International consensus statement on the diagnosis and management of phaeochromocytoma and paraganglioma in children and adolescents.

Phaeochromocytomas and paragangliomas (PPGL) are rare neuroendocrine tumours that arise not only in adulthood but also in childhood and adolescence. Up to 70-80% of childhood PPGL are hereditary, accounting for a higher incidence of metastatic and/or multifocal PPGL in paediatric patients than in adult patients. Key differences in the tumour biology and management, together with rare disease incidence and therapeutic challenges in paediatric compared with adult patients, mandate close expert cross-disciplinary teamwork. Teams should ideally include adult and paediatric endocrinologists, oncologists, cardiologists, surgeons, geneticists, pathologists, radiologists, clinical psychologists and nuclear medicine physicians. Provision of an international Consensus Statement should improve care and outcomes for children and adolescents with these tumours.

ERN GENTURIS clinical practice guidelines for the diagnosis, surveillance and management of people with Birt-Hogg-Dubé syndrome.

Birt-Hogg-Dubé syndrome (BHD syndrome) is an autosomal dominant multisystem disorder with variable expression due to pathogenic constitutional variants in the FLCN gene. Patients with BHD syndrome are predisposed to benign cutaneous fibrofolliculomas/trichodischomas, pulmonary cysts with an associated risk of spontaneous pneumothorax, and renal cell carcinoma. A requirement for updated International consensus recommendations for the diagnosis and management of BHD syndrome was identified. Based on a comprehensive literature review and expert consensus within the fields of respiratory medicine, urology, radiology, dermatology, clinical oncology and clinical genetics, updated recommendations for diagnosis, surveillance and management in BHD syndrome were developed. With the widespread availability of FLCN genetic testing, clinical scenarios in which a diagnosis should be considered and criteria for genetic testing were defined. Following a clinical and/or molecular diagnosis of BHD syndrome, a multidisciplinary approach to disease management is required. Regular renal cancer surveillance is recommended in adulthood and life-long, but the evidence base for additional tumour surveillance is limited and further research warranted. Recommendations for the treatment of cutaneous, pulmonary and renal manifestations are provided. Awareness of BHD syndrome needs to be raised and better knowledge of the clinical settings in which the diagnosis should be considered should enable earlier diagnosis. Further details, including areas for future research topics are available at: https://www.genturis.eu/l=eng/Guidelines-and-pathways/Clinical-practice-guidelines.html .

Characteristics, aetiology and implications for management of multiple primary renal tumours: a systematic review.

In a subset of patients with renal tumours, multiple primary lesions may occur. Predisposition to multiple primary renal tumours (MPRT) is a well-recognised feature of some inherited renal cancer syndromes. The diagnosis of MPRT should therefore provoke a thorough assessment for clinical and genetic evidence of disorders associated with predisposition to renal tumourigenesis. To better define the clinical and genetic characteristics of MPRT, a systematic literature review was performed for publications up to 3 April 2024. A total of 7689 patients from 467 articles were identified with MPRT. Compared to all patients with renal cell carcinoma (RCC), patients with MPRT were more likely to be male (71.8% versus 63%) and have an earlier age at diagnosis (<46 years, 32.4% versus 19%). In 61.1% of cases MPRT were synchronous. The proportion of cases with similar histology and the proportion of cases with multiple papillary renal cell carcinoma (RCC) (16.1%) were higher than expected. In total, 14.9% of patients with MPRT had a family history of cancer or were diagnosed with a hereditary RCC associated syndrome with von Hippel-Lindau (VHL) disease being the most common one (69.7%), followed by Birt-Hogg-Dubé (BHD) syndrome (14.2%). Individuals with a known or likely genetic cause were, on average, younger (43.9 years versus 57.1 years). In rare cases intrarenal metastatic RCC can phenocopy MPRT. We review potential genetic causes of MPRT and their implications for management, suggest an approach to genetic testing for individuals presenting with MPRT and considerations in cases in which routine germline genetic testing does not provide a diagnosis.

Wilms tumour resulting from paternal transmission of a TRIM28 pathogenic variant-A first report.

Wilms tumour (nephroblastoma) is a renal embryonal tumour that is frequently caused by constitutional variants in a small range of cancer predisposition genes. TRIM28 has recently been identified as one such gene. Previously, observational data strongly suggested a parent of origin effect, whereby Wilms tumour only occurred following maternal inheritance of a pathogenic genetic variant. However, here we report a child with bilateral Wilms tumour who had inherited a pathogenic TRIM28 variant from their father. This finding suggests that genetic counselling for paternally inherited pathogenic variants in TRIM28 should include discussion of a potential risk of Wilms tumour.

When should we offer antenatal sequencing for urinary tract malformations? A systematic review, cohort study and meta-analysis.

OBJECTIVE: Determine the incremental yield of prenatal exome sequencing (PES) over chromosome microarray (CMA) and/or karyotype for urinary tract malformations (UTMs). METHOD: A prospective cohort study encompassing data from the English Genomic Medicine Service North Thames Laboratory Hub for fetuses with bilateral echogenic kidneys (BEKs) was combined with data from a systematic review. MEDLINE, EMBASE, Web of Science, MedRxiv and GreyLit were searched from 01/2010-02/2023 for studies reporting on the yield of PES over CMA or karyotype in fetuses with UTMs. Pooled incremental yield was determined using a random effects model. PROSPERO CRD42023364544. RESULTS: Fourteen studies (410 cases) were included. The incremental yield for multisystem UTMs, any isolated UTMs, and BEKs was 31% [95% CI, 18%-46%; I2  = 78%], 16% [95% CI, 6%-26%; I2  = 80%] and 51% [95% CI, 27%-75%; I2  = 34%]. The most common clinical diseases and syndromes identified, based on the variant genes detected, were Bardet-Biedl syndrome (BBS genes), dominant and recessive polycystic kidney diseases (PKD1, PKD2 and PKHD1) and renal cysts and diabetes syndrome (HNF1B). CONCLUSION: There was a notable incremental genetic diagnostic yield when PES was applied to multisystem UTMs and BEKs. There was a modest incremental yield when this technique was used for UTMs other than BEKs.

Paediatric research sets new standards for therapy in paediatric and adult cholestasis.

Children with Alagille syndrome and progressive familial intrahepatic cholestasis (PFIC) experience debilitating pruritus, for which there have been few effective treatment options. In the past 2 years, the ileal bile acid transporter (IBAT) inhibitors maralixibat and odevixibat have been approved for the management of cholestatic pruritus in these individuals, representing an important step forward in improving their quality of life. Emerging data suggest these drugs might also improve event-free survival, therefore potentially altering the typical disease course currently seen in these disorders. This Review will discuss how genetic advances have clarified the molecular basis of cholestatic disorders, facilitating the development of new therapeutic options that have only been evaluated in children. We focus specifically on the newly licensed IBAT inhibitors for patients with Alagille syndrome and PFIC and explore the next steps for these drugs in relation to other paediatric and adult cholestatic disorders, recognising that they have the potential to benefit a wider group of patients with gastrointestinal and liver disease.

Management of phaeochromocytoma and paraganglioma in patients with germline SDHB pathogenic variants: an international expert Consensus statement.

Adult and paediatric patients with pathogenic variants in the gene encoding succinate dehydrogenase (SDH) subunit B (SDHB) often have locally aggressive, recurrent or metastatic phaeochromocytomas and paragangliomas (PPGLs). Furthermore, SDHB PPGLs have the highest rates of disease-specific morbidity and mortality compared with other hereditary PPGLs. PPGLs with SDHB pathogenic variants are often less differentiated and do not produce substantial amounts of catecholamines (in some patients, they produce only dopamine) compared with other hereditary subtypes, which enables these tumours to grow subclinically for a long time. In addition, SDHB pathogenic variants support tumour growth through high levels of the oncometabolite succinate and other mechanisms related to cancer initiation and progression. As a result, pseudohypoxia and upregulation of genes related to the hypoxia signalling pathway occur, promoting the growth, migration, invasiveness and metastasis of cancer cells. These factors, along with a high rate of metastasis, support early surgical intervention and total resection of PPGLs, regardless of the tumour size. The treatment of metastases is challenging and relies on either local or systemic therapies, or sometimes both. This Consensus statement should help guide clinicians in the diagnosis and management of patients with SDHB PPGLs.

Germline pathogenic variants in HNRNPU are associated with alterations in blood methylome.

HNRNPU encodes a multifunctional RNA-binding protein that plays critical roles in regulating pre-mRNA splicing, mRNA stability, and translation. Aberrant expression and dysregulation of HNRNPU have been implicated in various human diseases, including cancers and neurological disorders. We applied a next generation sequencing based assay (EPIC-NGS) to investigate genome-wide methylation profiling for >2 M CpGs for 7 individuals with a neurodevelopmental disorder associated with HNRNPU germline pathogenic loss-of-function variants. Compared to healthy individuals, 227 HNRNPU-associated differentially methylated positions were detected. Both hyper- and hypomethylation alterations were identified but the former predominated. The identification of a methylation episignature for HNRNPU-associated neurodevelopmental disorder (NDD) implicates HNPRNPU-related chromatin alterations in the aetiopathogenesis of this disorder and suggests that episignature profiling should have clinical utility as a predictor for the pathogenicity of HNRNPU variants of uncertain significance. The detection of a methylation episignaure for HNRNPU-associated NDD is consistent with a recent report of a methylation episignature for HNRNPK-associated NDD.

Epigenotype-genotype-phenotype correlations in SETD1A and SETD2 chromatin disorders.

Germline pathogenic variants in two genes encoding the lysine-specific histone methyltransferase genes SETD1A and SETD2 are associated with neurodevelopmental disorders (NDDs) characterized by developmental delay and congenital anomalies. The SETD1A and SETD2 gene products play a critical role in chromatin-mediated regulation of gene expression. Specific methylation episignatures have been detected for a range of chromatin gene-related NDDs and have impacted clinical practice by improving the interpretation of variant pathogenicity. To investigate if SETD1A and/or SETD2-related NDDs are associated with a detectable episignature, we undertook targeted genome-wide methylation profiling of > 2 M CpGs using a next-generation sequencing-based assay. A comparison of methylation profiles in patients with SETD1A variants (n = 6) did not reveal evidence of a strong methylation episignature. A review of the clinical and genetic features of the SETD2 patient group revealed that, as reported previously, there were phenotypic differences between patients with truncating mutations (n = 4, Luscan-Lumish syndrome; MIM:616831) and those with missense codon 1740 variants [p.Arg1740Trp (n = 4) and p.Arg1740Gln (n = 2)]. Both SETD2 subgroups demonstrated a methylation episignature, which was characterized by hypomethylation and hypermethylation events, respectively. Within the codon 1740 subgroup, both the methylation changes and clinical phenotype were more severe in those with p.Arg1740Trp variants. We also noted that two of 10 cases with a SETD2-NDD had developed a neoplasm. These findings reveal novel epigenotype-genotype-phenotype correlations in SETD2-NDDs and predict a gain-of-function mechanism for SETD2 codon 1740 pathogenic variants.

Clinical consensus guideline on the management of phaeochromocytoma and paraganglioma in patients harbouring germline SDHD pathogenic variants.

Patients with germline SDHD pathogenic variants (encoding succinate dehydrogenase subunit D; ie, paraganglioma 1 syndrome) are predominantly affected by head and neck paragangliomas, which, in almost 20% of patients, might coexist with paragangliomas arising from other locations (eg, adrenal medulla, para-aortic, cardiac or thoracic, and pelvic). Given the higher risk of tumour multifocality and bilaterality for phaeochromocytomas and paragangliomas (PPGLs) because of SDHD pathogenic variants than for their sporadic and other genotypic counterparts, the management of patients with SDHD PPGLs is clinically complex in terms of imaging, treatment, and management options. Furthermore, locally aggressive disease can be discovered at a young age or late in the disease course, which presents challenges in balancing surgical intervention with various medical and radiotherapeutic approaches. The axiom-first, do no harm-should always be considered and an initial period of observation (ie, watchful waiting) is often appropriate to characterise tumour behaviour in patients with these pathogenic variants. These patients should be referred to specialised high-volume medical centres. This consensus guideline aims to help physicians with the clinical decision-making process when caring for patients with SDHD PPGLs.

Update of penetrance estimates in Birt-Hogg-Dubé syndrome.

BACKGROUND: Birt-Hogg-Dubé (BHD) syndrome is a rare genetic syndrome caused by pathogenic or likely pathogenic germline variants in the FLCN gene. Patients with BHD syndrome have an increased risk of fibrofolliculomas, pulmonary cysts, pneumothorax and renal cell carcinoma. There is debate regarding whether colonic polyps should be added to the criteria. Previous risk estimates have mostly been based on small clinical case series. METHODS: A comprehensive review was conducted to identify studies that had recruited families carrying pathogenic or likely pathogenic variants in FLCN. Pedigree data were requested from these studies and pooled. Segregation analysis was used to estimate the cumulative risk of each manifestation for carriers of FLCN pathogenic variants. RESULTS: Our final dataset contained 204 families that were informative for at least one manifestation of BHD (67 families informative for skin manifestations, 63 for lung, 88 for renal carcinoma and 29 for polyps). By age 70 years, male carriers of the FLCN variant have an estimated 19% (95% CI 12% to 31%) risk of renal tumours, 87% (95% CI 80% to 92%) of lung involvement and 87% (95% CI 78% to 93%) of skin lesions, while female carriers had an estimated 21% (95% CI 13% to 32%) risk of renal tumours, 82% (95% CI 73% to 88%) of lung involvement and 78% (95% CI 67% to 85%) of skin lesions. The cumulative risk of colonic polyps by age 70 years old was 21% (95% CI 8% to 45%) for male carriers and 32% (95% CI 16% to 53%) for female carriers. CONCLUSIONS: These updated penetrance estimates, based on a large number of families, are important for the genetic counselling and clinical management of BHD syndrome.

Preferential MGMT hypermethylation in SDH-deficient wild-type GIST.

AIMS: Wild-type gastrointestinal stromal tumours (wtGIST) are frequently caused by inherited pathogenic variants, or somatic alterations in the succinate dehydrogenase subunit genes (SDHx). Succinate dehydrogenase is a key enzyme in the citric acid cycle. SDH deficiency caused by SDHx inactivation leads to an accumulation of succinate, which inhibits DNA and histone demethylase enzymes, resulting in global hypermethylation. Epigenetic silencing of the DNA repair gene MGMT has proven utility as a positive predictor of the therapeutic efficacy of the alklyating drug temozolomide (TMZ) in tumours such as glioblastoma multiforme. The aim of this study was to examine MGMT promoter methylation status in a large cohort of GIST. METHODS: MGMT methylation analysis was performed on 65 tumour samples including 47 wtGIST (33 SDH-deficient wtGIST and 11 SDH preserved wtGIST) and 21 tyrosine kinase (TK) mutant GIST. RESULTS: MGMT promoter methylation was detected in 8 cases of SDH-deficient (dSDH) GIST but in none of the 14 SDH preserved wild-type GIST or 21 TK mutant GIST samples analysed. Mean MGMT methylation was significantly higher (p 0.0449) and MGMT expression significantly lower (p<0.0001) in dSDH wtGIST compared with TK mutant or SDH preserved GIST. No correlation was identified between SDHx subunit gene mutations or SDHC epimutation status and mean MGMT methylation levels. CONCLUSION: MGMT promoter hypermethylation occurs exclusively in a subset of dSDH wtGIST. Data from this study support testing of tumour MGMT promoter methylation in patients with dSDH wtGIST to identify those patients who may benefit from most from TMZ therapy.

Cost-effectiveness model of renal cell carcinoma (RCC) surveillance in hereditary leiomyomatosis and renal cell carcinoma (HLRCC).

PURPOSE: To determine the cost-effectiveness of annual renal imaging surveillance (RIS) in hereditary leiomyomatosis and renal cell cancer (HLRCC). HLRCC is associated with a 21% risk to age 70 years of RCC. Presentations with advanced renal cell cancer (RCC) are associated with poor outcomes whereas RIS detects early-stage RCC; however, evidence for the cost-effectiveness of RIS is lacking. METHODS: We developed a decision-analytic model to compare, at different age starting points (11 years, 18 years, 40 years, 60 years), the costs and benefits of lifetime contrast-enhanced renal MRI surveillance (CERMRIS) vs no surveillance in HLRCC. Benefits were measured in life-years gained (LYG), quality-adjusted life years (QALYs) and costs in British Pounds Sterling (GBP). Net monetary benefit (NMB) was calculated using a cost-effectiveness threshold of £20 000/QALY. One-way sensitivity and probabilistic analyses were also performed. RESULTS: In the base-case 11-year age cohort, surveillance was cost-effective (Incremental_NMB=£3522 (95% CI -£2747 to £7652); Incremental_LYG=1.25 (95% CI 0.30 to 1.86); Incremental_QALYs=0.29 (95% CI 0.07 to 0.43)] at an additional mean discounted cost of £2185/patient (95% CI £430 to £4144). Surveillance was also cost-effective in other age cohorts and dominated a no surveillance strategy in the 40 year cohort [Incremental_NMB=£12 655 (95% CIs -£709 to £21 134); Incremental_LYG=1.52 (95% CI 0.30 to 2.26); Incremental_QALYs=0.58 (95% CI 0.12 to 0.87) with a cost saving of £965/patient (95% CI -£4202 to £2652). CONCLUSION: Annual CERMRI in HLRCC is cost-effective across age groups modelled.

UK recommendations for SDHA germline genetic testing and surveillance in clinical practice.

SDHA pathogenic germline variants (PGVs) are identified in up to 10% of patients with paraganglioma and phaeochromocytoma and up to 30% with wild-type gastrointestinal stromal tumours. Most SDHA PGV carriers present with an apparently sporadic tumour, but often the pathogenic variant has been inherited from parent who has the variant, but has not developed any clinical features. Studies of SDHA PGV carriers suggest that lifetime penetrance for SDHA-associated tumours is low, particularly when identified outside the context of a family history. Current recommended surveillance for SDHA PGV carriers follows an intensive protocol. With increasing implementation of tumour and germline large panel and whole-genome sequencing, it is likely more SDHA PGV carriers will be identified in patients with tumours not strongly associated with SDHA, or outside the context of a strong family history. This creates a complex situation about what to recommend in clinical practice considering low penetrance for tumour development, surveillance burden and patient anxiety. An expert SDHA working group was formed to discuss and consider this situation. This paper outlines the recommendations from this working group for testing and management of SDHA PGV carriers in clinical practice.

First step towards a consensus strategy for multi-locus diagnostic testing of imprinting disorders.

BACKGROUND: Imprinting disorders, which affect growth, development, metabolism and neoplasia risk, are caused by genetic or epigenetic changes to genes that are expressed from only one parental allele. Disease may result from changes in coding sequences, copy number changes, uniparental disomy or imprinting defects. Some imprinting disorders are clinically heterogeneous, some are associated with more than one imprinted locus, and some patients have alterations affecting multiple loci. Most imprinting disorders are diagnosed by stepwise analysis of gene dosage and methylation of single loci, but some laboratories assay a panel of loci associated with different imprinting disorders. We looked into the experience of several laboratories using single-locus and/or multi-locus diagnostic testing to explore how different testing strategies affect diagnostic outcomes and whether multi-locus testing has the potential to increase the diagnostic efficiency or reveal unforeseen diagnoses. RESULTS: We collected data from 11 laboratories in seven countries, involving 16,364 individuals and eight imprinting disorders. Among the 4721 individuals tested for the growth restriction disorder Silver-Russell syndrome, 731 had changes on chromosomes 7 and 11 classically associated with the disorder, but 115 had unexpected diagnoses that involved atypical molecular changes, imprinted loci on chromosomes other than 7 or 11 or multi-locus imprinting disorder. In a similar way, the molecular changes detected in Beckwith-Wiedemann syndrome and other imprinting disorders depended on the testing strategies employed by the different laboratories. CONCLUSIONS: Based on our findings, we discuss how multi-locus testing might optimise diagnosis for patients with classical and less familiar clinical imprinting disorders. Additionally, our compiled data reflect the daily life experiences of diagnostic laboratories, with a lower diagnostic yield than in clinically well-characterised cohorts, and illustrate the need for systematising clinical and molecular data.

HIRA loss transforms FH-deficient cells.

Fumarate hydratase (FH) is a mitochondrial enzyme that catalyzes the reversible hydration of fumarate to malate in the tricarboxylic acid (TCA) cycle. Germline mutations of FH lead to hereditary leiomyomatosis and renal cell carcinoma (HLRCC), a cancer syndrome characterized by a highly aggressive form of renal cancer. Although HLRCC tumors metastasize rapidly, FH-deficient mice develop premalignant cysts in the kidneys, rather than carcinomas. How Fh1-deficient cells overcome these tumor-suppressive events during transformation is unknown. Here, we perform a genome-wide CRISPR-Cas9 screen to identify genes that, when ablated, enhance the proliferation of Fh1-deficient cells. We found that the depletion of the histone cell cycle regulator (HIRA) enhances proliferation and invasion of Fh1-deficient cells in vitro and in vivo. Mechanistically, Hira loss activates MYC and its target genes, increasing nucleotide metabolism specifically in Fh1-deficient cells, independent of its histone chaperone activity. These results are instrumental for understanding mechanisms of tumorigenesis in HLRCC and the development of targeted treatments for patients.

Molecular Basis of Beckwith-Wiedemann Syndrome Spectrum with Associated Tumors and Consequences for Clinical Practice.

Beckwith-Wiedemann syndrome (BWS, OMIM 130650) is a congenital imprinting condition with a heterogenous clinical presentation of overgrowth and an increased childhood cancer risk (mainly nephroblastoma, hepatoblastoma or neuroblastoma). Due to the varying clinical presentation encompassing classical, clinical BWS without a molecular diagnosis and BWS-related phenotypes with an 11p15.5 molecular anomaly, the syndromic entity was extended to the Beckwith-Wiedemann spectrum (BWSp). The tumor risk of up to 30% depends on the molecular subtype of BWSp with causative genetic or epigenetic alterations in the chromosomal region 11p15.5. The molecular diagnosis of BWSp can be challenging for several reasons, including the range of causative molecular mechanisms which are frequently mosaic. The molecular basis of tumor formation appears to relate to stalled cellular differentiation in certain organs that predisposes persisting embryonic cells to accumulate additional molecular defects, which then results in a range of embryonal tumors. The molecular subtype of BWSp not only influences the overall risk of neoplasia, but also the likelihood of specific embryonal tumors.

Comparison of methylation episignatures in KMT2B- and KMT2D-related human disorders.

Aim & methods: To investigate peripheral blood methylation episignatures in KMT2B-related dystonia (DYT-KMT2B), the authors undertook genome-wide methylation profiling of ∼2 M CpGs using a next-generation sequencing-based assay and compared the findings with those in controls and patients with KMT2D-related Kabuki syndrome type 1 (KS1). Results: A total of 1812 significantly differentially methylated CpG positions (false discovery rate < 0.05) were detected in DYT-KMT2B samples compared with controls. Multi-dimensional scaling analysis showed that the 10 DYT-KMT2B samples clustered together and separately from 29 controls and 10 with pathogenic variants in KMT2D. The authors found that most differentially methylated CpG positions were specific to one disorder and that all (DYT-KMT2B) and most (Kabuki syndrome type 1) methylation alterations in CpG islands were gain of methylation events. Conclusion: Using sensitive methylation profiling methodology, the authors replicated recent reports of a methylation episignature for DYT-KMT2B. These findings will facilitate the development of episignature-based assays to improve diagnostic accuracy.

The authors compared the DNA methylation patterns in blood from individuals with two rare neurodevelopmental disorders (childhood-onset dystonia [DYT-KMT2B] and Kabuki syndrome type 1) and healthy control samples. These two disorders are associated with pathogenic variants in KMT2B and KMT2D, which encode proteins with related functions but cause distinct inherited disorders. Comparison of the methylation patterns in the two disorders showed that most DNA regions with altered methylation patterns differed between the two disorders and controls. These findings suggest that analyzing DNA methylation patterns could improve diagnostic testing for these disorders and might provide insights into how the clinical features of these disorders are caused.

Frequency of pathogenic germline variants in cancer susceptibility genes in 1336 renal cell carcinoma cases.

Renal cell carcinoma (RCC) occurs in a number of cancer predisposition syndromes, but the genetic architecture of susceptibility to RCC is not well defined. We investigated the frequency of pathogenic and likely pathogenic (P/LP) germline variants in cancer susceptibility genes (CSGs) within a large series of unselected RCC participants. Whole-genome sequencing data on 1336 RCC participants and 5834 controls recruited to the UK 100 000 Genomes Project, a nationwide multicentre study, was analyzed to identify rare P/LP short variants (single nucleotide variants and insertions/deletions ranging from 1 to 50 base pairs) and structural variants in 121 CSGs. Among 1336 RCC participants [mean: 61.3 years (±12 SD), range: 13-88 years; 64% male], 85 participants [6.4%; 95% CI (5.1, 7.8)] had one or more P/LP germline variant in a wider range of CSGs than previously recognized. A further 64 intragenic variants in CSGs previously associated with RCC were classified as a variant of uncertain significance (VUS) (24 'hot VUSs') and were considered to be of potential clinical relevance as further evaluation might results in their reclassification. Most patients with P variants in well-established CSGs known to predispose to renal cell carcinoma (RCC-CSGs) were aged <50 years. Burden test analysis for filtered variants in CSGs demonstrated a significant excess of CHEK2 variants in European RCC participants compared with the healthy European controls (P = 0.0019). Approximately, 6% of the patients with RCC unselected for family history have a germline variant requiring additional follow-up analysis. To improve diagnostic yield, we suggest expanding the panel of RCC-CSGs tested to include CHEK2 and all SDHx subunits and raising the eligibility criteria for age-based testing.