Somatic CAG repeat instability in intermediate alleles of the HTT gene and its potential association with a clinical phenotype.

Huntington disease (HD) is a neurodegenerative disorder caused by ≥36 CAGs in the HTT gene. Intermediate alleles (IAs) (27-35 CAGs) are not considered HD-causing, but their potential association with neurocognitive symptoms remains controversial. As HTT somatic CAG expansion influences HD onset, we hypothesised that IAs are somatically unstable, and that somatic CAG expansion may drive phenotypic presentation in some IA carriers. We quantified HTT somatic CAG expansions by MiSeq sequencing in the blood DNA of 164 HD subjects and 191 IA (symptomatic and control) carriers, and in the brain DNA of a symptomatic 33 CAG carrier. We also performed genotype-phenotype analysis. The phenotype of symptomatic IA carriers was characterised by motor (85%), cognitive (27%) and/or behavioural (29%) signs, with a late (58.7 ± 18.6 years), but not CAG-dependent, age at onset. IAs displayed somatic expansion that were CAG and age-dependent in blood DNA, with 0.4% and 0.01% of DNA molecules expanding by CAG and year, respectively. Somatic expansions of +1 and +2 CAGs were detected in the brain of the individual with 33 CAGs, with the highest expansion frequency in the putamen (10.3%) and the lowest in the cerebellum (4.8%). Somatic expansion in blood DNA was not different in symptomatic vs. control IA carriers. In conclusion, we show that HTT IAs are somatically unstable, but we found no association with HD-like phenotypes. It is plausible, however, that some IAs, close to the HD pathological threshold and with a predisposing genetic background, could manifest with neurocognitive symptoms.

Longitudinal changes in white matter as measured with diffusion tensor imaging in adult-onset myotonic dystrophy type 1.

Myotonic dystrophy type 1 is characterized by neuromuscular degeneration. Our objective was to compare change in white matter microstructure (fractional anisotropy, radial and axial diffusivity), and functional/clinical measures. Participants underwent yearly neuroimaging and neurocognitive assessments over three-years. Assessments encompassed full-scale intelligence, memory, language, visuospatial skills, attention, processing speed, and executive function, as well as clinical symptoms of muscle/motor function, apathy, and hypersomnolence. Mixed effects models were used to examine differences. 69 healthy adults (66.2% women) and 41 DM1 patients (70.7% women) provided 156 and 90 observations, respectively. There was a group by elapsed time interaction for cerebral white matter, where DM1 patients exhibited declines in white matter (all p<0.05). Likewise, DM1 patients either declined (motor), improved more slowly (intelligence), or remained stable (executive function) for functional outcomes. White matter was associated with functional performance; intelligence was predicted by axial (r = 0.832; p<0.01) and radial diffusivity (r = 0.291, p<0.05), and executive function was associated with anisotropy (r = 0.416, p<0.001), and diffusivity (axial: r = 0.237, p = 0.05 and radial: r = 0.300, p<0.05). Indices of white matter health are sensitive to progression in DM1. These results are important for clinical trial design, which utilize short intervals to establish treatment efficacy.

Masseter muscle volume as a disease marker in adult-onset myotonic dystrophy type 1.

The advent of clinical trials in myotonic dystrophy type 1 (DM1) necessitates the identification of reliable outcome measures to quantify different disease manifestations using minimal number of assessments. In this study, clinical correlations of mean masseter volume (mMV) were explored to evaluate its potential as a marker of muscle involvement in adult-onset DM1 patients. We utilised data from a preceding study, pertaining to 39 DM1 patients and 20 age-matched control participants. In this study participants had undergone MRI of the brain, completed various clinical outcome measures and had CTG repeats measured by small-pool PCR. Manual segmentation of masseter muscles was performed by a single rater to estimate mMV. The masseter muscle was atrophied in DM1 patients when compared to controls (p<0.001). Significant correlations were found between mMV and estimated progenitor allele length (p = 0.001), modal allele length (p = 0.003), disease duration (p = 0.009) and and the Muscle Impairment Rating Scale (p = 0.008). After correction for lean body mass, mMV was also inversely correlated with self-reported myotonia (p = 0.014). This study demonstrates that changes in mMV are sensitive in reflecting the underlying disease process. Quantitative MRI methods demonstrate that data concerning both central and peripheral disease could be acquired from MR brain imaging studies in DM1 patients.

Clinical and neuroradiological correlates of sleep in myotonic dystrophy type 1.

Abnormalities of sleep are common in myotonic dystrophy type 1 (DM1), but few previous studies have combined polysomnography with detailed clinical measures and brain imaging. In the present study, domiciliary polysomnography, symptom questionnaires and cognitive evaluation were undertaken in 39 DM1-affected individuals. Structural brain MRI was completed in those without contra-indication (n = 32). Polysomnograms were adequate for analysis in 36 participants. Sleep efficiency was reduced, and sleep architecture altered in keeping with previous studies. Twenty participants (56%) had moderate or severe sleep-disordered breathing (apnoea-hypopnoea index [AHI] ≥ 15). In linear modelling, apnoeas were positively associated with increasing age and male sex. AHI ≥ 15 was further associated with greater daytime pCO2 and self-reported physical impairment, somnolence and fatigue. Percentage REM sleep was inversely associated with cerebral grey matter volume, stage 1 sleep was positively associated with occipital lobe volume and stage 2 sleep with amygdala volume. Hippocampus volume was positively correlated with self-reported fatigue and somnolence. Linear relationships were also observed between measures of sleep architecture and cognitive performance. Findings broadly support the hypothesis that changes in sleep architecture and excessive somnolence in DM1 reflect the primary disease process in the central nervous system.

Cognitive Deficits, Apathy, and Hypersomnolence Represent the Core Brain Symptoms of Adult-Onset Myotonic Dystrophy Type 1.

Myotonic dystrophy type 1 is the most common form of muscular dystrophy in adults, and is primarily characterized by muscle weakness and myotonia, yet some of the most disabling symptoms of the disease are cognitive and behavioral. Here we evaluated several of these non-motor symptoms from a cross-sectional time-point in one of the largest longitudinal studies to date, including full-scale intelligence quotient, depression, anxiety, apathy, sleep, and cerebral white matter fractional anisotropy in a group of 39 adult-onset myotonic dystrophy type 1 participants (27 female) compared to 79 unaffected control participants (46 female). We show that intelligence quotient was significantly associated with depression (P < 0.0001) and anxiety (P = 0.018), but not apathy (P < 0.058) or hypersomnolence (P = 0.266) in the DM1 group. When controlling for intelligence quotient, cerebral white matter fractional anisotropy was significantly associated with apathy (P = 0.042) and hypersomnolence (P = 0.034), but not depression (P = 0.679) or anxiety (P = 0.731) in the myotonic dystrophy type 1 group. Finally, we found that disease duration was significantly associated with apathy (P < 0.0001), hypersomnolence (P < 0.001), IQ (P = 0.038), and cerebral white matter fractional anisotropy (P < 0.001), but not depression (P = 0.271) or anxiety (P = 0.508). Our results support the hypothesis that cognitive deficits, hypersomnolence, and apathy, are due to the underlying neuropathology of myotonic dystrophy type 1, as measured by cerebral white matter fractional anisotropy and disease duration. Whereas elevated symptoms of depression and anxiety in myotonic dystrophy type 1 are secondary to the physical symptoms and the emotional stress of coping with a chronic and debilitating disease. Results from this work contribute to a better understanding of disease neuropathology and represent important therapeutic targets for clinical trials.

Neurocognitive Features of Motor Premanifest Individuals With Myotonic Dystrophy Type 1.

OBJECTIVE: The goal of the study was to identify brain and functional features associated with premanifest phases of adult-onset myotonic dystrophy type 1 (i.e., PreDM1). METHODS: This cross-sectional study included 68 healthy adults (mean age = 43.4 years, SD = 12.9), 13 individuals with PreDM1 (mean age: 47.4 years, SD = 16.3), and 37 individuals with manifest DM1 (mean age = 45.2 years, SD = 9.3). The primary outcome measures included fractional anisotropy (FA), motor measures (Muscle Impairment Rating Scale, Grooved Pegboard, Finger-Tapping Test, and grip force), general cognitive abilities (Wechsler Adult Intelligence Scales), sleep quality (Scales for Outcomes in Parkinson's Disease-Sleep), and apathy (Apathy Evaluation Scale). RESULTS: Individuals with PreDM1 exhibited an intermediate level of white matter FA abnormality, where whole-brain FA was lower relative to healthy controls (difference of the estimated marginal mean [EMMdifference] = 0.02, 95% confidence interval (CI) 0.01-0.03, p < 0.001), but the PreDM1 group had significantly higher FA than did individuals with manifest DM1 (EMMdifference = 0.02, 95% CI 0.009-0.03, p < 0.001). Individuals with PreDM1 exhibited reduced performance on the finger-tapping task relative to control peers (EMMdifference = 5.70, 95% CI 0.51-11.00, p = 0.03), but performance of the PreDM1 group was better than that of the manifest DM1 group (EMMdifference = 5.60, 95% CI 0.11-11.00, p = 0.05). Hypersomnolence in PreDM1 was intermediate between controls (EMMdifference = -1.70, 95% CI -3.10-0.35, p = 0.01) and manifest DM1 (EMMdifference = -2.10, 95% CI -3.50-0.60, p = 0.006). CONCLUSIONS: Our findings highlight key CNS and functional deficits associated with PreDM1, offering insight in early disease course.

Associations Between Variant Repeat Interruptions and Clinical Outcomes in Myotonic Dystrophy Type 1.

OBJECTIVE: To assess the association between variant repeat (VR) interruptions in patients with myotonic dystrophy type 1 (DM1) and clinical symptoms and outcome measures after cognitive behavioral therapy (CBT) intervention. METHODS: Adult patients with DM1 were recruited within the OPTIMISTIC trial (NCT02118779). Disease-related history, current clinical symptoms and comorbidities, functional assessments, and disease- and health-related questionnaires were obtained at baseline and after 5 and 10 months. After genetic analysis, we assessed the association between the presence of VR interruptions and clinical symptoms' long-term outcomes and compared the effects of CBT in patients with and without VR interruptions. Core trial outcome measures analyzed were: 6-minute walking test, DM1-Activ-C, Checklist Individual Strength Fatigue Score, Myotonic Dystrophy Health Index, McGill-Pain questionnaire, and Beck Depression inventory-fast screen. Blood samples for DNA testing were obtained at the baseline visit for determining CTG length and detection of VR interruptions. RESULTS: VR interruptions were detectable in 21/250 patients (8.4%)-12 were assigned to the standard-of-care group (control group) and 9 to the CBT group. Patients with VR interruptions were significantly older when the first medical problem occurred and had a significantly shorter disease duration at baseline. We found a tendency toward a milder disease severity in patients with VR interruptions, especially in ventilation status, mobility, and cardiac symptoms. Changes in clinical outcome measures after CBT were not associated with the presence of VR interruptions. CONCLUSIONS: The presence of VR interruptions is associated with a later onset of the disease and a milder phenotype. However, based on the OPTIMISTIC trial data, the presence of VR interruptions was not associated with significant changes on outcome measures after CBT intervention. TRIAL REGISTRATION INFORMATION: ClinicalTrials.gov NCT02118779.

White matter microstructure relates to motor outcomes in myotonic dystrophy type 1 independently of disease duration and genetic burden.

Deficits in white matter (WM) integrity and motor symptoms are among the most robust and reproducible features of myotonic dystrophy type 1 (DM1). In the present study, we investigate whether WM integrity, obtained from diffusion-weighted MRI, corresponds to quantifiable motor outcomes (e.g., fine motor skills and grip strength) and patient-reported, subjective motor deficits. Critically, we explore these relationships in the context of other potentially causative variables, including: disease duration, elapsed time since motor symptom onset; and genetic burden, the number of excessive CTG repeats causing DM1. We found that fractional anisotropy (a measure of WM integrity) throughout the cerebrum was the strongest predictor of grip strength independently of disease duration and genetic burden, while radial diffusivity predicted fine motor skill (peg board performance). Axial diffusivity did not predict motor outcomes. Our results are consistent with the notion that systemic degradation of WM in DM1 mediates the relationship between DM1 progression and genetic burden with motor outcomes of the disease. Our results suggest that tracking changes in WM integrity over time may be a valuable biomarker for tracking therapeutic interventions, such as future gene therapies, for DM1.

A DM1 patient with CCG variant repeats: Reaching the diagnosis.

We report the case of a male patient presenting in his 50s with ptosis, facial and distal limb muscle weakness, clinical and electrical myotonia, and a prior history of cataract extraction. He had a dominant family history in keeping with a similar phenotype. Myotonic dystrophy type 1 was clinically suspected. Triplet-primed polymerase chain reaction in a diagnostic laboratory did not identify a typical CTG repeat expansion on two separate blood samples. However, subsequent genetic testing on a research basis identified a heterozygous repeat expansion containing CCG variant repeats. Our case highlights the point that variant repeats are not detectable on triplet-primed polymerase chain reaction and result in a milder phenotype of myotonic dystrophy. It is crucial to maintain a high clinical index of suspicion of this common neuromuscular condition.

Approaches to Sequence the HTT CAG Repeat Expansion and Quantify Repeat Length Variation.

BACKGROUND: Huntington's disease (HD) is an autosomal dominant neurodegenerative disorder caused by the expansion of the HTT CAG repeat. Affected individuals inherit ≥36 repeats and longer alleles cause earlier onset, greater disease severity and faster disease progression. The HTT CAG repeat is genetically unstable in the soma in a process that preferentially generates somatic expansions, the proportion of which is associated with disease onset, severity and progression. Somatic mosaicism of the HTT CAG repeat has traditionally been assessed by semi-quantitative PCR-electrophoresis approaches that have limitations (e.g., no information about sequence variants). Genotyping-by-sequencing could allow for some of these limitations to be overcome. OBJECTIVE: To investigate the utility of PCR sequencing to genotype large (>50 CAGs) HD alleles and to quantify the associated somatic mosaicism. METHODS: We have applied MiSeq and PacBio sequencing to PCR products of the HTT CAG repeat in transgenic R6/2 mice carrying ∼55, ∼110, ∼255 and ∼470 CAGs. For each of these alleles, we compared the repeat length distributions generated for different tissues at two ages. RESULTS: We were able to sequence the CAG repeat full length in all samples. However, the repeat length distributions for samples with ∼470 CAGs were biased towards shorter repeat lengths. CONCLUSION: PCR sequencing can be used to sequence all the HD alleles considered, but this approach cannot be used to estimate modal allele size or quantify somatic expansions for alleles ⪢250 CAGs. We review the limitations of PCR sequencing and alternative approaches that may allow the quantification of somatic contractions and very large somatic expansions.

Blood-Based Markers of Neuronal Injury in Adult-Onset Myotonic Dystrophy Type 1.

INTRODUCTION: The present study had four aims. First, neuronal injury markers, including neurofilament light (NF-L), total tau, glial fibrillary acidic protein (GFAP) and ubiquitin C-terminal hydrolase (UCH-L1), were compared between individuals with and without adult-onset myotonic dystrophy type 1 (DM1). Second, the impact of age and CTG repeat on brain injury markers was evaluated. Third, change in brain injury markers across the study period was quantified. Fourth, associations between brain injury markers and cerebral white matter (WM) fractional anisotropy (FA) were identified. METHODS: Yearly assessments, encompassing blood draws and diffusion tensor imaging on a 3T scanner, were conducted on three occasions. Neuronal injury markers were quantified using single molecule array (Simoa). RESULTS: The sample included 53 patients and 70 controls. NF-L was higher in DM1 patients than controls, with individuals in the premanifest phases of DM1 (PreDM1) exhibiting intermediate levels ( χ ( 2 ) 2 = 38.142, P < 0.001). Total tau was lower in DM1 patients than controls (Estimate = -0.62, 95% confidence interval [CI] -0.95: -0.28, P < 0.001), while GFAP was elevated in PreDM1 only (Estimate = 30.37, 95% CI 10.56:50.19, P = 0.003). Plasma concentrations of UCH-L1 did not differ between groups. The age by CTG interaction predicted NF-L: patients with higher estimated progenitor allelege length (ePAL) had higher NF-L at a younger age, relative to patients with lower CTG repeat; however, the latter exhibited faster age-related change (Estimate = -0.0021, 95% CI -0.0042: -0.0001, P = 0.045). None of the markers changed substantially over the study period. Finally, cerebral WM FA was significantly associated with NF-L (Estimate = -42.86, 95% CI -82.70: -3.02, P = 0.035). INTERPRETATION: While NF-L appears sensitive to disease onset and severity, its utility as a marker of progression remains to be determined. The tau assay may have low sensitivity to tau pathology associated with DM1.

Preliminary Findings on CTG Expansion Determination in Different Tissues from Patients with Myotonic Dystrophy Type 1.

Myotonic Dystrophy type 1 (DM1) is characterized by a high genetic and clinical variability. Determination of the genetic variability in DM1 might help to determine whether there is an association between CTG (Cytosine-Thymine-Guanine) expansion and the clinical manifestations of this condition. We studied the variability of the CTG expansion (progenitor, mode, and longest allele, respectively, and genetic instability) in three tissues (blood, muscle, and tissue) from eight patients with DM1. We also studied the association of genetic data with the patients' clinical characteristics. Although genetic instability was confirmed in all the tissues that we studied, our results suggest that CTG expansion is larger in muscle and skin cells compared with peripheral blood leukocytes. While keeping in mind that more research is needed in larger cohorts, we have provided preliminary evidence suggesting that the estimated progenitor CTG size in muscle could be potentially used as an indicator of age of disease onset and muscle function impairment.

Variant repeats within the DMPK CTG expansion protect function in myotonic dystrophy type 1.

OBJECTIVE: We tested the hypothesis that variant repeat interruptions (RIs) within the DMPK CTG repeat tract lead to milder symptoms compared with pure repeats (PRs) in myotonic dystrophy type 1 (DM1). METHODS: We evaluated motor, neurocognitive, and behavioral outcomes in a group of 6 participants with DM1 with RI compared with a case-matched sample of 12 participants with DM1 with PR and a case-matched sample of 12 unaffected healthy comparison participants (UA). RESULTS: In every measure, the RI participants were intermediate between UA and PR participants. For muscle strength, the RI group was significantly less impaired than the PR group. For measures of Full Scale IQ, depression, and sleepiness, all 3 groups were significantly different from each other with UA > RI > PR in order of impairment. The RI group was different from unaffected, but not significantly different from PR (UA > RI = PR) in apathy and working memory. Finally, in finger tapping and processing speed, RI did not differ from UA comparisons, but PR had significantly lower scores than the UA comparisons (UA = RI > PR). CONCLUSIONS: Our results support the notion that patients affected by DM1 with RI demonstrate a milder phenotype with the same pattern of deficits as those with PR indicating a similar disease process.

The Need for Establishing a Universal CTG Sizing Method in Myotonic Dystrophy Type 1.

The number of cytosine-thymine-guanine (CTG) repeats ('CTG expansion size') in the 3'untranslated region (UTR) region of the dystrophia myotonica-protein kinase (DMPK) gene is a hallmark of myotonic dystrophy type 1 (DM1), which has been related to age of disease onset and clinical severity. However, accurate determination of CTG expansion size is challenging due to its characteristic instability. We compared five different approaches (heat pulse extension polymerase chain reaction [PCR], long PCR-Southern blot [with three different primers sets-1, 2 and 3] and small pool [SP]-PCR) to estimate CTG expansion size in the progenitor allele as well as the most abundant CTG expansion size, in 15 patients with DM1. Our results indicated variability between the methods (although we found no overall differences between long PCR 1 and 2 and SP-PCR, respectively). While keeping in mind the limited sample size of our patient cohort, SP-PCR appeared as the most suitable technique, with an inverse significant correlation found between CTG expansion size of the progenitor allele, as determined by this method, and age of disease onset (r = -0.734, p = 0.016). Yet, in light of the variability of the results obtained with the different methods, we propose that an international agreement is needed to determine which is the most suitable method for assessing CTG expansion size in DM1.

Change over time in ability to perform activities of daily living in myotonic dystrophy type 1.

OBJECTIVE: The objective of this longitudinal, observational study was to investigate change over time in ability to perform activities of daily living in myotonic dystrophy type 1 (DM1). METHODS: Adults with genetically confirmed DM1 were recruited as part of the PhenoDM1 study in the UK. Data on activities of daily living were recorded through the DM1-ActivC at baseline and a follow-up visit after 12 (± 3) months. A subset of patients had advanced genetic testing to determine the size of the progenitor allele. RESULTS: Our sample comprised 150 patients with DM1 (mean age: 45 years; 52% female). Mean follow-up was 383 days. Mean DM1-ActivC total score at baseline was 71.24 (95% confidence interval 67.77-74.71) and at the follow-up visit 69.04 (65.54-72.54). Approximately 43% of patients had a lower score at the follow-up visit (indicating a decreased ability to perform activities of daily living), 24% a higher score (indicating an increased ability), and 33% the same score at baseline and follow-up. The mean annual change in the DM1-ActivC total score, estimated at - 2.06 (- 3.54 to - 0.59), was significantly related to patients' baseline score, but not sex, disease duration, timed test results, or cytosine-thymine-guanine repeat length. CONCLUSIONS: Change over time in ability to perform activities of daily living as recorded through the DM1-ActivC varies substantially between patients with DM1. Our data contribute to the understanding of the natural evolution of the disease, and should be helpful to inform the design of future trials based on the DM1-ActivC.

Towards development of a statistical framework to evaluate myotonic dystrophy type 1 mRNA biomarkers in the context of a clinical trial.

Myotonic dystrophy type 1 (DM1) is a rare genetic disorder, characterised by muscular dystrophy, myotonia, and other symptoms. DM1 is caused by the expansion of a CTG repeat in the 3'-untranslated region of DMPK. Longer CTG expansions are associated with greater symptom severity and earlier age at onset. The primary mechanism of pathogenesis is thought to be mediated by a gain of function of the CUG-containing RNA, that leads to trans-dysregulation of RNA metabolism of many other genes. Specifically, the alternative splicing (AS) and alternative polyadenylation (APA) of many genes is known to be disrupted. In the context of clinical trials of emerging DM1 treatments, it is important to be able to objectively quantify treatment efficacy at the level of molecular biomarkers. We show how previously described candidate mRNA biomarkers can be used to model an effective reduction in CTG length, using modern high-dimensional statistics (machine learning), and a blood and muscle mRNA microarray dataset. We show how this model could be used to detect treatment effects in the context of a clinical trial.

Activities of daily living in myotonic dystrophy type 1.

OBJECTIVES: The objective of this cross-sectional, observational study was to investigate performance of activities of daily living in patients with myotonic dystrophy type 1 (DM1). MATERIALS AND METHODS: Adults with genetically confirmed DM1 were recruited from Newcastle University (Newcastle upon Tyne, UK) and University College London Hospitals NHS Foundation Trust (London, UK). Data on activities of daily living were recorded through the DM1-ActivC (scale scores range between 0 and 100, where a higher/lower score indicates a higher/lower ability). RESULTS: Our sample comprised 192 patients with DM1 (mean age: 46 years; 51% female). Patients reported most difficulties with running, carrying and putting down heavy objects, and standing on one leg, and least difficulties with eating soup, washing upper body, and taking a shower. Irrespective of the disease duration (mean: 20 years), most patients were able to perform basic and instrumental activities of daily living (eg personal hygiene and grooming, showering, eating, cleaning and shopping), with the exception of functional mobility/transfer tasks (eg walking uphill and running). The mean DM1-ActivC total score was estimated at 71 (95% CI: 68-74). Estimated progenitor cytosine-thymine-guanine repeat length and age explained 27% of the variance in DM1-ActivC total scores (P < .001). CONCLUSIONS: We show that DM1 impairs performance of activities of daily living, in particular those requiring a high degree of muscle strength, stability and coordination. Yet, across the evolution of the disease, the majority of patients will still be able to independently perform most basic and instrumental activities of daily living.

A DM1 family with interruptions associated with atypical symptoms and late onset but not with a milder phenotype.

Carriage of interruptions in CTG repeats of the myotonic dystrophy protein kinase gene has been associated with a broad spectrum of myotonic dystrophy type 1 (DM1) phenotypes, mostly mild. However, the data available on interrupted DM1 patients and their phenotype are scarce. We studied 49 Spanish DM1 patients, whose clinical phenotype was evaluated in depth. Blood DNA was obtained and analyzed through triplet-primed polymerase chain reaction (PCR), long PCR-Southern blot, small pool PCR, AciI digestion, and sequencing. Five patients of our registry (10%), belonging to the same family, carried CCG interruptions at the 3'-end of the CTG expansion. Some of them presented atypical traits such as very late onset of symptoms ( > 50 years) and a severe axial and proximal weakness requiring walking assistance. They also showed classic DM1 symptoms including cardiac and respiratory dysfunction, which were severe in some of them. Sizes and interrupted allele patterns were determined, and we found a contraction and an expansion in two intergenerational transmissions. Our study contributes to the observation that DM1 patients carrying interruptions present with atypical clinical features that can make DM1 diagnosis difficult, with a later than expected age of onset and a previously unreported aging-related severe disease manifestation.

A genetic association study of glutamine-encoding DNA sequence structures, somatic CAG expansion, and DNA repair gene variants, with Huntington disease clinical outcomes.

BACKGROUND: Huntington disease (HD) is caused by an unstable CAG/CAA repeat expansion encoding a toxic polyglutamine tract. Here, we tested the hypotheses that HD outcomes are impacted by somatic expansion of, and polymorphisms within, the HTT CAG/CAA glutamine-encoding repeat, and DNA repair genes. METHODS: The sequence of the glutamine-encoding repeat and the proportion of somatic CAG expansions in blood DNA from participants inheriting 40 to 50 CAG repeats within the TRACK-HD and Enroll-HD cohorts were determined using high-throughput ultra-deep-sequencing. Candidate gene polymorphisms were genotyped using kompetitive allele-specific PCR (KASP). Genotypic associations were assessed using time-to-event and regression analyses. FINDINGS: Using data from 203 TRACK-HD and 531 Enroll-HD participants, we show that individuals with higher blood DNA somatic CAG repeat expansion scores have worse HD outcomes: a one-unit increase in somatic expansion score was associated with a Cox hazard ratio for motor onset of 3·05 (95% CI = 1·94 to 4·80, p = 1·3 × 10-6). We also show that individual-specific somatic expansion scores are associated with variants in FAN1 (pFDR = 4·8 × 10-6), MLH3 (pFDR = 8·0 × 10-4), MLH1 (pFDR = 0·004) and MSH3 (pFDR = 0·009). We also show that HD outcomes are best predicted by the number of pure CAGs rather than total encoded-glutamines. INTERPRETATION: These data establish pure CAG length, rather than encoded-glutamine, as the key inherited determinant of downstream pathophysiology. These findings have implications for HD diagnostics, and support somatic expansion as a mechanistic link for genetic modifiers of clinical outcomes, a driver of disease, and potential therapeutic target in HD and related repeat expansion disorders. FUNDING: CHDI Foundation.

Genetic determinants of disease severity in the myotonic dystrophy type 1 OPTIMISTIC cohort.

OBJECTIVE: To evaluate the role of genetic variation at the DMPK locus on symptomatic diversity in 250 adult, ambulant patients with myotonic dystrophy type 1 (DM1) recruited to the Observational Prolonged Trial in Myotonic Dystrophy Type 1 to Improve Quality of Life-Standards, a Target Identification Collaboration (OPTIMISTIC) clinical trial. METHODS: We used small pool PCR to correct age at sampling biases and estimate the progenitor allele CTG repeat length and somatic mutational dynamics, and AciI digests and repeat primed PCR to test for the presence of variant repeats. RESULTS: We confirmed disease severity is driven by progenitor allele length, is further modified by age, and, in some cases, sex, and that patients in whom the CTG repeat expands more rapidly in the soma develop symptoms earlier than predicted. We revealed a key role for variant repeats in reducing disease severity and quantified their role in delaying age at onset by approximately 13.2 years (95% confidence interval 5.7-20.7, 2-tailed t test t = -3.7, p = 0.0019). CONCLUSIONS: Careful characterization of the DMPK CTG repeat to define progenitor allele length and presence of variant repeats has increased utility in understanding clinical variability in a trial cohort and provides a genetic route for defining disease-specific outcome measures, and the basis of treatment response and stratification in DM1 trials.