Cognitive Battery

The cognitive state of participants was assessed by a comprehensive neuropsychological battery of the Enroll-HD consisting of the following tests: SDMT, SWRT, SCNT, SIT; CLF, LVF; TMT-A, TMT-B and MMSE. In the following, these neuropsychological tests will be explained in more detail.

Symbol Digit Modalities Test

The Symbol Digit Modalities Test (SDMT) (Smith, 1982) assessed the psychomotor speed and working memory. The first digits (1-9) assigned to symbols were presented to the participants during this assessment. In the second step, the participants were required to write below each presented symbol the correct corresponding digit. The number of correct matches completed in 90 seconds was recorded.

Stroop Word Reading Test

This assessment has very good properties for detecting a cognitive decline in the early stages of HD as it measures attention and psychomotor speed. This assessment is a subtest of the Stroop tests (Stroop, 1935, Golden, 1978). During this assessment, the participant was asked to read words indicating colours (red, blue, green) printed in black ink as quickly as possible; the time limit was 45 seconds. Total correct answers (correctly named items), total errors and total self-corrections performed in 45 seconds were recorded.

75

Stroop Color Naming Test

The Stroop Colour Naming Test (SCNT), as part of the Stroop tests, has very good properties for detecting a cognitive decline in the early stages of HD as it measures attention and psychomotor speed (Stroop, 1935, Golden, 1978). During this assessment, the participant was asked to read the colour of ink patches on the template (e.g., green, blue, red) as quickly as possible; the time limit was 45 seconds. Total correct answers (correctly named items), total errors and total self- corrections performed in 45 seconds were recorded.

Stroop Interference Test

The Stroop Interference Test (SIT), as the third subtest, is used to measure response inhibition (Golden, 1978). The participants see words indicating colours (red, blue, green), each written in red, blue or green ink, incongruent to the colour indicated by the letters (e.g., the word ‘red’ written in blue ink). Participants have to inhibit uttering the words read and instead have to name the colour of the ink in which the incongruent colour names are printed. The time limit is 45 seconds. Total correct answers (correctly named items), total errors and total self-corrections performed in 45 seconds were recorded.

Letter Fluency Test

The Letter Fluency Test (LFT) measures mental flexibility and semantic knowledge (Benton and Hamsher, 1976). In the LFT, participants were requested to utter as many words as possible, starting with a particular letter within one minute (response is speech motor output). LFT consists of three sub-tests, each using a different letter (i.g. S, M, T in German), tapping into the categories of high, medium and low frequency in the lexicon of the respective language (Senft, 2008). The total score as the sum of the correct, unique words of all sub-tests was recorded. The repetitive words count only once.

Categorial Fluency Test

In the Categorial Fluency Test (CFT), which is used to measure verbal learning and recall processes, participants are asked to name as many different words from the category "animals " as possible in one minute (Benton and Hamsher, 1976). The total of correctly named words (animals) was recorded.

76

Trail Making Test – Part A

The Trail Making Test (TMT) consists of A and B tasks, drawing straight lines connecting appropriate circles (Reitan, 1955). The TMT-A measures psychomotor speed. The rules for connecting circles labelled with numbers or letters differ between the two tasks: In the TMT-A, participants must connect given numbers in ascending order (i.e., 1-2-3-4), from 1 till 25. The time to complete the task is measured as a primary outcome, the number of failures or self-correction is also reported.

TMT-A is an ideal measure of psychomotor speed.

Trail Making Test – Part B

The Trail Making Test – Part B (TMT-B) is the second task on the TMT with drawing straight lines connecting appropriate circles labelled with numbers and letters (Reitan, 1955). In the TMT-B, participants must connect given numbers and letters in alternating order (i.e., 1-A-2-B-3-C;).

Thereby, the TMT-B taps executive and visual-perceptual functions, such as cognitive flexibility and selective attention abilities (Tombaugh, 2004). The time to complete the task is recorded as a primary outcome, the number of failures or self-correction is also reported.

Mini-Mental Status Examination

The Mini-Mental State Examination (MMSE) is a short screening questionnaire consisting of eleven tasks measuring five areas of cognitive function: orientation, registration, attention and calculation, recall and language. The MMSE is a 30-point test, quick in the administration (5 minutes). It is widely used in clinical and research settings to assess general cognitive functioning and screen for pronounced cognitive decline. Higher scores indicate better cognition. The MMSE is applied as per the standard operating procedure of the original publication (Folstein et al., 1975).

Statistical Analyses

All statistical tests were performed using “IBM SPSS Statistics Software” for Windows, Version 22.0.

The level of significance was set at α = 0.05 (two-tailed). Normality of data was assessed using visual inspection of Quantile-Quantile plots (Q-Q plots) and Shapiro-Wilk tests. The significance of these results was used to decide whether parametric or non-parametric tests were to be performed subsequently. Continuous variables are expressed as mean, SD and range, while categorical variables are expressed as percentages and ordinal variables as medians. Age and education in years were characterised by mean and SDs, and gender was characterized as a binomial or nominal variable in percentages in relation to the whole sample. The descriptive statistical analysis was

77 performed to evaluate the number of study participants in both cohorts, in the manifest patient group (HD) and healthy controls (NC) and to assess their demographic characteristics. For all proportions calculated, 95% confidence intervals were also reported.

Spearman correlation coefficients (r) were used to evaluate the strength and direction of the linear relationships between two or more ordinal variables. The strength and direction of correlations between two or more continuous variables were calculated using Pearson correlation coefficients.

In particular, Pearson’s correlation coefficients were used to describe the strength and direction of the linear relationships between the test measures in the battery and the demographic variables of age and education.

The Pearson chi-square (χ2) test was used for dichotomous variables. To evaluate differences among groups, independent-samples t-tests were applied. Effect sizes were reported in the form of Eta- Squared (η²) and Cohen’s d (as proposed by (Cohen, 1988). The Bonferroni correction for multiple comparisons was applied in between groups. Receiver Operating Characteristic (ROC) curves with area under the curve (AUC) and 95% Confidence Intervals (CIs) were calculated and used to compare the diagnostic accuracy of the neuropsychological tests included in the cognitive battery to assess cognitive impairment in HD. A one-way between-groups analysis of variance (ANOVA) was conducted to explore the impact of cognitive impairment in HD on the discriminative potential of each test included in the cognitive battery.