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Allogeneic hematopoietic stem cell transplantation (HSCT) is a potentially curative treatment for children with life-threatening hematological diseases, but the procedure comprises a high risk of infertility. We have previously shown that two-thirds of male survivors had azoospermia median 18 years after pediatric HSCT [1]. However, some patients with azoospermia may produce sperm that do not reach the ejaculate, giving hope for biological paternity. Microdissection testicular sperm extraction (mTESE) is a surgical procedure for men with non-obstructive azoospermia [2]. During mTESE, the testicular tissue is systematically examined using an operating microscope to identify dilated seminiferous tubules that may contain active sperm production. Biopsies from such areas are examined for the presence of mature spermatozoa that, if found, are cryopreserved for assisted reproduction. mTESE is not routinely offered to men with azoospermia after HSCT, and the literature is, to our knowledge, limited to 15 case reports worldwide [3,4,5]. Here we present a case series of five male survivors of pediatric allogeneic HSCT who, due to their wish of biological paternity, underwent mTESE for the purpose of future assisted reproduction.

Background: Exon deletions are generally considered pathogenic, particularly when they are located out of frame. Here, we describe a pediatric, female patient presenting with hypercalcemia and a small cell carcinoma of the ovary, hypercalcemic type, and carrying a germline de novo SMARCA4 exon 14 deletion. Methods: The SMARCA4 deletion was identified by whole genome sequencing, and the effect on the RNA level was examined by gel- and capillary electrophoresis and nanopore sequencing. Results: The deletion was in silico predicted to be truncating, but RNA analysis revealed two major transcripts with deletion of exon 14 alone or exon 14 through 15, where the latter was located in-frame. Because the patient's phenotype matched that of other patients with pathogenic germline variants in SMARCA4, the deletion was classified as likely pathogenic. Conclusion: We propose to include RNA analysis in classification of single-exon deletions, especially if located outside of known functional domains, as this can identify any disparate effects on the RNA and DNA level, which may have implications for variant classification using the American College of Medical Genetics and Genomics guidelines.

In this editorial we present an overview and insights of the management of hereditary polyposis syndromes. The primary focus was on familial adenomatous polyposis, juvenile polyposis syndrome and Peutz-Jegher syndrome. Genetic testing has become increasingly available and is easier than ever to integrate into clinical practice. Furthermore, several genes have been added to the expanding list of genes associated with hereditary polyposis syndromes, allowing for precise diagnostics and tailored follow-up. Endoscopic evaluation of patients with hereditary polyposis syndromes is paramount in the surveillance strategies. Current endoscopic procedures include both diagnostic procedures and surveillance as well as therapeutic interventions. Recommendations for endoscopic procedures in the upper and lower gastrointestinal canal were described. Surgery is still a key component in the management of patients with hereditary polyposis syndromes. The increased cancer risk in these patients often render prophylactic procedures or intended curative procedures in the case of cancer development. Surgical interventions in the upper and lower gastrointestinal canal were described with relevant considerations. Development of chemopreventive medications is ongoing. Few drugs have been investigated, including nonsteroidal anti-inflammatory drugs. It has been demonstrated that cyclooxygenase-2 inhibitors may lower the number of polyps. Other medications are currently under investigation, but none have, to date, consistently been able to prevent development of disease.

Background: Guidelines from the European Hereditary Tumor Group as well as The Danish National Guidelines for Peutz–Jeghers Syndrome (PJS) state that both prenatal diagnosis and preimplantation genetic testing for monogenic disorders (PGT-M) should be offered to patients with PJS. However, only a few cases resulting in viable pregnancies have been published. Objective: We present two cases of PJS patients going through PGT-M for PJS. We highlight the awareness of this possibility and discuss the technical and ethical challenges of performing PGT-M for PJS. Methods and results: Case 1: A 36-year-old male with PJS and his partner were referred for genetic counseling. The patient carried a pathogenic de novo variant in STK11. After a terminated pregnancy of a fetus carrying the same pathogenic variant, microsatellite polymorphic marker analysis was established, and the patient was offered PGT-M. The female partner of the patient gave birth to a healthy boy after five years of fertility treatment. Case 2: A 35-year-old female with PJS and her partner were referred for genetic counseling. She carried an inherited pathogenic STK11 variant. The couple was offered PGT-M. Genetic testing of the embryos was performed using microsatellite polymorphic markers. After two rounds of oocyte extraction a blastocyst predicted not to be affected by PJS was identified. The blastocyst was transferred; however, this did not result in a viable pregnancy. Conclusions: PGT-M can be offered to patients with PJS. The process may be long and filled with ethical dilemmas requiring patients to be motivated and persistent.

Background: Studies suggest that Wilms tumours (WT) are caused by underlying genetic (5%-10%) and epigenetic (2%-29%) mechanisms, yet studies covering both aspects are sparse. Methods: We performed prospective whole-genome sequencing of germline DNA in Danish children diagnosed with WT from 2016 to 2021, and linked genotypes to deep phenotypes. Results: Of 24 patients (58% female), 3 (13%, all female) harboured pathogenic germline variants in WT risk genes (FBXW7, WT1 and REST). Only one patient had a family history of WT (3 cases), segregating with the REST variant. Epigenetic testing revealed one (4%) additional patient (female) with uniparental disomy of chromosome 11 and Beckwith-Wiedemann syndrome (BWS). We observed a tendency of higher methylation of the BWS-related imprinting centre 1 in patients with WT than in healthy controls. Three patients (13%, all female) with bilateral tumours and/or features of BWS had higher birth weights (4780 g vs 3575 g; p=0.002). We observed more patients with macrosomia (>4250 g, n=5, all female) than expected (OR 9.98 (95% CI 2.56 to 34.66)). Genes involved in early kidney development were enriched in our constrained gene analysis, including both known (WT1, FBXW7) and candidate (CTNND1, FRMD4A) WT predisposition genes. WT predisposing variants, BWS and/or macrosomia (n=8, all female) were more common in female patients than male patients (p=0.01). Conclusion: We find that most females (57%) and 33% of all patients with WT had either a genetic or another indicator of WT predisposition. This emphasises the need for scrutiny when diagnosing patients with WT, as early detection of underlying predisposition may impact treatment, follow-up and genetic counselling.

BACKGROUND: The etiology of central nervous system (CNS) tumors in children is largely unknown and population-based studies of genetic predisposition are lacking.METHODS: In this prospective, population-based study, we performed germline whole-genome sequencing in 128 children with CNS tumors, supplemented by a systematic pedigree analysis covering 3,543 close relatives.RESULTS: Thirteen children (10%) harbored pathogenic variants in known cancer genes. These children were more likely to have medulloblastoma (OR 5.9, CI 1.6-21.2) and develop metasynchronous CNS tumors (p=0.01). Similar carrier frequencies were seen among children with low-grade glioma (12.8%) and high-grade tumors (12.2%). Next, considering the high mortality of childhood CNS tumors throughout most of human evolution, we explored known pediatric-onset cancer genes, showing that they are more evolutionarily constrained than genes associated with risk of adult-onset malignancies (p=5e-4) and all other genes (p=5e-17). Based on this observation, we expanded our analysis to 2 986 genes exhibiting high evolutionary constraint in 141 456 humans. This analysis identified eight directly causative loss-of-functions variants, and showed a dose-response association between degree of constraint and likelihood of pathogenicity - raising the question of the role of other highly constrained gene alterations detected.CONCLUSIONS: ∽10% of pediatric CNS tumors can be attributed to rare variants in known cancer genes. Genes associated with high risk of childhood cancer show evolutionary evidence of constraint.

Hereditary polyposis syndromes are characterized by a large number and/or histopathologically specific polyps in the gastrointestinal tract and a high risk of both colorectal cancer and extracolonic cancer at an early age. While the genes responsible for some of the syndromes, eg, APC in familial adenomatous polyposis and STK11 in Peutz-Jeghers syndrome, have been known for decades, novel genetic causes have recently been detected that have shed light on the broader clinical spectrum of syndromes. Genetic diagnoses are important because they can facilitate a personalized surveillance program. Furthermore, at-risk members of the patient’s family can be tested and enrolled in surveillance as needed. In some cases, prenatal diagnostics should be offered. In this paper, we describe the development in germline genetics of the hereditary polyposis syndromes over the last 10–12 years, their clinical characteristics, as well as how to implement genetic analyses in the diagnostic pipeline.

Historically, cancer predisposition syndromes (CPSs) were rarely established for children with cancer. This nationwide, population-based study investigated how frequently children with cancer had or were likely to have a CPS. Children (0-17 years) in Denmark with newly diagnosed cancer were invited to participate in whole-genome sequencing of germline DNA. Suspicion of CPS was assessed according to Jongmans'/McGill Interactive Pediatric OncoGenetic Guidelines (MIPOGG) criteria and familial cancer diagnoses were verified using population-based registries. 198 of 235 (84.3%) eligible patients participated, of whom 94/198 (47.5%) carried pathogenic variants (PVs) in a CPS gene or had clinical features indicating CPS. Twenty-nine of 198 (14.6%) patients harbored a CPS, of whom 21/198 (10.6%) harbored a childhood-onset and 9/198 (4.5%) an adult-onset CPS. In addition, 23/198 (11.6%) patients carried a PV associated with biallelic CPS. Seven of the 54 (12.9%) patients carried two or more variants in different CPS genes. Seventy of 198 (35.4%) patients fulfilled the Jongmans' and/or MIPOGG criteria indicating an underlying CPS, including two of the 9 (22.2%) patients with an adult-onset CPS versus 18 of the 21 (85.7%) patients with a childhood-onset CPS (p = 0.0022), eight of the additional 23 (34.8%) patients with a heterozygous PV associated with biallelic CPS, and 42 patients without PVs. Children with a central nervous system (CNS) tumor had family members with CNS tumors more frequently than patients with other cancers (11/44, p = 0.04), but 42 of 44 (95.5%) cases did not have a PV in a CPS gene. These results demonstrate the value of systematically screening pediatric cancer patients for CPSs and indicate that a higher proportion of childhood cancers may be linked to predisposing germline variants than previously supposed.

Background With the implementation of a research project providing whole genome sequencing (WGS) to all pediatric cancer patients in Denmark (2016–2019), we sought to investigate healthcare professionals' views on WGS as it was actively being implemented in pediatric oncology. Methods Semistructured interviews were carried out with pediatric oncologists, clinical geneticists, and research coordinating nurses (N = 17), followed by content analysis of transcribed interviews. Interviews were supplemented by ethnographic observations on Danish pediatric oncology wards. Additionally, questionnaires were distributed to healthcare professionals concerning when they found it appropriate to approach families regarding WGS. The response rate was 74%. Results Healthcare professionals see imbalances in doctor–patient relationship, especially the double role doctors have as clinicians and researchers. Some were concerned that it might not be possible to obtain meaningful informed consent from all families following diagnosis. Still, 94% of respondents found it acceptable to approach families during the first 4 weeks from the child's diagnosis. Views on the utility of WGS, treatment adaptation, and surveillance differed among interviewees. Conclusion Overall, healthcare professionals see dilemmas arising from WGS in the pediatric oncology clinic, and some advocate for further educational sessions with families and healthcare professionals. Despite concerns, healthcare professionals overwhelmingly supported early approach of families regarding WGS. Interviewees disagree on the benefits of surveillance based on genetic findings.

Michael D Kappelman,1 Aksel Lange,2 Rachel L Randell,3 Patricia V Basta,4 Robert S Sandler,5 Kristina Laugesen,2 Anna Byrjalsen,2 Tina Christensen,2 Trine Frøslev,2 Rune Erichsen2,6 1Department of Pediatrics, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA; 2Department of Clinical Epidemiology,Aarhus University Hospital, Aarhus, Denmark; 3Department of Pediatrics, Duke University School of Medicine, Duke University, Durham, NC, USA; 4Department of Epidemiology, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA; 5Department of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA; 6Surgical Department, Horsens Regional Hospital, Horsens, Denmark Background: Epidemiologic studies combining exposure and outcome data with the collection of biosamples are needed to study gene–environment interactions that might contribute to the etiology of complex diseases such as pediatric Crohn’s disease (CD). Nationwide registries, including those in Denmark and other Scandinavian countries, provide efficient and reliable sources of data for epidemiological studies evaluating the environmental determinants of disease. We performed a pilot study to test the feasibility of collecting salivary DNA to augment registry data in established cases of pediatric CD and randomly selected, population-based controls.Subjects and methods: Cases of CD born after 1995 and residing in the central region of Denmark were identified through the Danish National Patient Registry and confirmed by using standard diagnostic criteria. Age- and gender-matched controls were selected at random through the civil registration system. Cases and controls were contacted by mail and telephone and invited to submit a saliva sample. DNA was extracted and genotyped for six CD-associated single-nucleotide polymorphisms (SNPs).Results: A total of 53 cases of pediatric CD were invited, and 40 contributed a saliva sample (75% response rate

With an impending introduction of genome sequencing into paediatric oncology to facilitate personalised medicine, this study examines parent perspectives on participating in whole genome sequencing (WGS) research in the difficult weeks following diagnosis. As an embedded part of Sequencing Tumor and Germline DNA—Implications and National Guidelines (STAGING), a project aiming to implement WGS of all newly diagnosed paediatric cancer patients in Denmark, a parent perspective study was conducted by a clinical geneticist and anthropologist to document pragmatic, social and ethical dilemmas. Following genetic counselling, systematic debriefings were held and the anthropologist carried out in‐depth parent interviews (N = 30 parents to 15 patients). Parents were approached about STAGING 2–28 days after diagnosis. The majority of interviewed parents reported that an early approach had been feasible for them, a few found it too early. Participation was explained in terms of altruism and a desire to learn more about why their child had developed cancer. A number of parents openly disagreed about the amount of information they wanted reported back. Enrolment in WGS research around the time of diagnosis is feasible, however, flexibility from researchers is essential. Notwithstanding high participation rates and a tendency to choose full disclosure, caution as regards the consequences of participating in WGS research is warranted.

Germ line mutations causing paediatric cancer predisposition syndromes (PCPSs) are more common than previously anticipated and are now recognised as a significant contributor to the incidence of childhood cancer. Advances in and increased clinical application of next-generation sequencing technologies have led to a rise in paediatric patients undergoing whole genome sequencing (WGS). This review focuses on the potential syndromes/diagnoses, which WGS may reveal in patients with childhood cancers, and highlights the clinical and psychosocial impact of PCPSs.

BRCA1, c.4096+3A>G was identified in a consanguineous Danish family with several cases of breast/ovarian cancer. In silico analysis and splicing assays indicated that the variant caused aberrant splicing. However, based on segregation data and the finding of a healthy homozygous carrier, we classify the BRCA1 c.4096+3A>G variant as likely benign.