopurinol, 187 mg/m2/week compared to 379 (p<0.001) for the weeks before allopurinol. Conclusions Addition of allopurinol to standard MT with 6MP and methotrexate leads to substantially increased DNA-TG levels. DNA-TG/e-TGN ratio was unchanged indicating that DNA incorporation of TGN was not significantly affected during allopurinol treatment. The 46 % rise in DNA-TG, together with our previously published data with higher proportion of ANC levels within target, lower ALT and no increase in SAE, indicate that addition of allopurinol could be an effective strategy to optimize ALL MT.

Detection of relapse with ctDNA analysis 9 months prior to clinical presentation in a patient (patient 2) with ALK p.R1275Q-positive neuroblastoma treated with lorlatinib. A, MIBG scans. B, Levels of ctDNA. C, NSE. CS, Curie score; ND, not detected; TID, temozolomide, irinotecan and dinutuximab; URL, upper reference limit (16.3 µg/L).

BackgroundChildhood cancer predisposition (ChiCaP) syndromes are increasingly recognized as contributing factors to childhood cancer development. Yet, due to variable availability of germline testing, many children with ChiCaP might go undetected today. We report results from the nationwide and prospective ChiCaP study that investigated diagnostic yield and clinical impact of integrating germline whole-genome sequencing (gWGS) with tumor sequencing and systematic phenotyping in children with solid tumors.MethodsgWGS was performed in 309 children at diagnosis of CNS (n = 123, 40%) or extracranial (n = 186, 60%) solid tumors and analyzed for disease-causing variants in 189 known cancer predisposing genes. Tumor sequencing data were available for 74% (227/309) of patients. In addition, a standardized clinical assessment for underlying predisposition was performed in 95% (293/309) of patients.FindingsThe prevalence of ChiCaP diagnoses was 11% (35/309), of which 69% (24/35) were unknown at inclusion (diagnostic yield 8%, 24/298). A second-hit and/or relevant mutational signature was observed in 19/21 (90%) tumors with informative data. ChiCaP diagnoses were more prevalent among patients with retinoblastomas (50%, 6/12) and high-grade astrocytomas (37%, 6/16), and in those with non-cancer related features (23%, 20/88), and ≥2 positive ChiCaP criteria (28%, 22/79). ChiCaP diagnoses were autosomal dominant in 80% (28/35) of patients, yet confirmed de novo in 64% (18/28). The 35 ChiCaP findings resulted in tailored surveillance (86%, 30/35) and treatment recommendations (31%, 11/35).InterpretationOverall, our results demonstrate that systematic phenotyping, combined with genomics-based diagnostics of ChiCaP in children with solid tumors is feasible in large-scale clinical practice and critically guides personalized care in a sizable proportion of patients.FundingThe study was supported by the Swedish Childhood Cancer Fund and the Ministry of Health and Social Affairs.

The ovarian cortical reserve of follicles is vital for fertility. Some medical treatments are toxic to follicles, leading to premature ovarian insufficiency. Ovarian tissue cryopreservation is an established method to preserve fertility in adults and even applied in prepuberty despite unproven efficacy. Here, we analyze transcriptomes of 120 cortical follicles from children and adults for detailed comparison. We discover heterogeneity with two main types of follicles in both age groups: one with expected oocyte-granulosa profiles and another with predicted role in signaling. Transcriptional changes during growth to the secondary stage are similar overall in children and adults, but variations related to extracellular matrix, theca cells, and miRNA profiles are found. Notably, cyclophosphamide dose correlates with interferon signaling in child follicles. Additionally, morphology alone is insufficient for follicle categorization suggesting a need for additional markers. Marker genes for early follicle activation are determined. These findings will help refine follicular classification and fertility preservation techniques across critical ages.

Early detection of relapse and emergence of an HRAS p.Q61L mutation during lorlatinib treatment in a patient (patient 5) with ALK p.F1174L-positive neuroblastoma. A, MIBG scans at initial diagnosis, at the first relapse, after 4 months of lorlatinib (partial response), and at second relapse after 10.5 months of lorlatinib treatment. B, Allele frequency of HRAS p.Q61L in tumor tissue at different time points. C, Levels of ctDNA. D, Clinical tumor markers, values are normalized to the upper reference limit (URL) for each marker. CGA, chromogranin A; Lorla, lorlatinib; ND, not detected.

Lasting response to lorlatinib in a patient (patient 1) with relapsed neuroblastoma harboring ALK p.R1275Q. A, Chest computed tomography (top) and MIBG scans (bottom) at time of initial diagnosis, at relapse, and after 2 months of lorlatinib treatment. Red arrows denote tumors. B, Levels of ctDNA during and after lorlatinib treatment. ND, not detected. C, Chromogranin A. URL, upper reference limit (102 µg/L). NSE, urine VMA and urine HVA were within normal ranges at time of relapse and were therefore not further analyzed. HVA, homovanillic acid; VMA, vanillylmandelic acid.

STUDY QUESTION Can human pre- and peri-pubertal testicular cells obtained from childhood cancer patients, previously treated with chemotherapy, form testicular organoids (TOs)?SUMMARY ANSWER Organoid formation from testicular tissue collected from childhood cancer patients positively correlates with SRY-Box transcription factor 9 (SOX9) expression in Sertoli cells, which in turn negatively correlates with previous exposure to alkylating chemotherapy.WHAT IS KNOWN ALREADY Pre- and peri-pubertal boys exposed to highly gonadotoxic therapies can only safeguard their fertility potential through testicular tissue cryopreservation. Today, there is no established clinical tool to restore fertility using these testicular samples. Organoids hold promise in providing fundamental early insights in creating such platforms. However, the generation of TOs that closely resemble the innate testis, to enable a thorough monitoring of the necessary steps for germ cell differentiation and somatic functionalities, remains a challenge.STUDY DESIGN, SIZE, DURATION We used a Matrigel-based three-layer gradient culture system to generate human TOs and to reveal whether chemotherapy exposure affects TO formation capacity and the functionality of pre- and peri-pubertal testicular somatic cells. Testicular cells of 11 boys (aged 7.7 +/- 4.1 (mean +/- SD) years) were assessed for TO formation in relation to previous chemotherapy exposure and SOX9 expression in histological sections of paraffin-embedded testicular tissue samples collected on the day of biopsy and compared with testicular tissue samples obtained from 28 consecutive patients (aged 6.9 +/- 3.8 (mean +/- SD) years). All 39 patients were part of the fertility preservation project NORDFERTIL; an additional 10 samples (from boys aged 5.5 +/- 3.5 (mean +/- SD) years, without an underlying pathology) in an internal biobank collection were used as controls.PARTICIPANTS/MATERIALS, SETTING, METHODS We obtained 49 testicular tissue samples from boys aged 0.8-13.4 years. Fresh samples (n = 11) were dissociated into single-cell suspensions and applied to a three-layer gradient culture system for organoid formation. Histological sections of another 28 samples obtained as part of the fertility preservation project NORDFERTIL, and 10 samples from a sample collection of a pathology biobank were used to evaluate the effects of prior exposure to alkylating agents on testicular samples. Testicular organoid formation was defined based on morphological features, such as compartmentalized structures showing cord formation, and protein expression of testicular cell-specific markers for germ and somatic cells was evaluated via immunohistochemical staining. Hormone secretion was analysed by specific enzyme-linked immunosorbent assays for testosterone and anti-M & uuml;llerian hormone (AMH) production.MAIN RESULTS AND THE ROLE OF CHANCE Our results revealed that 4 out of 11 prepubertal testicular samples formed TOs that showed compartmentalized cord-like structures surrounded by interstitial-like areas and increasing levels of both testosterone as well as AMH over a 7-day culture period. We observed that SOX9 expression was correlated positively with TO formation. Moreover, exposure to alkylating agents before biopsy was inversely correlated with SOX9 expression (P = 0.006).LARGE SCALE DATA N/A.LIMITATIONS, REASONS FOR CAUTION Due to the limited amount of material available, only 11 out of the 39 pre- and peri-pubertal testicular tissue samples could be used for the organoid formation experiments. The testicular tissue samples obtained from a sample collection of the internal biobank of Department of Pathology, Karolinska University Hospital were considered normal and included in the study if no testicular pathology was reported. However, detailed information regarding previous medical treatments and/or testicular volumes of the patients included in this biobank was not available.WIDER IMPLICATIONS OF THE FINDINGS Our observations suggest that SOX9 expression may serve as a putative indicator of TO formation, indicating a critical role of Sertoli cells in promoting organoid formation, seminiferous tubule integrity, and testicular function in pre- and peri-pubertal testicular tissue.STUDY FUNDING/COMPETING INTEREST(S) This study was supported by grants from the Swedish Childhood Cancer Foundation (PR2019-0123; PR2022-0115; TJ2020-0023) (J.-B.S.), Finnish Cancer Society (K.J.), Finnish Foundation for Paediatric Research (K.J.), Swedish Research Council (2018-03094; 2021-02107) (J.-B.S.), and Birgitta and Carl-Axel Rydbeck's Research Grant for Paediatric Research (2020-00348; 2020-00335; 2021-00073; 2022-00317) (J.-B.S. and K.J.). Y.C. and Y.Y. received a scholarship from the Chinese Scholarship Council. J.P.A-L. was supported by a Starting Grant in Medicine and Health (2022-01467) from the Swedish Research Council. R.T.M. was supported by a UKRI Future Leaders Fellowship (MR/S017151/1). The MRC Centre for Reproductive Health was supported by an MRC Centre Grant (MR/N022556/1). The authors declare no competing interests.

BackgroundAdvances in treatment of childhood malignancies have improved overall cure rates to 80%. Nevertheless, cancer is still the most common cause of childhood mortality in Sweden. The prognosis is particularly poor for relapse of high-risk malignancies. In the international INFORM registry, tumor tissue from patients with relapsed, refractory, or progressive pediatric cancer as well as from very-high risk primary tumors is biologically characterized using next-generation sequencing to identify possible therapeutic targets. We analyzed data from Swedish children included in the INFORM registry concerning patient characteristics, survival, sequencing results and whether targeted treatment was administered to the children based on the molecular findings.MethodsA registry-based descriptive analysis of 184 patients included in the INFORM registry in Sweden during 2016–2021.ResultsThe most common diagnoses were soft tissue and bone sarcomas followed by high grade gliomas [including diffuse intrinsic pontine glioma (DIPG)]. Complete molecular analysis was successful for 203/212 samples originating from 184 patients. In 88% of the samples, at least one actionable target was identified. Highly prioritized targets, according to a preset scale, were identified in 48 (24%) samples from 40 patients and 24 of these patients received matched targeted treatment but only six children within a clinical trial. No statistically significant benefit in terms of overall survival or progression free survival was observed between children treated with matched targeted treatment compared to all others.ConclusionThis international collaborative study demonstrate feasibility regarding sequencing of pediatric high-risk tumors providing molecular data regarding potential actionable targets to clinicians. For a few individuals the INFORM analysis was of utmost importance and should be regarded as a new standard of care with the potential to guide targeted therapy.