Publications by authors named "Clifton Dalgard"

85 Publications

Apolipoprotein e (APOE) ε4 genotype influences memory performance following remote traumatic brain injury in U.S. military service members and veterans.

Brain Cogn 2021 Sep 3;154:105790. Epub 2021 Sep 3.

Tramatic Brain Injury Center of Excellence, Silver Spring, MD, USA; Walter Reed National Military Medical Center, Bethesda, MD, USA; National Intrepid Center of Excellence, Bethesda, MD, USA; Uniformed Services University of the Health Sciences, Bethesda, MD, USA.

The purpose of this study was to examine the association between the apolipoprotein E (APOE) ε4 allele and neurocognitive functioning following traumatic brain injury (TBI) in military service members and veterans (SMVs). Participants included 176 SMVs with a history of remote TBI (≥1 year post-injury), categorized into mild (n = 100), moderate (n = 40), and severe (n = 36) TBI groups. Participants completed a neuropsychological assessment and APOE genotyping (n = 46 ε4+, n = 130 ε4-). Neurocognitive composite scores representing memory, executive functioning, and visual processing speed were computed. ANCOVAs adjusting for race, education, combat exposure, and PTSD symptom severity showed a significant main effect of ε4 on the memory composite, such that ε4+ SMVs exhibited poorer memory performance than ε4- SMVs. When ε2 allele carriers were removed from the analyses, associations with memory were strengthened, demonstrating a possible protective effect of the ε2 allele. No main effect of TBI group was identified on any cognitive composite, nor were there any significant TBI group × ε4 status interactions for any cognitive composite. Future studies with larger samples are needed to verify these findings, but our results suggest an important relationship between ε4 status and memory functioning following remote TBI of all severities.
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http://dx.doi.org/10.1016/j.bandc.2021.105790DOI Listing
September 2021

Association of Variants in the SPTLC1 Gene With Juvenile Amyotrophic Lateral Sclerosis.

JAMA Neurol 2021 Aug 30. Epub 2021 Aug 30.

Maurice Wohl Clinical Neuroscience Institute, Institute of Psychiatry, Psychology and Neuroscience, King's College London, London, United Kingdom.

Importance: Juvenile amyotrophic lateral sclerosis (ALS) is a rare form of ALS characterized by age of symptom onset less than 25 years and a variable presentation.

Objective: To identify the genetic variants associated with juvenile ALS.

Design, Setting, And Participants: In this multicenter family-based genetic study, trio whole-exome sequencing was performed to identify the disease-associated gene in a case series of unrelated patients diagnosed with juvenile ALS and severe growth retardation. The patients and their family members were enrolled at academic hospitals and a government research facility between March 1, 2016, and March 13, 2020, and were observed until October 1, 2020. Whole-exome sequencing was also performed in a series of patients with juvenile ALS. A total of 66 patients with juvenile ALS and 6258 adult patients with ALS participated in the study. Patients were selected for the study based on their diagnosis, and all eligible participants were enrolled in the study. None of the participants had a family history of neurological disorders, suggesting de novo variants as the underlying genetic mechanism.

Main Outcomes And Measures: De novo variants present only in the index case and not in unaffected family members.

Results: Trio whole-exome sequencing was performed in 3 patients diagnosed with juvenile ALS and their parents. An additional 63 patients with juvenile ALS and 6258 adult patients with ALS were subsequently screened for variants in the SPTLC1 gene. De novo variants in SPTLC1 (p.Ala20Ser in 2 patients and p.Ser331Tyr in 1 patient) were identified in 3 unrelated patients diagnosed with juvenile ALS and failure to thrive. A fourth variant (p.Leu39del) was identified in a patient with juvenile ALS where parental DNA was unavailable. Variants in this gene have been previously shown to be associated with autosomal-dominant hereditary sensory autonomic neuropathy, type 1A, by disrupting an essential enzyme complex in the sphingolipid synthesis pathway.

Conclusions And Relevance: These data broaden the phenotype associated with SPTLC1 and suggest that patients presenting with juvenile ALS should be screened for variants in this gene.
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http://dx.doi.org/10.1001/jamaneurol.2021.2598DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8406220PMC
August 2021

X-linked recessive TLR7 deficiency in ~1% of men under 60 years old with life-threatening COVID-19.

Sci Immunol 2021 08;6(62)

Specialized Immunology Laboratory of Dr. Shahrooei, Sina Medical Complex, Ahvaz, Iran.

Autosomal inborn errors of type I IFN immunity and autoantibodies against these cytokines underlie at least 10% of critical COVID-19 pneumonia cases. We report very rare, biochemically deleterious X-linked variants in 16 unrelated male individuals aged 7 to 71 years (mean: 36.7 years) from a cohort of 1,202 male patients aged 0.5 to 99 years (mean: 52.9 years) with unexplained critical COVID-19 pneumonia. None of the 331 asymptomatically or mildly infected male individuals aged 1.3 to 102 years (mean: 38.7 years) tested carry such variants ( = 3.5 × 10). The phenotypes of five hemizygous relatives of index cases infected with SARS-CoV-2 include asymptomatic or mild infection (=2, 5 and 38 years), or moderate (=1, 5 years), severe (=1, 27 years), or critical (=1, 29 years) pneumonia. Two boys (aged 7 and 12 years) from a cohort of 262 male patients with severe COVID-19 pneumonia (mean: 51.0 years) are hemizygous for a deleterious TLR7 variant. The cumulative allele frequency for deleterious variants in the male general population is < 6.5x10 We also show that blood B cell lines and myeloid cell subsets from the patients do not respond to TLR7 stimulation, a phenotype rescued by wild-type The patients' blood plasmacytoid dendritic cells (pDCs) produce low levels of type I IFNs in response to SARS-CoV-2. Overall, X-linked recessive TLR7 deficiency is a highly penetrant genetic etiology of critical COVID-19 pneumonia, in about 1.8% of male patients below the age of 60 years. Human TLR7 and pDCs are essential for protective type I IFN immunity against SARS-CoV-2 in the respiratory tract.
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http://dx.doi.org/10.1126/sciimmunol.abl4348DOI Listing
August 2021

RNA-sequencing of AVPV and ARH reveals vastly different temporal and transcriptomic responses to estradiol in the female rat hypothalamus.

PLoS One 2021 18;16(8):e0256148. Epub 2021 Aug 18.

Dept of Neurobiology, and the Laboratory of Neuroendocrinology of the Brain Research Institute, UCLA DGSOM, Los Angeles, California, United States of America.

In females, estrogens have two main modes of action relating to gonadotropin secretion: positive feedback and negative feedback. Estrogen positive and negative feedback are controlled by different regions of the hypothalamus: the preoptic area/anterior portion (mainly the anteroventral periventricular nucleus, AVPV) of the hypothalamus is associated with estrogen positive feedback while the mediobasal hypothalamus (mainly the arcuate nucleus of the hypothalamus, ARH), is associated with estrogen negative feedback. In this study, we examined the temporal pattern of gene transcription in these two regions following estrogen treatment. Adult, ovariectomized, Long Evans rats received doses of estradiol benzoate (EB) or oil every 4 days for 3 cycles. On the last EB priming cycle, hypothalamic tissues were dissected into the AVPV+ and ARH+ at 0 hrs (baseline/oil control), 6 hrs, or 24 hrs after EB treatment. RNA was extracted and sequenced using bulk RNA sequencing. Differential gene analysis, gene ontology, and weighted correlation network analysis (WGCNA) was performed. Overall, we found that the AVPV+ and ARH+ respond differently to estradiol stimulation. In both regions, estradiol treatment resulted in more gene up-regulation than down-regulation. S100g was very strongly up-regulated by estradiol in both regions at 6 and 24 hrs after EB treatment. In the AVPV+ the highest number of differentially expressed genes occurred 24 hrs after EB. In the ARH+, the highest number of genes differentially expressed by EB occurred between 6 and 24 hrs after EB, while in the AVPV+, the fewest genes changed their expression between these time points, demonstrating a temporal difference in the way that EB regulates transcription these two areas. Several genes strongly implicated in gonadotropin release were differentially affected by estradiol including Esr1, encoding estrogen receptor-α and Kiss1, encoding kisspeptin. As an internal validation, Kiss1 was up-regulated in the AVPV+ and down-regulated in the ARH+. Gene network analysis revealed the vastly different clustering of genes modulated by estradiol in the AVPV+ compared with the ARH+. These results indicate that gene expression in these two hypothalamic regions have specific responses to estradiol in timing and direction.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0256148PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8372949PMC
August 2021

Apolipoprotein E e4 is associated with worse self-reported neurobehavioral symptoms following uncomplicated mild traumatic brain injury in U.S. military service members.

Behav Brain Res 2021 Oct 29;415:113491. Epub 2021 Jul 29.

Traumatic Brain Injury Center of Excellence, 1335 E W Hwy, Silver Spring, MD, 20910, United States; Walter Reed National Military Medical Center, 4494 Palmer Rd N, Bethesda, MD, 20814, United States; National Intrepid Center of Excellence, Palmer Rd S, Bethesda, MD, 20814, United States; Uniformed Services University of the Health Sciences, 4301 Jones Bridge Rd, Bethesda, MD, 20814, United States. Electronic address:

Past research has found a relationship between the apolipoprotein E (APOE) e4 allele and worse neurobehavioral functioning following mild traumatic brain injury (MTBI) in civilian populations. The purpose of this study was to examine this relationship in service members and veterans (SMVs) following MTBI. Participants were 151 SMVs (103 uncomplicated MTBI; 48 Injured Controls [IC]) prospectively enrolled in the DVBIC-TBICoE 15-Year Longitudinal TBI Study. Participants completed a battery of self-reported neurobehavioral symptom measures on average 76.2 months post-injury (SD = 31.8). APOE genotyping was undertaken using non-fasting blood samples. Participants were classified into four subgroups based on injury (MTBI vs. IC) and APOE e4 allele status (e4 present/absent). In the IC group, there were no significant differences across APOE e4 status subgroups for all measures. In the MTBI group, participants with the APOE e4 allele had significantly worse scores on measures of depression, pain, anxiety, grief, positive well-being, social participation, and resilience compared to those without the e4 allele (d = .44 to d = .69). When comparing the number of 'clinically elevated' neurobehavioral measures simultaneously, the MTBI/e4 present subgroup consistently had a higher number of elevated measures compared to the MTBI/e4 absent, IC/e4 present, and IC/e4 absent subgroups. The APOE e4 allele was associated with poorer neurobehavioral outcome in SMVs in the chronic phase of recovery following MTBI. APOE e4 could be incorporated into screening tools to predict SMVs at risk for poor long-term neurobehavioral outcome in an effort to provide early intervention to improve long-term clinical outcome.
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http://dx.doi.org/10.1016/j.bbr.2021.113491DOI Listing
October 2021

Genetic regulation of nonsense-mediated decay underlies association with risk of severe COVID-19.

medRxiv 2021 Jul 13. Epub 2021 Jul 13.

Genomic regions have been associated with COVID-19 susceptibility and outcomes, including the chr12q24.13 locus encoding antiviral proteins OAS1-3. Here, we report genetic, functional, and clinical insights into genetic associations within this locus. In Europeans, the risk of hospitalized vs. non-hospitalized COVID-19 was associated with a single 19Kb-haplotype comprised of 76 variants included in a 95% credible set within a large genomic fragment introgressed from Neandertals. The risk haplotype was also associated with impaired spontaneous but not treatment-induced SARS-CoV-2 clearance in a clinical trial with pegIFN-λ1. We demonstrate that two exonic variants, rs10774671 and rs1131454, affect splicing and nonsense-mediated decay of . We suggest that genetically-regulated loss of expression contributes to impaired spontaneous clearance of SARS-CoV-2 and elevated risk of hospitalization for COVID-19. Our results provide the rationale for further clinical studies using interferons to compensate for impaired spontaneous SARS-CoV-2 clearance, particularly in carriers of the risk haplotypes.
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http://dx.doi.org/10.1101/2021.07.09.21260221DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8288155PMC
July 2021

Personalized Single-Cell Proteogenomics to Distinguish Acute Myeloid Leukemia from Non-Malignant Clonal Hematopoiesis.

Blood Cancer Discov 2021 07 25;2(4):319-325. Epub 2021 May 25.

Laboratory of Myeloid Malignancies, Hematology Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892 United States.

Genetic mutations associated with acute myeloid leukemia (AML) also occur in age-related clonal hematopoiesis, often in the same individual. This makes confident assignment of detected variants to malignancy challenging. The issue is particularly crucial for AML post-treatment measurable residual disease monitoring, where results can be discordant between genetic sequencing and flow cytometry. We show here, that it is possible to distinguish AML from clonal hematopoiesis and to resolve the immunophenotypic identity of clonal architecture. To achieve this, we first design patient-specific DNA probes based on patient's whole-genome sequencing, and then use them for patient-personalized single-cell DNA sequencing with simultaneous single-cell antibody-oligonucleotide sequencing. Examples illustrate AML arising from and mutated clones as well as independently. The ability to personalize single-cell proteogenomic assessment for individual patients based on leukemia-specific genomic features has implications for ongoing AML precision medicine efforts.
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http://dx.doi.org/10.1158/2643-3230.BCD-21-0046DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8265308PMC
July 2021

Survival motor neuron deficiency slows myoblast fusion through reduced myomaker and myomixer expression.

J Cachexia Sarcopenia Muscle 2021 Aug 11;12(4):1098-1116. Epub 2021 Jun 11.

Department of Anatomy, Physiology, and Genetics, Uniformed Services University of the Health Sciences, F. Edward Hébert School of Medicine, Bethesda, MD, USA.

Background: Spinal muscular atrophy is an inherited neurodegenerative disease caused by insufficient levels of the survival motor neuron (SMN) protein. Recently approved treatments aimed at increasing SMN protein levels have dramatically improved patient survival and have altered the disease landscape. While restoring SMN levels slows motor neuron loss, many patients continue to have smaller muscles and do not achieve normal motor milestones. While timing of treatment is important, it remains unclear why SMN restoration is insufficient to fully restore muscle size and function. We and others have shown that SMN-deficient muscle precursor cells fail to efficiently fuse into myotubes. However, the role of SMN in myoblast fusion is not known.

Methods: In this study, we show that SMN-deficient myoblasts readily fuse with wild-type myoblasts, demonstrating fusion competency. Conditioned media from wild type differentiating myoblasts do not rescue the fusion deficit of SMN-deficient cells, suggesting that compromised fusion may primarily be a result of altered membrane dynamics at the cell surface. Transcriptome profiling of skeletal muscle from SMN-deficient mice revealed altered expression of cell surface fusion molecules. Finally, using cell and mouse models, we investigate if myoblast fusion can be rescued in SMN-deficient myoblast and improve the muscle pathology in SMA mice.

Results: We found reduced expression of the muscle fusion proteins myomaker (P = 0.0060) and myomixer (P = 0.0051) in the muscle of SMA mice. Suppressing SMN expression in C2C12 myoblast cells reduces expression of myomaker (35% reduction; P < 0.0001) and myomixer, also known as myomerger and minion, (30% reduction; P < 0.0001) and restoring SMN levels only partially restores myomaker and myomixer expression. Ectopic expression of myomixer improves myofibre number (55% increase; P = 0.0006) and motor function (35% decrease in righting time; P = 0.0089) in SMA model mice and enhances motor function (82% decrease in righting time; P < 0.0001) and extends survival (28% increase; P < 0.01) when administered in combination with an antisense oligonucleotide that increases SMN protein levels.

Conclusions: Here, we identified reduced expression of muscle fusion proteins as a key factor in the fusion deficits of SMN-deficient myoblasts. This discovery provides a novel target to improve SMA muscle pathology and motor function, which in combination with SMN increasing therapy could enhance clinical outcomes for SMA patients.
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http://dx.doi.org/10.1002/jcsm.12740DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8350220PMC
August 2021

Prospective Assessment of SARS-CoV-2 Seroconversion (PASS) study: an observational cohort study of SARS-CoV-2 infection and vaccination in healthcare workers.

BMC Infect Dis 2021 Jun 9;21(1):544. Epub 2021 Jun 9.

Infectious Disease Clinical Research Program, Department of Preventive Medicine and Biostatistics, Uniformed Services University of the Health Sciences, Bethesda, MD, USA.

Background: SARS-CoV-2 is a recently emerged pandemic coronavirus (CoV) capable of causing severe respiratory illness. However, a significant number of infected people present as asymptomatic or pauci-symptomatic. In this prospective assessment of at-risk healthcare workers (HCWs) we seek to determine whether pre-existing antibody or T cell responses to previous seasonal human coronavirus (HCoV) infections affect immunological or clinical responses to SARS-CoV-2 infection or vaccination.

Methods: A cohort of 300 healthcare workers, confirmed negative for SARS-CoV-2 exposure upon study entry, will be followed for up to 1 year with monthly serology analysis of IgM and IgG antibodies against the spike proteins of SARS-CoV-2 and the four major seasonal human coronavirus - HCoV-OC43, HCoV-HKU1, HCoV-229E, and HCoV-NL63. Participants will complete monthly questionnaires that ask about Coronavirus Disease 2019 (COVID-19) exposure risks, and a standardized, validated symptom questionnaire (scoring viral respiratory disease symptoms, intensity and severity) at least twice monthly and any day when any symptoms manifest. SARS-CoV-2 PCR testing will be performed any time participants develop symptoms consistent with COVID-19. For those individuals that seroconvert and/or test positive by SARS-CoV-2 PCR, or receive the SARS-CoV-2 vaccine, additional studies of T cell activation and cytokine production in response to SARS-CoV-2 peptide pools and analysis of Natural Killer cell numbers and function will be conducted on that participant's cryopreserved baseline peripheral blood mononuclear cells (PBMCs). Following the first year of this study we will further analyze those participants having tested positive for COVID-19, and/or having received an authorized/licensed SARS-CoV-2 vaccine, quarterly (year 2) and semi-annually (years 3 and 4) to investigate immune response longevity.

Discussion: This study will determine the frequency of asymptomatic and pauci-symptomatic SARS-CoV-2 infection in a cohort of at-risk healthcare workers. Baseline and longitudinal assays will determine the frequency and magnitude of anti-spike glycoprotein antibodies to the seasonal HCoV-OC43, HCoV-HKU1, HCoV-229E, and HCoV-NL63, and may inform whether pre-existing antibodies to these human coronaviruses are associated with altered COVID-19 disease course. Finally, this study will evaluate whether pre-existing immune responses to seasonal HCoVs affect the magnitude and duration of antibody and T cell responses to SARS-CoV-2 vaccination, adjusting for demographic covariates.
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http://dx.doi.org/10.1186/s12879-021-06233-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8188741PMC
June 2021

Tensin 1 (TNS1) is a modifier gene for low body mass index (BMI) in homozygous [F508del]CFTR patients.

Physiol Rep 2021 Jun;9(11):e14886

The Collaborative Health Initiative Research Program, Uniformed Services University of the Health Sciences, Bethesda, MD, USA.

Cystic fibrosis (CF) is a life-limiting autosomal recessive genetic disease caused by variants in the CFTR gene, most commonly by the [F508del] variant. Although CF is a classical Mendelian disease, genetic variants in several modifier genes have been associated with variation of the clinical phenotype for pulmonary and gastrointestinal function and urogenital development. We hypothesized that whole genome sequencing of a well-phenotyped CF populations might identify novel variants in known, or hitherto unknown, modifier genes. Whole genome sequencing was performed on the Illumina HiSeq X platform for 98 clinically diagnosed cystic fibrosis patient samples from the Adult CF Clinic at the University of California San Diego (UCSD). We compared protein-coding, non-silent variants genome wide between CFTR [F508del] homozygotes vs CFTR compound heterozygotes. Based on a single variant score test, we found 3 SNPs in common variants (MAF >5%) that occurred at significantly different rates between homozygous [F508del]CFTR and compound heterozygous [F508del]CFTR patients. The 3 SNPs were all located in one gene on chromosome 2: Tensin 1 (TNS1: rs3796028; rs2571445: and rs918949). We observed significantly lower BMIs in homozygous [F508del]CFTR patients who were also homozygous for Tensin 1 rs918949 (T/T) (p = 0.023) or rs2571445 (G/G) (p = 0.02) variants. The Tensin 1 gene is thus a potential modifier gene for low BMI in CF patients homozygous for the [F508del]CFTR variant.
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http://dx.doi.org/10.14814/phy2.14886DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8176904PMC
June 2021

Accelerating Medicines Partnership: Parkinson's Disease. Genetic Resource.

Mov Disord 2021 08 7;36(8):1795-1804. Epub 2021 May 7.

Center for Alzheimer's and Related Dementias, National Institute on Aging, Bethesda, Maryland, USA.

Background: Whole-genome sequencing data are available from several large studies across a variety of diseases and traits. However, massive storage and computation resources are required to use these data, and to achieve sufficient power for discoveries, harmonization of multiple cohorts is critical.

Objectives: The Accelerating Medicines Partnership Parkinson's Disease program has developed a research platform for Parkinson's disease (PD) that integrates the storage and analysis of whole-genome sequencing data, RNA expression data, and clinical data, harmonized across multiple cohort studies.

Methods: The version 1 release contains whole-genome sequencing data derived from 3941 participants from 4 cohorts. Samples underwent joint genotyping by the TOPMed Freeze 9 Variant Calling Pipeline. We performed descriptive analyses of these whole-genome sequencing data using the Accelerating Medicines Partnership Parkinson's Disease platform.

Results: The clinical diagnosis of participants in version 1 release includes 2005 idiopathic PD patients, 963 healthy controls, 64 prodromal subjects, 62 clinically diagnosed PD subjects without evidence of dopamine deficit, and 705 participants of genetically enriched cohorts carrying PD risk-associated GBA variants or LRRK2 variants, of whom 304 were affected. We did not observe significant enrichment of pathogenic variants in the idiopathic PD group, but the polygenic risk score was higher in PD both in nongenetically enriched cohorts and genetically enriched cohorts. The population analysis showed a correlation between genetically enriched cohorts and Ashkenazi Jewish ancestry.

Conclusions: We describe the genetic component of the Accelerating Medicines Partnership Parkinson's Disease platform, a solution to democratize data access and analysis for the PD research community. © 2021 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society. This article is a U.S. Government work and is in the public domain in the USA.
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http://dx.doi.org/10.1002/mds.28549DOI Listing
August 2021

Proteogenomic landscape of uterine leiomyomas from hereditary leiomyomatosis and renal cell cancer patients.

Sci Rep 2021 04 30;11(1):9371. Epub 2021 Apr 30.

Gynecologic Cancer Center of Excellence, Department of Gynecologic Surgery and Obstetrics, Uniformed Services University of the Health Sciences and Walter Reed National Military Medical Center, 8901 Wisconsin Avenue, Bethesda, MD, 20889, USA.

Pathogenic mutations in fumarate hydratase (FH) drive hereditary leiomyomatosis and renal cell cancer (HLRCC) and increase the risk of developing uterine leiomyomas (ULMs). An integrated proteogenomic analysis of ULMs from HLRCC (n = 16; FH-mutation confirmed) and non-syndromic (NS) patients (n = 12) identified a significantly higher protein:transcript correlation in HLRCC (R = 0.35) vs. NS ULMs (R = 0.242, MWU p = 0.0015). Co-altered proteins and transcripts (228) included antioxidant response element (ARE) target genes, such as thioredoxin reductase 1 (TXNRD1), and correlated with activation of NRF2-mediated oxidative stress response signaling in HLRCC ULMs. We confirm 185 transcripts previously described as altered between HLRCC and NS ULMs, 51 co-altered at the protein level and several elevated in HLRCC ULMs are involved in regulating cellular metabolism and glycolysis signaling. Furthermore, 367 S-(2-succino)cysteine peptides were identified in HLRCC ULMs, of which sixty were significantly elevated in HLRCC vs. NS ULMs (LogFC = 1.86, MWU p < 0.0001). These results confirm and define novel proteogenomic alterations in uterine leiomyoma tissues collected from HLRCC patients and underscore conserved molecular alterations correlating with inactivation of the FH tumor suppressor gene.
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http://dx.doi.org/10.1038/s41598-021-88585-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8087684PMC
April 2021

Linkage of Alzheimer disease families with Puerto Rican ancestry identifies a chromosome 9 locus.

Neurobiol Aging 2021 08 28;104:115.e1-115.e7. Epub 2021 Feb 28.

John P. Hussman Institute for Human Genomics, University of Miami Miller School of Medicine, Miami, FL, USA; Dr. John T. Macdonald Foundation Department of Human Genetics, University of Miami Miller School of Medicine, Miami, FL, USA.

The genetic admixture of Caribbean Hispanics provides an opportunity to discover novel genetic factors in Alzheimer disease (AD). We sought to identify genetic variants for AD through a family-based design using the Puerto Rican (PR) Alzheimer Disease Initiative (PRADI). Whole-genome sequencing (WGS) and parametric linkage analysis were performed for 100 individuals from 23 multiplex PRADI families. Variants were prioritized by minor allele frequency (<0.01), functional potential [combined annotation dependent depletion score (CADD) >10], and co-segregation with AD. Variants were further ranked using an independent PR case-control WGS dataset (PR10/66). A genome-wide significant linkage peak was found in 9p21 with a heterogeneity logarithm of the odds score (HLOD) >5.1, which overlaps with an AD linkage region from two published independent studies. The region harbors C9orf72, but no expanded repeats were observed in the families. Seven variants prioritized by the PRADI families also displayed evidence for association in the PR10/66 (p < 0.05), including a missense variant in UNC13B. Our study demonstrated the importance of family-based design and WGS in genetic study of AD.
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http://dx.doi.org/10.1016/j.neurobiolaging.2021.02.019DOI Listing
August 2021

Transcriptomic Analysis of Mouse Brain After Traumatic Brain Injury Reveals That the Angiotensin Receptor Blocker Candesartan Acts Through Novel Pathways.

Front Neurosci 2021 22;15:636259. Epub 2021 Mar 22.

Graduate Program in Neuroscience, Uniformed Services University of the Health Sciences, Bethesda, MD, United States.

Traumatic brain injury (TBI) results in complex pathological reactions, where the initial lesion is followed by secondary inflammation and edema. Our laboratory and others have reported that angiotensin receptor blockers (ARBs) have efficacy in improving recovery from traumatic brain injury in mice. Treatment of mice with a subhypotensive dose of the ARB candesartan results in improved functional recovery, and reduced pathology (lesion volume, inflammation and gliosis). In order to gain a better understanding of the molecular mechanisms through which candesartan improves recovery after controlled cortical impact injury (CCI), we performed transcriptomic profiling on brain regions after injury and drug treatment. We examined RNA expression in the ipsilateral hippocampus, thalamus and hypothalamus at 3 or 29 days post injury (dpi) treated with either candesartan (0.1 mg/kg) or vehicle. RNA was isolated and analyzed by bulk mRNA-seq. Gene expression in injured and/or candesartan treated brain region was compared to that in sham vehicle treated mice in the same brain region to identify genes that were differentially expressed (DEGs) between groups. The most DEGs were expressed in the hippocampus at 3 dpi, and the number of DEGs reduced with distance and time from the lesion. Among pathways that were differentially expressed at 3 dpi after CCI, candesartan treatment altered genes involved in angiogenesis, interferon signaling, extracellular matrix regulation including integrins and chromosome maintenance and DNA replication. At 29 dpi, candesartan treatment reduced the expression of genes involved in the inflammatory response. Some changes in gene expression were confirmed in a separate cohort of animals by qPCR. Fewer DEGs were found in the thalamus, and only one in the hypothalamus at 3 dpi. Additionally, in the hippocampi of sham injured mice, 3 days of candesartan treatment led to the differential expression of 384 genes showing that candesartan in the absence of injury had a powerful impact on gene expression specifically in the hippocampus. Our results suggest that candesartan has broad actions in the brain after injury and affects different processes at acute and chronic times after injury. These data should assist in elucidating the beneficial effect of candesartan on recovery from TBI.
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http://dx.doi.org/10.3389/fnins.2021.636259DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8019829PMC
March 2021

Transcriptomics of Long-Term Meditation Practice: Evidence for Prevention or Reversal of Stress Effects Harmful to Health.

Medicina (Kaunas) 2021 Mar 1;57(3). Epub 2021 Mar 1.

Department of Physiology and Health, Maharishi International University, Fairfield, IA 52556, USA.

Stress can overload adaptive mechanisms, leading to epigenetic effects harmful to health. Research on the reversal of these effects is in its infancy. Early results suggest some meditation techniques have health benefits that grow with repeated practice. This study focused on possible transcriptomic effects of 38 years of twice-daily Transcendental Meditation (TM) practice. First, using Illumina BeadChip microarray technology, differences in global gene expression in peripheral blood mononuclear cells (PBMCs) were sought between healthy practitioners and tightly matched controls ( = 12, age 65). Second, these microarray results were verified on a subset of genes using quantitative polymerase chain reaction (qPCR) and were validated using qPCR in larger TM and control groups ( = 45, age 63). Bioinformatics investigation employed Ingenuity Pathway Analysis (IPA), DAVID, Genomatix, and R packages. The 200 genes and loci found to meet strict criteria for differential expression in the microarray experiment showed contrasting patterns of expression that distinguished the two groups. Differential expression relating to immune function and energy efficiency were most apparent. In the TM group, relative to the control, all 49 genes associated with inflammation were downregulated, while genes associated with antiviral and antibody components of the defense response were upregulated. The largest expression differences were shown by six genes related to erythrocyte function that appeared to reflect a condition of lower energy efficiency in the control group. Results supporting these gene expression differences were obtained with qPCR-measured expression both in the well-matched microarray groups and in the larger, less well-matched groups. These findings are consistent with predictions based on results from earlier randomized trials of meditation and may provide evidence for stress-related molecular mechanisms underlying reductions in anxiety, post-traumatic stress disorder (PTSD), cardiovascular disease (CVD), and other chronic disorders and diseases.
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http://dx.doi.org/10.3390/medicina57030218DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8001870PMC
March 2021

Genome sequencing analysis identifies new loci associated with Lewy body dementia and provides insights into its genetic architecture.

Nat Genet 2021 03 15;53(3):294-303. Epub 2021 Feb 15.

Reta Lila Weston Institute, UCL Queen Square Institute of Neurology, University College London, London, UK.

The genetic basis of Lewy body dementia (LBD) is not well understood. Here, we performed whole-genome sequencing in large cohorts of LBD cases and neurologically healthy controls to study the genetic architecture of this understudied form of dementia, and to generate a resource for the scientific community. Genome-wide association analysis identified five independent risk loci, whereas genome-wide gene-aggregation tests implicated mutations in the gene GBA. Genetic risk scores demonstrate that LBD shares risk profiles and pathways with Alzheimer's disease and Parkinson's disease, providing a deeper molecular understanding of the complex genetic architecture of this age-related neurodegenerative condition.
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http://dx.doi.org/10.1038/s41588-021-00785-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7946812PMC
March 2021

Response of human macrophages to gamma radiation is mediated via expression of endogenous retroviruses.

PLoS Pathog 2021 02 8;17(2):e1009305. Epub 2021 Feb 8.

Department of Pharmacology & Molecular Therapeutics, Uniformed Services University of the Health Sciences, Bethesda, Maryland, United States of America.

Ionizing radiation-induced tissue damage recruits monocytes into the exposed area where they are differentiated to macrophages. These implement phagocytic removal of dying cells and elicit an acute inflammatory response, but can also facilitate tumorigenesis due to production of anti-inflammatory cytokines. Using primary human monocyte-derived macrophages (MDMs) and the THP1 monocytic cell line, we demonstrate that gamma radiation triggers monocyte differentiation toward the macrophage phenotype with increased expression of type I interferons (IFN-I) and both pro- and anti-inflammatory macrophage activation markers. We found that these changes correlate with significantly upregulated expression of 622 retroelements from various groups, particularly of several clades of human endogenous retroviruses (HERVs). Elevated transcription was detected in both sense and antisense directions in the HERV subgroups tested, including the most genetically homogeneous clade HML-2. The level of antisense transcription was three- to five-fold higher than of the sense strand levels. Using a proximity ligation assay and immunoprecipitation followed by RNA quantification, we identified an increased amount of the dsRNA receptors MDA-5 and TLR3 bound to an equivalent number of copies of sense and antisense chains of HERVK HML-2 RNA. This binding triggered MAVS-associated signaling pathways resulting in increased expression of IFN-I and inflammation related genes that enhanced the cumulative inflammatory effect of radiation-induced senescence. HML-2 knockdown was accompanied with reduced expression and secretion of IFNα, pro-inflammatory (IL-1β, IL-6, CCL2, CCL3, CCL8, and CCL20) and anti-inflammatory (IL10) modulators in irradiated monocytes and MDMs. Taken together, our data indicate that radiation stress-induced HERV expression enhances the IFN-I and cytokine response and results in increased levels of pro-inflammatory modulators along with expression of anti-inflammatory factors associated with the macrophage tumorigenic phenotype.
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http://dx.doi.org/10.1371/journal.ppat.1009305DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7895352PMC
February 2021

Increased APOE ε4 expression is associated with the difference in Alzheimer's disease risk from diverse ancestral backgrounds.

Alzheimers Dement 2021 07 1;17(7):1179-1188. Epub 2021 Feb 1.

John P. Hussman Institute for Human Genomics, Miller School of Medicine, University of Miami, Miami, Florida, USA.

Introduction: Apolipoprotein E (APOE) ε4 confers less risk for Alzheimer's disease (AD) in carriers with African local genomic ancestry (ALA) than APOE ε4 carriers with European local ancestry (ELA). Cell type specific transcriptional variation between the two local ancestries (LAs) could contribute to this disease risk differences.

Methods: Single-nucleus RNA sequencing was performed on frozen frontal cortex of homozygous APOE ε4/ε4 AD patients: seven with ELA, four with ALA.

Results: A total of 60,908 nuclei were sequenced. Within the LA region (chr19:44-46Mb), APOE was the gene most differentially expressed, with ELA carriers having significantly more expression (overall P < 1.8E ) in 24 of 32 cell clusters. The transcriptome of one astrocyte cluster, with high APOE ε4 expression and specific to ELA, is suggestive of A1 reactive astrocytes.

Discussion: AD patients with ELA expressed significantly greater levels of APOE than ALA APOE ε4 carriers. These differences in APOE expression could contribute to the reduced risk for AD seen in African APOE ε4 carriers.
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http://dx.doi.org/10.1002/alz.12287DOI Listing
July 2021

Broadly effective metabolic and immune recovery with C5 inhibition in CHAPLE disease.

Nat Immunol 2021 02 4;22(2):128-139. Epub 2021 Jan 4.

Clinical Genomics Program, NIAID, NIH, Bethesda, MD, USA.

Complement hyperactivation, angiopathic thrombosis and protein-losing enteropathy (CHAPLE disease) is a lethal disease caused by genetic loss of the complement regulatory protein CD55, leading to overactivation of complement and innate immunity together with immunodeficiency due to immunoglobulin wasting in the intestine. We report in vivo human data accumulated using the complement C5 inhibitor eculizumab for the medical treatment of patients with CHAPLE disease. We observed cessation of gastrointestinal pathology together with restoration of normal immunity and metabolism. We found that patients rapidly renormalized immunoglobulin concentrations and other serum proteins as revealed by aptamer profiling, re-established a healthy gut microbiome, discontinued immunoglobulin replacement and other treatments and exhibited catch-up growth. Thus, we show that blockade of C5 by eculizumab effectively re-establishes regulation of the innate immune complement system to substantially reduce the pathophysiological manifestations of CD55 deficiency in humans.
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http://dx.doi.org/10.1038/s41590-020-00830-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7856263PMC
February 2021

Genetic determinants of survival in progressive supranuclear palsy: a genome-wide association study.

Lancet Neurol 2021 02 17;20(2):107-116. Epub 2020 Dec 17.

Department of Clinical and Movement Neurosciences, University College London Queen Square Institute of Neurology, London, UK; Movement Disorders Centre, University College London Queen Square Institute of Neurology, London, UK. Electronic address:

Background: The genetic basis of variation in the progression of primary tauopathies has not been determined. We aimed to identify genetic determinants of survival in progressive supranuclear palsy (PSP).

Methods: In stage one of this two stage genome-wide association study (GWAS), we included individuals with PSP, diagnosed according to pathological and clinical criteria, from two separate cohorts: the 2011 PSP GWAS cohort, from brain banks based at the Mayo Clinic (Jacksonville, FL, USA) and in Munich (Germany), and the University College London PSP cohort, from brain banks and the PROSPECT study, a UK-wide longitudinal study of patients with atypical parkinsonian syndromes. Individuals were included if they had clinical data available on sex, age at motor symptom onset, disease duration (from motor symptom onset to death or to the date of censoring, Dec 1, 2019, if individuals were alive), and PSP phenotype (with reference to the 2017 Movement Disorder Society criteria). Genotype data were used to do a survival GWAS using a Cox proportional hazards model. In stage two, data from additional individuals from the Mayo Clinic brain bank, which were obtained after the 2011 PSP GWAS, were used for a pooled analysis. We assessed the expression quantitative trait loci (eQTL) profile of variants that passed genome-wide significance in our GWAS using the Functional Mapping and Annotation of GWAS platform, and did colocalisation analyses using the eQTLGen and PsychENCODE datasets.

Findings: Data were collected and analysed between Aug 1, 2016, and Feb 1, 2020. Data were available for 1001 individuals of white European ancestry with PSP in stage one. We found a genome-wide significant association with survival at chromosome 12 (lead single nucleotide polymorphism rs2242367, p=7·5 × 10, hazard ratio 1·42 [95% CI 1·22-1·67]). rs2242367 was associated with survival in the individuals added in stage two (n=238; p=0·049, 1·22 [1·00-1·48]) and in the pooled analysis of both stages (n=1239; p=1·3 × 10, 1·37 [1·25-1·51]). An eQTL database screen revealed that rs2242367 is associated with increased expression of LRRK2 and two long intergenic non-coding RNAs (lncRNAs), LINC02555 and AC079630.4, in whole blood. Although we did not detect a colocalisation signal for LRRK2, analysis of the PSP survival signal and eQTLs for LINC02555 in the eQTLGen blood dataset revealed a posterior probability of hypothesis 4 of 0·77, suggesting colocalisation due to a single shared causal variant.

Interpretation: Genetic variation at the LRRK2 locus was associated with survival in PSP. The mechanism of this association might be through a lncRNA-regulated effect on LRRK2 expression because LINC02555 has previously been shown to regulate LRRK2 expression. LRRK2 has been associated with sporadic and familial forms of Parkinson's disease, and our finding suggests a genetic overlap with PSP. Further functional studies will be important to assess the potential of LRRK2 modulation as a disease-modifying therapy for PSP and related tauopathies.

Funding: PSP Association, CBD Solutions, Medical Research Council (UK).
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http://dx.doi.org/10.1016/S1474-4422(20)30394-XDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7116626PMC
February 2021

Genome-Wide Analysis of Off-Target CRISPR/Cas9 Activity in Single-Cell-Derived Human Hematopoietic Stem and Progenitor Cell Clones.

Genes (Basel) 2020 12 13;11(12). Epub 2020 Dec 13.

Cellular and Molecular Therapeutics Branch, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA.

CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9)-mediated genome editing holds remarkable promise for the treatment of human genetic diseases. However, the possibility of off-target Cas9 activity remains a concern. To address this issue using clinically relevant target cells, we electroporated Cas9 ribonucleoprotein (RNP) complexes (independently targeted to two different genomic loci, the locus on chromosome 2 and the locus on chromosome 19) into human mobilized peripheral blood-derived hematopoietic stem and progenitor cells (HSPCs) and assessed the acquisition of somatic mutations in an unbiased, genome-wide manner via whole genome sequencing (WGS) of single-cell-derived HSPC clones. Bioinformatic analysis identified >20,000 total somatic variants (indels, single nucleotide variants, and structural variants) distributed among Cas9-treated and non-Cas9-treated control HSPC clones. Statistical analysis revealed no significant difference in the number of novel non-targeted indels among the samples. Moreover, data analysis showed no evidence of Cas9-mediated indel formation at 623 predicted off-target sites. The median number of novel single nucleotide variants was slightly elevated in Cas9 RNP-recipient sample groups compared to baseline, but did not reach statistical significance. Structural variants were rare and demonstrated no clear causal connection to Cas9-mediated gene editing procedures. We find that the collective somatic mutational burden observed within Cas9 RNP-edited human HSPC clones is indistinguishable from naturally occurring levels of background genetic heterogeneity.
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http://dx.doi.org/10.3390/genes11121501DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7762975PMC
December 2020

Pathogenic Huntingtin Repeat Expansions in Patients with Frontotemporal Dementia and Amyotrophic Lateral Sclerosis.

Neuron 2021 02 26;109(3):448-460.e4. Epub 2020 Nov 26.

Molecular Markers Laboratory, IRCCS Istituto Centro San Giovanni di Dio Fatebenefratelli, Brescia 25125, Italy; MAC Memory Clinic, IRCCS Istituto Centro San Giovanni di Dio Fatebenefratelli, Brescia 25125, Italy.

We examined the role of repeat expansions in the pathogenesis of frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) by analyzing whole-genome sequence data from 2,442 FTD/ALS patients, 2,599 Lewy body dementia (LBD) patients, and 3,158 neurologically healthy subjects. Pathogenic expansions (range, 40-64 CAG repeats) in the huntingtin (HTT) gene were found in three (0.12%) patients diagnosed with pure FTD/ALS syndromes but were not present in the LBD or healthy cohorts. We replicated our findings in an independent collection of 3,674 FTD/ALS patients. Postmortem evaluations of two patients revealed the classical TDP-43 pathology of FTD/ALS, as well as huntingtin-positive, ubiquitin-positive aggregates in the frontal cortex. The neostriatal atrophy that pathologically defines Huntington's disease was absent in both cases. Our findings reveal an etiological relationship between HTT repeat expansions and FTD/ALS syndromes and indicate that genetic screening of FTD/ALS patients for HTT repeat expansions should be considered.
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http://dx.doi.org/10.1016/j.neuron.2020.11.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7864894PMC
February 2021

Mutational Analysis of Known ALS Genes in an Italian Population-Based Cohort.

Neurology 2021 01 18;96(4):e600-e609. Epub 2020 Nov 18.

From "Rita Levi Montalcini" Department of Neuroscience (M.G., A. Calvo, C.M., A. Canosa, U.M., R.V., A. Chiò), University of Turin, Italy; Biocomputational Group (J.D., R.J.G.) and Neuromuscular Diseases Research Section (M.G., R.C., B.J.T.), Laboratory of Neurogenetics, National Institute on Aging, NIH, Porter Neuroscience Research Center, Bethesda, MD; Laboratory of Genetics, Department of Clinical Pathology (M. Brunetti, M. Barberis, L.S.), Azienda Ospedaliero Universitaria Città della Salute e della Scienza di Torino, Turin; Department of Health Sciences Interdisciplinary Research Center of Autoimmune Diseases (L.C., S.D.), "Amedeo Avogadro" University of Eastern Piedmont; ALS Center (L.M.), Department of Neurology, Azienda Ospedaliera Universitaria Maggiore della Carità, Novara, Italy; Neurodegenerative Diseases Research Unit, Laboratory of Neurogenetics (S.W.S.), National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda; Department of Neurology (S.W.S., B.J.T.), Johns Hopkins University Medical Center; Department of Anatomy, Physiology & Genetics (C.D.), and The American Genome Center, Collaborative Health Initiative Research Program (C.D.), Uniformed Services University of the Health Sciences, Bethesda, MD; and Institute of Cognitive Sciences and Technologies (A. Chiò), National Council of Research, Rome, Italy.

Objective: To assess the burden of rare genetic variants and to estimate the contribution of known amyotrophic lateral sclerosis (ALS) genes in an Italian population-based cohort, we performed whole genome sequencing in 959 patients with ALS and 677 matched healthy controls.

Methods: We performed genome sequencing in a population-based cohort (Piemonte and Valle d'Aosta Registry for ALS [PARALS]). A panel of 40 ALS genes was analyzed to identify potential disease-causing genetic variants and to evaluate the gene-wide burden of rare variants among our population.

Results: A total of 959 patients with ALS were compared with 677 healthy controls from the same geographical area. Gene-wide association tests demonstrated a strong association with , whose rare variants are the second most common cause of disease after expansion. A lower signal was observed for , proving that its effect on our cohort is driven by a few known causal variants. We detected rare variants in other known ALS genes that did not surpass statistical significance in gene-wise tests, thus highlighting that their contribution to disease risk in our cohort is limited.

Conclusions: We identified potential disease-causing variants in 11.9% of our patients. We identified the genes most frequently involved in our cohort and confirmed the contribution of rare variants in disease risk. Our results provide further insight into the pathologic mechanism of the disease and demonstrate the importance of genome-wide sequencing as a diagnostic tool.
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http://dx.doi.org/10.1212/WNL.0000000000011209DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7905787PMC
January 2021

Identification of a pathogenic intronic KIF5A mutation in an ALS-FTD kindred.

Neurology 2020 12 19;95(22):1015-1018. Epub 2020 Oct 19.

From the Neuromuscular Diseases Research Section (S.S-A, B.J.T.), Laboratory of Neurogenetics, NIA, NIH; Department of Anatomy (C.L.D.), Physiology & Genetics, Uniformed Services University of the Health Sciences; The American Genome Center (C.L.D., C.A., D.N.H., M.D.W.), Collaborative Health Initiative Research Program, Uniformed Services University of the Health Sciences; Computational Biology Core (J.D.), Laboratory of Neurogenetics, NIA, NIH, Bethesda, MD; Rita Levi Montalcini Department of Neuroscience (A.C.), University of Turin, Italy and AOU Città della Salute e della Scienza, Turin, Italy; Division of Neurology and Neurobiology (R. Bowser), Barrow Neurological Institute, Phoenix, AZ; Department of Neurology (E.P.P.), Neuromuscular Center, Neurological Institute, Cleveland Clinic, OH; Department of Neurology (R. Bedlack), Duke University and the Durham VA Medical Center, NC; and Department of Neurology (B.J.T.), Johns Hopkins University, Baltimore, MD.

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http://dx.doi.org/10.1212/WNL.0000000000011064DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7734922PMC
December 2020

Torin 2 Derivative, NCATS-SM3710, Has Potent Multistage Antimalarial Activity through Inhibition of Phosphatidylinositol 4-Kinase ( PI4KIIIβ).

ACS Pharmacol Transl Sci 2020 Oct 11;3(5):948-964. Epub 2020 Sep 11.

Department of Microbiology and Immunology, Uniformed Services University of the Health Sciences, Bethesda, Maryland 20814, United States.

Drug resistance is a constant threat to malaria control efforts making it important to maintain a good pipeline of new drug candidates. Of particular need are compounds that also block transmission by targeting sexual stage parasites. Mature sexual stages are relatively resistant to all currently used antimalarials except the 8-aminoquinolines that are not commonly used due to potential side effects. Here, we synthesized a new Torin 2 derivative, NCATS-SM3710 with increased aqueous solubility and specificity for and demonstrate potent activity against all life cycle stages. NCATS-SM3710 also has low nanomolar ECs against cultured asexual parasites (0.38 ± 0.04 nM) and late stage gametocytes (5.77 ± 1 nM). Two independent NCATS-SM3710/Torin 2 resistant parasite lines produced by growth in sublethal Torin 2 concentrations both had genetic changes in PF3D7_0509800, annotated as a phosphatidylinositol 4 kinase ( PI4KIIIβ). One line had a point mutation in the putative active site (V1357G), and the other line had a duplication of a locus containing  PI4KIIIβ. Both lines were also resistant to other  PI4K inhibitors. In addition NCATS-SM3710 inhibited purified  PI4KIIIβ with an IC of 2.0 ± 0.30 nM. Together the results demonstrate that  PI4KIIIβ is the target of Torin 2 and NCATS-SM3710 and provide new options for potent multistage drug development.
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http://dx.doi.org/10.1021/acsptsci.0c00078DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7551716PMC
October 2020

Inborn errors of type I IFN immunity in patients with life-threatening COVID-19.

Science 2020 10 24;370(6515). Epub 2020 Sep 24.

Laboratory of Virology and Infectious Disease, The Rockefeller University, New York, NY, USA.

Clinical outcome upon infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) ranges from silent infection to lethal coronavirus disease 2019 (COVID-19). We have found an enrichment in rare variants predicted to be loss-of-function (LOF) at the 13 human loci known to govern Toll-like receptor 3 (TLR3)- and interferon regulatory factor 7 (IRF7)-dependent type I interferon (IFN) immunity to influenza virus in 659 patients with life-threatening COVID-19 pneumonia relative to 534 subjects with asymptomatic or benign infection. By testing these and other rare variants at these 13 loci, we experimentally defined LOF variants underlying autosomal-recessive or autosomal-dominant deficiencies in 23 patients (3.5%) 17 to 77 years of age. We show that human fibroblasts with mutations affecting this circuit are vulnerable to SARS-CoV-2. Inborn errors of TLR3- and IRF7-dependent type I IFN immunity can underlie life-threatening COVID-19 pneumonia in patients with no prior severe infection.
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http://dx.doi.org/10.1126/science.abd4570DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7857407PMC
October 2020

Distinguishing Smoking-Related Lung Disease Phenotypes Via Imaging and Molecular Features.

Chest 2021 02 16;159(2):549-563. Epub 2020 Sep 16.

Department of Medicine, Division of Pulmonary and Critical Care Medicine, Brigham and Women's Hospital, Boston, MA; Applied Chest Imaging Laboratory, Brigham and Women's Hospital, Boston, MA.

Background: Chronic tobacco smoke exposure results in a broad range of lung pathologies including emphysema, airway disease and parenchymal fibrosis as well as a multitude of extra-pulmonary comorbidities. Prior work using CT imaging has identified several clinically relevant subgroups of smoking related lung disease, but these investigations have generally lacked organ specific molecular correlates.

Research Question: Can CT imaging be used to identify clinical phenotypes of smoking related lung disease that have specific bronchial epithelial gene expression patterns to better understand disease pathogenesis?

Study Design And Methods: Using K-means clustering, we clustered participants from the COPDGene study (n = 5,273) based on CT imaging characteristics and then evaluated their clinical phenotypes. These clusters were replicated in the Detection of Early Lung Cancer Among Military Personnel (DECAMP) cohort (n = 360), and were further characterized using bronchial epithelial gene expression.

Results: Three clusters (preserved, interstitial predominant and emphysema predominant) were identified. Compared to the preserved cluster, the interstitial and emphysema clusters had worse lung function, exercise capacity and quality of life. In longitudinal follow-up, individuals from the emphysema group had greater declines in exercise capacity and lung function, more emphysema, more exacerbations, and higher mortality. Similarly, genes involved in inflammatory pathways (tumor necrosis factor-α, interferon-β) are more highly expressed in bronchial epithelial cells from individuals in the emphysema cluster, while genes associated with T-cell related biology are decreased in these samples. Samples from individuals in the interstitial cluster generally had intermediate levels of expression of these genes.

Interpretation: Using quantitative CT imaging, we identified three groups of individuals in older ever-smokers that replicate in two cohorts. Airway gene expression differences between the three groups suggests increased levels of inflammation in the most severe clinical phenotype, possibly mediated by the tumor necrosis factor-α and interferon-β pathways.

Clinical Trial Registration: COPDGene (NCT00608764), DECAMP-1 (NCT01785342), DECAMP-2 (NCT02504697).
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http://dx.doi.org/10.1016/j.chest.2020.08.2115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8039011PMC
February 2021

Magnitude and Dynamics of the T-Cell Response to SARS-CoV-2 Infection at Both Individual and Population Levels.

medRxiv 2020 Sep 17. Epub 2020 Sep 17.

Immunotherapy, Cell Therapy and Biobank (ITCB), Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS, Meldola, FC, Italy.

T cells are involved in the early identification and clearance of viral infections and also support the development of antibodies by B cells. This central role for T cells makes them a desirable target for assessing the immune response to SARS-CoV-2 infection. Here, we combined two high-throughput immune profiling methods to create a quantitative picture of the T-cell response to SARS-CoV-2. First, at the individual level, we deeply characterized 3 acutely infected and 58 recovered COVID-19 subjects by experimentally mapping their CD8 T-cell response through antigen stimulation to 545 Human Leukocyte Antigen (HLA) class I presented viral peptides (class II data in a forthcoming study). Then, at the population level, we performed T-cell repertoire sequencing on 1,815 samples (from 1,521 COVID-19 subjects) as well as 3,500 controls to identify shared "public" T-cell receptors (TCRs) associated with SARS-CoV-2 infection from both CD8 and CD4 T cells. Collectively, our data reveal that CD8 T-cell responses are often driven by a few immunodominant, HLA-restricted epitopes. As expected, the T-cell response to SARS-CoV-2 peaks about one to two weeks after infection and is detectable for at least several months after recovery. As an application of these data, we trained a classifier to diagnose SARS-CoV-2 infection based solely on TCR sequencing from blood samples, and observed, at 99.8% specificity, high early sensitivity soon after diagnosis (Day 3-7 = 85.1% [95% CI = 79.9-89.7]; Day 8-14 = 94.8% [90.7-98.4]) as well as lasting sensitivity after recovery (Day 29+/convalescent = 95.4% [92.1-98.3]). These results demonstrate an approach to reliably assess the adaptive immune response both soon after viral antigenic exposure (before antibodies are typically detectable) as well as at later time points. This blood-based molecular approach to characterizing the cellular immune response has applications in clinical diagnostics as well as in vaccine development and monitoring.
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http://dx.doi.org/10.1101/2020.07.31.20165647DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7418734PMC
September 2020

MutEnricher: a flexible toolset for somatic mutation enrichment analysis of tumor whole genomes.

BMC Bioinformatics 2020 Jul 31;21(1):338. Epub 2020 Jul 31.

The American Genome Center, Collaborative Health Initiative Research Program, Uniformed Services University of the Health Sciences, Bethesda, MD, USA.

Background: Analysis of somatic mutations from tumor whole exomes has fueled discovery of novel cancer driver genes. However, ~ 98% of the genome is non-coding and includes regulatory elements whose normal cellular functions can be disrupted by mutation. Whole genome sequencing (WGS), on the other hand, allows for identification of non-coding somatic variation and expanded estimation of background mutation rates, yet fewer computational tools exist for specific interrogation of this space.

Results: We present MutEnricher, a flexible toolset for investigating somatic mutation enrichment in both coding and non-coding genomic regions from WGS data. MutEnricher contains two distinct modules for these purposes that provide customizable options for calculating sample- and feature-specific background mutation rates. Additionally, both MutEnricher modules calculate feature-level and local, or "hotspot," somatic mutation enrichment statistics.

Conclusions: MutEnricher is a flexible software package for investigating somatic mutation enrichment that is implemented in Python, is freely available, can be efficiently parallelized, and is highly configurable to researcher's specific needs. MutEnricher is available online at https://github.com/asoltis/MutEnricher .
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http://dx.doi.org/10.1186/s12859-020-03695-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7393734PMC
July 2020

Deficiency of Adenosine Deaminase 2 (DADA2): Hidden Variants, Reduced Penetrance, and Unusual Inheritance.

J Clin Immunol 2020 08 8;40(6):917-926. Epub 2020 Jul 8.

Metabolic, Cardiovascular and Inflammatory Disease Genomics Branch, National Human Genome Research Institute (NHGRI), Bethesda, MD, USA.

Purpose: Deficiency of adenosine deaminase 2 (DADA2) is an autosomal recessive disorder that manifests with fever, early-onset vasculitis, strokes, and hematologic dysfunction. This study aimed to identify disease-causing variants by conventional Sanger and whole exome sequencing in two families suspected to have DADA2 and non-confirmatory genotypes. ADA2 enzymatic assay confirmed the clinical diagnosis of DADA2. Molecular diagnosis was important to accurately identify other family members at risk.

Methods: We used a variety of sequencing technologies, ADA2 enzymatic testing, and molecular methods including qRT-PCR and MLPA.

Results: Exome sequencing identified heterozygosity for the known pathogenic variant ADA2: c.1358A>G, p.Tyr453Cys in a 14-year-old female with a history of ischemic strokes, livedo, and vasculitis. No second pathogenic variant could be identified. ADA2 enzymatic testing in combination with quantitative RT-PCR suggested a loss-of-function allele. Subsequent genome sequencing identified a canonical splice site variant, c.-47+2T>C, within the 5'UTR of ADA2. Two of her unaffected siblings were found to carry the same two pathogenic variants. A homozygous 800-bp duplication comprising exon 7 of ADA2 was identified in a 5-year-old female with features consistent with Diamond-Blackfan anemia (DBA). The duplication was missed by Sanger sequencing of ADA2, chromosomal microarray, and exome sequencing but was detected by MLPA in combination with long-read PCR sequencing. The exon 7 duplication was also identified in her non-symptomatic father and younger sister.

Conclusions: ADA2 pathogenic variants may not be detected by conventional sequencing and genetic testing and may require the incorporation of additional diagnostic methods. A definitive molecular diagnosis is crucial for all family members to make informed treatment decisions.
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http://dx.doi.org/10.1007/s10875-020-00817-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7416912PMC
August 2020
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