Germline Variants In Genes Research Articles

Clonal Hematopoiesis in Relatives of Adults with Myeloid Neoplasms with Germline Predisposition

Introduction: Approximately 10% of adults with myelodysplastic neoplasms (MDS) or acute myeloid leukemia (AML) carry germline variants in genes predisposing to hematologic neoplasms (HNGP). Clonal hematopoiesis (CH), whether as a pre-malignant event or as compensating mechanism, is a focus of interest in the field of germline predisposition in adults. However, systematic and longitudinal studies on HNGP patients' healthy relatives, critical to understand how CH and germline predisposition interact, remain limited. Our study aimed to analyze the prevalence, dynamics, and impact of CH in relatives of HNPG adult patients who carry the causative germline abnormality. Methods: In 2018, the Catalan public healthcare system established criteria for germline testing when diagnosing a hematological neoplasm. These criteria, updated annually, include early-onset, syndromic phenotype, family history of thrombocytopenia or hematological neoplasms, and the presence of a potential germline variant in the initial HN workout. For each patient, a HNG-driven NGS virtual panel was performed using DNA extracted from hair follicles. Additionally, if the physical examination revealed characteristics related to Telomere Biology Disorders (TBD), telomere length was assessed by flow-FISH. First and second-degree relatives of germline variant carriers were offered predictive testing, using Sanger sequencing on peripheral blood (PB). In cases of TBD without an identified molecular cause, flow-FISH was used to select those at-risk relatives. Family carriers underwent NGS for genes with recurrent acquired mutations in myeloid malignancies in PB at the time of diagnosis and after a three-year follow-up period. Results: From 2019 to 2024, we studied 97 patients who met the criteria for germline testing and identified 21 HNPG index cases (21.6%). HNPG patients were defined by the presence of a pathogenic/likely pathogenic variant in ETV6 (n=3), GATA2 (n=2), DDX41 (n=6), ELANE (n=1), CHEK2 (n=1), RUNX1 (n=1), ATM (n=1), TP53 (n=1) or a TBD diagnosis (n=5). Among these last, in 2 we identified a causative variant (RTEL1 and TINF2) and the remaining 3 reached the diagnosis due to a telomere length <1%. These 21 index cases were diagnosed as follows: 11 MDS with multilineage dysplasia, 2 MDS with ring sideroblasts, 2 hypoplastic MDS, 2 MDS with excess blasts, and 4 AML. The median age at diagnosis was 48 years (range: 19-76). Thirty-four out of 60 studied relatives (56.6%) carried the germline variant or telomere abnormality. Twenty-seven (79.4%) were studied using a somatic NGS panel in PB. Among these cases, CH was detected at first time-point in 7 of the carrier relatives (26%) across 4 different families. One family was notable, as 3 of the studied healthy ETV6 carriers had developed CH: a 71-year-old male with normal CBCs and variants in TP53 (VAF 4%; c.394A>G) and SRSF2 (VAF 1.5%;c.284C>T), a 76-year-old female with mild thrombocytopenia and variants in JAK2 (VAF 16%;c.1849G>T), NFE (VAF 10%;c.768_769dup), DNMT3A (VAF 3.2%;c.2693T>A) and PTPN11 (VAF 5%;c.785T>A), and a 48-year-old female with normal CBCs and one variant in CBL (VAF 2.3%; c.1150T>C). Two relatives from one TBD family presented CH: a 57-year-old male with no CBC alteration and a variant in PPMD1 (VAF 4.8%; c.1451T>A) and a 45-year-old female with normal CBC and a variant in U2AF1 (VAF 4.7%; c.101C>T). Another TBD family carrier had a variant in DNMT3A (VAF 1.7%; c.1228G>A). One family carrier from a DDX41 family, a 79-year-old male with normal CBCs, presented a variant in DNMT3A (VAF 3.1%; c.1015-3C>G). We performed a 3 year follow-up NGS in 8 relatives, of which 5 (62%) showed clonal evolution, evidenced by an increase in VAF (n=2) or the acquisition of new variants (n=3). In a second ETV6 family, a heathy carrier relative acquired a KRAS variant VAF (1.3%, c.34G>C) not detected at the initial study. No carrier relatives developed a hematologic neoplasm during the follow-up period. Conclusion: In one of the largest series studied to date of HNGP relatives, we found a 26% of CH development in carriers of the causative predisposition abnormality. That percentage is approximately 10x higher than in the age-adjusted population group, prompting us to extend the longitudinal follow-up of these families. We describe 2 ETV6 families, in which CH was defined in 4 healthy relatives by genes associated with myeloproliferative diseases, a molecular pattern to be confirmed.

Genotype‐phenotype associations in individuals with Diamond Blackfan anaemia

AbstractIntroductionDiamond Blackfan anaemia (DBA) is a rare disorder characterized by failure of red blood cell production, congenital abnormalities and cancer predisposition, primarily caused by pathogenic germline variants in genes encoding ribosomal proteins.MethodsWe conducted a genotype‐phenotype and outcome study of 121 patients with DBA spanning the 20‐year history of the National Cancer Institute's Inherited Bone Marrow Failure Syndromes study. Patient phenotypes were compared by large versus small ribosomal protein genes, across genes with >5 cases (RPS19, RPS29, RPS26 and RPL35A) and by type of pathogenic variants (hypomorphic versus null, large deletions versus others).ResultsA pathogenic germline variant was identified in 71% of patients (n = 86/121) from 54 families. After adjusting for multiple testing, we found that patients with RPS29 variants were least likely to need treatment for anaemia while those with large ribosomal protein subunit variants had a higher proportion of intellectual disability and gastrointestinal abnormalities compared with small ribosomal protein subunit variants (p < 3.5 × 10−4). There were no statistically significant differences in overall survival or cancer incidence among patients with large or small ribosomal subunit genes.ConclusionThis detailed genotype‐phenotype study of DBA improves our understanding of the role of germline genetics in the clinical manifestations that may help guide the management of people with DBA.

Atypical hemolytic uremic syndrome during induction chemotherapy in neuroblastoma, a rare phenomenon or common congenital predisposition?

Atypical hemolytic uremic syndrome (aHUS) is a complement-mediated thrombotic microangiopathy sometimes associated with germline variants in genes of the complement system. Clinical findings of microangiopathic hemolytic anemia, thrombocytopenia, and acute kidney injury arise due to aberrant complement protein activation in the circulation. A 13-month-old boy with metastatic neuroblastoma (NB) developed aHUS during his first cycle of induction chemotherapy with germline testing revealing a complement factor H(CFH) gene mutation, currently classified as a variant of uncertain significance (VUS). Now he is in disease remission after successful complement blockade therapy, thus highlighting a unique presentation of aHUS in a patient with newly diagnosed NB.

Understanding familial risk of pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is a deadly disease that is the result of an accumulation of sequential genetic alterations. These genetic alterations can either be inherited, such as pathogenic germline variants that are associated with an increased risk of cancer, or acquired, such as somatic mutations that occur during the lifetime of an individual. Understanding the genetic basis of inherited risk of PDAC is essential to advancing patient care and outcomes through improved clinical surveillance, early detection initiatives, and targeted therapies. In this review we discuss factors associated with an increased risk of PDAC, the prevalence of genetic variants associated with an increased risk in patients with PDAC, estimates of PDAC risk in carriers of pathogenic germline variants in genes associated with an increased risk of PDAC. The role of common variants in pancreatic cancer risk will also be discussed.

Abstract NG01: Germline-mediated immunoediting sculpts breast cancer subtypes and metastatic proclivity

Abstract NG01: Germline-mediated immunoediting sculpts breast cancer subtypes and metastatic proclivity

Polygenic Modifiers of Expressivity in Telomere Biology Disorders

Telomere biology disorders (TBDs) are caused by pathogenic germline variants in genes involved in telomere maintenance resulting in very short telomeres for age and high risks of bone marrow failure, certain cancers, as well as pulmonary and liver disease. Interestingly, some pathogenic TBD mutations have variable expressivity and incomplete penetrance, even within families. Variant carriers can present with early manifestations such as the mucocutaneous triad and bone marrow failure or may not manifest until later in life with adult-onset idiopathic pulmonary fibrosis or liver disease, or have no associated clinical phenotype. The underlying causes of this heterogeneity are poorly understood. We hypothesized that common genetic variants contributing to inter-individual variation in telomere length in the general population may combine with the effects of monogenic TBD-causing variants to underlie variable clinical manifestations in TBDs. To examine this, we developed polygenic scores (PGS) to predict telomere length using genome-wide SNPs from the UK Biobank. We then applied the most predictive polygenic score to the out-of-sample All of Us cohort, as well as the National Cancer Institute's longitudinal cohort of individuals with inherited bone marrow failure syndromes, including those with TBDs (ClinicalTrials.gov Identifier: NCT00027274). If monogenic mutations and polygenic predisposition to short telomeres both contribute to the clinical severity of TBDs and likelihood of early-onset manifestations, the distribution of genetically predicted telomere length in this ascertained cohort would be shifted towards shorter telomeres compared with the population average. Consistent with this, individuals with TBDs enriched for early-onset bone marrow failure phenotypes (n = 92) have a median polygenic score 0.32 standard deviations (SDs) shorter than the UK Biobank (p=0.002), and 0.31 SDs shorter than the external All of Us cohort (p=0.003) ( Figure). Individuals with TBDs also have shorter genetically predicted telomeres compared to individuals with Fanconi anemia (n = 45) and Diamond Blackfan anemia (n = 49) (p=0.008), indicating that this phenomenon is specific to TBDs. We binned the PGS into quintiles and show that the lowest quintile is associated with an odds ratio of TBD of 1.89 [1.19,2.86], while being in the highest quintile is associated with an odds ratio of 0.54 [0.29,1.02]. Interestingly, TBD cases with no identified mutation have a median PGS 0.1 SDs towards shorter telomeres than patients with a known mutation. Collectively, these results indicate that common, genome-wide determinants of telomere length are significant modifiers of the early-onset manifestations of TBDs. We hypothesized that the converse should also be true: TBD causal variants should be present in adult population cohorts, and adult individuals with a pathogenic variant who avoided the severe childhood-onset manifestations of TBDs, should not have polygenically predicted short telomere length. To test this hypothesis, we filtered the UK Biobank for TBD causal variants annotated in ClinVar, and identified 71 putative pathogenic cases. Importantly, among these cases there were no individuals with bone marrow failure, but these individuals did have shorter telomeres compared with the UK Biobank. Interestingly, these 71 putative cases had a population-normal telomere length PGS, and significantly longer PGS predicted telomere length than the NCI cohort (p=0.025). We also find that being classified as among these pathogenic cases is associated with an odds ratio for developing pulmonary fibrosis of 10.79 [4.34,26.8]. In summary, despite normal PGS and absence of bone marrow failure, these putative cases have short telomeres and increased risk of adult manifestations of TBDs, consistent with the idea that the pathogenic mutations and common genetic variation affecting telomere length combine to impact expressivity. Collectively, these findings indicate that polygenic effects on telomere length can modify the impacts of monogenic mutations in TBD-associated genes. These results have implications not only for clinical risk assessment in patients with TBDs, but also for our general understanding and interpretation of variable penetrance/expressivity in presumed monogenic disorders.

A common germline variant in CYP11B1 is associated with adverse clinical outcome of treatment with abiraterone or enzalutamide

Extragonadal androgens play a pivotal role in prostate cancer disease progression on androgen receptor signaling inhibitors (ARSi), including abiraterone and enzalutamide. We aimed to investigate if germline variants in genes involved in extragonadal androgen synthesis contribute to resistance to ARSi and may predict clinical outcomes on ARSi. We included ARSi naive metastatic prostate cancer patients treated with abiraterone or enzalutamide and determined 18 germline variants in six genes involved in extragonadal androgen synthesis. Variants were tested in univariate and multivariable analysis for the relation with overall survival (OS) and time to progression (TTP) by Cox regression, and PSA response by logistic regression. A total of 275 patients were included. From the investigated genes CYP17A1, HSD3B1, CYP11B1, AKR1C3, SRD5A1 and SRD5A2, only rs4736349 in CYP11B1 in homozygous form (TT), present in 54 patients (20%), was related with a significantly worse OS (HR = 1.71, 95% CI 1.09 – 2.68, p = 0.019) and TTP (HR = 1.50, 95% CI 1.08 – 2.09, p = 0.016), and was related with a significantly less frequent PSA response (OR = 0.48, 95% CI 0.24 – 0.96, p = 0.038) on abiraterone or enzalutamide in a multivariable analysis. The frequent germline variant rs4736349 in CYP11B1 is, as homozygote, an independent negative prognostic factor for treatment with abiraterone or enzalutamide in ARSi naive metastatic prostate cancer patients. Our findings warrant prospective investigation of this potentially important predictive biomarker.

Clinical Consequences of Rare Germline Variation in Genes Associated with Coagulation across 451,958 Individuals

Introduction Despite deep biological insights over the preceding decades, our ability to assess for inherited hypercoagulable states remains limited. In light of these challenges and the increasing relevance of precision medicine and next-generation sequencing (NGS) to clinical practice, the identification of rare germline variants in coagulation-focused NGS panels is increasingly becoming accepted practice. However, the discovery of variants of unknown significance (VUS) remains a common clinical conundrum in molecular testing, particularly because evidence for the thrombosis risk associated with many genes derives from small, uncontrolled studies, case reports, and pedigree analysis. Large-scale evaluation of the clinical consequences associated with loss of function in individual coagulation-related genes would contribute to the clinical assessment of patients presenting with heterozygous VUS in poorly characterized genes. Objectives Determine the risk of four thrombosis phenotypes associated with rare, germline loss-of-function variants in a major clinical coagulation-focused NGS panel. Methods The UK Biobank (UKBB) contains paired clinical and whole exome sequencing data for over 450,000 participants. We identified rare (minor allele frequency <0.1%) germline variants that were predicted in silico to alter protein activity (functional impact score ≥0.8) within the 54 evaluable genes of the Yale NGS coagulation panel. For each gene, parallel multivariable Firth's logistic regression models were generated to assess the association between rare qualifying variants and four thrombotic phenotypes as well as coagulopathic bleeding in a gene-level collapsing analysis (N=270 independent models). We adjusted for age, sex, ancestry, and sequencing batch. Phenotypes of interest included venous thromboembolism (VTE), myocardial infarction (MI), stroke (CVA), peripheral artery disease (PAD), and coagulopathic bleeding. The Bonferroni corrected p-value for each phenotype was 0.001. We validated functional impact score predictions in seven secreted proteins measured in a subset of 47,923 UKBB participants who underwent plasma proteomic assessment (Olink Explore 1536 panel). Results A total of 451,958 individuals in the UKBB with whole exome sequencing data were evaluated. Among the 54 genes in the Yale NGS coagulation panel, there were 18,975 qualifying variants (98.4% heterozygous). Function-altering variants in 4 of 54 genes were associated with a statistically significant increased risk of thrombosis after adjusting for multiple comparisons: PROC (OR = 5.7, 95% CI: 3.2 - 9.6, p = 1.6x10 -7), SERPINC1 (OR = 5.0, 95% CI: 2.7 - 8.7, p = 4.0x10 -6), PROS1 (OR = 3.2, 95% CI: 2.0 - 4.8, p = 6.0x10 -6), and STAB2 (OR = 1.4, 95% CI: 1.2 - 1.7, p = 5.3x10 -4) (Figure 1). Nominal significance was achieved for 7 genes in the VTE phenotype, 1 gene in CVA, 3 genes in MI, 5 genes in PAD, and 0 genes in coagulopathic bleeding. Variants in a total of 37 genes did not appear to be significantly associated with any of the evaluated phenotypes. To assess the reliability of variant functional impact scores derived in silico, plasma levels for 13 of the 53 coagulation genes were analyzed. Of these, 8 were identified as secreted proteins amenable to assessment by plasma levels, including ADAMTS13 ( ADAMTS13), factor IX ( F9), factor VII ( F7), tissue plasminogen activator ( PLAT), urokinase ( PLAU), protein C ( PROC), plasminogen activator inhibitor 1 ( SERPINE1), and von Willebrand factor ( VWF). For these proteins, average plasma levels consistently demonstrated a negative correlation with functional impact score (Pearson r = -0.98, P=0.0009), indicating that our in silico assessments in this context are broadly reliable (Figure 2). Conclusion Within the commonly encountered clinical context of heterozygous loss of function, most genes contained in coagulation-focused NGS panels likely have fairly weak effects on disease risk that are limited to venous thrombosis. However, our findings do not rule out the possibility of more profound effects from homozygous or compound heterozygous loss of function in these same genes. By defining the clinical consequence of heterozygous loss of function in specific coagulation-related genes across a large population, our findings may help inform the interpretation of molecular testing results in which patients are found to have a VUS.

Molecular and cellular evidence for the impact of a hypertrophic cardiomyopathy-associated RAF1 variant on the structure and function of contractile machinery in bioartificial cardiac tissues

Noonan syndrome (NS), the most common among RASopathies, is caused by germline variants in genes encoding components of the RAS-MAPK pathway. Distinct variants, including the recurrent Ser257Leu substitution in RAF1, are associated with severe hypertrophic cardiomyopathy (HCM). Here, we investigated the elusive mechanistic link between NS-associated RAF1S257L and HCM using three-dimensional cardiac bodies and bioartificial cardiac tissues generated from patient-derived induced pluripotent stem cells (iPSCs) harboring the pathogenic RAF1 c.770 C > T missense change. We characterize the molecular, structural, and functional consequences of aberrant RAF1–associated signaling on the cardiac models. Ultrastructural assessment of the sarcomere revealed a shortening of the I-bands along the Z disc area in both iPSC-derived RAF1S257L cardiomyocytes and myocardial tissue biopsies. The aforementioned changes correlated with the isoform shift of titin from a longer (N2BA) to a shorter isoform (N2B) that also affected the active force generation and contractile tensions. The genotype-phenotype correlation was confirmed using cardiomyocyte progeny of an isogenic gene-corrected RAF1S257L-iPSC line and was mainly reversed by MEK inhibition. Collectively, our findings uncovered a direct link between a RASopathy gene variant and the abnormal sarcomere structure resulting in a cardiac dysfunction that remarkably recapitulates the human disease.

Detection of germline variants with pathogenic potential in 48 patients with familial colorectal cancer by using whole exome sequencing

BackgroundHereditary genetic mutations causing predisposition to colorectal cancer are accountable for approximately 30% of all colorectal cancer cases. However, only a small fraction of these are high penetrant mutations occurring in DNA mismatch repair genes, causing one of several types of familial colorectal cancer (CRC) syndromes. Most of the mutations are low-penetrant variants, contributing to an increased risk of familial colorectal cancer, and they are often found in additional genes and pathways not previously associated with CRC. The aim of this study was to identify such variants, both high-penetrant and low-penetrant ones.MethodsWe performed whole exome sequencing on constitutional DNA extracted from blood of 48 patients suspected of familial colorectal cancer and used multiple in silico prediction tools and available literature-based evidence to detect and investigate genetic variants.ResultsWe identified several causative and some potentially causative germline variants in genes known for their association with colorectal cancer. In addition, we identified several variants in genes not typically included in relevant gene panels for colorectal cancer, including CFTR, PABPC1 and TYRO3, which may be associated with an increased risk for cancer.ConclusionsIdentification of variants in additional genes that potentially can be associated with familial colorectal cancer indicates a larger genetic spectrum of this disease, not limited only to mismatch repair genes. Usage of multiple in silico tools based on different methods and combined through a consensus approach increases the sensitivity of predictions and narrows down a large list of variants to the ones that are most likely to be significant.

Frequency of pathogenic germline variants in cancer susceptibility genes in 841 renal cell carcinoma cases.

e16551 Background: Hereditary Renal cell carcinoma (RCC) syndrome accounts for approximately 5-10% of all kidney cancers. However, the genetic architecture of susceptibility to RCC is not well defined. In this study, we investigated the prevalence and spectrum of pathogenic/likely pathogenic germline variants of cancer susceptibility genes in unselected RCC patients in the real-world Chinese population. Methods: We retrospectively profiled the germline mutations of 841 unselected RCC patients using a next-generation sequencing panel consisting of 808 cancer-related genes. Interpretation of sequence variations was classified according to the American College of Medical Genetics and Genomics guidelines. Genomic alterations include single nucleotide variations (SNV), short insertions/deletions (INDEL) and copy number variations (CNV). Results: Within 841 unselected RCC patients included in this study, 132 germline pathogenic/likely pathogenic (P/LP) variants were identified in 125 patients (14.9%), of which 114 (13.6%) were mutations of SNV/INDEL and 11 (1.3%) were CNV. Overall, 46 patients (5.5%) carried mutations of hereditary RCC syndrome-associated genes such as VHL (13, 1.5%), FH (13, 1.5%) and FLCN (8, 1.0%). Interestingly, variants in other cancer-predisposition genes not traditionally associated with RCC occurred in 9.4% of the unselected series, including 55 (6.5%) patients carried mutations in DNA damage (DDR)-related genes (e.g., MUTYH, CHEK2 and BRCA1/2) and the other 24 (2.9%) had germline mutations of non-DDR-related genes (e.g., SPINK1 and APC). The median age of patients with mutations of RCC syndrome-associated genes was 38 (range 19-77), while the median age of patients with mutations of other cancer-predisposition genes was 52(range 22-82). Furthermore, in patients older than 46 years, the prevalence of P/LP variants in RCC-associated and non–RCC-associated genes were 1.9% and 6.7%, respectively. Notably, among 125 patients with identified germline mutations, nearly half (56, 44.8%) did not meet the recommended criteria for genetic testing. Conclusions: Our findings confirm a high prevalence of pathogenic germline variants in genes associated with both hereditary RCC and other cancer predispositions in the unselected Chinese RCC population. Current guidelines recommendation for genetic testing seemed not sensitive enough to identify patients with hereditary RCC susceptibility. It is rational to promote genetic testing and expand the range of genes in RCC population with advanced age.

EDITORIAL COMMENT

Paragangliomas and pheochromocytomas (PGL/PCC) are rare tumors that are derived from neural crest chromaffin cells either from neuroendocrine tissues distributed along the spine or from within the adrenal gland respectively. Approximately 40% of cases are associated with pathogenic germline variants in genes associated to cancer susceptibility syndromes, such as MAX, NF1, RET, SDHA, and SDHAF2, including several gene that associated with inherited kidney cancer, such as VHL, SDHB, SDHC, SDHD, and TMEM127. 1 Shuch B Ricketts CJ Metwalli AR Pacak K Linehan WM. The genetic basis of pheochromocytoma and paraganglioma: implications for management. Urology. 2014; 83: 1225-1232 Google Scholar ,2 Dahia PL. Pheochromocytoma and paraganglioma pathogenesis: learning from genetic heterogeneity. Nat Rev Cancer. 2014; 14: 108-119 Google Scholar A recent addition to this list is germline alteration of the FH gene, known to cause hereditary leiomyomatosis and renal cell cancer (HLRCC). 3 Richter S Gieldon L Pang Y et al. Metabolome-guided genomics to identify pathogenic variants in isocitrate dehydrogenase, fumarate hydratase, and succinate dehydrogenase genes in pheochromocytoma and paraganglioma. Genet Med. 2019; 21: 705-717 Google Scholar , 4 Clark GR Sciacovelli M Gaude E et al. Germline FH mutations presenting with pheochromocytoma. J Clin Endocrinol Metab. 2014; 99: E2046-E2050 Google Scholar , 5 Muller M Ferlicot S Guillaud-Bataille M et al. Reassessing the clinical spectrum associated with hereditary leiomyomatosis and renal cell carcinoma syndrome in French FH mutation carriers. Clin Genet. 2017; 92: 606-615 Google Scholar

An Association Study of Germline Variants in Bladder Cancer-Related Genes with the Prognosis of Non-Muscle Invasive Bladder Cancer.

Various germline genetic variants are associated with the prognosis of non-muscle invasive bladder cancer (NMIBC). Germline variants in genes frequently somatically mutated in bladder cancer have not been studied thoroughly in relation to risk of recurrence or progression in NMIBC. To identify germline DNA variants in bladder carcinogenesis-related genes associated with recurrence or progression in NMIBC. We analysed associations between single-nucleotide polymorphisms (SNPs) and NMIBC recurrence and progression using data from the Nijmegen Bladder Cancer Study (NBCS, 1,443 patients). We included 5,053 SNPs within 46 genes known to have mutation, overexpression or amplification in bladder cancer. We included all recurrences in the statistical analysis and performed both single variant analysis and gene-based analysis. SNPs and genes that showed significant or suggestive association (false discovery rate P value < 20%) were followed-up in independent cohorts for replication analysis, through eQTL analysis and tests for association of tumour expression levels with NMIBC recurrence and progression. Single variant analysis showed no statistically significant associations with recurrence or progression. In gene-based analysis, the aggregate effect of the 25 SNPs in the Cyclin D1 gene (CCND1) was statistically significantly associated with NMIBC recurrence (Punadj = 0.001, PFDR = 0.046), but not with progression (Punadj = 0.17, PFDR = 0.54). Validation analysis in independent cohorts did not confirm the association of CCND1 with NMIBC recurrence. We could not identify reproducible associations between common germline variants in bladder carcinogenesis-related genes and NMIBC recurrence or progression.

Circulating MicroRNAs Identify Early Phenotypic Changes in Sarcomeric Hypertrophic Cardiomyopathy.

Hypertrophic cardiomyopathy (HCM) is the most common genetic cardiomyopathy. Pathogenic germline variation in genes encoding the sarcomere is the predominant cause of disease. However diagnostic features, including unexplained left ventricular hypertrophy, typically do not develop until late adolescence or after. The early stages of disease pathogenesis and the mechanisms underlying the transition to a clinically overt phenotype are not well understood. In this study, we investigated if circulating microRNAs (miRNAs) could stratify disease stage in sarcomeric HCM. We performed arrays for 381 miRNAs using serum from HCM sarcomere variant carriers with and without a diagnosis of HCM and healthy controls. To identify differentially expressed circulating miRNAs between groups, multiple approaches were used including random forest, Wilcoxon rank sum test, and logistic regression. The abundance of all miRNAs was normalized to miRNA-320. Of 57 sarcomere variant carriers, 25 had clinical HCM and 32 had subclinical HCM with normal left ventricular wall thickness (21 with early phenotypic manifestations and 11 with no discernible phenotypic manifestations). Circulating miRNA profile differentiated healthy controls from sarcomere variant carriers with subclinical and clinical disease. Additionally, circulating miRNAs differentiated clinical HCM from subclinical HCM without early phenotypic changes; and subclinical HCM with and without early phenotypic changes. Circulating miRNA profiles did not differentiate clinical HCM from subclinical HCM with early phenotypic changes, suggesting biologic similarity between these groups. Circulating miRNAs may augment the clinical stratification of HCM and improve understanding of the transition from health to disease in sarcomere gene variant carriers.

Intravesical BCG in patients with non-muscle invasive bladder cancer induces trained immunity and decreases respiratory infections

BackgroundBCG is recommended as intravesical immunotherapy to reduce the risk of tumor recurrence in patients with non-muscle invasive bladder cancer (NMIBC). Currently, it is unknown whether intravesical BCG application induces...

P071: Germline variants in genes associated with parkinsonism in cancer patients: A case report

P071: Germline variants in genes associated with parkinsonism in cancer patients: A case report

Somatic mosaicism in STAG2-associated cohesinopathies: Expansion of the genotypic and phenotypic spectrum.

STAG2 is a component of the large, evolutionarily highly conserved cohesin complex, which has been linked to various cellular processes like genome organization, DNA replication, gene expression, heterochromatin formation, sister chromatid cohesion, and DNA repair. A wide spectrum of germline variants in genes encoding subunits or regulators of the cohesin complex have previously been identified to cause distinct but phenotypically overlapping multisystem developmental disorders belonging to the group of cohesinopathies. Pathogenic variants in STAG2 have rarely been implicated in an X-linked cohesinopathy associated with undergrowth, developmental delay, and dysmorphic features. Here, we describe for the first time a mosaic STAG2 variant in an individual with developmental delay, microcephaly, and hemihypotrophy of the right side. We characterized the grade of mosaicism by deep sequencing analysis on DNA extracted from EDTA blood, urine and buccal swabs. Furthermore, we report an additional female with a novel de novo splice variant in STAG2. Interestingly, both individuals show supernumerary nipples, a feature that has not been reported associated to STAG2 before. Remarkably, additional analysis of STAG2 transcripts in both individuals showed only wildtype transcripts, even after blockage of nonsense-mediated decay using puromycin in blood lymphocytes. As the phenotype of STAG2-associated cohesinopathies is dominated by global developmental delay, severe microcephaly, and brain abnormalities, we investigated the expression of STAG2 and other related components of the cohesin complex during Bioengineered Neuronal Organoids (BENOs) generation by RNA sequencing. Interestingly, we observed a prominent expression of STAG2, especially between culture days 0 and 15, indicating an essential function of STAG2 in early brain development. In summary, we expand the genotypic and phenotypic spectrum of STAG2-associated cohesinopathies and show that BENOs represent a promising model to gain further insights into the critical role of STAG2 in the complex process of nervous system development.

Avascular Necrosis and Minimal Trauma Bone Fractures in Patients with Dyskeratosis Congenita

Introduction: Dyskeratosis congenita (DC) and related telomere biology disorders (TBD) result from germline variants in genes required for telomere maintenance. DC is traditionally characterized by a triad of nail dysplasia, oral leukoplakia, and reticular skin pigmentation. Significant complications include bone marrow failure (BMF), pulmonary fibrosis, liver disease, and hematologic and solid organ malignancies. Previous reports described avascular necrosis (AVN) of the femoral head or humerus in DC/TBD. However, these findings were limited to a few individual case reports. Herein, we present a study of AVN and minimal trauma fractures (MTF) in a large cohort of patients with DC/TBD. Methods: We reviewed medical records of all patients with a confirmed diagnosis of DC/TBD enrolled in the IRB-approved Inherited Bone Marrow Failure Syndromes study (NCT00027274) between 2002 - 2019 for the history of AVN and/or MTF. Data were compiled on DC/TBD characteristics, treatments, as well as body mass index (BMI), lipid profile, bone mineral density (BMD), and 25-hydroxyvitamin D (25[OH]D) levels. We used stratified random sampling to match DC/TBD patients with AVN and/or MTF by age, gender, inheritance pattern and genotype with a control group of DC/TBD patients with no AVN/MTF. Distributional variance was assessed for all t-tests and two-sided p values <0.05 were considered significant. Statistical analysis was carried out using RStudio (Ver 1.4.1106). Results: Forty-two of 233 patients (18%) with DC/TBDs experienced at least one AVN and/or MTF at a median age of 21.9 years (range 4 to 58.7); 15/42 were females (Table 1). Twenty-three patients had AVN alone, 6 had MTF alone, and 13 developed both AVN and MTF (Figure 1). The median age at first AVN was 24.1 years (range 6.3 - 58.7) and at first MTF 13.6 years (range 4 - 44.3). AVN was more common in patients with autosomal recessive/X-linked recessive (AR/XLR)/TINF2 DC (n=20, 21.3%) and developed at a younger age (median 18.8 years, range 6.3 - 47.8) than in patients with autosomal dominant (AD) disease (12.5%; median age 29.9 years, range 11.1 - 58.7); p=0.013. MTF occurred almost exclusively in patients with AR/XLR/TINF2 DC (n=17, 19%) compared with AD DC/TBD (n=1, .9%); p<.0001. There were 68 AVN events in 36 patients (median 2 events/patient, range 1 - 5). The most frequent sites were femoral heads (33/36) followed by humeral heads (5/36) and knees (3/36); 22/33 had bilateral femoral head AVN (Figure 1). Twenty-two of 26 patients with severe BMF received androgen/steroids within 5 years of AVN and 8 received hematopoietic cell transplantation (HCT). Compared with the matched control group, significantly more DC patients with AVN had severe BMF and received androgen/corticosteroids (p<0.02). Importantly, 12/36 DC/TBD patients (33%) developed AVN without prior HCT, corticosteroid, or androgen treatment. 24/36 patients received treatment for AVN, including total hip arthroplasty (n=15) and core decompression (n=5) or other medical intervention (n=4). Median time from diagnosis with AVN to treatment was 0.81 years (range 0.03 - 2.7). We identified 38 MTF events in 19 DC patients; the most frequent fracture site was the femoral diaphysis (11 events in 8 patients) (Figure 1). Fractures occurred spontaneously or during routine activities such as entering a vehicle, running in the playground, or low-impact dancing as per patient reports. Seventeen patients (89%) with MTF had severe BMF, 12 (63%) had received HCT (7 had taken androgens/steroids). Compared with matched controls, significantly more DC patients with MTF had received HCT (p=0.02). Other factors such as BMD, elevated cholesterol, overweight/obesity, and 25[OH]D levels were not significantly associated with AVN or MTF in our cohort. Conclusions: AVN and MTF are highly prevalent in patients with DC/TBDs. AVN developed across the entire DC/TBD spectrum but was significantly associated with severe BMF and AR/XLR/TINF2 DC. MTFs occurred almost exclusively in young patients with severe disease indicated by AR/XLR/TINF2 mode of inheritance. Improved screening and effective clinical management of symptoms for patients at higher risk of developing AVN/MTF is needed. Furthermore, these findings signal the need for future research exploring the mechanism underlying AVN and MTF in patients with DC/TBDs. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

LBMON174 Germline Molecular Genetics Of Pituitary Adenomas

Abstract Introduction Pituitary adenomas (PA) are common. Considered to be mostly sporadic tumors for a long time, it has become clear over the last decade that a significant proportion of PA carry germline and/or somatic mutations. This study investigated the germline genetics of a large series of patients (pts) with PA using whole-exome sequencing (WES) and found a high rate of germline mutations in a number of genes including genes that were only known to be somatically mutated in PA. Methods After obtaining ethical approval and informed consent, we randomly included 128 pts (65 Females and 63 males, median age 33 years, IQ range 25-44) with PA (34 microadenomas and 94 macroadenomas). The diagnosis includes acromegaly in 16 pts, Cushing disease in 10 pts, prolactinomas in 50 pts, non-functioning PA in 37 pts, TSHoma in 1 pt, and gonadotrophinoma in 14 pts. None of those pts had a family history of PA or known syndromes. We isolated DNA from peripheral blood and performed WES using Illumina platform (Illumina NovaSeq 6000). For bioinformatics analysis, we removed all intronic and ncRNA variants and focused on exonic, exonic-splicing and splicing variants. We applied strict quality filters within the variant calling pipeline to exclude artifacts arising from the DNA sequencing and subsequent steps. We also removed all synonymous variants and variants marked as benign or likely benign by either Clinvar or ACMG Intervar databases. We followed the ACMG criteria in assigning pathogenicity levels to the variants. We focused on genes that were reported before to be involved in the germline, mosaic or somatic genetics of PA (AIP, GNAS, MEN1, DICER1, CDKN1B, PIK3CA, NF1, USP48 and USP8, GPR101, PRKARIA, PRKACB, MAX, CABLES1, SF3B1 and the SDHx group). Results We found 35 different variants in the PA-associated genes in 60 pts (46.9%). Some of these variants occurred in more than one patient with a total number of occurrences of 80. Most of these variants are mono-allelic (96.25%). These variants were found in the following genes: GNAS (10 variants in 39 pts), AIP (one variant in 1 patient), CDKN1B (2 variants in 2 pts), DICER1 (1 variant in 1 patient), MEN1 (2 variants in 3 pts), NF1 (3 variants in 3 pts), PIK3CA (1 variant in 1 patient), SDHA (3 variants in 3 pts), SDHAF2 (2 variants in 5 pts), SDHB (4 variants in 3 pts), SDHD (3 variants in 3 pts), USP48 (2 variants in 8 pts) and USP8 (1 variant in 1 patient). These variants included missense, non-sense, and small indels. Conclusions Germline variants in genes previously reported to be associated with PA are common in apparently sporadic PA occurring in about 47% of cases. For the first time, we report a significant occurrence of germline variants in genes that were previously known to be only somatically mutated (e. g., USP8, USP48, GNAS). Presentation: Monday, June 13, 2022 12:30 p.m. - 2:30 p.m.

Genetic screening of hypertensive patients with aldosterone hypersecretion under conditions of stress.

Although ACTH is considered a secondary regulator of aldosterone production, patients with apparent essential hypertension have been treated with mineralocorticoid receptor antagonists (MRAs). In this study, we aimed to identify potentially damaging variants that might be implicated in the phenotype of a well-characterized cohort of 21 hypertensive patients without PA but with stress-induced aldosterone hypersecretion. The patients' blood pressure was normalized though MRA administration. Genetic screening was performed through whole-exome sequencing (WES), and variants in PA-associated or in ion-channels of aldosterone-regulating genes were prioritized. Variants with population frequency < 0.01, predicted to alter protein structure and classified as likely pathogenic by in silico tools, were retained. Qualifying variants were identified in nine of the 21 patients screened. Seven patients were carriers of six potentially damaging variants in six genes associated with PA (KCNK9, KCNK5, ATP13A3, SLC26A2, CACNA1H, and CACNA1D). A novel variant in the KCNK9 gene (p.V221M) is reported. Our analysis revealed two variants in two novel susceptibility genes for aldosterone hypersecretion, namely, KCNK16 (p.P255H) and CACNA2D3 (p.V557I). WES revealed potentially damaging germline variants in genes participating in aldosterone synthesis/regulating pathways in 9/21 patients of our cohort. The variants identified might play a role in aldosterone hypersecretion under conditions of stress. The potential pathogenicity of these variants should be examined in future functional studies.