The Clinical Impact of CLIR Tools toward Rapid Resolution of Post-Newborn Screening Confirmatory Testing for X-Linked Adrenoleukodystrophy in California

Since the start of X-linked adrenoleukodystrophy (ALD) newborn screening in California, more than half of the diagnosed cases were found to have an ATP binding cassette subfamily D member 1 (ABCD1) gene variant of uncertain significance (VUS). To determine retrospectively the likelihood that these were true positive cases, we used a web-based post-analytical tool in Collaborative Laboratory Integrated Reports (CLIR). Confirmatory plasma very long-chain fatty-acids (VLCFA) profiles for ALD screen positive infant boys were run through the CLIR ALD tool. We compared the distribution by ABCD1 variant classification (pathogenic, likely pathogenic, VUS, and no variant) with the CLIR tool score interpretation (non-informative, possibly ALD, likely ALD, and very likely ALD) and the current case diagnosis. The study showed that CLIR tool positive interpretations were consistent with 100% of the pathogenic and likely pathogenic variants on the ABCD1 gene if a more conservative guideline was used. The tool interpretations were also consistent with screened cases that were determined to not have disease (our no-disorder group). The CLIR tool identified 19 diagnosed ALD cases with VUS to be potential false positives, representing a 40% reduction among all diagnosed ALD cases with VUS. The reduction could be extended to 65% if a more aggressive threshold was used. Identifying such preventable false positives could alleviate the follow-up burden for patients, their families, and California Special Care Centers.

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Adrenoleukodystrophy Newborn Screening in California Since 2016: Programmatic Outcomes and Follow-Up

International Journal of Neonatal Screening ◽

10.3390/ijns7020022 ◽

2021 ◽

Vol 7 (2) ◽

pp. 22

Author(s):

Jamie Matteson ◽

Stanley Sciortino ◽

Lisa Feuchtbaum ◽

Tracey Bishop ◽

Richard S. Olney ◽

...

Keyword(s):

Newborn Screening ◽

Program Implementation ◽

Screening Program ◽

Birth Prevalence ◽

Variant Of Uncertain Significance ◽

Implementation Challenges ◽

Screening Programs ◽

Uncertain Significance ◽

Long Term Follow Up

X-linked adrenoleukodystrophy (ALD) is a recent addition to the Recommended Uniform Screening Panel, prompting many states to begin screening newborns for the disorder. We provide California’s experience with ALD newborn screening, highlighting the clinical and epidemiological outcomes observed as well as program implementation challenges. In this retrospective cohort study, we examine ALD newborn screening results and clinical outcomes for 1,854,631 newborns whose specimens were received by the California Genetic Disease Screening Program from 16 February 2016 through 15 February 2020. In the first four years of ALD newborn screening in California, 355 newborns screened positive for ALD, including 147 (41%) with an ABCD1 variant of uncertain significance (VUS) and 95 males diagnosed with ALD. After modifying cutoffs, we observed an ALD birth prevalence of 1 in 14,397 males. Long-term follow-up identified 14 males with signs of adrenal involvement. This study adds to a growing body of literature reporting on outcomes of newborn screening for ALD and offering a glimpse of what other large newborn screening programs can expect when adding ALD to their screening panel.

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Revealing the Mutational Spectrum in Southern Africans With Amyotrophic Lateral Sclerosis

Neurology Genetics ◽

10.1212/nxg.0000000000000654 ◽

2022 ◽

Vol 8 (1) ◽

pp. e654

Author(s):

Melissa Nel ◽

Amokelani C. Mahungu ◽

Nomakhosazana Monnakgotla ◽

Gerrit R. Botha ◽

Nicola J. Mulder ◽

...

Keyword(s):

Amyotrophic Lateral Sclerosis ◽

Population Group ◽

Genetic Ancestry ◽

Familial Case ◽

Data Sets ◽

Variant Of Uncertain Significance ◽

Pathogenic Variants ◽

Sporadic Als ◽

Uncertain Significance ◽

Lateral Sclerosis

Background and ObjectivesTo perform the first screen of 44 amyotrophic lateral sclerosis (ALS) genes in a cohort of African genetic ancestry individuals with ALS using whole-genome sequencing (WGS) data.MethodsOne hundred three consecutive cases with probable/definite ALS (using the revised El Escorial criteria), and self-categorized as African genetic ancestry, underwent WGS using various Illumina platforms. As population controls, 238 samples from various African WGS data sets were included. Our analysis was restricted to 44 ALS genes, which were curated for rare sequence variants and classified according to the American College of Medical Genetics guidelines as likely benign, uncertain significance, likely pathogenic, or pathogenic variants.ResultsThirteen percent of 103 ALS cases harbored pathogenic variants; 5 different SOD1 variants (N87S, G94D, I114T, L145S, and L145F) in 5 individuals (5%, 1 familial case), pathogenic C9orf72 repeat expansions in 7 individuals (7%, 1 familial case) and a likely pathogenic ANXA11 (G38R) variant in 1 individual. Thirty individuals (29%) harbored ≥1 variant of uncertain significance; 10 of these variants had limited pathogenic evidence, although this was insufficient to permit confident classification as pathogenic.DiscussionOur findings show that known ALS genes can be expected to identify a genetic cause of disease in >11% of sporadic ALS cases of African genetic ancestry. Similar to European cohorts, the 2 most frequent genes harboring pathogenic variants in this population group are C9orf72 and SOD1.

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Germline pathologic mutations and cancer family history in prostate cancer patients.

Journal of Clinical Oncology ◽

10.1200/jco.2018.36.6_suppl.378 ◽

2018 ◽

Vol 36 (6_suppl) ◽

pp. 378-378

Author(s):

Marcus Marie Moses ◽

Elisa Ledet ◽

Emma M. Ernst ◽

Patrick Cotogno ◽

Joshua Schiff ◽

...

Keyword(s):

Prostate Cancer ◽

Family History ◽

San Francisco ◽

Treatment Options ◽

Distant Metastases ◽

Cancer Center ◽

Chi Square ◽

Variant Of Uncertain Significance ◽

Pathogenic Variants ◽

Uncertain Significance

378 Background: Prostate cancer (PCa) patients (pts) with metastases and/or strong family history (FH) of cancer (Ca) are at higher risk of a germline mutation. The identification of alterations in PCa pts may be important for risk stratification as well as personalizing treatment options. The goal of this study was characterization of FH and pathogenic variants (PV) detected in PCa pts, with both localized and metastatic disease. Methods: 300 PCa pts from Tulane Cancer Center underwent germline testing. 265 Caucasian (C) and 35 African-Americans (AA) were tested and met the NCCN criteria for testing and/or had distant metastases (mets). Germline genetic testing was done via commercial panels (30-80 genes) (Invitae. San Francisco, Ca). PCa pts had extensive FH screening. Clinical annotation included age at diagnosis (dx), race, and presence of mets at any time. Chi square tests were used to compare clinical correlates and PVs. Results: Of the 300 pts tested, 182 pts (60.6%) had mets and 118 (39.4%) did not. 41 pts (13.6%) had ≥ 1 germline pathogenic variant (PV) and 161 pts (53.6%) had ≥ 1 germline variant of uncertain significance (VUS). PVs were detected in BRCA2 (n = 10), MUTYH (n = 8), CHEK2 (n = 6), BRCA1 (n = 4), ATM (n = 4), TP53 (n = 3), PMS2 (n = 2), BLM (n = 2), MITF (n = 2), NBN (n = 1), and RAD51D (n = 1). MUTYH and MITF are not known to be linked to prostate cancer. There was no significant relationships in FH PCa and FH non-PCa in regard to likelihood of a PV (p = .86 and p = .18). Of the 300 pts tested, 136 pts (45.3%) had PCa FH, 131 pts (43.6%) had breast Ca FH, 61 pts (20.3%) had lung Ca FH, 61 pts (20.3%) had colon Ca FH, 37 pts (12.3%) had pancreatic Ca FH, and 32 pts (10.6%) had ovarian Ca FH. 45.6% of C men (n = 121) and 42.8% of AA men (n = 15) had PCa FH. Pts with a non-PCa FH (n = 255) were 1.37 times more likely to develop mets (p = .01168). The median age of dx were 61 for PV pts, 62 for VUS pts, and 61 for negative pts (non-significant). 21/182 pts with mets (11.5%) had a PV; 8/182 (4.4%) pts with mets had a BRCA2 PV. Conclusions: In metastatic patients, FH of prostate cancer alone cannot predict those with PV. The most common Cas observed in these pts were breast, lung, colon and pancreatic. A larger cohort is needed to fully characterize and understand the co-segregation of PCa with other Cas.

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A Biochemical Platform to Define the Relative Specific Activity of IDUA Variants Identified by Newborn Screening

International Journal of Neonatal Screening ◽

10.3390/ijns6040088 ◽

2020 ◽

Vol 6 (4) ◽

pp. 88

Author(s):

Seok-Ho Yu ◽

Laura Pollard ◽

Tim Wood ◽

Heather Flanagan-Steet ◽

Richard Steet

Keyword(s):

Newborn Screening ◽

Single Cell ◽

Specific Activity ◽

Relative Specific Activity ◽

Variants Of Uncertain Significance ◽

Pathogenic Variants ◽

Scheie Syndrome ◽

Cell Clones ◽

Uncertain Significance ◽

Specific Activities

The lysosomal storage disorder, mucopolysaccharidosis I (MPSI), results from mutations in IDUA, the gene that encodes the glycosaminoglycan-degrading enzyme α-L-iduronidase. Newborn screening efforts for MPSI have greatly increased the number of novel IDUA variants identified, but with insufficient experimental evidence regarding their pathogenicity, many of these variants remain classified as variants of uncertain significance (VUS). Defining pathogenicity for novel IDUA variants is critical for decisions regarding medical management and early intervention. Here, we describe a biochemical platform for the characterization of IDUA variants that relies on viral delivery of IDUA DNA into IDUA-deficient HAP1 cells and isolation of single cell expression clones. The relative specific activity of wild-type and variant α-iduronidase was determined using a combination of Western blot analysis and α-iduronidase activity assays. The specific activity of each variant enzyme was consistent across different single cell clones despite variable IDUA expression and could be accurately determined down to 0.05–0.01% of WT α-iduronidase activity. With this strategy we compared the specific activities of known pseudodeficiency variants (p.His82Gln, p.Ala79Thr, p.Val322Glu, p.Asp223Asn) or pathogenic variants (p.Ser633Leu, p.His240Arg) with variants of uncertain significance (p.Ser586Phe, p.Ile272Leu). The p.Ser633Leu and p.His240Arg variants both show very low activities consistent with their association with Scheie syndrome. In our experiments, however, p.His240Arg exhibited a specific activity five times higher than p.Ser633Leu in contrast to other reports showing equivalent activity. Cell clones expressing the p.Ser586Phe and p.Ile272Leu variants had specific activities in the range of other pseudodeficiency variants tested. Our findings show that pseudodeficiency and pathogenic variants can be distinguished from each other with regard to specific activity, and confirms that all the pseudodeficiency variants variably reduce α-iduronidase activity. We envision this platform will be a valuable resource for the rigorous assessment of the novel IDUA variants emerging from the expansion of newborn screening efforts.

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Exome Sequencing Highlights a Potential Role for Concealed Cardiomyopathies in Youthful Sudden Cardiac Death

Circulation Genomic and Precision Medicine ◽

10.1161/circgen.121.003497 ◽

2021 ◽

Author(s):

Raquel Neves ◽

David J. Tester ◽

Michael A. Simpson ◽

Elijah R. Behr ◽

Michael J. Ackerman ◽

...

Keyword(s):

Exome Sequencing ◽

Strong Evidence ◽

Susceptibility Gene ◽

Susceptibility Genes ◽

Medical Genetics ◽

Variant Of Uncertain Significance ◽

Pathogenic Variants ◽

Genetic Test Results ◽

Uncertain Significance ◽

Genetics And Genomics

Background: Sudden cardiac arrest (SCA) and sudden unexplained death (SUD) are feared sequelae of many genetic heart diseases. In rare circumstances, pathogenic variants in cardiomyopathy-susceptibility genes may result in electrical instability leading to SCA/SUD before any structural manifestations of underlying cardiomyopathy are evident. Methods: Collectively, 38 unexplained SCA survivors (21 males; mean age at SCA 26.4±13.1 years), 68 autopsy-inconclusive SUD cases (49 males; mean age at death 20.4±9.0 years) without disease-causative variants in the channelopathy genes, and 973 ostensibly healthy controls were included. Following exome sequencing, ultrarare (minor allele frequency ≤0.00005 in any ethnic group within Genome Aggregation Database [gnomAD, n=141 456 individuals]) nonsynonymous variants identified in 24 ClinGen adjudicated definitive/strong evidence cardiomyopathy-susceptibility genes were analyzed. Eligible variants were adjudicated as pathogenic, likely pathogenic, or variant of uncertain significance in accordance with current American College of Medical Genetics and Genomics guidelines. Results: Overall, 7 out of 38 (18.4%) SCA survivors and 14 out of 68 (20.5%) autopsy-inconclusive, channelopathic-negative SUD cases had at least one pathogenic/likely pathogenic or a variant of uncertain significance nonsynonymous variant within a strong evidence, cardiomyopathy-susceptibility gene. Following American College of Medical Genetics and Genomics criterion variant adjudication, a pathogenic or likely pathogenic variant was identified in 3 out of 38 (7.9%; P =0.05) SCA survivors and 8 out of 68 (11.8%; P =0.0002) autopsy-inconclusive SUD cases compared to 20 out of 973 (2.1%) European controls. Interestingly, the yield of pathogenic/likely pathogenic variants was significantly greater in autopsy-inconclusive SUD cases with documented interstitial fibrosis (4/11, 36%) compared with only 4 out of 57 (7%, P <0.02) SUD cases without ventricular fibrosis. Conclusion: Our data further supports the inclusion of strongevidence cardiomyopathy-susceptibility genes on the genetic testing panels used to evaluate unexplained SCA survivors and autopsy-inconclusive/negative SUD decedents. However, to avoid diagnostic miscues, the careful interpretation of genetic test results in patients without overt phenotypes is vital.

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Classification of the canonical splice alteration MUTYH c.934-2A>G is likely benign based on RNA and clinical data

Molecular Case Studies ◽

10.1101/mcs.a006152 ◽

2021 ◽

pp. mcs.a006152

Author(s):

Felicia Hernandez ◽

Blair Rene Conner ◽

Marcy E. Richardson ◽

Holly LaDuca ◽

Elizabeth Chao ◽

...

Keyword(s):

Amino Acids ◽

Colonic Polyps ◽

Autosomal Recessive Disorder ◽

Splice Defect ◽

Critical Domain ◽

Variant Of Uncertain Significance ◽

Aberrant Transcript ◽

Pathogenic Variants ◽

Uncertain Significance ◽

Exon 11

MUTYH-associated polyposis (MAP) is an autosomal recessive disorder characterized by the development of multiple adenomatous colonic polyps and an increased lifetime risk of colorectal cancer. Germline biallelic pathogenic variants in MUTYH are responsible for MAP. The MUTYH c.934-2A>G (NM_001128425.1) variant, which is also known as c.850-2A>G for NM_001048174.2, has been identified in our laboratory in more than 800 patients, including homozygous and compound heterozygote carriers. The variant was initially classified as a variant of uncertain significance (VUS) due to lack of a MAP phenotype in biallelic carriers. In two unrelated female patients who were heterozygous carriers of this variant, further testing by RNA sequencing identified an aberrant transcript with a deletion of 9 nucleotides at the start of exon 11 (MUTYH r.934_942del9). This event is predicted to lead to an in-frame loss of 3 amino acids in a non-critical domain of the protein. This was the only splice defect identified in these patients which was not present in the controls and the aberrant transcript is derived exclusively from the variant allele, strongly supporting the cause of this splice defect as being the intronic variant, MUTYH c.934-2A>G. The splicing analysis demonstrating a small in-frame skipping of 3 amino acids in a non-critical domain along with the absence of a MAP phenotype in our internal cohort of biallelic carriers provides evidence that the variant is likely benign and not of clinical significance.

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Comprehensive molecular-genetic analysis of mid-frequency sensorineural hearing loss

Scientific Reports ◽

10.1038/s41598-021-01876-1 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Zuzana Pavlenkova ◽

Lukas Varga ◽

Silvia Borecka ◽

Miloslav Karhanek ◽

Miloslava Huckova ◽

...

Keyword(s):

Hearing Loss ◽

Sensorineural Hearing Loss ◽

Genetic Disorder ◽

Molecular Genetic ◽

Molecular Genetic Analysis ◽

Sensorineural Hearing ◽

Variant Of Uncertain Significance ◽

Pathogenic Variants ◽

Causal Genes ◽

Uncertain Significance

AbstractThe genetic heterogeneity of sensorineural hearing loss (SNHL) is a major hurdle to the detection of disease-causing variants. We aimed to identify underlying causal genes associated with mid-frequency hearing loss (HL), which contributes to less than about 1% of SNHL cases, by whole exome sequencing (WES). Thirty families segregating mid-frequency SNHL, in whom biallelic GJB2 mutations had been previously excluded, were selected from among 851 families in our DNA repository of SNHL. DNA samples from the probands were subjected to WES analysis and searched for candidate variants associated with SNHL. We were able to identify the genetic aetiology in six probands (20%). In total, we found three pathogenic and three likely pathogenic variants in four genes (COL4A5, OTOGL, TECTA, TMPRSS3). One more proband was a compound heterozygote for a pathogenic variant and a variant of uncertain significance (VUS) in MYO15A gene. To date, MYO15A and TMPRSS3 have not yet been described in association with mid-frequency SNHL. In eight additional probands, eight candidate VUS variants were detected in five genes (DIAPH1, MYO7A, TECTA, TMC1, TSPEAR). Seven of these 16 variants have not yet been published or mentioned in the available databases. The most prevalent gene was TECTA, identified in 23% of all tested families. Furthermore, we confirmed the hypothesis that a substantive portion of cases with this conspicuous audiogram shape is a consequence of a genetic disorder.

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Interpretation of Variants of Uncertain Significance in the Clinical Setting: A Case of Treatable Ataxia

10.21203/rs.3.rs-55670/v1 ◽

2020 ◽

Author(s):

Miguel Wilken ◽

Malco Rossi ◽

Marcelo Merello

Keyword(s):

Response To Treatment ◽

Autosomal Dominant Cerebellar Ataxia ◽

Spinocerebellar Ataxia Type ◽

Clinical Approach ◽

Variant Of Uncertain Significance ◽

Pathogenic Variants ◽

Whole Exome ◽

Limb Ataxia ◽

Uncertain Significance ◽

Negative Effect

Abstract Background: Spinocerebellar ataxia type 38 (SCA38) is an autosomal dominant cerebellar ataxia caused by pathogenic variants in the elongation of very long chain fatty acids-like 5 gene (ELOVL5). Improvement of ataxia with a docosahexaenoic acid (DHA) replacement therapy has been reported.Case presentation: A 73-year-old man of Hispanic descent presented with gait and limb ataxia, dysarthria, slow and hypometric saccades, hearing loss, mild cognitive impairment, and hypopalesthesia. The initial scale for the assessment and rating of ataxia (SARA) score was 11. After a negative routine workout for ataxia and testing for common forms due to expanded repeats, whole-exome sequencing (WES) identified a heterozygous variant (c.327+1G>A) in the ELOVL5 gene that was predicted to have a negative effect on splicing but was categorized as a Variant of Uncertain Significance (VUS). The patient was started on DHA 600 mg/day. Four months later, the patient showed a considerable reduction in the scale for the assessment and rating of ataxia (SARA) score, from 11 to 5 points, with a clear improvement in gait and limb ataxia that was sustained at 24 months of follow-up.Conclusions: We illustrate the case of a patient presenting with a variant considered genetically and biochemically of uncertain significance. Despite being a VUS, its location in a gene that is known to cause ataxia (SCA38), as well as a compatible phenotype, led to the interpretation of this variant as probably pathogenic from a clinical practice standpoint, especially considering prior reports that showed clinical improvement with a specific, over-the-counter, pharmacological treatment. A further satisfactory response to treatment supported our clinical approach.

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Four Novel Variants in POU4F3 Cause Autosomal Dominant Nonsyndromic Hearing Loss

Neural Plasticity ◽

10.1155/2020/6137083 ◽

2020 ◽

Vol 2020 ◽

pp. 1-12

Author(s):

Tian-Yi Cui ◽

Xue Gao ◽

Sha-Sha Huang ◽

Yan-Yan Sun ◽

Si-Qi Zhang ◽

...

Keyword(s):

Hearing Loss ◽

Autosomal Dominant ◽

Asian Population ◽

Nonsyndromic Hearing Loss ◽

Hereditary Hearing Loss ◽

Variant Of Uncertain Significance ◽

Targeted Next Generation Sequencing ◽

Pathogenic Variants ◽

Novel Variants ◽

Uncertain Significance

Hereditary hearing loss is one of the most common sensory disabilities worldwide. Mutation of POU domain class 4 transcription factor 3 (POU4F3) is considered the pathogenic cause of autosomal dominant nonsyndromic hearing loss (ADNSHL), designated as autosomal dominant nonsyndromic deafness 15. In this study, four novel variants in POU4F3, c.696G>T (p.Glu232Asp), c.325C>T (p.His109Tyr), c.635T>C (p.Leu212Pro), and c.183delG (p.Ala62Argfs∗22), were identified in four different Chinese families with ADNSHL by targeted next-generation sequencing and Sanger sequencing. Based on the American College of Medical Genetics and Genomics guidelines, c.183delG (p.Ala62Argfs∗22) is classified as a pathogenic variant, c.696G>T (p.Glu232Asp) and c.635T>C (p.Leu212Pro) are classified as likely pathogenic variants, and c.325C>T (p.His109Tyr) is classified as a variant of uncertain significance. Based on previous reports and the results of this study, we speculated that POU4F3 pathogenic variants are significant contributors to ADNSHL in the East Asian population. Therefore, screening of POU4F3 should be a routine examination for the diagnosis of hereditary hearing loss.

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Genomic Observations of a Rare/Pathogenic SMAD3 Variant in Loeys–Dietz Syndrome 3 Confirmed by Protein Informatics and Structural Investigations

Medicina ◽

10.3390/medicina55050137 ◽

2019 ◽

Vol 55 (5) ◽

pp. 137

Author(s):

John E. Richter ◽

Ayesha Samreen ◽

Charitha Vadlamudi ◽

Haytham Helmi ◽

Ahmed N. Mohammad ◽

...

Keyword(s):

Connective Tissue ◽

Autosomal Dominant ◽

Connective Tissue Disorders ◽

Structural Investigations ◽

Variant Of Uncertain Significance ◽

Common Features ◽

Pathogenic Variants ◽

Uncertain Significance ◽

History Of ◽

Arterial Aneurysms

Background and objectives: Loeys–Dietz syndrome 3, also known as aneurysms-–osteoarthritis syndrome, is an autosomal dominant genetic connective tissue disease caused by pathogenic variants in SMAD3, a transcription factor involved in TGF-β signaling. This disorder is characterized by early-onset osteoarthritis and arterial aneurysms. Common features include scoliosis, uvula abnormalities, striae, and velvety skin. Materials and Methods: The pathogenicity of a variant of uncertain significance in the SMAD3 gene was evaluated (variant c.220C > T) through personalized protein informatics and molecular studies. Results: The case of a 44-year-old male, who was originally presumed to have Marfan syndrome, is presented. An expanded gene panel determined the probable cause to be a variant in SMAD3, c.220C > T (p.R74W). His case was complicated by a history of stroke, but his phenotype was otherwise characteristic for Loeys–Dietz syndrome 3. Conclusion: This case emphasizes the importance of comprehensive genetic testing to evaluate patients for connective tissue disorders, as well as the potential benefit of utilizing a protein informatics platform for the assessment of variant pathogenicity.

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