Rybp (DEDAF) is a member of the Rybp/Yaf2 protein family and has been shown to encode pro-apoptotic functions and to be essential for mouse embryogenesis. The related Yaf2 protein has not been ...studied extensively at the cellular or organismal levels. Here we describe zebrafish yaf2 (zyaf2) and show that it is widely expressed during early embryogenesis, with subsequent enrichment of transcripts in the anterior head region. Depletion of zYaf2 during embryogenesis using specific morpholinos activates a wide-spread program of apoptosis and causes developmental arrest before the one somite stage. Partial depletion of Yaf2, achieved by injecting lower dosages of morpholino, circumvents the early arrest but leads to CNS degeneration associated with excessive apoptosis. These phenotypes can be rescued by co-injection of human YAF2 mRNA with the morpholinos or by treatment with a pan-caspase inhibitor or a caspase 8-specific inhibitor. Finally, the observed activation of caspase 8 in the morphants is in accord with the ability of Yaf2 to inhibit caspase 8-mediated apoptosis in cultured cells. Our findings implicate Yaf2 as a survival factor during early zebrafish development and organogenesis. This may suggest that Yaf2 and Rybp can encode opposing functions in the regulation of apoptosis.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Introduction: Severe aplastic anemia (SAA) is primarily an immune-mediated disease, but a subset of patients have a germline genetic predisposition. Germline genetic testing for patients with SAA is ...recommended to guide patient treatment, including adjustment for hematopoietic cell transplantation (HCT) modalities. Hemophagocytic lymphohistiocytosis (HLH) is a life threatening hyperinflammatory condition often associated with cytopenias. HLH etiology is genetically variable with X-linked or autosomal recessive (XLR or AR) modes of inheritance. The role of monoallelic pathogenic/likely pathogenic (P/LP) variants in recessive HLH-associated genes in SAA etiology or outcomes is unclear. In this study, we determined the frequency of monoallelic HLH-associated gene variants in a large cohort of patients with acquired SAA and evaluated their association with HCT outcomes. Methods: The Transplant Outcomes in Aplastic Anemia (TOAA) project is a collaboration between the NCI and the Center for International Blood and Marrow Transplant Research and consists of genomic and clinical data from 824 patients with SAA who received HCT between 1989-2015. We excluded 140 patients with inherited bone marrow failure syndrome (92 with a clinical diagnosis and 48 genetically identified; McReynolds, Blood, 2022). We used exome sequencing data from the remaining 684 non-inherited SAA patients and used ACMG/AMP criteria to curate variants in 14 HLH-associated genes (AR: PRF1, UNC13D, STX11, STXBP2, ITK, CD27, LYST, RAB27A, AP3B1, BLOC1S6, SLC7A7; and XLR: XIAP, SH2D1A, MAGT1). Variants were deemed deleterious variants of uncertain significance (del-VUS) if there were damaging predictions in >3/5 in-silico meta-predictors (BayesDel, REVEL, CADD, MetaSVM and EIGEN). Follow-up started at the day of HCT and ended at outcome of interest, death, loss to follow-up, or end of study period (August 2017). We used the Kaplan-Meier estimator to calculate the probability of overall survival (OS) after HCT, using a log-rank test for statistical comparisons. For other HCT outcomes, we used cumulative incidence calculator accounting for competing risk of death. Results: There were 57 HLH variants in 49 of the 684 (7.2%) patients: 17 variants present in 19 patients (2.8%) were pathogenic or likely pathogenic (P/LP) and 40 were del-VUS. Eight of 17 P/LP variants were loss of function (3 frameshift, 3 splice site loss, and 2 nonsense). Among the 19 patients with P/LP variants, 16 (84%) had monoallelic variants in genes with AR inheritance, and PRF1 was the most frequently affected (in 8/19 patients). The median age at HCT was comparable in patients with (median=20 years; range=5-62) and without (median=20 years; range=0.2-77) P/LP HLH variants. We did not observe statistically significant differences in other demographics or transplant-related factors in the two groups, except for a higher frequency of male in patients with P/LP variants (79% vs. 55%, in those with and without HLH variants, respectively; p=0.04). All patients with P/LP HLH variants achieved neutrophil engraftment by day 30 compared with 92% of patients without HLH variants. The cumulative incidence of neutrophil engraftment at day 20 was 89%, 95% CI=70-99% for patients with HLH variants and 64%, 95% CI=60-67% in those without variants (p=0.01) (Figure 1A). No statistically significant associations between HLH variants and other key HCT outcomes were noted (p>0.05). The 5-year OS probability in all patients with P/LP variants was 89% (95% CI=72-99) compared to 66% (95% CI=63-70) in those without variants (p=0.06) (Figure 1B). When restricting the analysis to patients receiving reduced intensity/non-myeloablative regimen, the 5-year OS was 92% (95% CI=72-100%) in those with P/LP HLH variants versus 75% (95% CI=70-79%) in those without variants. Conclusions: We found that 7.2% of patients undergoing HCT for acquired SAA carried potentially deleterious variants in HLH genes, with 2.8% carrying a P/LP variant. HCT outcomes in patients with monoallelic P/LP variants was similar to those without P/LP HLH variants. Our data suggest that identification of monoallelic P/LP variants in HLH genes does not influence HCT outcomes, and no special treatment consideration, other than that used for SAA, is warranted.
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Dyskeratosis congenita and related telomere biology disorders (DC/TBDs) are multisystem illnesses caused by germline pathogenic variants (PV) in telomere biology genes resulting in very short ...telomeres. The inheritance varies and includes autosomal dominant (AD), autosomal recessive (AR) and X-linked (XLR) disease. Young patients may present with bone marrow failure (BMF) and the mucocutaneous triad of dysplastic nails, abnormal skin pigmentation, and oral leukoplakia, whereas adults may be identified with isolated BMF, pulmonary fibrosis (PF), or liver disease. In addition to BMF, DC/TBDs are associated with high risk of acute myeloid leukemia (AML), myelodysplastic syndrome (MDS), head and neck squamous cell cancer (HNSCC) and many other medical problems. Hematopoietic cell transplant (HCT), lung, and/or liver transplantation may be required. A limited number of genotype-phenotype studies suggest the mode of inheritance to be associated with phenotypes.
We studied 229 individuals (144 males, 85 females) with DC/TBDs enrolled in an IRB-approved longitudinal cohort study of inherited BMF syndromes (NCT00027274) between 01/2002 and 05/2019. Questionnaires and medical records were reviewed and a subset of 93 patients (40.6%) was systematically evaluated at the NIH Clinical Center (CC) during a 5-day visit. The median age at diagnosis for the entire cohort was 23.4 (0-82.2) years and median follow-up time was 4.2 (0-36.2) years. The PV was identified in 180 (78.6%) patients: 98 had AD disease (TERT, TERC, RTEL1, ACD, or PARN), 58 had XLR or AR (TERT, RTEL1, ACD, WRAP53, PARN, or DKC1) and 24 TINF2 (AD or de novo).
In the entire cohort of 229 patients, 69 (30.1%) required HCT for BMF median age 14.9 (0.9-63.1) years and 58% of these had AR/XLR or TINF2-associated disease. Eight patients received lung transplant for PF median age 51.4 (13.1-62.4) years. Liver transplant was required in 5 patients median age 25.8 (9.9-56.7) years. Twenty-seven non-HCT patients developed cancer median age of 42.1 (18.2-67.5) years. In 5 patients, cancer occurred after HCT median time since HCT=4.6 (1.8-14.6) years and 3 developed post-HCT lymphoproliferative disease.
Ninety-three patients were systematically phenotyped at the CC; 52 (55.9%) had >=1 triad feature and 33 (35.5%) had severe BMF. Three non-HCT patients had a history of cancer. PF was present in 6 patients and 3 had severe liver disease (portal hypertension and/or liver cirrhosis/fibrosis). Esophageal strictures were present in 7 and 5 had hip avascular necrosis (AVN, non-HCT). Median follow-up time after CC visit was 7.1 (0-17.1) years with median age at last follow-up of 29.6 (2.2-79.5) years . During follow-up, 9 patients progressed to severe BMF, and 8 developed cancer (4 post-HCT). Ten developed PF (5 post-HCT), and 9 advanced to pulmonary arteriovenous malformations (8 post-HCT). Eight patients progressed to severe liver disease (7 post-HCT). Gastrointestinal telangiectasias were seen in 6 individuals. Four patients developed esophageal stricture and 10 AVN (7 after radiotherapy and/or HCT).
At last follow up of the entire cohort, 92 (40.2%) of 229 patients were deceased. The main causes of death were lung disease (28.3%), treatment-related complications (19.6%), cancer (16.3%), or BMF (14.1%). The median overall survival was 54.2 years 95% confidence interval (CI) 47.3-59.8 and HCT-free median survival was 46.9 years (95% CI 37.9-54.2). Overall survival was significantly better in patients with AD versus XLR/AR or TINF2 disease (p<0.0001, log-rank test, median survival 66.5 versus 33.2 years, respectively).
Our study systematically quantified the presence and progression of DC/TBDs clinical manifestations and illustrates the significant morbidity faced by patients with these multisystem disorders. Long term survival is poor overall, but significantly better in patients with AD disease compared with other inheritance patterns. Although HCT has improved outcomes related to BMF, this study illustrates the pressing need to develop therapies for patients with DC/TBDs. These data also form the foundation for incorporating the mode of inheritance into evidence-based clinical care guidelines, risk stratification, and possibly treatment decisions in patients with DC/TBDs.
No relevant conflicts of interest to declare.
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•Pre-HCT mosaicism is related to increased relapse risk and lower survival after unrelated HCT, independent of cytogenetics at diagnosis.•Pre-HCT mosaicism could be a useful clinical tool to guide ...risk stratification in acute lymphoblastic leukemia patients.
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Objective The zebrafish is an established model system for studying the embryonic emergence of tissues and organs, including the hematopoietic system. We hypothesized that key signaling pathways ...controlling embryonic hematopoiesis continue to be important in the adult, and we sought to develop approaches to test this in zebrafish, focused on the bone morphogenetic protein (BMP) signaling pathway. Functions for this pathway in adult hematopoiesis have been challenging to probe in other models. Materials and Methods Several approaches tested the function of BMP signaling during adult zebrafish hematopoiesis. First, we evaluated steady-state hematopoiesis in adult fish that are heterozygous for mutant alleles of Smad5, or are homozygous for mutant alleles, and rescued to adulthood by injection of RNA encoding Smad5. Second, we tested the relative ability of smad5 mutant fish to recover from hemolytic anemia. Third, we generated a transgenic line that targets the expression of a dominant-negative BMP receptor to adult-stage Gata1+ progenitor cells. Results Adult fish with a strong mutant smad5 allele are anemic at steady state and, in addition, respond to hemolytic anemia with kinetics that are altered compared to wild-type fish. Fish expressing a mutant BMP receptor in early Gata1+ definitive progenitors generate excessive eosinophils. Conclusions Our study provides proof of principle that regulation of adult hematopoiesis can be studied in zebrafish by altering specific pathways. We show that the BMP signaling pathway is relevant for adult hematopoiesis to maintain steady state erythropoiesis, control the erythropoietic response following stress anemia, and to generate normal numbers of eosinophils.
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Methane is a potent greenhouse gas and its release to the atmosphere is widely believed to contribute to global warming. Ruminal enteric CH4 production represents a loss of 2–15% of the animal's ...gross energy (GE) intake and contributes nearly 20% of US CH4 emissions. Studies have evaluated the CH4 inhibiting potential of select short chain nitrocompounds, such as nitroethane, but results demonstrating their effects on ruminant exhaled CH4 emissions are lacking. Our study determined effects of oral nitroethane administration on CH4 emissions, accumulations of volatile fatty acids (VFA) and on ruminal CH4 producing activity in steers fed a forage based diet containing 8.8MJ/kg of metabolizable energy on a dry matter (DM) basis. Effects of nitroethane administration on ruminal nitroethane reducing activity were also determined. Holstein steers (n=24) of 317±6.5kg body weight (BW) were assigned to 4 treatments that included: 0, 30, 60 and 120mgnitroethane/kgBW/d. Treatments were administered via oral gavage twice daily at 08:00 and 16:00h for 8d. DM intake decreased quadratically as level of nitroethane increased with steers administered 60 and 120mgnitroethane/kgBW consuming 14 and 7% lower DM, respectively, than steers administered 0 or 30mgnitroethane/kgBW. Methane emissions as a proportion of GE intake and ruminal CH4 producing activity both decreased linearly (P<0.001) as level of nitroethane increased. Compared to control steers, daily administration of nitroethane at 60 and 120mg/kg BW reduced CH4 emissions as a proportion of GE intake 9–22% and ruminal CH4 producing activity 24–26%. Ruminal VFA accumulations were unaffected by nitroethane treatment. Results demonstrate that short term oral administration of nitroethane may be an effective anti-methanogenic compound in steers fed high forage diets. Further research is warranted to determine if strategies using nitroethane lower enteric CH4 emissions in ruminants long term. Ultimately, nitrocompounds which can be reduced by rumen microbes to yield compounds with nutritional value for the host, such as amino acids, would be preferred.
This paper is part of the special issue entitled: Greenhouse Gases in Animal Agriculture – Finding a Balance between Food and Emissions, Guest Edited by T.A. McAllister, Section Guest Editors: K.A. Beauchemin, X. Hao, S. McGinn and Editor for Animal Feed Science and Technology, P.H. Robinson.
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Shwachman-Diamond syndrome (SDS) is an inherited bone marrow failure syndrome (IBMFS) with gastrointestinal manifestations (pancreatic insufficiency) and cytopenias, primarily neutropenia. Skeletal ...dysplasias and short stature are frequent. Patients with SDS are at significantly increased risk of myelodysplastic syndrome and acute myeloid leukemia. More than 90% of patients have autosomal recessive inheritance of germline pathogenic variants in SBDS, a ribosome biogenesis gene. The most common variants are c.258+2T>C (disrupts the donor splice site of intron 2, leading to an 8bp deletion and a premature protein truncation due to a frameshift); and c.183_184TA>CT (introduces an in-frame stop codon). To date, the vast majority of pathogenic alleles reported have been single nucleotide variants (SNVs) or small insertions/deletions. Rare cases have been reported with SBDS exon 3 deletions.
The National Cancer Institute's IBMFS study is a longitudinal cohort study with 521 families enrolled, including 54 SDS or SDS-like families. Through clinical testing or whole exome sequencing all but nine families have had their disease-causing alleles identified. Three of the nine families had a known single pathogenic variant in SBDS. Array comparative genomic hybridization (aCGH) was uninformative in all but one.
We initially focused on the family with a known SBDS c.258+2T>C and potential deletion on aCGH. The 27 year-old (yo) male proband was diagnosed with SDS at 5yo due to a history of malabsorption, requiring pancreatic enzyme supplementation, failure to thrive (FTT), short stature and neutropenia. He also has metaphyseal dysplasia, scoliosis, psychomotor retardation and learning disability. A bone marrow (BM) biopsy at 21yo showed hypoplasia with mild dysplastic changes and a del20q clone. aCGH showed a possible large deletion, but the resolution was too low to determine exact breakpoints. Therefore, long-range Single Molecule, Real-Time (SMRT) Sequencing (PacBio Systems) was undertaken. This identified a read spanning a 19kb deletion with exact coordinates (hg19: chr7:66,436,397-66,455,294), and was confirmed with polymerase chain reaction and Sanger sequencing. This deletion removes part of intron 4, all of exon 5 and the 3'UTR of SBDS. Western blotting for SBDS showed significant decrease in protein production from cultured fibroblasts. This deletion in combination with the known SBDS variant and western blot is consistent with the SDS phenotype in this patient.
We then designed a targeted SMRT sequencing panel consisting of the four published SDS genes: SBDS, EFL1, DNAJC21 and SRP54 to evaluate the remaining families. The second proband evaluated was a 19yo female heterozygous for the SBDS c.258+2T>C variant. She had a history of malabsorption requiring enzyme supplementation and FTT in childhood. She has short stature, deformity of the distal metaphysis of the ulnae, and ongoing neutropenia with a hypocellular BM but no cytogenetic clones. Targeted SMRT sequencing identified a deletion in SBDS which deletes all of exon 3 and part of the surrounding introns (872bp, hg19: chr7: 66,457,992-66,458,863). Western blotting for SBDS showed a decrease in protein production in cultured fibroblasts consistent with a diagnosis of SDS. Sequencing of the family members indicated the deletion was paternal in origin, while the SNV was maternal.
A deletion of SRP54 was identified in two affected siblings from a third family; both siblings have a history of neutropenia and mild BM dysplasia. The deletion encompasses exon 8 of SRP54, which encodes part of the G-domain, critical for SRP54 cotranslational function, and is near other reported autosomal dominant pathogenic variants. One affected individual in this family also has a single SBDS c.183_184TA>CT variant, but had normal SBDS protein levels by western blot. In this family, the SBDS variant is likely not disease causing as individuals with heterozygous pathogenic variants of SBDS are generally asymptomatic, but rather the dominant SRP54 deletion is the disease causing genetic alteration in this SDS-like family.
No large deletions or insertions were identified in DNAJC21. Analysis of EFL1 is on-going. These cases illustrate the advantages of using long-read sequencing methodologies to identify large deletions which are not readily found using short-read sequencing, and identify large deletions as a novel mechanism of SDS inheritance.
No relevant conflicts of interest to declare.
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Background: It is important to identify inherited bone marrow failure syndromes (IBMFS) in patients with aplastic anemia in order to provide appropriate therapy. IBMFS are diagnosed through genetic ...or other diagnostic testing but can also go unrecognized, particularly in adults who may lack classic IBMFS features. This is particularly critical in light of increasing identification of individuals with variants of uncertain significance (VUS) and those with single pathogenic variants in an autosomal recessive (AR) gene or X-linked recessive (XLR) gene in females (SPVR) (“carriers”) during evaluation. In this study of patients diagnosed with acquired severe aplastic anemia (SAA), we evaluated putatively causal variants in IBMFS genes to determine the frequency of patients with an unrecognized IBMFS or SPVR and assessed their association with outcomes after hematopoietic cell transplant (HCT).
Methods: We used pre-HCT blood samples and clinical data from the Transplant Outcomes in Aplastic Anemia study (TOAA; a collaboration between the National Cancer Institute and the Center for International Blood and Marrow Transplant Research). Germline whole exome sequencing was performed on 732 patients with acquired SAA who received HCT between 1991-2015. A total of 104 IBMFS genes, (51 autosomal dominant (AD), 46 AR, 3 both AR and AD, 4 XLR) were evaluated for both single nucleotide (SNV) and copy number variants (CNV). All variants were curated using ACMG/AMP criteria, and a subset were validated by Sanger sequencing. Variants classified as VUS according to ACMG/AMP criteria and with a damaging score prediction in 3 of 5 in silico meta-predictors were categorized as deleterious VUS. Patients were divided into 3 groups based on known inheritance patterns of identified genes into those with 1) unrecognized IBMFS 2) SPVR or 3) neither (presumed acquired SAA). For telomere biology genes with AD/AR inheritance, we used telomeres <10 th percentile for age (by qPCR) to distinguish IBMFS from SPVR. For statistical analysis, we used the Kaplan-Meier estimator to calculate the probability of overall survival. The log-rank test was used to compare the survival distribution across patient categories. Cox proportional hazard models were used for multivariable analysis.
Results: We identified 309 variants of them 156 were pathogenic or likely pathogenic (P/LP) (112 (71.8%) loss of function SNVs, and 10 CNVs), and 153 were deleterious VUS. Patients with deleterious VUS did not have different survival compared with those with no variants, and these were not considered for designating unrecognized IBMFS and SPVR cases
Our analysis showed that 6.7% (N=49/732) of patients had variants consistent with an unrecognized IBMFS; 22 were AR (21 compound heterozygous, 1 homozygous), 26 AD and 1 XLR. Approximately half of those patients (22/49, 45%) had P/LP variants in hematopoiesis genes, and 31% in telomere biology genes (Figure 1A). We identified 79 patients with an SPVR, most in SBDS and FANCM.
Patients with an unrecognized IBMFS had worse overall survival when compared with patients with presumed acquired SAA (log-rank p=0.0098) (Figure 1B). Multivariable analysis confirmed this association (HR=2.11, 95% confidence interval (CI)=1.38-3.22, p=0.001). The observed survival difference was not mitigated by lower conditioning regimen intensity (HR=2.1, p=0.01 in myeloablative condition (MAC), and HR=2.3, p=0.016, in reduced intensity regimens (RIC)).
Patients with an SPVR had no post-HCT survival difference than presumed acquired SAA regardless of conditioning regimen (overall HR=0.96, p=0.85; MAC HR=1.4, p=0.27; RIC HR=0.3, p=0.1).
Conclusions: A sizable subset of patients (6.7%) with reported immune mediated acquired SAA had unrecognized inherited disorder and 33% were adults at HCT. Unrecognized IBMFS was associated with statistically significant poorer survival after HCT. In contrast, post-HCT survival in patients with an SPVR (“carriers”) was not different than those with acquired SAA. This work underscores the importance of identifying SAA patients with clinically meaningful underlying inherited disorders to enable the use therapeutic approaches to minimize regimen toxicity and late complications. It further highlights that identification of a single pathogenic variant in an autosomal recessive gene or X-linked gene in females and VUS are associated with post-HCT survival similar to acquired SAA.
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Paczesny: Medical University of South Carolina: Patents & Royalties: inventor on the ST2 bispecific antibody patent application. Lee: Kadmon: Research Funding; National Marrow Donor Program: Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding; JANSSEN: Other; AstraZeneca: Research Funding; Takeda: Research Funding; Novartis: Other: clinical trials, Research Funding; Syndax: Research Funding; Pfizer: Research Funding; Incyte: Research Funding.
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Genetic testing has been increasingly used to assist with differential diagnosis of acquired vs inherited bone marrow failure syndromes (IBMFS), a group of rare and heterogeneous diseases. However, ...the assay is still costly and not routinely available for many hematologists. To improve decision-making for genetic testing, we developed a genomic-based machine-learning model based on a two-step data-driven clustering and classification process to predict the likelihood of BMF patients having either an acquired or inherited disease based on 27 clinical and laboratory variables recorded at initial clinical encounter.
Clinical records from two independent cohorts of patients screened for pathogenic variants in genes associated with IBMFS were included in this study: the NIH cohort with 441 consecutive patients followed at the NHLBI and NCI, and the USP cohort with 172 consecutive patients from the Medical School of Ribeirão Preto/USP. In a binary target classification, cases were labeled as inherited if they had a pathogenic/likely pathogenic disease-causing variant and as acquired when they had benign or likely benign variants or negative genetic test, regardless of patients' clinical diagnoses. K-means clustering was first applied to resolve our highly dimensional data into two main clusters (Clusters A and B). An optimized bootstrap aggregation ensemble Cluster A specific was trained with cases from the NIH cohort (n=359). The model was then validated with Cluster A cases from the external USP cohort (n=127). The binary classification task was utilized to predict the etiology of BMF cases, labeled as acquired or inherited depending on patients' genomic data.
At first, unsupervised clustering separately grouped datasets into Cluster A, the largest group mostly represented by aplastic anemia (AA), and Cluster B, those underrepresented in our cohort including some classical IBMFS at early disease onset. The ensemble model Cluster A-specific was accurate to predict the BMF etiology in 88% of cases, correctly predicting inherited and likely immune BMF in 72% and 92% of cases, respectively. Out of the 27 initial clinical variables included in the model, 25 were found to be important for prediction. Telomere length (TL), age, and clinical variables were most important for the model's predictive accuracy, highlighting that a comprehensive history and physical examination encompassing all organ systems is imperative. Based on our model, genetic testing must be considered for patients in Cluster A predicted to have inherited disease and also for patients in Cluster B as no specific model was available but they were more likely to have IBMFS in comparison to Cluster A (50% vs 30%). We also recommend genetic screening in patients from Cluster A predicted to have acquired disease who are children (age <18 years who may not have clinical signs of IBMFS), have consanguinity in the family, have a diagnosis of myelodysplastic syndromes with or without suspicion for familial predisposition to myeloid malignancies (all cases where the model had limited prediction). A model without TL, an assay that can also be limited in low-resource centers, underperformed for prediction of inherited cases with sensitivity of 55%, highlighting the importance of TL measurement for the model's performance.
Our machine-learning model reproduced the clinical knowledge used by clinicians specialized in BMF and accurately predicted BMF etiology in 88% of cases. The model was particularly accurate for differential diagnosis of immune AA in adults, which may allow for selections of patients in whom rapidly starting immunosuppression rather than waiting weeks for genetic results is preferable. Clinical variables were strong predictors and adult patients with severe AA rarely had an inherited disease without a positive family history, a suggestive phenotype of IBMFS, or consanguinity being present. The generalizability of our model indicates that this tool can be used by hematologists not specialized in BMF to prioritize patients that would benefit from genetic testing. TL was a top predictor and a key variable for this model's accuracy. Implementation of TL measurement may be critical for differential diagnosis of BMF, especially in low-resource centers where genetic testing is not feasible or readily available. We plan to continue adding to the model to better predict IBMFS cases that were underrepresented in the current cohort.
Calado: Instituto Butantan: Consultancy; Agios: Membership on an entity's Board of Directors or advisory committees; Alexion Brasil: Consultancy; Novartis Brasil: Honoraria; Team Telomere, Inc.: Membership on an entity's Board of Directors or advisory committees; AA&MDS International Foundation: Research Funding. Young: Novartis: Research Funding.
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Abstract
Telomere biology disorders (TBDs) are cancer-prone syndromes associated with increased risk of acute myeloid leukemia (AML), head and neck squamous cell carcinoma (HNSCC), bone marrow ...failure, pulmonary fibrosis, and liver disease. Germline variants in at least 15 different telomere biology genes have been implicated (ACD, CTC1, TERT, TERC, STN1, NAF1, NOP10, NHP2, TINF2, RTEL1, PARN, ZCCHC8, DKC1, WRAP53, POT1) with autosomal dominant (AD), autosomal recessive (AR), or X-linked (XLR) inheritance as well as de novo occurrence. We hypothesize that TBDs may be more common than the currently estimated 1:1,000,000.
We analyzed the prevalence of germline variants in the 15 TBD-associated genes using 1) The Genome Aggregation Database gnomAD v2.1.1 non-cancer and 2) The Cancer Genome Atlas (TCGA). Germline TCGA variants were called using HaplotypeCaller, Freebayes, and UnifiedGenotyper. All variants with minor allele frequency (MAF) <1% in gnomAD non-cancer were considered. They were classified as potentially deleterious based on ClinVar and/or loss of function classification and/or stringent in silico predictions utilizing REVEL, MetaSVM, CADD, BayesDel, Eigen for missense and a combination of spliceAI, spidex, and dbscSNV for splice site variants. For TERC (RNA telomerase template) variants in the pseudoknot/template region were considered deleterious. The known disease-causing inheritance pattern (AD, AR, XLR) for each gene was used to identify TCGA cases with a probable TBD.
There were 1215 potentially deleterious variants in TBD associated genes in the gnomAD v2.1.1 non-cancer dataset (n=134,187 samples) for a combined prevalence of 0.9%, without accounting for zygosity. In genes with AD/AR inheritance, variants were most common in: RTEL1 (249), TERT (104), and PARN (104). CTC1 and WRAP53, both associated with solely AR TBDs, also showed high variant frequencies (280 and 102, respectively). In the TCGA dataset, 9089 cancer cases (32 solid tumors and AML) were evaluated. We identified 227 cases (2.5%) with 161 mono- or biallelic rare, potentially deleterious variants in the 15 genes. Ninety-two cases had a probable underlying TBD 1% of total, 41 male, 51 female, median age at cancer diagnosis 46.5 (1-92) years. Cancer diagnoses in these 92 cases included 24 solid tumors. The frequency of individuals with deleterious TBD-associated variants ranged from 0.4-2.8% in each of the 24 affected cancers. Notably, the heterozygous frequencies in lung squamous cell carcinoma and liver hepatocellular carcinoma were 1.6% and 2.2%, respectively. Our results show that the prevalence of TBDs may be noticeably higher than previously estimated, specifically in individuals with cancer. TBD-associated pathogenic germline variants may be implicated in the etiology of more solid tumor entities than previously recognized, warranting further studies.
Citation Format: Marena R. Niewisch, Jung Kim, Judith C. Lunger, Lisa J. McReynolds, Sharon A. Savage. Understanding the role of telomere biology gene variation in cancer etiology abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4108.
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