Lesion Evolution and Neurodegeneration in RVCL-S a Monogenic Microvasculopathy

ARTICLE OPEN ACCESS Lesion evolution and neurodegeneration in RVCL-S A monogenic microvasculopathy

Andria L. Ford, MD, MSCI, Victoria W. Chin, BS, Slim Fellah, PhD, Michael M. Binkley, PhD, Allie M. Bodin, BS, Correspondence Vamshi Balasetti, MD, Yewande Taiwo, MS, Peter Kang, MD, Doris Lin, MD, Joanna C. Jen, MD, PhD, Dr. Ford M. Gilbert Grand, MD, Madonna Bogacki, AA, M. Kathryn Liszewski, BA, Dennis Hourcade, PhD, [email protected] Yasheng Chen, DSc, Jason Hassenstab, PhD, Jin-Moo Lee, MD, PhD, Hongyu An, DSc, Jonathan J. Miner, MD, PhD, and John P. Atkinson, MD Neurology® 2020;95:e1918-e1931. doi:10.1212/WNL.0000000000010659

Abstract RELATED ARTICLE Objective Editorial Neuroimaging and To characterize lesion evolution and neurodegeneration in retinal vasculopathy with cerebral fi leukoencephalopathy and systemic manifestations (RVCL-S) using multimodal MRI. cognitive pro le in RVCL-S: Possible new insights into pathogenesis? Methods We prospectively performed MRI and cognitive testing in RVCL-S and healthy control cohorts. Page 611 Gray and white matter volume and disruption of white matter microstructure were quantified. Asymmetric spin echo acquisition permitted voxel-wise oxygen extraction fraction (OEF) MORE ONLINE calculation as an in vivo marker of microvascular ischemia. The RVCL-S cohort was included in CME Course a longitudinal analysis of lesion subtypes in which hyperintense lesions on fluid-attenuated NPub.org/cmelist inversion recovery (FLAIR), T1-postgadolinium, and diffusion-weighted imaging were delineated and quantified volumetrically.

Results Twenty individuals with RVCL-S and 26 controls were enrolled. White matter volume and microstructure declined faster in those with RVCL–S compared to controls. White matter atrophy in RVCL-S was highly linear (ρ = −0.908, p < 0.0001). Normalized OEF was elevated in RVCL-S and increased with disease duration. Multiple cognitive domains, speciﬁcally those measuring working memory and processing speed, were impaired in RVCL-S. Lesion volumes, regardless of subtype, progressed/regressed with high variability as a function of age, while FLAIR lesion burden increased near time to death (p < 0.001).

Conclusion RVCL-S is a monogenic microvasculopathy aﬀecting predominantly the white matter with regard to atrophy and cognitive impairment. White matter volumes in RVCL-S declined linearly, providing a potential metric against which to test the eﬃcacy of future therapies. Progressive elevation of white matter OEF suggests that microvascular ischemia may underlie neurodegeneration in RVCL-S.

From the Department of Neurology (A.L.F., V.W.C., S.F., M.B.M., A.M.B., V.B., Y.T., P.K., Y.C., J.H., J.-M.L.), Mallinckrodt Institute of Radiology (A.L.F., J.-M.L., H.A.), Department of Ophthalmology (M.G.G.), and Department of Medicine (M.B., M.K.L., D.H., J.J.M., J.P.A.), Division of Rheumatology, Washington University School of Medicine, St. Louis, MO; Department of Radiology (D.L.), The Johns Hopkins University School of Medicine, Baltimore, MD; and Department of Neurology (J.C.J.), Icahn School of Medicine at Mount Sinai, New York, NY.

Go to Neurology.org/N for full disclosures. Funding information and disclosures deemed relevant by the authors, if any, are provided at the end of the article.

The Article Processing Charge was funded by NIH. This is an open access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivatives License 4.0 (CC BY-NC-ND), which permits downloading and sharing the work provided it is properly cited. The work cannot be changed in any way or used commercially without permission from the journal. e1918 Copyright © 2020 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Academy of Neurology. Glossary CADASIL = cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy; CBF = cerebral blood flow; cSVD = cerebral small vessel disease; DSST = Digit Symbol Substitution Test; DWI =diffusion-weighted imaging; FA = fractional anisotropy; FCSRT = Free and Cued Selective Reminding Test; FLAIR = fluid-attenuated inversion recovery; MD = mean diffusivity; MoCA = Montreal Cognitive Assessment; NAWM = normal-appearing white matter; nCBF = CBF normalized to gray matter; nOEF = OEF normalized to gray matter; OEF = oxygen extraction fraction; RVCL-S = retinal vasculopathy with cerebral leukoencephalopathy and systemic manifestations.

Retinal vasculopathy with cerebral leukoencephalopathy and effect due to vasogenic edema. Imaging studies in RVCL-S have – systemic manifestations (RVCL-S) is a familial micro- been limited to small case series without controls,8 12 including a vasculopathy leading to early vision loss and progressive neu- recent study of pseudotumor lesions in 6 patients that quantified the rocognitive deterioration in the fourth to sixth decades, duration of diffusion restriction and ring enhancement across serial ultimately fatal within ≈10 years of symptom onset.1 The au- MRI scans.11 tosomal dominant disease is caused by a C-terminal frameshift mutation in TREX1 (a 39–59 repair exonuclease 1),2 with 40 Defining pathogenesis in RVCL-S has been challenged by the families identified worldwide (M.K.L. and J.P.A., unpublished, inability of animal models to recapitulate the human disease phe- 5 2018-2020). In 2016, a multicenter study defined the genetic notype. Brain MRI findings give some insight into mechanisms: ff and clinicopathologic spectra of the disease on the basis of punctate di usion-restricting lesions support a role for microvas- aggregate data in 78 TREX1 mutation carriers from 11 unrelated cular ischemia, while nodular and ring-enhancing lesions suggest families, proposing diagnostic criteria to increase recognition disruption of the blood-brain barrier. In support of ischemia, brain and to facilitate future studies.3 pathology in RVCL-S demonstrates a microvasculopathy associated with multilaminated basement membranes, mural thickening fi Despite recent progress, disease mechanisms in RVCL-S remain and hyalinization, luminal stenosis, and brinoid necrosis, which fi can resemble radiation necrosis.13 Chronic inflammatory cells may poorly de ned. The TREX1 frameshift mutations produce a trun- 3,13 cated TREX1 protein that lacks its anchor to the endoplasmic re- be seen near the microvasculature or rarely in the parenchyma. ticulum.2 A recent study described interactions between the TREX1 In postmortem brain tissue of patients with RVCL-S, TREX1 is carboxy-terminal region and oligosaccharide transferase at the en- readily detectable within microglia surrounding the white matter microvasculature, suggesting that TREX1 mislocalization may play doplasmic reticulum (organelle that promotes normal N-linked gly- 7 4 a role in white matter vulnerability. While immune serologies are cosylation). Experiments with knock-in mice and human cell lines normal in patients, mouse models of RVCL-S induced a non- demonstrated that expression of the truncated TREX1 protein dys- nuclear autoimmunity that was suppressed by a chemotherapeutic regulated the oligosaccharide transferase complex, resulting in free agent, suggesting that immune activation may contribute to path- glycan release, immune activation, and autoantibody production.4,5 ogenesis or progression.4,5 Alternatively, the relatively small nuclear proportion of the truncated TREX1 protein, which harbors a functional 39 DNase, is increased 6 In this study, we evaluated the temporal-spatial dynamics of after apoptotic or genotoxic stress ; thus, genome stability may be neuroimaging features and cognitive performance from a pro- compromised by unregulated activity of the misplaced DNase in its spective cohort of patients with RVCL-S compared to healthy untethered state. Despite these potential disease mechanisms, it is controls. We sought to characterize neurodegeneration and lesion ff unclear why the microvasculature is most markedly a ected in evolution to enable testing of future treatments against this natural RVCL-S. Immunohistochemistry studies in postmortem brain tissue history, to quantify domain-specific cognitive impairment, and to from patients with RVCL-S and patients with stroke without RVCL- apply advanced MRI methods to reveal pathogenic mechanisms, S demonstrated that TREX1 localizes predominantly to white matter hypothesizing that decreased cerebral blood flow (CBF) and in- Iba1+ microglia in association with the microvasculature near ische- creased oxygen extraction fraction (OEF) in RVCL-S would 7 mic lesions, suggesting a role for TREX1 in response to ischemia. support the role of chronic microvascular ischemia.

Due to the rarity of RVCL-S, it has been challenging to evaluate therapeutic efficacy when only a few patients can be tested and Methods without a contemporaneous placebo arm. From the aggregate data above,3 cognitive and psychological impairments have been defined Standard protocol approvals, registrations, as present or absent by history and examination. MRI findings and patient consents demonstrated (1) small/medium T2 hyperintense white matter The studies were approved by the institutional review board. lesions in all but 1 patient (97%), noting that some were associated with diffusion restriction or postcontrast enhancement, and (2) ring- Study participants enhancing pseudotumor lesions in 84%, which are frequently ob- The studies included a prospective cohort with a standard served with advanced disease and may cause life-threatening mass imaging protocol and an older prospective cohort without

Neurology.org/N Neurology | Volume 95, Number 14 | October 6, 2020 e1919 standard imaging (figure 1 shows enrollment overview). For Cognitive and psychological assessment the recent cohort with standard imaging, we enrolled adults On the same day as the MRI scan, cognitive assessment was with genetically confirmed RVCL-S and age-matched con- performed in participants undergoing the standard MRI trols. Exclusion criteria included contraindications to MRI protocol. Tests were selected on the basis of demonstrated and pregnancy. Controls were also excluded for history of reliability and sensitivity to decline in neurodegenerative neurologic or chronic illness. While controls had a single MRI diseases and included the Montreal Cognitive Assessment scan, individuals with RVCL-S had 1 to 5 MRI scans during (MoCA), a global cognitive screening measure sensitive to the study period, typically at 6- to 12-month intervals. Vital executive impairment14; the Wechsler Adult Intelligence signs and laboratory samples, history of medications, and in- Scale–Revised Digit Symbol Substitution Test (DSST),15,16 terim illness were obtained with each scan. Image analyses to for working memory and processing speed; category – measure atrophy, white matter microstructure, and metrics of fluency17 19 (animal and vegetable naming) and the Bushcke blood flow and oxygen metabolism were performed only on and Grober Free and Cued Selective Reminding Test scans acquired with the standard imaging protocol described (FCSRT),20,21 for semantic and episodic memory, re- below. The older cohort without standard imaging included spectively; and Gait Speed,22 a measure of global cognitive individuals with RVCL-S who had been followed up at the and motor processing. Levels of depression and anxiety were Washington University RVCL Research Center (rvcl-re- assessed with the Geriatric Depression Scale23 and the State- search.wustl.edu/) over the past 2 decades in whom MRIs Trait Anxiety Inventory,24 respectively. were obtained as part of standard clinical care. Longitudinal analyses evaluating lesion subtypes over time included all MRI Image protocol and processing scans in patients with RVCL-S (those performed with and without standard imaging). MRI acquisition, segmentation, and coregistration For the standard imaging protocol, controls and participants Common disease features in patients with RVCL-S were with RVCL-S underwent brain MRI on a Siemens 3T Tim assessed by review of the patient’s medical record and were Trio or 3T Biograph mMR scanner (Erlangen, Germany) present if documented by the treating physician at the Wash- with a 12-channel head coil. Standard 3D magnetization- ington University RVCL Research Center. Common neuro- prepared rapid gradient-echo T1 (echo time/repetition time radiologic features were present if described in the radiology 2.95/1,800 milliseconds, inversion time 1,000 milliseconds, report. Systemic features of RVCL-S were defined as: (1) liver flip angle 8°, 1.0 × 1.0 × 1.0 mm), fluid-attenuated inversion disease if alanine transaminase, aspartate transaminase, or al- recovery (FLAIR) (echo time/repetition time 93/9,000 mil- kaline phosphatase was elevated beyond the laboratory refer- liseconds, inversion time 2,500 milliseconds, 1.0 × 0.9 × ence range; (2) anemia if hemoglobin fell below the laboratory 3 mm). Magnetization-prepared rapid gradient-echo images reference range; (3) nephropathy if serum creatinine or urine were segmented into gray and white matter with Statistical protein was elevated; (4) hypertension if the patient required Parametric Mapping version 12.25 Using FMRIB Software daily antihypertensive medications; (5) hypothyroidism if the Library (FSL, Oxford University, Oxford, UK), we segmented patient required thyroid replacement; and (6) Reynaud phe- subcortical nuclei with the FMRIB Integrated Registration nomenon if listed as a chart diagnosis. and Segmentation Tool26 and coregistered images obtained

Figure 1 Participant enrollment scheme

Participants with retinal vasculopathy with cerebral leukoencephalopathy and systemic manifestations (RVCL-S) and healthy controls were prospectively enrolled with a standard imaging protocol. These participants had advanced MRI analyses to evaluate atrophy, white matter microstructure, and metrics of tissue ischemia (CBF, OEF). This RVCL-S cohort was enlarged to include patients with scans dating farther back (without standard imaging), who had been followed over many years. Given that these scans did not have a standard imaging protocol, only the lesion segmentation analysis was performed in the larger cohort. FA = fractional anisotropy; MD = mean diffusivity.

e1920 Neurology | Volume 95, Number 14 | October 6, 2020 Neurology.org/N within a scan session with the FMRIB Linear Image Regis- Statistical analysis tration Tool,27,28 with accuracy confirmed by visual in- Statistical analyses were performed with SAS version 9.4 (SAS spection. Segmented brain volumes were normalized to total Institute Inc, Cary, NC). Data are described with median intracranial volume to adjust for head size, defined as the sum (interquartile range). A Wilcoxon rank-sum test compared of gray matter, white matter, and CSF volumes.29 RVCL-S and control cohorts for several neuroimaging outcomes: gray matter volume, white matter volume, FA, MD, Diffusion tensor imaging nCBF, and nOEF. For all imaging analyses except lesion ff Di usion tensor imaging was acquired as follows: echo time/ subtype analysis (described below) in which patients con- repetition time 89/10,100 milliseconds and isotropic 2.0 × 2.0 × tributed multiple scan time points, the median imaging value fi 2 2.0 mm. Twenty- ve directions were acquired; b = 0 s/mm and and the corresponding median age from the scan time points 2 fi ff b = 1,400 s/mm ( eld of view 224 mm). Di usion tensor para- were used. Raw p values were reported with significance noted metric maps including fractional anisotropy (FA) and mean dif- after adjustment with the Benjamini-Hochberg procedure to ff fusivity (MD) were calculated with the FSL di usion toolbox maintain a family-wise error rate <0.05. To evaluate the in- 30 (Oxford University). Individual mean FA and MD were mea- dependent effects of age, disease, and their interaction on sured in normal-appearing white matter (NAWM) after exclusion neuroimaging outcomes, linear regression was performed. of voxels within the manually delineated FLAIR lesion mask (de- When criteria for normality were not met, data underwent scribed below), such that only nonlesioned tissue was included. natural logarithmic transformation, and multivariate normality was retested. The interaction term was included if signif- CBF and OEF image acquisition and processing icant at p < 0.05; otherwise, the interaction term was removed. We used a pseudocontinuous arterial spin labeling sequence (echo time/repetition time 12/3,780 milliseconds, 3.0 × 3.0 × For cognitive and psychological analyses, a Wilcoxon rank- 5.0 mm, 22 slices, number of averages 80, labeling duration 2,000 sum test compared the RVCL-S and control cohorts. Age at milliseconds, postlabel delay 1,500 milliseconds) to measure testing and years of education were compared. For partici- CBF.31 Blood T1 for CBF calculation was measured in the su- 32,33 pants with RVCL-S with multiple cognitive assessments perior sagittal sinus of each participant as previously described. (performed before each MRI scan), the most recent test result An asymmetric spin echo sequence measured tissue deoxy- 34,35 was used. Because underlying depression and anxiety may hemoglobin, permitting OEF quantification. CBF and OEF ff 37–39 36 a ect cognitive performance, we evaluated whether were calculated as previously described. Individual mean CBF RVCL-S predicted cognitive performance after adjusting for and OEF within gray matter and white matter were calculated. To the Geriatric Depression Scale and State-Trait Anxiety In- ff limit partial volume e ects, voxels with <0.7 probability of being ventory scores. Two linear regression models, one adjusting fi classi ed correctly as gray or white matter after T1 segmentation for depression and one adjusting for anxiety, as predictors of were removed, and a 1-voxel morphologic erosion between tissue each cognitive domain were performed separately (maximum types was applied. To account for participant-related systemic 2 covariates per model) to avoid model overfitting. effects on whole-brain CBF and OEF such as anemia or age, white matter CBF and OEF were normalized to gray matter (nCBF and Only the participants with RVCL-S with at least 3 scan time nOEF, respectively). points over at least 30 months were included for the lesion subtype analysis. Of these 7 patients, 5 had known age at Lesion subtype segmentation death, and 2 were still alive with age at death imputed as the Five lesion subtypes were evaluated for hyperintensity: (1) all median value from all patients with RVCL-S who had died. FLAIR lesions, (2) FLAIR lesions excluding pseudotumors, (3) Repeated patient data were included as a random effect in all FLAIR lesions including only pseudotumors, (4) all T1 post- models. For models not meeting normality assumptions, data gadolinium lesions, and (5) all diffusion-weighted imaging (DWI) underwent transformation before regression with the natural lesions. Hyperintense lesions were manually outlined by a vascular logarithmic function or nonlinear fits, as needed. Across all neurology fellow and board-certified neurologist (V.B.) and study analyses, results were presented only if valid without medical student (V.W.C.) and then reviewed and edited by a collinearity and with normal residuals. board-certified vascular neurologist (A.L.F.) using Medical Image Processing, Analysis, and Visualization software (mipav.cit.nih. Data availability fi gov/). Pseudotumors were de ned as a central T1 ring-enhancing Data are available on request to the corresponding author. lesion located near the lateral ventricles that was associated with surrounding vasogenic edema on FLAIR. While FLAIR lesions were separated into 2 lesion subtypes based on being a pseudo- Results tumor or not (to separate out the effect of vasogenic edema on lesion volume), T1 postgadolinium lesions, also referred to as Cohort characteristics nodular contrast-enhancing lesions, were all included as a single For the prospective study with the standard MRI protocol, 14 lesion group because the volumes between the ring-enhancing individuals with RVCL-S and 26 age-matched healthy con- central lesion of a pseudotumor and other enhancing lesions trols were enrolled. An additional 6 patients with RVCL-S did throughout the white matter do not differ significantly. not have any standard imaging, and this larger cohort (n = 20)

Neurology.org/N Neurology | Volume 95, Number 14 | October 6, 2020 e1921 Table 1 Participant characteristics

Prospective cohort with standard imaging Prospective cohort with and protocol without standard imaging

Healthy controls Patients with RVCL-S Patients with (n = 26) (n = 14) RVCL-S (n = 20)

Age at baseline scan, ya 50 (38, 54) 48 (37, 54) 46 (40, 54)

Age at last scan, y —— 50 (42, 55)

Age at death, y — 59 (59, 59) 56 (53, 60)

Female sex, n (%) 18 (69) 9 (64) 14 (70)

White, n (%)b 16 (62) 14 (100) 20 (100)

No. of scansb 1.0 (1.0, 1.0) 2.0 (1.0, 2.8) 2 (1.0, 4.2)

Duration between baseline and last scan, y — 1.4 (1.0, 1.9) 3.1 (2.6, 5.7)

Common disease features, n (%)c

Retinopathy (retinal examination not performed in one) 9 (69) 15 (79)

Focal neurologic deficitsd 5 (36) 10 (50)

Cognitive impairment 7 (50) 13 (65)

Psychiatric disturbance 9 (64) 15 (75)

Seizures 1 (7) 3 (15)

Migraine headache 7 (50) 12 (60)

Common neuroradiologic features, n (%)e

All FLAIR lesions 11 (79) 17 (85)

Punctate white matter lesions on FLAIR 11 (79) 17 (85)

Pseudotumor ring-enhancing lesions with vasogenic edema on FLAIR 4 (29) 10 (50)

Contrast-enhancing lesions 11 (79) 17 (85)

DWI lesions 8 (57) 14 (70)

Other systemic manifestations, n (%)f

Liver disease 5/11 (45) 9/16 (56)

Anemia 4/14 (29) 7/19 (37)

Nephropathy 6/14 (43) 10/19 (53)

Hypertension requiring medications 7/14 (50) 10/20 (50)

Hypothyroidism requiring medication 4/14 (29) 4/19 (21)

Raynaud phenomenon 3/14 (21) 3/20 (15)

Abbreviations: DWI = diffusion-weighted imaging; FLAIR = fluid-attenuated inversion recovery; RVCL-S = retinal vasculopathy with cerebral leukoencephalopathy and systemic manifestations. Data are shown as median (25th, 75th percentile) as appropriate. a For controls, there was a single scan; thus, only baseline scan age is reported. For patients with RVCL-S with standard imaging, there could be single or multiple scans, and the median imaging values were used. Thus, the median age across scan time points is reported as the age at baseline scan. For the larger RVCL-S cohort with and without standard imaging that was used for the longitudinal lesion analysis, individual scan data were used, so both the age at baseline and age at last scans are reported. b There were no significant differences between patients with RVCL-S and healthy controls except for more Whites (p = 0.002) and a larger number of MRI scans in the RVCL-S group (p < 0.001). c Common disease features were present if documented within the medical record by the treating physician at the Washington University RVCL Research Center. d Focal neurologic deficits included any report of language, visual-spatial, motor, sensory, cranial nerve, coordination, balance, or gait deficits in the chart. The exception was that any vision deficits were not included in this category but were included only in the retinopathy category. e Common neuroradiologic features were present if described in the radiology report or identified on image review by a board-certified vascular neurologist (A.L.F.). f Other systemic manifestations were present as follows: liver disease if alanine transaminase, aspartate transaminase, or alkaline phosphatase was elevated beyond the laboratory reference range; anemia if either hemoglobin or hematocrit was low; nephropathy if either serum creatinine or urine protein was elevated; hypertension if the patient required daily home antihypertensive medications; hypothyroidism if the patient was taking thyroid replacement medication; and Raynaud phenomenon if listed as a clinical diagnosis. Smaller denominators are due to unavailability of laboratory values in some participants.

e1922 Neurology | Volume 95, Number 14 | October 6, 2020 Neurology.org/N Figure 2 White matter, but not gray matter, atrophy is accelerated in RVCL-S

(A) Normalized gray matter (GM) volumes did not differ between healthy controls (CTLs) and individuals with retinal vasculopathy with cerebral leukoencephalopathy and systemic manifestations (RVCL-S). (B) Normal- ized white matter (WM) volumes were significantly lower in those with RVCL- S compared to CTLs (p < 0.0001). (C) Rate of GM atrophy did not differ between CTLs and individuals with RVCL- S. (D) Rate of WM atrophy was faster in individuals with RVCL-S compared to CTLs (p = 0.0002 for interaction of age and RVCL-S status). was used for the longitudinal lesion analysis. Baseline char- lesion volume for each participant’s scan to the white matter acteristics for the study cohorts are shown in table 1. Indi- volume to account for this potential misclassification (patient viduals from 6 unrelated families with RVCL-S were example of misclassification is shown in figure S1, data avail- represented, with 18 of 20 individuals carrying the most able from Dryad, dryad.org/10.5061/dryad.stqjq2c0j). This common TREX1 mutation, V235fs, while 2 individuals carried was performed in participants with RVCL-S and controls the T250Nfs mutation.3 There were no differences in age or alike, although the control participants were free of risk sex between the controls and participants with RVCL-S; factors and were not elderly and therefore had very small however, there was a greater percentage of Whites (p = 0.002) lesion burden (median white matter lesion volume 0.0 and more MRI scans performed (p < 0.001) in the RVCL-S [0.0–0.05] cm3). After doing so, we continued to find re- cohort. duced normalized white matter volumes in participants with RVCL-S compared to controls (0.283 vs 0.316, respectively; White matter volume and microstructure p < 0.0001). decline faster in RVCL-S While atrophy in RVCL-S has been noted qualitatively on We next evaluated the relationship of tissue-type atrophy as a brain imaging, tissue-type atrophy and disruption in white function of age and disease status. The rate of decline in white matter microstructure have not previously been quantified in matter volume was faster in participants with RVCL-S com- RVCL-S. Because lesions occur most commonly, yet not ex- pared to controls (p = 0.0002 for the interaction of age and clusively, in white matter, we hypothesized that white matter RVCL-S status), while rate of gray matter atrophy did not would demonstrate greater atrophy in RVCL-S, even after differ between the 2 cohorts (figure 2, C and D and table 2). accounting for lesion volume and that rate of decline would be Furthermore, the decline in normalized white matter volume greater than that in age-matched controls. White matter vol- in the RVCL-S cohort (n = 14) was highly linear (Spearman umes, normalized to intracranial volume, were reduced in ρ = −0.908, p < 0.0001) compared to controls (ρ = −0.051, p = patients with RVCL-S compared to controls (0.281 vs 0.316, 0.806). In figure 3, 3 patient examples with sequential images respectively; p < 0.0001), whereas normalized gray matter spanning 5 to 10 years demonstrate cerebral atrophy quali- volumes did not differ between the 2 cohorts (0.466 in pa- tatively; however, by visual inspection, it is unclear whether tients vs 0.480 in controls; p = 0.340) (figure 2, A and B). atrophy affects gray matter, white matter, or both. For these 3 Because tissue segmentation techniques are based on T1 patients, quantitative white matter volumes decreased over imaging and may misclassify lesioned tissue within white time (relative decrease 23.5%–29.6% from baseline) com- matter as gray matter, we compared white matter volumes pared to gray matter (relative decrease −4.4% to 6.4% from between patients with RVCL-S and controls after adding the baseline).

Neurology.org/N Neurology | Volume 95, Number 14 | October 6, 2020 e1923 Table 2 White matter atrophy, disruption of white matter microstructure, and white matter OEF are increased in RVCL-S

Dependent variable Predictor β Coefficient (standard error) p Value Log-transformeda Normala

Atrophy

nGM volume RVCL-S status −0.002 (0.012) 0.8385 No Yes

Age −0.0023 (0.000) <0.0001

nWM volume RVCL-S status −0.213 (0.081) 0.0126 Yes Yes

Age −0.006 (0.001) 0.0001

Interaction 0.007 (0.002) 0.0002

WM microstructure (DTI metrics)

FA_NAWM RVCL-S status −0.0095 (0.030) 0.0036 No Yes

Age −0.001 (0.000) 0.0138

Interactionb 0.002 (0.001) 0.0060

MD_NAWM RVCL-S status 0.153 (0.042) 0.0009 No Yes

Age 0.002 (0.001) 0.0228

Interactionb −0.003 (0.001) 0.0011

CBF

nCBF_NAWM RVCL-S status 0.0009 (0.024) 0.707 No Yes

Age 0.002 (0.001) 0.041

OEF

nOEF_NAWM RVCL-S status 0.078 (0.038) 0.0468 Yes Yes

Age 0.003 (0.001) 0.0003

Interactionb −0.002 (0.001) 0.0103

Abbreviations: CBF = cerebral blood flow; DTI = diffusion tensor imaging; FA = fractional anisotropy; MD = mean diffusivity; NAWM = normal-appearing white matter; nCBF = normalized CBF; nGM = normalized gray matter; nOEF = normalized OEF; nWM = normalized WM; OEF = oxygen extraction fraction; RVCL-S = retinal vasculopathy with cerebral leukoencephalopathy and systemic manifestations; WM = white matter. Linear regression evaluated age, RVCL-S disease status, and their interaction as predictors of brain volume, WM microstructure, CBF, and OEF. a Data transformation was performed when normality assumptions were not met. After transformation, normality assumptions were retested. b Interaction term was included as the product of age and RVCL-S status. If the interaction term was nonsignificant, then it was removed from the model.

Next, we evaluated the decrease in FA and increase in MD with white matter CBF may be decreased while OEF would be age as metrics of disrupted white matter microstructure and that increased in patients with RVCL-S compared to healthy con- have been found to precede the development of white matter trols. Representative CBF and OEF maps in a patient with hyperintensities in sporadic cerebral small vessel disease RVCL-S are shown in figure 4A. While nCBF in NAWM did (cSVD).40,41 We measured the effect of RVCL-S disease status not differ between the 2 cohorts, nOEF in NAWM was higher on FA and MD in NAWM after adjusting for age. The reduction in patients with RVCL-S compared to controls (p = 0.009; in FA (p = 0.0060) and increase in MD (p = 0.0011) were faster figure 4B). We further evaluated whether nCBF decreased and in participants with RVCL-S compared to controls (p values nOEF increased over time, suggestive of progressive white given for interaction of age and disease status) (table 2). matter ischemia with disease duration. The interaction of age and RVCL-S status predicted increasing OEF in NAWM (p = White matter OEF is elevated in RVCL-S and 0.0103), while the interaction did not predict change in CBF increases with disease duration (figure 4, C and D, and table 2). The progressive elevation in While the mechanisms underlying retinal and neuro- white matter OEF with disease duration may indicate chronic degeneration in RVCL-S are unclear, pathologic findings con- small vessel ischemia as an underlying disease mechanism. sistently demonstrate a small vessel vasculopathy associated with multilaminated basement membranes, mural thickening Cognitive and psychological assessment and hyalinization, luminal stenosis, and fibrinoid necrosis.3,13 in RVCL-S To investigate in vivo signs of small vessel ischemia in RVCL-S, While individuals with RVCL-S may develop lacunar we measured voxel-wise CBF and OEF. We hypothesized that stroke with acute focal neurologic deficits, more e1924 Neurology | Volume 95, Number 14 | October 6, 2020 Neurology.org/N Figure 3 Progression in cerebral atrophy and evolution of lesion subtypes in RVCL-S

Top row shows fluid-attenuated inversion recovery (FLAIR) images; middle row shows diffusion-weighted imaging (DWI) images; bottom row shows T1 postgadolinium images. (A) A 45-year-old woman with retinal vasculopathy with cerebral leukoencephalopathy and systemic manifestations (RVCL-S) demonstrated a pseudotumor adjacent to the right frontal horn. The pseudotumor demonstrates an underlying ring-enhancing lesion on T1 postgadolinium with central hyperintensity on DWI and surrounding vasogenic edema on FLAIR. MRI scan a decade later demonstrates that the previous pseudotumor is inactive without central diffusion restriction, contrast enhancement, or significant vasogenic edema on FLAIR. Diffuse central > peripheral atrophy is prominent, and additional white matter lesions have developed a decade after the baseline MRI. (B) MRI scan of a 54-year-old woman with RVCL-S shows multiple subcortical and periventricular white matter lesions, several which are hyperintense on DWI and a few of which show nodular enhancement on T1 postgadolinium. Five years later, her MRI scan just before death demonstrates a relative decrease/regression in number and overall burden of white matter lesions on FLAIR, DWI, and T1 postgadolinium, with greater lesion burden near the ventricles and less within the subcortical white matter; however, substantial atrophy (central and peripheral) has progressed over the 5 years before death. Sequential neuroimaging on this patient never revealed any pseudotumors. (C) A 43-year-old woman with RVCL-S demonstrates periventricular and subcortical white matter lesions on FLAIR imaging, several, but not all, of which are bright on DWI and show nodular enhancement on T1 postgadolinium. Follow-up MRI scan 7 years later near the time of death shows substantial progression in FLAIR lesion burden with overall decrease in DWI hyperintense lesions. Furthermore, nodular enhancement of white matter lesions has diminished, but a ring-enhancing pseudotumor lesion has appeared adjacent to the right frontal horn. Notably, there is substantial progression of atrophy (central > peripheral) over the 7-year interval. commonly, individuals experience subclinical cognitive decline.3 performance, we evaluated whether RVCL-S was an independent It has been unclear which cognitive domains are most affected in predictor of cognitive impairment after adjusting for the Geriatric RVCL-S and how underlying depression and anxiety affect cog- Depression Scale and State-Trait Anxiety Inventory scores in nitive performance. A standard cognitive assessment was per- separate models. When adjusted for depression, RVCL-S formed in 12 and 11 participants in the RVCL-S and control remained an independent predictor of semantic fluency (β cohorts, respectively (table 3). There were no differences in age at [95% confidence intervals] = −12.3 [−5.3 to −19.3], p = 0.002) testing (p = 0.740) and years of education (p = 0.379) in the and processing speed (DSST score) (β = −13.1 [−1.4 to −24.8]; p cohorts undergoing cognitive testing. Of the items tested, se- = 0.030) but did not independently predict MoCA score, FCSRT mantic memory (tested as category fluency or the number of score, or gait speed. When adjusted for anxiety, RVCL-S remained animals and vegetables named in 2 minutes) demonstrated the independently predictive of semantic memory (β = −12.9 [−6.4 to greatest difference between cohorts, with a median of 27 words in −19.4]; p = 0.0006), DSST score (β = −16.4 [−5.1 to −27.8]; p = participants with RVCL-S and 42 words in controls (p = 0.0009). 0.007), and gait speed (β = −0.266[−0.025 to −0.506]; p = 0.032) Participants with RVCL-S scored lower on processing speed but did not independently predict MoCA or FCSRT score. measured by DSST (p = 0.005) and gait speed (p = 0.006). On psychological evaluation, the Geriatric Depression Scale score was Burden of FLAIR lesions in RVCL-S increases higher (p = 0.008) with a nonsignificant increase in State-Trait near time to death Anxiety Inventory scores (p = 0.056) in those with RVCL-S. To evaluate the temporal evolution of lesion subtypes in Given that increased depression and anxiety affect cognitive RVCL-S, MRI scans from participants with RVCL-S (n = 20)

Neurology.org/N Neurology | Volume 95, Number 14 | October 6, 2020 e1925 Figure 4 White matter OEF increases with disease duration and may indicate progressive microvascular ischemia

(A) Representative cerebral blood flow (CBF), oxygen extraction fraction (OEF), and fluid-attenuated inversion recovery (FLAIR) maps in an individual with retinal vasculopathy with cerebral leukoencephalopathy and systemic manifestations (RVCL-S). (B) Normalized white matter OEF (nOEF_ NAWM), but not normalized white matter CBF (nCBF_NAWM), was significantly elevated in patients with RVCL-S compared to healthy controls (CTLs). (C) nOEF_NAWM progressively increased in participants with RVCL-S compared to CTLs (p = 0.0103); however, rate of change in nCBF_NAWM was not different in those with RVCL-S compared to CTLs.

were evaluated for hyperintensity across 5 lesion types: (1) all analysis, with repeated patient data included as a random effect. FLAIR lesions, (2) FLAIR lesions excluding pseudotumors, Seven participants and 60 MRI scans were included with a (3) FLAIR pseudotumor lesions only, (4) T1 postgadolinium median (interquartile range) between scans of 0.5 (0.4–1.0) lesions, and (5) DWI lesions. Figure 3 shows 3 individuals years and total duration of imaging follow-up of 5 (3–6.5) with RVCL-S and MRIs spanning 5 to 10 years, including years. Age as a predictor of lesion volume did not produce valid representative maps for FLAIR (row 1), DWI (row 2), and T1 models with or without data transformation or nonlinear postgadolinium (row 3). The examples qualitatively demon- methods (data available from Dryad, figure S2, dryad.org/10. strate how FLAIR, DWI, and T1 gadolinium-enhancing le- 5061/dryad.stqjq2c0j). However, time to death was a signifi- sions progress and regress over time. cant predictor of lesion volume for all FLAIR lesions and for FLAIR lesions excluding pseudotumors (figure 5). FLAIR le- To evaluate whether total lesion volume changed with age or sions including pseudotumors increased at a rate of 22%/y near time to death, individuals with ≥3 scan time points across closer to death (p < 0.0001). Similarly, FLAIR lesions excluding a minimum of 30 months were included in linear regression pseudotumors increased at a rate of 15%/y closer to death (p < e1926 Neurology | Volume 95, Number 14 | October 6, 2020 Neurology.org/N Table 3 Cognitive and psychological testing in patients with RVCL-S compared to healthy controls

Healthy controls (n = 11) Patients with RVCL-Sa (n = 12) p Valueb

Age at testing, y 54 (50, 56) 52 (42, 58) 0.740

Education, y 16 (15, 16) 15 (12, 16) 0.379

MoCA score 29 (26, 29) 25 (24, 27) 0.055

DSST, No. of symbols completed in 90 s 52 (47, 54) 32 (21, 48) 0.005a,b,c

Category fluency, No. of words in 2 min 42 (39, 45) 27 (24, 32) 0.0009a,b,c

FCSRT: free recall score 37 (34, 39) 33 (28, 34) 0.032b

Gait speed, m/s 1.32 (1.23, 1.40) 1.15 (0.90, 1.2) 0.006a,b

Geriatric Depression Scale score 2 (0, 4) 16 (5, 18) 0.008b

State-Trait Anxiety Inventory score 54 (47, 64) 78 (59, 92) 0.056

Abbreviations: DSST = Digit Symbol Substitution Test; FCSRT = Bushcke and Grober Free and Cued Selective Reminding Test; MoCA = Montreal Cognitive Assessment; RVCL-S = retinal vasculopathy with cerebral leukoencephalopathy and systemic manifestations. Data are shown as median (25th, 75th percentile). a RVCL-S disease status remained a significant predictor of cognitive test score after adjustment for State-Trait Anxiety Inventory score using multivariable linear regression modeling. b Significance after adjustment with the Benjamini-Hochberg procedure to maintain a family-wise error rate <0.05. c RVCL-S disease status remained a significant predictor of cognitive test score after adjustment for Geriatric Depression Scale score using multivariable linear regression modeling.

0.0001). Models including time to death as a predictor of Prior studies have focused on lesion characterization in pseudotumors only, contrast-enhancing lesions, or DWI lesion RVCL-S without attention to tissue-type atrophy. Our work burden were invalid without meeting normality assumptions reveals that white matter atrophy is a prominent feature in even after data transformation. Of all lesion subtypes, DWI and RVCL-S. Indeed, the decline in white matter volumes oc- contrast-enhancing lesions were the most common to regress. curred earlier than expected, appearing to diverge from controls between 30 and 40 years of age, before substantial lesion To evaluate the onset of lesion appearance in RVCL-S, development (figure 2 and table 2), suggesting earlier onset of lesion volumes were plotted across lesion subtypes as a microstructural disease than previously known. Moreover, the function of age for the entire RVCL-S cohort (data avail- decline in white matter volume with age was highly linear, able from Dryad, figure S3, dryad.org/10.5061/dryad. suggesting that normalized white matter volume could pro- stqjq2c0j). Individuals were more likely to have brain le- vide a metric to demonstrate slowed neurodegeneration in sions after 40 years of age regardless of lesion subtype; future clinical trials. Such a neuroimaging outcome has pre- however, data are limited at younger ages. Five patients viously been unavailable for the therapies that have been (including 9 MRI scans) at <40 years of age showed min- suggested or tried.3,4,42 imal FLAIR lesion burden (median [interquartile range] 0.16 [0.02–0.37] cm3), with 2 individuals having no brain Prior studies have reported the prevalence of cognitive (56%) lesions. Lesions were common after 40 years of age (me- and psychological (42%) impairment by chart review.3 Our dian [interquartile range] 4.9 [1.5–18.3] cm3); 67 scans findings demonstrate impairment in specific cognitive do- demonstrated ≥1 FLAIR lesions. mains, with the largest impairments identified for timed tests of working memory and processing speed (table 3). It is likely that the white matter injury observed in RVCL-S may be Discussion contributing to deterioration in processing and psychomotor speed, as has been shown in other neurodegenerative diseases ff – RVCL-S is devastating and untreatable. Identifying e ective characterized by white matter changes.43 46 The Geriatric therapies will likely require an in-depth understanding of Depression Scale score was significantly higher in RVCL-S, molecular mechanisms. The current study provides the early which negatively affects processing speed, although semantic stages of a neuroradiologic natural history of RVCL-S. By fluency and processing speed continued to be affected by fi pro ling participants across a wide age range, we modeled the RVCL-S even after adjustment for underlying depression and rate of change in key neuroradiologic features such as cerebral anxiety. atrophy, disruption of white matter microstructure, and lesion subtypes. Furthermore, we obtained a standard cognitive The largest cohort of RVCL-S has previously been described battery in patients with RVCL-S and controls to determine with a high prevalence of both small/medium FLAIR hyper- which domains are affected most by the disease. intensities and ring-enhancing pseudotumors.3 We in

Neurology.org/N Neurology | Volume 95, Number 14 | October 6, 2020 e1927 Figure 5 Lesions in RVCL-S, regardless of subtype, progress/regress with high interparticipant variability

Each line represents an individual patient. As time to death decreases, fluid-attenuated inversion recovery (FLAIR) lesion burden accelerates. To evaluate the temporal evolution of lesion subtypes in retinal vasculopathy with cerebral leukoencephalopathy and systemic manifestations (RVCL-S), MRI scans from 7 patients with RVCL-S who contributed ≥3 time points over at least 30 months were evaluated across 5 lesion subtypes. Time to death was evaluated as a predictor of lesion volume in linear regression analysis with repeated patient data included as a random effect. As time to death decreased, FLAIR lesion volumes increased, including (A) a 22% increase in lesion volume per year closer to death for all FLAIR lesions (p < 0.0001) and (B) a 15% increase in lesion volume per year closer to death for FLAIR lesions excluding pseudotumors. Time to death as a predictor of (C) pseudotumor lesion volume on FLAIR, (D) contrast-enhancing lesion volume, and (E) diffusion-weighted imaging (DWI) hyperintense lesion volume produced invalid models. Significant fluctuation in lesion burden for these last 3 lesion subtypes is evident, suggesting high interparticipant variability and limited sample size.

addition found that regardless of pseudotumors, patients a likely pathogenic mechanism. Regional elevation in ce- demonstrate nodular contrast-enhancing lesions in 84% and rebral OEF measured by brain MRI or PET has provided a DWI hyperintense lesions in 68%. The appearance of small, biomarker of ischemic vulnerability in several clinical set- subcortical DWI hyperintense and contrast-enhancing lesions tingssuchascarotidocclusivedisease,48 acute ischemic that appear and regress over time may suggest chronic, pro- stroke,49 and sickle cell anemia.36,50 Along with other ad- gressive microvascular ischemia. While the contrast- vanced MRI methods that provide biomarkers of in vivo enhancing and DWI lesion burden fluctuated over time and pathophysiology such as quantifying cerebrovascular re- was not predicted by age or time to death, the FLAIR lesions activity and white matter microstructure, we propose that accumulated significantly and nonlinearly as time to death regional OEF may provide a metric of microvascular health decreased (figure 5). Assessing the rate of FLAIR lesion in RVCL-S and other forms of cSVD. In support of our progression may assist in determining the prognosis or ap- findings implicating chronic microvascular ischemia, propriate treatments targeting later stages of disease. Finally, pathologic studies have described a small vessel vascul- we found that FLAIR lesion volumes were very low before 40 opathy in RVCL-S associated with multilaminated base- years of age (data available from Dryad, figure S3, dryad.org/ ment membranes, mural thickening and hyalinization, 10.5061/dryad.stqjq2c0j). This finding is limited by having luminal stenosis, and fibrinoid necrosis. Consistent with a only 5 patients imaged before 40 years of age; however, low selective vulnerability of white matter in RVCL-S, Kothari lesion volumes were also identified in a cohort of 33 mutation et al.7 demonstrated numerous TREX1-expressing micro- carriers from the Netherlands in whom FLAIR lesion volume glia in association with the microvasculature in both le- in individuals <40 years of age (n = 11) was 0.19 cm3, similar sioned white matter and NAWM, but much less so in gray to our median volume of 0.16 cm3.47 matter. In support of an endothelial activation phenotype in RVCL-S, von Willebrand factor antigen and propeptide, White matter OEF was elevated in the RVCL-S cohort, which and angiopoietin-2 were elevated in 3 Dutch families with increased over time (figure 4), suggesting chronic ischemia as RVCL-S.e51 Another study evaluated vascular reactivity by e1928 Neurology | Volume 95, Number 14 | October 6, 2020 Neurology.org/N measuring flow-mediated dilation of the brachial artery, standardize neuroimaging protocols across RVCL-S centers finding a reduction in percent change in arterial diameter worldwide may help expand and refine our findings. For ex- in patients with RVCL-S.e52 Therefore, increasing evi- ample, in addition to white matter atrophy, there may be an dence suggests endothelial damage and ischemia as un- increase in the rate of gray matter atrophy that may be de- derlying pathogenic mechanisms, with a potential role for tectable in larger cohorts. Furthermore, larger longitudinal TREX1-expressing microglia or macrophages.7 imaging studies in RVCL-S cohorts will allow evaluation of the phenotypic variability such as how sex, kindred, mutation, Unexpectedly, we found that white matter CBF was not lower in and other vascular comorbid conditions affect the onset and patients with RVCL-S compared to controls. We postulate that progression of neurodegeneration and lesion evolution. ischemia early in disease leads to regions of impaired vascular autoregulation that prevent the tissue from finely regulating Acknowledgment CBF, leading to greater fluctuations in CBF but not a net lower The authors thank the patients and families with RVCL-S and CBF. Local inability to regulate CBF may lead to intermittent healthy controls who have participated in this research. ischemia when systemic blood pressure fluctuations drop local CBF, leading to a compensatory elevation in regional OEF and Study funding eventually tissue infarction. Similar to our findings, in sporadic This work was supported by Clayco Foundation, Energy 4 A Cure cSVD, CBF in NAWM has typically not been low compared to Foundation, Cure CRV Research, NIH: National Heart, Lung, and controls.e53,e54 Measures of degree of impairment in cerebral Blood Institute (R01HL129241, A.L.F. and H.A.), NIH: National vasoreactivity, but not baseline CBF, have been predictive of Institute of Neurological Disorders and Stroke (R01NS116565, disease progression in sporadic and in another inherited cSVD, A.L.F. and H.A.), and Washington University St. Louis Clinical and cerebral autosomal dominant arteriopathy with subcortical in- Translational Science Award (UL1 TR000448, A.L.F.). − farcts and leukoencephalopathy (CADASIL).e55 e57 Disclosure As one of several monogenic microvasculopathies, RVCL-S The authors report no disclosures relevant to the manuscript. may provide an additional cSVD model to study vascular Go to Neurology.org/N for full disclosures. contributions to dementia as the second leading cause of dementia and a key contributor to Alzheimer dementia.e58 Publication history Rapid neurodegeneration in RVCL-S may facilitate studies of Received by Neurology October 10, 2019. Accepted in final form the mechanisms and treatments for sporadic cSVD, which April 10, 2020. progresses later in life and over a longer duration. Microvas- cular pathology in genetic forms of cSVD such as RVCL-S and CADASIL has demonstrated findings qualitatively similar to Appendix Authors those seen in sporadic cSVD but with more severe pathology seen in the hereditary forms,e59 suggesting that cSVD of di- Name Location Contribution fi verse origins may have a common nal pathway toward Andria L. Washington University Conceived and designed the microvasculopathy and white matter degeneration. Ford, MD, School of Medicine, St. study, acquired the data, MSCI Louis, MO performed the analysis, and drafted the manuscript Given the rarity of RVCL-S with <200 individuals identified Victoria W. Washington University Acquired the data and worldwide, our work provides a relatively large cohort scan- Chin, BS School of Medicine, St. performed the analysis ned with standard imaging at multiple time points. However, Louis, MO there are limitations to consider. First, rates of white matter Slim Fellah, Washington University Acquired the data, performed volume, DTI, CBF, and OEF change were obtained by PhD School of Medicine, St. the analysis, and edited the regressing median values per individual by age and disease Louis, MO manuscript status. This analysis was chosen (1) to reduce the noise that Michael M. Washington University Acquired the data and occurs in the setting of sequential imaging over a short du- Binkley, PhD School of Medicine, St. performed the statistical Louis, MO analysis ration and (2) because only single imaging sessions were obtained in the control cohort. We plan to acquire more data Allie M. Washington University Acquired the data and fi Bodin, BS School of Medicine, St. performed the analysis in both cohorts over a longer duration to con rm and improve Louis, MO the accuracy of our findings. Another limitation is that voxel- Vamshi University of Missouri Acquired the data and wise measurements of CBF using pseudocontinuous arterial Balasetti, School of Medicine, performed the analysis spin labeling have low signal-to-noise ratio in the deep white MD Columbia, MO ff matter, which may limit our ability to detect di erences in Yewande Washington University Acquired the data and white matter CBF between the 2 cohorts. Previous work has Taiwo, MS School of Medicine, St. performed the analysis investigated this limitation, finding that the signal remains Louis, MO e60 adequate for the majority of white matter. Moreover, we Peter Kang, Washington University Acquired the data, performed used the recommended parameters that resulted from this MD School of Medicine, St. the analysis, and edited the Louis, MO manuscript work to optimize white matter signal. Future efforts to Continued Neurology.org/N Neurology | Volume 95, Number 14 | October 6, 2020 e1929 11. Hedderich DM, Lummel N, Deschauer M, et al. Magnetic resonance imaging char- Appendix (continued) acteristics of retinal vasculopathy with cerebral leukoencephalopathy and systemic manifestations. Clin Neuroradiol 2020;30:229–236. 12. Dhamija R, Schiff D, Lopes MB, Jen JC, Lin DD, Worrall BB. Evolution of brain Name Location Contribution lesions in a patient with TREX1 cerebroretinal vasculopathy. Neurology 2015;85: 1633–1634. Doris Lin, Johns Hopkins Acquired the data, performed 13. Kolar GR, Kothari PH, Khanlou N, Jen JC, Schmidt RE, Vinters HV. 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Frontal assessment battery oxygen extraction: stages of cerebral haemodynamic impairment revisited. Brain as a useful tool to differentiate mild cognitive impairment due to subcortical ischemic 2002;125:595–607. vascular disease from Alzheimer disease. Dement Geriatr Cogn Disord 2016;42: 49. Liu Z, Li Y. Cortical cerebral blood flow, oxygen extraction fraction, and metabolic 331–341. rate in patients with middle cerebral artery stenosis or acute stroke. AJNR Am J 45. Jacobs HI, Leritz EC, Williams VJ, et al. Association between white matter micro- Neuroradiol 2016;37:607–614. structure, executive functions, and processing speed in older adults: the impact of 50. Fields ME, Guilliams KP, Ragan DK, et al. Regional oxygen extraction predicts border vascular health. Hum Brain Mapp 2013;34:77–95. zone vulnerability to stroke in sickle cell disease. Neurology 2018;90:e1134–e1142. 46. Turken A, Whitfield-Gabrieli S, Bammer R, Baldo JV, Dronkers NF, Gabrieli JD. Cognitive processing speed and the structure of white matter pathways: convergent References e51-e60 are available at Dryad, https://doi.org/10.5061/dryad.stqjq2c0j.

Neurology.org/N Neurology | Volume 95, Number 14 | October 6, 2020 e1931 Lesion evolution and neurodegeneration in RVCL-S: A monogenic microvasculopathy Andria L. Ford, Victoria W. Chin, Slim Fellah, et al. Neurology 2020;95;e1918-e1931 Published Online before print September 4, 2020 DOI 10.1212/WNL.0000000000010659

This information is current as of September 4, 2020

Updated Information & including high resolution figures, can be found at: Services http://n.neurology.org/content/95/14/e1918.full

References This article cites 50 articles, 10 of which you can access for free at: http://n.neurology.org/content/95/14/e1918.full#ref-list-1 Citations This article has been cited by 5 HighWire-hosted articles: http://n.neurology.org/content/95/14/e1918.full##otherarticles Subspecialty Collections This article, along with others on similar topics, appears in the following collection(s): All Cerebrovascular disease/Stroke http://n.neurology.org/cgi/collection/all_cerebrovascular_disease_strok e MRI http://n.neurology.org/cgi/collection/mri Vascular dementia http://n.neurology.org/cgi/collection/vascular_dementia Errata An erratum has been published regarding this article. Please see next page or: /content/96/19/919.1.full.pdf Permissions & Licensing Information about reproducing this article in parts (figures,tables) or in its entirety can be found online at: http://www.neurology.org/about/about_the_journal#permissions Reprints Information about ordering reprints can be found online: http://n.neurology.org/subscribers/advertise

Neurology ® is the official journal of the American Academy of Neurology. Published continuously since 1951, it is now a weekly with 48 issues per year. Copyright Copyright © 2020 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Academy of Neurology.. All rights reserved. Print ISSN: 0028-3878. Online ISSN: 1526-632X. Disputes & Debates: Editors’ Choice

Steven Galetta, MD, FAAN, Editor Aravind Ganesh, MD, DPhil, FRCPC, Deputy Editor Ariane Lewis, MD, Deputy Editor James E. Siegler III, MD, Deputy Editor

Editors’ Note: Association of Initial Imaging Modality and Futile Recanalization After Thrombectomy In “Association of Initial Imaging Modality and Futile Recanalization After Thrombec- tomy,” Meinel et al. reported that the rate of futile recanalization (defined as 90-day modified Rankin Scale (mRS) score 4–6 despite successful recanalization) among patients enrolled in the BEYOND-SWIFT multicenter, retrospective observational registry was significantly higher in patients selected for thrombectomy based on a CT scan as compared with patients selected for thrombectomy based on an MRI. Siegler and Thon cautioned that these findings should not dissuade against (1) use of a CT scan to determine eligibility for thrombectomy because this could lead to unnecessary delays in care or (2) performance of thrombectomy after CT scans because the imaging modality itself does not directly affect outcome and 40% of patients selected for thrombectomy based on a CT scan had a good 90-day mRS score (which represents a 22%–27% absolute increase compared with patients treated with medical management in the DAWN and DEFUSE-3 trials). Meinel et al agreed that it is imperative to avoid delays before thrombectomy and reinforced that their findings should not influence the selection of imaging modality for potential thrombectomy candidates. They also pointed out that implementation of high-speed MRI protocols should be considered to obtain more detailed information than a CT scan can provide while avoiding lengthy delays between imaging and groin puncture.

Ariane Lewis, MD, and Steven Galetta, MD Neurology® 2021;96:915. doi:10.1212/WNL.0000000000011904

Reader Response: Association of Initial Imaging Modality and Futile Recanalization After Thrombectomy

James E. Siegler (Camden, NJ) and Jesse M. Thon (Camden, NJ) Neurology® 2021;96:915–916. doi:10.1212/WNL.0000000000011898

We read with interest the results of the BEYOND-SWIFT retrospective observational registry by Meinel et al.1 Determination of candidacy for acute stroke intervention is a rapidly evolving and highly controversial ﬁeld because we have seen in recently published trials, which are expanding eligibility criteria.

We would respectfully caution the authors to avoid using language such as “there is a need to reduce [futile recanalization]” in the opening of their article and to reiterate it verbatim in the Discussion because these are misleading proclamations. Certainly, it remains a priority among all clinicians to provide cost-eﬀective care with the aim of improving functional outcomes and/or satisfying end-of-life preferences. However, to leverage “futile recanalization” to justify whether CT or MRI-based modalities be used in selecting thrombectomy candidates will undoubtedly bias clinicians in their imaging recommendations (leading to delays in care, as the investigators have shown). It may also falsely inﬂuence a clinician’s determination of thrombectomy eligibility. To conclude that CT-based selection of thrombectomy candidates results in a higher rate of futile

Author disclosures are available upon request ([email protected]).

Neurology.org/N Neurology | Volume 96, Number 19 | May 11, 2021 915 Copyright © 2021 American Academy of Neurology. Unauthorized reproduction of this article is prohibited. thrombectomy when compared with MRI-based methods (although accurate) is inherently misleading. We do not question the eﬀect of imaging modality on treatment-related outcome. However, even among CT-based selection of patients from BEYOND-SWIFT, 40% of patients achieved functional independence by 90 days. This is a marginal 5%–9% decrement from what was observed in DAWN2 and DEFUSE-3,3 and (more importantly) a 22%–27% absolute increase in the rate of functional achievement of these patients when compared to medically managed controls from these trials.

To suggest that these patients are “futile recanalizers” is a misrepresentation and decontextu- alization of the data. Such claims could impede the care of many treatment responders, potentially growing the number of disabled stroke survivors or even resulting in medicolegal consequences for clinicians who justify treatment deferral based on these ﬁndings. It is worth- while to consider CT versus MRI in predicting the long-term outcome after thrombectomy, but knowing that time is a critical determinant of functional outcomes in treated large vessel oc- clusion4 should encourage clinicians to rely on the quickest imaging assessment—in this case, CT.

1. Meinel TR, Kaesmacher J, Mosimann PJ, et al. Association of initial imaging modality and futile recanalization after thrombectomy. Neurology 2020;95:e2331–e2342. 2. Nogueira RG, Jadhav AP, Haussen DC, et al. Thrombectomy 6 to 24 hours after stroke with a mismatch between deﬁcit and infarct. N Engl J Med 2018;378:11–21. 3. Albers GW, Marks MP, Kemp S, et al. Thrombectomy for stroke at 6 to 16 hours with selection by perfusion imaging. N Engl J Med 2018;378:708–718. 4. Jahan R, Saver JL, Schwamm LH, et al. Association between time to treatment with endovascular reperfusion therapy and outcomes in patients with acute ischemic stroke treated in clinical practice. JAMA 2019;322:252–263.

Author Response: Association of Initial Imaging Modality and Futile Recanalization After Thrombectomy

Thomas Raphael Meinel (Bern, Switzerland), Johannes Kaesmacher (Bern, Switzerland), and Urs Fischer (Bern, Switzerland) Neurology® 2021;96:916–917. doi:10.1212/WNL.0000000000011900

We thank the authors for their comment in response to our article.1 In their letter, they point out an important issue, namely, the questionable term “futile intervention.” This term has been used as successful recanalization despite an unfavorable outcome at 3 months.2,3 However, the deﬁnition of an unfavorable outcome has varied, and patient and caregiver perspectives are quite heterogeneous regarding what to consider an unfavorable outcome.

We fully agree with the authors that time delays seen in MR imaging are to be avoided and further efforts are necessary to close the gap in the onset-to-groin time between CT and MRI patients. Given the vast amount of high-speed MRI protocols published, this seems feasible. However, we do not agree that our article suggests that we generally favor MRI over CT for suspected thrombectomy candidates. In fact, we did question the effect of the imaging modality on the overall outcome, stating that “the main aim of this study was to sensitize stroke physicians that apparently the imaging modality influences their decisions regarding which patients to treat by MT. Whether this results in an overall better, worse, or equal outcome can only be judged by upcoming RCTs on this issue.”

Furthermore, we do not think that it is appropriate to compare the results of this cohort including early time-window patients to the DAWN and DEFUSE-3 population. Neither did we call only CT patients as “futile recanalizers.” Rather, we considered both MRI and CT patients that had a modiﬁed Rankin Scale of 4–6 at 3 months with a sensitivity analysis

Author disclosures are available upon request ([email protected]).

916 Neurology | Volume 96, Number 19 | May 11, 2021 Neurology.org/N Copyright © 2021 American Academy of Neurology. Unauthorized reproduction of this article is prohibited. considering only mRS 5–6 at 3 months as futile. This was done because there may be a higher consensus among physicians and also patients that mRS 5–6 despite successful reperfusion constitutes a futile treatment (5-Year QUALE 0.05).4 Given the fact that current guidelines judge MRI and CT equivalent,5 we doubt that an established MRI workﬂow will lead to medicolegal consequences.

We fully agree that time delays should be avoided and a potentially life-saving treatment should be offered to any patient fulfilling criteria for endovascular stroke treatment. Clinicians should be aware that the initial imaging modality might affect their treatment decisions. Whether MR imaging as compared with CT imaging leads to over-selection and time delays or that it harbors the potential to reduce futile interventions without over-excluding patients can only be evaluated in upcoming randomized controlled trials. Until then, our study should not influence clinicians to change the imaging modality for suspected thrombectomy candidates.

1. Meinel TR, Kaesmacher J, Mosimann PJ, et al. Association of initial imaging modality and futile recanalization after thrombectomy. Neurology 2020;95:e2331–e2342. 2. Espinosa de Rueda M, Parrilla G, Manzano-Fern´andez S, et al. Combined multimodal computed tomography score correlates with futile recanalization after thrombectomy in patients with acute stroke. Stroke 2015;46:2517–2522. 3. Hussein HM, Georgiadis AL, Vazquez G, et al. Occurrence and predictors of futile recanalization following endovascular treatment among patients with acute ischemic stroke: a multicenter study. AJNR Am J Neuroradiol 2010;31:454–458. 4. Ganesh A, Luengo-Fernandez R, Pendlebury ST, Rothwell PM. Weights for ordinal analyses of the modiﬁed Rankin Scale in stroke trials: a population-based cohort study. EClinicalMedicine 2020;23:100415. 5. Powers WJ, Rabinstein AA, Ackerson T, et al. Guidelines for the early management of patients with acute ischemic stroke: 2019 update to the 2018 guidelines for the early management of acute ischemic stroke: a guideline for Healthcare Professionals from the American Heart Association/American Stroke Association. Stroke 2019;50:e344–e418.

Editors’ Note: Comparison of Ice Pack Test and Single-Fiber EMG Diagnostic Accuracy in Patients Referred for Myasthenic Ptosis In “Comparison of Ice Pack Test and Single-Fiber EMG Diagnostic Accuracy in Patients Referred for Myasthenic Ptosis,” Giannoccaro et al. compared the diagnostic accuracy of the ice pack test (IPT) with single-fiber EMG (SF-EMG) on the orbicularis oculi muscle in 155 patients, with ptosis being evaluated for ocular myasthenia defined as a positive response to the edrophonium test, presence of acetylcholine-receptor antibodies, a decrement of >10% of the third to fifth compound muscle action potential after repetitive nerve stimulation, or unequivocal response to oral steroids or acetylcholinesterase inhibitors for at least 3 months. They found that the IPT and SF-EMG had similar diagnostic accuracy in this patient population. Silvestri noted interest that (1) the IPT, a simple and cheap bedside assessment, is comparable with a complicated test like the SF-EMG; (2) the results of the IPT and SF-EMG were discordant for 10% of subjects (mostly in the setting of mild or isolated ptosis); and (3) the utility of the IPT was not evaluated in patients with isolated diplopia. Giannoccaro and Liguori responded that the discordance in results generally occurred in the setting of a negative IPT, which may be related to the lack of repetition of the test. They also cited previously published data that the IPT is useful in patients with isolated diplopia, particularly because SF-EMG may be negative in these patients.

Ariane Lewis, MD, and Steven Galetta, MD Neurology® 2021;96:917. doi:10.1212/WNL.0000000000011905

Author disclosures are available upon request ([email protected]).

Neurology.org/N Neurology | Volume 96, Number 19 | May 11, 2021 917 Copyright © 2021 American Academy of Neurology. Unauthorized reproduction of this article is prohibited. Reader Response: Comparison of Ice Pack Test and Single-Fiber EMG Diagnostic Accuracy in Patients Referred for Myasthenic Ptosis

Nicholas Silvestri (Buffalo) Neurology® 2021;96:918. doi:10.1212/WNL.0000000000011901

Giannoccaro et al.1 reported their findings on the sensitivity and specificity of the ice pack test and single-fiber electromyography (SF-EMG) in the diagnosis of ocular myasthenia gravis. It is interesting—and perhaps humbling—that such a straightforward test as cooling muscle with ice has very similar diagnostic accuracy compared with a technically complex and sophisticated test as SF-EMG in patients presenting with eyelid ptosis. These findings are largely in line with previous research evaluating the positive and negative predictive values of the ice pack test in ocular myasthenia gravis.2-5

One concern is the discordance in the results of the 2 tests in 10% of the study participants, although this occurred more often in cases of mild and isolated ptosis. Another limitation is that the authors did not assess the utility of the ice pack test in patients with isolated diplopia, a fairly common clinical manifestation in those ultimately diagnosed with ocular myasthenia.

Overall, the results of this study are encouraging given the simplicity of the ice pack test and a relatively limited availability of SF-EMG, and they highlight the continued importance of the physical examination in neurologic diagnosis in an age of extensive and often “shotgun” approach to diagnostic testing.

1. Giannoccaro MP, Paolucci M, Zenesini C, et al. Comparison of ice pack test and single-ﬁber EMG diagnostic accuracy in patients referred for myasthenic ptosis. Neurology 2020;95:e1800–e1806. 2. Ellis FD, Hoyt CS, Ellis FJ, Jeﬀery AR, Sondhi N. Extraocular muscle responses to orbital cooling (ice test) for ocular myasthenia gravis diagnosis. J AAPOS 2000;4:271–281. 3. Ertas M, Arac N, Kumral K, Tuncbay T. Ice test as a simple diagnostic aid for myasthenia gravis. Acta Neurol Scand 1994;89:227–229. 4. Lertchavanakul A, Gamnerdsiri P, Hirunwiwatkul P. Ice test for ocular myasthenia gravis. J Med Assoc Thai 2001;84(suppl 1): S131–S136. 5. Fakiri MO, Tavy DLJ, Hama-Amin AD, Wirtz PW. Accuracy of the ice test in the diagnosis of myasthenia gravis in patients with ptosis. Muscle Nerve 2013;48:902–904.

Author Response: Comparison of Ice Pack Test and Single-Fiber EMG Diagnostic Accuracy in Patients Referred for Myasthenic Ptosis

Maria Pia Giannoccaro (Bologna, Italy) and Rocco Liguori (Bologna, Italy) Neurology® 2021;96:918–919. doi:10.1212/WNL.0000000000011902

We thank Dr. Silvestri for the comment on our article1 and agree that a relatively high proportion of patients showed discordant results between the ice pack test and single-ﬁber electromyography (SF-EMG) because they were observed in 15% of patients with a ﬁnal diagnosis of ocular myasthenia (OM). Most of discordant cases were related to the negativity of the ice pack test in the presence of an altered SF-EMG result. One explanation could be the lack of repetition of the ice pack test, as we outlined in the discussion. Indeed, a previous study showed that repeated ice pack tests improved sensitivity by 34.6% compared with a single test.2 We also found that the repetition of the ice pack test in some patients increased its sensitivity and, therefore, the number of patients showing concordant results (unpublished data).

Author disclosures are available upon request ([email protected]).

918 Neurology | Volume 96, Number 19 | May 11, 2021 Neurology.org/N Copyright © 2021 American Academy of Neurology. Unauthorized reproduction of this article is prohibited. We also acknowledged that we did not investigate the usefulness of the ice pack test in patients with isolated diplopia. Nevertheless, Chatzistefanou et al.3 reported a sensitivity of 76.9% and a speciﬁcity of 98.3%, suggesting that the ice pack test is similarly useful in the assessment of myasthenic diplopia. This could be particularly relevant because we showed that diagnostic tests for OM—including SF-EMG—are frequently negative in patients with this clinical presentation.4

1. Giannoccaro MP, Paolucci M, Zenesini C, et al. Comparison of ice pack test and single-fiber EMG diagnostic accuracy in patients referred for myasthenic ptosis. Neurology 2020;95:e1800–e1806. 2. Park JY, Yang HK, Hwang JM. Diagnostic value of repeated ice tests in the evaluation of ptosis in myasthenia gravis. PLoS One 2017;12: e0177078. 3. Chatzistefanou KI, Kouris T, Iliakis E, et al. The ice pack test in the differential diagnosis of myasthenic diplopia. Ophthalmology 2009;116:2236–2243. 4. Giannoccaro MP, Di Stasi V, Zanesini C, et al. Sensitivity and specificity of single-fibre EMG in the diagnosis of ocular myasthenia varies accordingly to clinical presentation. J Neurol 2020;267:739–745.

CORRECTIONS Lesion Evolution and Neurodegeneration in RVCL-S A Monogenic Microvasculopathy Neurology® 2021;96:919. doi:10.1212/WNL.0000000000011323

In the article “Lesion Evolution and Neurodegeneration in RVCL-S: A Monogenic Micro- vasculopathy” by Ford et al.,1 the following sentence was omitted from the Study Funding statement: “This work was supported by Clayco Foundation, Energy 4A Cure Foundation, Cure CRV Research and The Foundation for Barnes Jewish Hospital (MB, MKL, DH, JJM, JPA).” The authors regret the omission.

Reference 1. Ford AL, Chin VW, Fellah S, et al. Lesion evolution and neurodegeneration in RVCL-S: a monogenic microvasculopathy. Neurology 2020;95:e1918–e1931.

Apathy and Risk of Probable Incident Dementia Among Community-Dwelling Older Adults Neurology® 2021;96:919. doi:10.1212/WNL.0000000000011756

In the article “Apathy and Risk of Probable Incident Dementia Among Community-Dwelling Older Adults” by Bock et al.,1 the Study Funding should read: “This research was supported by National Institute on Aging (NIA) Contracts N01-AG-6-2101; N01-AG-6-2103; N01-AG-6- 2106; NIA grant R01-AG028050, and NINR grant R01-NR012459. This research was funded in part by the Intramural Research Program of the NIH, National Institute on Aging. It is also supported through NIA K24 AG-31155.” The authors regret the omission.

Reference 1. Bock MA, Bahorik A, Brenowitz WD, Yaﬀe K. Apathy and risk of probable incident dementia among community-dwelling older adults. Neurology 2020;95:e3280–e3287.

Author disclosures are available upon request ([email protected]).