Neurodegeneration Trajectory in Pediatric and Adult/Late DM1: a Follow-up MRI Study Across a Decade

RESEARCH ARTICLE Neurodegeneration trajectory in pediatric and adult/late DM1: A follow-up MRI study across a decade Garazi Labayru 1,2,3 , Antonio Jimenez-Marin 4,5 , Esther Fern andez 6 , Jorge Villanua 6 , Miren Zulaica 2,3 , Jesus M. Cortes 4,7,8 , Ibai D ıez 9,10,11 , Jorge Sepulcre 9,10 , Adolfo L opez de Munain 2,3,12,13 & Andone Sistiaga 1,2,3 1 Personality, Assessment and psychological treatment department; Psychology Faculty, University of the Basque Country (UPV/EHU), San Sebasti an, Gipuzkoa, Spain 2 Neuroscience Area, Biodonostia Research Institute, Osakidetza, Donostia-San Sebasti an, Gipuzkoa, Spain 3 Centro de Investigaci on Biom edica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Institute Carlos III, Madrid, Spain 4 Biocruces-Bizkaia Health Research Institute, Barakaldo, Bizkaia, Spain 5 Biomedical Research Doctorate Program, University of the Basque Country (UPV/EHU), Leioa, Spain 6 Osatek, Donostia University Hospital, DonostiaSan Sebasti an, Gipuzkoa, Spain 7 Cell Biology and Histology Department, University of the Basque Country (UPV/EHU), Leioa, Spain 8 IKERBASQUE, The Basque Foundation for Science, Bilbao, Spain 9 Gordon Center for Medical Imaging, Department of Radiology, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 10 Athinoula A. Martinos Center for Biomedical Imaging, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts 11 Neurotechnology Laboratory, Tecnalia Health Department, Derio, Spain 12 Neurology Department, Donostia University Hospital, DonostiaSan Sebasti an, Gipuzkoa, Spain 13 Neuroscience Department, University of the Basque Country (UPV/EHU), Donostia-San Sebasti an, Gipuzkoa, Spain Correspondence Garazi Labayru, University of the Basque Country, Psychology Faculty, Avda. Tolosa, 70. 20018 DonostiaSan Sebasti an, Gipuzkoa, Spain. Tel: +00 34 94 301 8382; Fax: +34 94 301 5670; E-mail: [email protected] Received: 9 April 2020; Revised: 24 July 2020; Accepted: 25 July 2020 Annals of Clinical and Translational Neurology 2020; 7(10): 1802–1815 Funding Information CIBERNED609 Eusko JaurlaritzaPRE_ 2016_1_0187PRE_2019_1_0070SAIO08PE08BF01 Institute of Health Carlos III cofounded by Fondo Europeo de Desarrollo Regional-FEDERPI17/01231 PI17/01841 doi: 10.1002/acn3.51163 Abstract Objective: To characterize the progression of brain structural abnormalities in adults with pediatric and adult/late onset DM1, as well as to examine the potential predictive markers of such progression. Methods: 21 DM1 patients (pediatric onset: N =9; adult/late onset: N =12) and 18 healthy controls (HC) were assessed longitudinally over 9.17 years through brain MRI. Additionally, patients underwent neuropsychological, genetic, and muscular impairment assessment. Inter-group comparisons of total and voxel-level regional brain volume were conducted through Voxel Based Morphometry (VBM); cross-sectionally and longitudinally, analyzing the associations between brain changes and demographic, clinical, and cognitive outcomes. Results: The percentage of GM loss did not significantly differ in any of the groups compared with HC and when assessed independently, adult/late DM1 patients and their HC group suffered a significant loss in WM volume. Regional VBM analyses revealed subcortical GM damage in both DM1 groups, evolving to frontal regions in the pediatric onset patients. Muscular impairment and the outcomes of certain neuropsychological tests were significantly associated with follow-up GM damage, while visuoconstruction, attention, and executive function tests showed sensitivity to WM degeneration over time. Interpretation: Distinct patterns of brain atrophy and its progression over time in pediatric and adult/late onset DM1 patients are suggested. Results indicate a possible neurodevelopmental origin of the brain abnormalities in DM1, along with the possible existence of an additional neurodegenerative process. Fronto-subcortical networks appear to be involved in the disease progression at young adulthood in pediatric onset DM1 patients. The involvement of a multimodal integration network in DM1 is discussed. 1802 ª2020 The Authors. Annals of Clinical and Translational Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made. Introduction Myotonic Dystrophy type 1 (DM1) is the most common form of adult muscular dystrophy. It is an autosomal dominant disorder affecting multiple systems. The severity of symptoms varies according to age of onset, with earlier onset patients being more severely affected. Aside from the known progressive nature of the muscular impairment, many of the clinical symptoms have been suggested to be part of an accelerated aging process. CNS studies with a focus on brain pathology have defined DM1 as a combination of tauopathy, spliceopathy, and RNAopathy, all of which contribute to neurodegeneration. 1 Furthermore, from a neuropsychological perspective, recent findings suggest that cognitive functions suffer a decline in several areas, beyond that expected in normal aging. 2–6 Although neuroimaging data are thought to support the previously suggested hypothesis of neurodegenerative processes in DM1, to the best of our knowledge, there have been only two previous attempts to longitudinally study a hypothesized progressive impairment in brain structures. While Gliem et al 7 did not find a greater volume loss over time in DM1 for either gray or white matter tissue, Conforti et al. found a progression in white matter lesions along with greater brain atrophy assessed by the ventricular/brain ratio. 8 The aim of this study was to longitudinally assess the structural brain changes of DM1 patients over a period of more than 9 years and to delineate the pathway of disease progression. Additionally, we aimed to examine the potential genetic, muscular, clinical, and cognitive markers of this progression. Materials and Methods Participants The DM1 patients analyzed in this work were selected from those attending the Neurology Department of the Donostia University Hospital (Gipuzkoa, Spain). Healthy controls (HC) included accompanying relatives and nonrelatives of DM1 patients using the service. Inclusion criteria for DM1 patients included being older than 18 years with molecular confirmation of the clinical diagnosis. Patients were excluded if any of the following criteria were met both at baseline and at followup: congenital form, history of major psychiatric or somatic disorder, acquired brain damage or alcohol or drug abuse, presence of corporal paramagnetic body devices that could impede an MRI study, and the presence of cerebral anomalies that could affect the volumetric analysis. HC participants were required to satisfy the same inclusion criteria, except for the clinical diagnosis. The DM1 participants were classified into two groups according to their age of onset based on the most recently proposed classification (OMMYD-4): adults with pediatric onset DM1 when age of onset was between 1-18 years old, and adults with adult and late onset DM1 (from now on adult/late DM1) when patients had either adult onset (18-40 years old) or late onset (>40 years old). Figure 1 shows flow chart of the recruitment process. From the healthy volunteers with a valid MRI at baseline, only those whose age, gender and years of education were equal or closely similar to any re-scanned patient were invited to participate in order to form demographically equivalent groups. The first 20 participants that agreed to be re-scanned were suitable for forming two groups equivalent in sex, age, and years of education with a valid sample size for purposes of comparison with each of the DM1 groups. A final group of 21 DM1 patients (9 pediatric and 12 adult/late) and one of 18 HC were included for the analysis. The 18 HC controls were then subdivided to form the two comparison groups: one for comparisons with pediatric onset DM1 (HC-pediatric: N =12) and one for comparisons with adult/late onset DM1 (HC-adult/late: N=14). The healthy volunteers were included to form each control group in a controlled selection process, ensuring gender equivalence, and mean age differences between groups of less than 4 years, which were non-significant and with small effect sizes. All participants were informed of the objectives and details of the study and signed an informed consent form. The study was approved by the Ethics Committee of the Donostia University Hospital. Clinical and neuropsychological assessment Clinical data were extracted from medical records. Additionally, both at baseline and at follow-up, all patients were clinically examined by a neurologist with the Muscular Impairment Rating Scale (MIRS) 9 and underwent a neuropsychological assessment. All patients were examined by an experienced neuropsychologist who was blind to the patient’s clinical condition (CTG expansion size, clinical form, maternal or paternal inheritance pattern, muscular impairment, and MRI results). Neuropsychological assessment included the following subtests from the Wechsler Adult Intelligence Scale III (WAIS III) 10 : Block design, Digit span, Object assembly, Arithmetic, Similarities and Vocabulary. An estimated IQ score was calculated from a two subtest short form (Block design and Vocabulary) with high reliability (r xx =.93) and validity (r=.87) based on Sattler and Ryan. 11 Other cognitive tests used were: ª2020 The Authors. Annals of Clinical and Translational Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association 1803 G. Labayru et al.Neurodegeneration in DM1: A Follow-up MRI Study Stroop test, 12 California Computerized Assessment Package (CALCAP), 13 Rey Auditory Verbal Learning Test (RAVLT), 14 phonemic (P) and semantic (animals) verbal fluency test, 15,16 Rey-Osterrieth Complex Figure test (ROCF), 17 Raven’s progressive matrices, 18 Benton’s Judgement of Line Orientation, 19 and the Wisconsin Card Sorting Test. 20 Raw scores were converted into standardized T values according to Spanish norms for each test. MRI acquisition and data preprocessing MR scanning was conducted on a 1.5 Tesla scanner (Achieva Nova, Philips). The current results are based on a high-resolution volumetric “turbo field echo” (TFE) series (Sagital 3D T1 weighted acquisition, TR =7.2, TE =3.3, flip angle =8, matrix =256 x 232, slice thickness 1mm, voxel dimensions of 1mm x 1mm x 1mm, NSA =1, no slices 160, gap =0, total scan duration 5´ 34¨). All the scans, both at baseline and at follow-up, were acquired on the same MR scanner. To study voxel-based GM volume loss in DM1 patients and its association with different clinical and neuropsychological outcomes, FSL (version 6.01) Voxel Based Morphometry (VBM) was used, 21 which is an optimized VBM protocol 22 carried out with FSL tools. 23 First, structural images were brain-extracted and GM-segmented before being registered to the MNI 152 standard space using non-linear registration. 24 The resulting images were averaged and flipped along the x-axis to create a left-right symmetric, study-specific GM template. Second, all native GM images were non-linearly registered to this studyspecific template and "modulated" to correct for local expansion (or contraction) due to the non-linear component of the spatial transformation. The modulated GM images were then smoothed with an isotropic Gaussian kernel with a sigma of 3. The same VBM procedure was applied to the WM-segmented images. To estimate global GM and WM brain tissue volume, normalized for subject head size, the SIENAX tool was used. 25 Figure 1. Flow-chart of sample recruitment from baseline to follow-up. DM1 =Myotonic Dystrophy Type 1; HC =healthy controls. 1804 ª2020 The Authors. Annals of Clinical and Translational Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association Neurodegeneration in DM1: A Follow-up MRI Study G. Labayru et al. Statistical analysis For all statistical analyses listwise deletion was used to deal with missing values. Demographic (sex, age at baseline, age at follow-up, years of education) and clinical (CTG repeats, inheritance pattern, and MIRS score) data were analyzed using the SPSS (IBM SPSS Statistics 24) statistical package. Inter-group comparisons were conducted to compare DM1 patients and HC, as well as the two DM1 groups, using contingency analysis (Chisquare) for categorical data and parametric (t-test) or non-parametric (Mann–Whitney U) for interval data, where appropriate. In order to dismiss the possibility of a higher functioning sample in the re-tested groups (selectiveattrition),thesameanalyseswereemployedto compare those patients that failed to re-test at followup and those who were retested. Intra-group analysis of the longitudinal evolution of clinical and neuropsychological data was conducted using the Wilcoxon signedrank test. In order to address the general objective of this study, three main analyses were carried out (note that no direct statistical comparison was made between pediatric onset and adult/late onset DM1 groups): Total GM and WM volume analyses: Crosssectional and longitudinal: Intra-group and intergroup comparisons Total GM and WM volume comparisons, both intergroup and intra-group, were analyzed using the SPSS statistical package. All analyses were conducted by correcting the volume with the head size. Inter-group cross-sectional comparisons of total GM and WM volumes were conducted to compare DM1 patients and HC at baseline and at follow-up, using a parametric ttest (the data met the assumptions for parametric tests). Longitudinal analyses of GM and WM volumes were separately assessed for DM1 patients and HC using the Wilcoxon signed-rank test. Finally, the percentage of volume loss from baseline to follow-up in each group was calculated and comparisons were made between each DM1 group and their HC group, using a univariate ANOVA test corrected for time between the baseline and the follow-up scan. Regional GM and WM volume analysis from baseline to follow-up: VBM inter-group analysis A general linear model was used to compute the intergroup statistical analysis using FSL, controlling for age and head size. All the results were obtained using twotailed tests and corrected for multiple comparisons using the Monte Carlo simulation cluster-wise correction, as implemented in the AFNI software (version 19.3.00) (https://afni.nimh.nih.gov/) with 10,000 iterations to estimate the probability of false positive clusters with a P value <0.05. Only clusters of more than 50 contiguous voxels were reported. Association between brain volume and demographic, clinical, and neuropsychological outcomes: VBM intra-group analysis To study the potential predictive capacity of demographic, clinical, and neuropsychological variables at baseline, three separate analyses were conducted. First, in order to assess the predictive capacity of these variables at baseline (CTG, MIRS, years of education, disease inheritance, and neuropsychological scores) for volume loss (volume variations between follow-up and baseline), partial correlation analyses controlling for age, head size, and time span from baseline to follow-up scan were conducted. The results were further corrected by multiple comparisons using the false discovery rate (FDR) strategy. These analyses were conducted separately for pediatric and adult/late onset DM1 groups. Second, in order to assess the predictive capacity of the same variables for the image at follow-up, partial correlation analyses were conducted, controlling for age, head size and time from baseline assessment to follow-up scan, between predictive variables and global GM and WM volume at follow-up. These analyses were conducted separately for pediatric and adult/late onset DM1 groups. Finally, to assess the association between the previous variables and regional GM volume at follow-up, a general linear model analysis was applied to evaluate the relationship between GM volume and CTG, MIRS clinical scale, years of education, and inheritance pattern of the disease, as well as the outcomes of neuropsychological tests. The results were controlled for age, head size and time span between the baseline scan and the follow-up (except for neuropsychological variables, which were adjusted for time between the baseline neuropsychological assessment and the follow-up scan). The statistical tests were twotailed and corrected for multiple comparisons using Monte Carlo simulations with cluster-wise correction after 10,000 iterations to estimate the probability of false positive clusters with a Pvalue <0.05. After corrections, a mask was applied using the results of the group difference to look for any overlap with the regions where DM1 patients show a significant GM volume loss compared with controls. Only clusters of more than 50 contiguous voxels were reported. Analyses were ª2020 The Authors. Annals of Clinical and Translational Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association 1805 G. Labayru et al.Neurodegeneration in DM1: A Follow-up MRI Study conducted separately for pediatric and adult/late onset DM1 groups. Results Statistically significant differences were found between the excluded and the included DM1 patients for the following variables: CTG expansion size at baseline (t(34) =3.276; p= .002, d=1.1) with more repetitions in the excluded DM1 (mean =1002.88, SD =432.88) than the included DM1 (mean =534.4, SD =421.21) and MIRS score at baseline (U=45.5; P=0.001; r=0.58), with the excluded DM1 presenting greater muscular impairment (mean =3.62, SD =0.87, mean rank =24.5) than the included DM1 (mean =2.29, SD =0.96 mean rank =13.17). No difference was found between the excluded and the included DM1 patients in terms of sex, inheritance pattern, years of education, age, and IQ estimate. The demographic and clinical characteristics of the sample are summarized in Tables 1 and 2. None of the DM1 patient groups differed from the corresponding HC groups in terms of sex, age at baseline, age at follow-up or years of education. Pediatric onset DM1 patients were significantly younger and had a greater CTG expansion at both baseline and follow-up compared with adult/late onset DM1 patients. Both groups showed a significant increase in MIRS score and CTG repeat size from baseline to follow up. The mean time from baseline to follow-up was 9.17 (SD =0.47) years for the complete sample. Neuropsychological outcomes of the DM1 groups are shown in Supplementary Table S1. Table 1. Baseline and follow-up demographic characteristics of the sample divided into 4 subgroups: pediatric DM1, adult/late DM1, HC-pediatric, HC-adult/late. Pediatric DM1 (N =9) HC-pediatric (N =12) Statistic PEffect sizeMean/N(%) (SD) Mean/N(%) (SD) Sex Male 4 (44.4%) 5 (41.7%) X 2 =.016 0.899 V=.028 Female 5 (55.6%) 7 (58.3%) Age at baseline 30 (6.59) 33.5 (8.32) t=1.039 0.312 d=.46 Age at follow-up 39.67 (6.61) 42.5 (8.06) t=0.858 0.401 d=.36 Years of education 14.22 (4.26) 18.89 (6.49) U=22.5 0.111 r=.35 Adult/late DM1 (N =12) HC-adult/late (N =14) Statistic PEffect sizeMean/N(%) (SD) Mean/N(%) (SD) Sex Male 6 (50%) 8 (57.1%) X 2 =0.133 0.716 V=0.071 Female 6 (50%) 6 (42.9%) Age at baseline 45.83 (9.05) 43.64 (8.11) t=0.651 0.521 d=0.26 Age at follow-up 55.17 (8.89) 52.71 (8.27) t=0.728 0.473 d=0.29 Years of education 13.42 (6.68) 17.64 (5.93) t=0.976 0.340 d=0.41 Pediatric DM1 (N =9) Adult/late DM1 (N =12) Statistic PEffect sizeMean/N(%) (SD) Mean/N(%) (SD) Sex Male 4 (44.4%) 6 (50%) X 2 =0.064 0.801 V=0.055 Female 5 (55.6%) 6 (50%) Age at baseline 30 (6.59) 45.83 (9.05) t=4.427 0.000 d=1.95 Age at follow-up 39.67 (6.61) 55.17 (8.89) t=4.387 0.000 d=1.93 Inheritance Maternal 6 (66.7%) 3 (27.3%) X 2 =3.104 0.078 V=0.394 Paternal 3 (33.3%) 8 (72.7%) CTG at baseline 851.89 (444.77) 362.83 (325.54) t=2.916 0.009 d=1.29 CTG at follow-up 1044.44 (444.43) 500.75 (504.03) U=21.500 0.021 r=0.05 MIRS at baseline 2,56 (0.88) 2.08 (1) U=41.000 0.382 r=0.22 MIRS at follow-up 3.11 (1.17) 2.67 (1.37) U=46.500 0.602 r=0.12 Note: DM1: Myotonic Dystrophy Type 1; HC: Healthy controls; SD: Standard Deviation. Descriptive data are shown as mean and SD for age at baseline, age at follow-up and years of education. Frequency (N) and percentage (%) are shown only for sex. 1806 ª2020 The Authors. Annals of Clinical and Translational Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association Neurodegeneration in DM1: A Follow-up MRI Study G. Labayru et al. Total GM and WM volume analyses: crosssectional and longitudinal: Intra-group and inter-group analyses With regard to cross-sectional comparisons of brain volume, patients in the pediatric onset group presented lower volumes of both GM and WM compared with controls, both at baseline and at follow up. Conversely, the adult and late onset group obtained lower volumes only for GM when compared with controls (Table 3). Longitudinal analyses of GM and WM volume loss over time showed that none of the groups suffered a significant decrease in GM volume, and only adult/late DM1 patients and their HC group suffered a significant loss in WM volume. Nonetheless, when comparing the longitudinal variations between groups, the percentage of volume loss did not differ between pediatric onset DM1 and their controls, or between adult/late onset DM1 and their controls. However, the global GM volume decrease in the pediatric group reached 3.86% compared with a 0.63% decrease in HC, and, despite being non-significant, the effect size was large (ES =0.94). Similarly, a medium effect size (ES =0.57) was found for the difference between the percentage of GM volume loss in adult/late onset DM1 (2.71%) and HC-adult/late (0.85%). Accordingly, moderate effect sizes were found in patients’ GM volume loss when intra-group analyses were conducted for both pediatric and adult/late onset patients (r =0.43 and r =0.4, respectively). Regional GM and WM volume analysis from baseline to follow-up: VBM inter-group analysis Results from the GM VBM analyses are depicted in Figure 2 (and Supplementary Tables S2, S3, and S4 for a detailed report on significant clusters). At baseline, areas where pediatric onset DM1 showed a decrease in GM volumes compared with the HC-pediatric group were the bilateral thalamus, caudate, putamen, parahippocampal gyrus, right hippocampus, right lingual gyrus, left Rolandic operculum, left middle and superior temporal gyrus, left Heschl’s gyrus, left parietal operculum, left supramarginal gyrus, and left postcentral gyrus. At follow-up, these areas were still decreased in patients and, additionally, new cortical areas were involved, particularly the left precentral gyrus, bilateral inferior frontal gyrus (triangular part and orbital part), right inferior frontal gyrus (opercular part), bilateral parietal operculum, and left insula. Adult/late onset DM1 patients at baseline showed GM decrease in the bilateral caudate, putamen and thalamus and left insula in comparison with the HCadult/late group, and at follow-up, these areas expanded to adjacent regions such as the right hippocampus and para-hippocampal gyrus, together with the cerebellum. Additionally, a decrease was found in the HC-adult/late group compared with adult/late onset DM1 patients, located at baseline in the following areas of the left hemisphere: inferior, middle and superior temporal gyri, middle and superior temporal poles, fusiform gyrus, inferior and middle frontal gyri (orbital part), parahippocampal gyrus, and the cerebellum. At follow-up, a significant decrease remained in only some of these areas in the HCadult/late group compared with adult/late DM1 patients, again in the following regions of the left hemisphere: the middle and superior temporal lobes, fusiform gyrus, and the parahippocampal gyrus. Results from WM VBM analysis are shown in Supplementary Tables S5 and S6. At baseline, areas where pediatric onset DM1 showed a decrease in WM volumes compared with the HC-pediatric group were the bilateral posterior limb of internal capsule, left retrolenticular part of internal capsule and bilateral cerebral peduncle. At follow-up, these areas were still decreased in patients and, additionally, new areas were involved: left anterior corona radiata, left sagittal stratum, left fornix and stria terminalis and right retrolenticular part of internal capsule. Adult/late onset DM1 patients at baseline showed WM decrease in the right anterior corona radiata and the genu of corpus callosum. At follow-up, these areas were still Table 2. Longitudinal evolution of clinical features in pediatric DM1 and adult/late DM1 groups. N Baseline Follow-up Statistic P Effect sizeMean (SD) Mean (SD) Pediatric DM1 CTG 9 851.89 (444.77) 1044.44 (444.43) t=3.181 0.013 d=1.5 MIRS 9 2.56 (0.88) 3.11 (1.17) t=3.162 0.013 d=.75 Adult/late DM1 CTG 12 362.83 (325.54) 500.75 (504.03) Z=2.31 0.021 r=.47 MIRS 12 2.08 (0.99) 2.67 (1.37) Z=2.33 0.020 r=.48 Note: DM1: Myotonic Dystrophy Type 1; HC: Healthy controls; SD: Standard Deviation. ª2020 The Authors. Annals of Clinical and Translational Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association 1807 G. Labayru et al.Neurodegeneration in DM1: A Follow-up MRI Study Table 3. Cross-sectional (inter-group) and longitudinal (intra-group and inter-group) comparisons for GM and WM volume. N Mean SD DM1 versus HC Mean SD DM1 versus HC Intra-group Followup - Baseline Intergroup comparison of percentage of volume loss ZPMean FPg Hedges Baseline Follow-up Pediatric DM1 versus HC-pediatric GM volume t=4.208 t=5.071 DM1 9 728492.55 (59435.88) p=0.000 703854.60 (53407.41) P=0.000 1.836 0.066 3.86 ¶ 3.497 0.078 0.94 HC 12 829069.52 (50050.01) d=1.86 819800.71 (50693.01) d=2.24 1.569 0.117 0.63 ¶ WM volume DM1 9 656340.98 (45830.39) t=4.334 658010.04 (28468.26) t=4.643 0.296 0.767 0.25 ¶ 0.097 0.759 0.16 HC 12 721993.51 (22597.64) p=0.000 712342.09 (25040.87) p=0.000 1.412 0.158 0.77 ¶ d=1.91 d=2.05 Adult/late DM1 versus HC-adult/late GM volume t=2.172 t=2.324 DM1 12 756500.85 (42058.37) p=.040 737651.22 (52119.48) p=0.029 1.961 0.050 2.71 $ 1.721 0.203 0.57 HC 14 787456.64 (30440.24) d=.85 779936.16 (40616.75) d=0.91 1.475 0.140 0.85 $ WM volume DM1 12 703532.11 (40638.08) t=1.582 685715.24 (36237.65) t=1.256 3.059 0.002 2.63 $ 0.106 0.748 0.14 HC 14 727201.95 (35684.99) p=0.127 704947.47 (41037.58) p=0.221 2.856 0.004 2.97 $ d=0.62 d=0.49 Note: DM1: Myotonic Dystrophy Type 1; HC: Healthy controls; SD: Standard Deviation. Longitudinal inter-group comparisons of percentage of volume loss are calculated using time to follow-up as a covariate. ¶ Covariate value: 9.1624. $ Covariate value: 9.0973. 1808 ª2020 The Authors. Annals of Clinical and Translational Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association Neurodegeneration in DM1: A Follow-up MRI Study G. Labayru et al. decreased in patients and, additionally, new areas were involved: left anterior corona radiata, bilateral superior corona radiata, body of corpus callosum and left superior longitudinal fasciculus. Association between brain volume and demographic, clinical, and neuropsychological outcomes: VBM intragroup analysis For pediatric and adult/late onset DM1 patients, none of the clinical (MIRS score, CTG expansion size, inheritance pattern) or demographic (years of education) variables at baseline were significantly associated with the percentage of GM and WM volume loss from baseline to follow-up. However, among the neuropsychological tests, the ROCF copy and two CALCAP measures (Sequential 1 reaction time (RT) and Sequential 2 RT) were positively correlated with the percentage of WM volume loss only in adult/late onset DM1 patients (Figure 3). Lower scores on neuropsychological tests were associated with a greater percentage of total WM volume loss. No association was found between any clinical, demographic, or neuropsychological variable with the total GM and WM volume at follow-up in either DM1 group. However, the outcomes of some neuropsychological tests were significantly correlated with specific regions of reduced GM and WM at follow-up. In the pediatric onset DM1 group the results were inconclusive due to the reduced sample size available in most of the neuropsychological tests, which strongly affected the sensitivity of the statistical procedure. Only the results of the Stroop task (color-word and interference) were regionally correlated with decreased GM at follow-up (data not shown). Results in the adult/late onset group for GM are shown in Supplementary Figure S1 (see Supplementary Table S7 for a detailed report on significant clusters) and for WM in Supplementary Table S8. To focus on the association between neuropsychological outcomes and affected brain GM areas, the regions defined by the follow-up mask of group differences between adult/late DM1 and HC-adult/ late are indicated by black transparent shading. The MIRS scale was the only clinical variable that significantly correlated with lower GM and WM volume at follow-up. The specific areas where lower GM volumes at follow-up were associated with the MIRS score were the bilateral putamen, thalamus, left caudate, right amygdala, hippocampus, left olfactory cortex, right anterior cingulum, left Rolandic operculum, left superior temporal gyrus, left Heschl’s gyrus, left parietal operculum (particularly the OP4 area), left supramarginal gyrus, left postcentral gyrus, right medial and superior frontal orbital gyrus Figure 2. Voxel-Based Morphometry analyses showing significantly decreased regions in patients compared with HC at both baseline (blue) and follow-up (red). The depicted regions are those that survived multiple comparisons adjusted for age and brain size. Panel 2A) shows the masks where pediatric onset patients obtained lower gray matter values than their corresponding HC-pediatric group. Panel 2B) shows the masks where adult/late onset patients had lower gray matter values than their corresponding HC-adult/late group. The mask at baseline is represented with transparency in order to visualize the areas of overlap between baseline and follow-up (dark blue and dark brown) and the non-overlapping areas (light blue and light brown). ª2020 The Authors. Annals of Clinical and Translational Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association 1809 G. Labayru et al.Neurodegeneration in DM1: A Follow-up MRI Study and bilateral gyrus rectus. The specific areas where lower WM volumes at follow-up were associated with the MIRS score were the left anterior corona radiata, bilateral superior corona radiata, body of corpus callosum and right superior longitudinal fascisulus (see Figure 4 and Supplementary Table S9 for a detailed report on significant clusters on GM and Supplementary Table S10 for WM). Discussion This study tests the hypothesis of a neurodegenerative process in DM1, examining for the first time, the different profiles of structural brain involvement in pediatric and adult/ late DM1. For this purpose, the same patients and controls were followed across a timespan of almost a decade. Figure 3. Partial correlation analyses of neuropsychological T scores with percentage of WM volume loss from baseline to follow-up in adult/late onset DM1 patients. Only the results that survived a false discovery rate (FDR) correction are presented. Figure 4. Gray matter (GM) volume decrease at follow-up associated with MIRS score of adult/late DM1 patients at baseline. The T-statistic showing the relationship between GM volume and MIRS score is displayed. Only results surviving multiple comparisons are shown, correcting for age, head size, and time from baseline assessment to follow-up scan. A black-transparency mask is displayed to show the damage mask of adult/ late onset DM1 compared with HC-adult/late at follow-up. 1810 ª2020 The Authors. Annals of Clinical and Translational Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association Neurodegeneration in DM1: A Follow-up MRI Study G. Labayru et al.