Vitamin D to Improve Outcomes by Leveraging Early Treatment: Long-term Brain Outcomes in Vitamin D Deficient Patients (VIOLET-BUD)
Conditions: Cognitive Decline
Sponsor: Vanderbilt University Medical Center

Jin Han, MD, MSc 
 
 
VIOLET-BUD  
 
 
1 
Protocol Date:  04/12/2019 
 
 
 
 
 
 
 
 
 
 
Vitamin D to Improve Outcomes by Leveraging Early Treatment: Long-term Brain 
Outcomes in Vitamin D Deficient Patients (VIOLET-BUD) 
 
 
 
 
 
 
Jin H. Han, MD, MSc 
 
Associate Professor 
Vanderbilt University Medical Center  
Department of Emergency Medicine  
 
1313 21st Avenue South, 312 Oxford House  
Nashville, TN 37232-4700 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

Jin Han, MD, MSc 
 
 
VIOLET-BUD  
 
 
2 
Protocol Date:  04/12/2019 
 
Table of Contents: 
 
Study Schema 
 
1.0 
Background 
 
2.0 
Rationale and Specific Aims 
 
3.0 
Inclusion/Exclusion Criteria 
 
4.0 
Enrollment/Randomization 
 
5.0 
Study Procedures 
 
6.0 
Risks  
 
7.0 
Reporting of Adverse Events or Unanticipated Problems involving Risk to 
Participants or Others 
 
8.0 
Study Withdrawal/Discontinuation 
 
9.0 
Statistical Considerations 
 
10.0 
Privacy/Confidentiality Issues 
 
11.0 
Follow-up and Record Retention 
 
12.0 
References 
 
 

Jin Han, MD, MSc 
 
 
VIOLET-BUD  
 
 
3 
Protocol Date:  04/12/2019 
 
1.0 
Background 
Forty percent of patients who develop acute respiratory distress syndrome (ARDS) will 
die during hospitalization.1 Among those who survive, 75% will develop long-term 
cognitive impairment (LTCI).2 LTCI is a growing public health problem and is defined as 
a new deficit or a worsening pre-existing deficit in cognition that remains persistent after 
an acute illness. Its reach extends beyond ARDS. Our research group observed that 
34% of intensive care unit (ICU) survivors, regardless of ARDS status, develop cognitive 
impairment similar in severity to moderate traumatic brain injury and 24% develop even 
more severe impairments similar to mild Alzheimer’s disease.3 A substantial proportion 
of ICU survivors will also develop impairments in executive function,3 which leads to 
increased disability, worsening quality of life, and reduced employment.4,5 Interventions 
that preserve long-term cognition and executive function after critical illness are lacking.  
 
Early, high-dose oral Vitamin D repletion could potentially preserve long-term cognition 
in critically ill patients. Vitamin D is a pleiotropic secosteroid hormone that modulates 
systemic and central nervous system (CNS) inflammatory responses.6-9 Inflammation in 
response to an acute illness plays a prominent role in dementia pathogenesis.10-15  In 
addition to down-regulating systemic inflammation,7-9 Vitamin D also exerts its anti-
inflammatory effects directly in the brain. Vitamin D receptors are located on microglial 
cells (CNS macrophages) which play a key role in neuroinflammation.15-19 
 
An abundance of recent observational data suggest Vitamin D deficiency is associated 
with poorer long-term cognition20,21 and increased risk of Alzheimer’s dementia in 
community-dwelling adults.21 Based upon our preliminary studies, Vitamin D deficiency 
is also associated with accelerated cognitive decline in hospitalized, acutely ill patients. 
To our knowledge, no studies have evaluated if Vitamin D repletion improves these 
cognitive outcomes in the setting of an acute or critical illness.  
 
2.0 
Rationale and Specific Aims 
A significant proportion of ICU survivors will acquire LTCI and executive dysfunction, 
especially in those who develop ARDS during hospitalization. These patients suffer from 
significant disability, a reduction in employment, and poorer quality of life.4,5,22-29 Because 
inflammation is likely to be an underpinning of LTCI pathophysiology, Vitamin D 
treatment can potentially improve long-term cognition by attenuating systemic and CNS 
inflammatory responses. Numerous observational cohort studies and our preliminary 
data have reported that Vitamin D deficiency is associated with LTCI and executive 
dysfunction. Randomized control trials designed to determine if Vitamin D repletion 
improves long-term cognitive outcomes are needed. 
 
We propose this ancillary study to a parent double-blinded, placebo-controlled 
randomized control trial (RCT) evaluating how a single, high-dose (540,000 IU) oral 
Vitamin D3 treatment affects 90-day mortality in patients who are at high risk for ARDS 
and have Vitamin D deficiency (plasma 25-hydroxyvitamin D < 20 ng/ml). The parent 
RCT (Vitamin D to Improve Outcomes by Leveraging Early Treatment [VIOLET], 
NCT03096314) is part of the Clinical Trials Network for the Prevention and Early 
Treatment of Acute Lung Injury (PETAL) sponsored by the NHLBI. The VIOLET trial 
completed enrollment in July 2018 with 1,360 randomized patients. Our proposed 
ancillary study will provide additional funding to perform comprehensive 
neuropsychological (cognitive) evaluations, which were not part of the parent trial.  
These neuropsychological evaluations will be conducted 8 to 26 months after 

Jin Han, MD, MSc 
 
 
VIOLET-BUD  
 
 
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Protocol Date:  04/12/2019 
 
randomization among a subset of 140 survivors enrolled in VIOLET. This ancillary study 
will conducted in 7 (out of 42) sites. Our study, entitled VIOLET: Long-term Brain 
Outcomes in Vitamin D deficient patients (VIOLET-BUD), has the following hypothesis 
and specific aim: 
 
Aim: Determine if early administration of a single high-dose (540,000 IU) oral 
vitamin D3 (cholecalciferol) treatment improves long-term global cognition and 
executive function as determined by comprehensive neuropsychological testing 
in 140 critically ill patients with Vitamin D deficiency at enrollment.  
Hypothesis: Critically ill patients with Vitamin D deficiency who are treated with a 
single high-dose of Vitamin D3 will have significantly better long-term global 
cognition and executive function than those treated with placebo.  
 
If Vitamin D is found to improve long-term (8 to 26 months after randomization) global 
cognition and executive function, then we will have discovered a novel therapy that 
would allow ICU survivors to keep their cognitive abilities and help them resume their 
lives as they did prior to the critical illness. Because oral Vitamin D is simple, 
inexpensive, and safe, this intervention could be feasibly implemented in ICUs 
worldwide.  
 
3.0 
Inclusion/Exclusion Criteria 
All patients enrolled in the VIOLET parent study will be considered for enrollment in 
VIOLET-BUD. VIOLET-BUD has the following exclusions: 
1) Deaf or blind 
2) Non-English speaking 
Patients who are deaf or blind will be excluded, because the neuropsychological testing 
have auditory and visual components. Non-English speaking patients will be excluded 
because the neuropsychological raters can only perform the assessments in English 
only. 
 
4.0 
Enrollment/Randomization 
Because VIOLET-BUD is an ancillary study to the VIOLET parent study, randomization 
to the Vitamin D3 versus placebo group has been completed. The PETAL Clinical 
Coordinating Center performed computerized randomization using 1:1 allocation 
stratified by enrolling hospital. For VIOLET-BUD, the site research and clinical staff, 
patient and authorized surrogate will be blinded to treatment assignment.  
 
VIOLET-BUD will enroll patients from 7 (out of 42) PETAL sites that participated in 
VIOLET. The local site will contact all patients who were enrolled in VIOLET to recruit 
patients for VIOLET-BUD. As part of the VIOLET parent study’s informed consent 
document, participants agreed to be contacted for future studies. Although the patient 
contact will typically occur by phone, the initial method of contact may occur by e-mail, 
letter, or text depending on patient preferences or availability. To maximize the feasibility 
of the study, the long-term follow-up assessments conducted between 8 and 26 months) 
after randomization will be allowable. Patients enrolled in VIOLET will be contacted 
consecutively. 
 
Once a VIOLET patient has been contacted, the local site study coordinator will use a 
phone or e-mail script (Appendix A). He/she will describe VIOLET-BUD’s study protocol 
in lay terminology over the phone. It will be emphasized that the data collected will be for 
research purposes. The research staff will inform the patient that there is no obligation to 

Jin Han, MD, MSc 
 
 
VIOLET-BUD  
 
 
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Protocol Date:  04/12/2019 
 
participate in the study. If the patient refuses to participate in VIOLET-BUD or died prior 
to the long-term follow-up, then this will be recorded. If the patient cannot follow-up 
because they reside in a skilled nursing facility or has significant mobility issues, lives 
too far from the local site, cannot obtain transportation to the local site, is incarcerated, 
or is too cognitive impaired to participate, then this will be recorded. 
 
 
If the patient agrees to participate, then the local staff will review the informed consent 
document with the patient. Because patients may have LTCI significant enough to affect 
capacity, there is a possibility that the patient may not be capable of provide informed 
consent. In these cases, consent will be obtained from an authorized surrogate. The 
determination of whether or not a patient can provide informed consent will be 
determined by the study coordinator or investigators. They will be trained to determine 
who is not consentable through clinical judgment and direct patient interaction 
(Appendix A).  Patients will be deemed capable of consenting if: 1) they are able to 
carry a normal adult conversation and 2) they are able to recall aspects of the consent 
(e.g. Can you tell me what the purpose and risks of the study are?”).  The patient will be 
asked questions about the study to ensure they comprehend study procedures.  If they 
unable to adequately answer these questions, then consent will be obtained from an 
authorized surrogate to also be contacted by phone. 
 
If the patient or their authorized surrogate provides consent to participate in VIOLET-
BUD, then the local site coordinator will setup an appointment for the patient to receive 
their outcome assessment at the local site. Patients will be contacted from 8 to 26 
months after randomization in the parent study. Because the VIOLET parent study has 
completed enrollment, the wide enrollment window will facilitate recruitment and help 
achieve our sample size. After the appointment is setup, the local site study coordinator 
will send the patient or their authorized surrogate a copy of the informed consent 
document via their preferred method of contact. A letter (Appendix B) with the local site 
study coordinator’s name and contact information, the appointment date, time, location, 
and a map with directions to the videophone neuropsychological testing area will also be 
enclosed or attached. On the day of the appointment, the patient or authorized surrogate 
will be provided an opportunity to ask additional questions. If they agree to participate in 
VIOLET-BUD, then they will sign the informed consent document. A copy of the 
informed consent document will be provided to the patient or their authorized surrogate. 
Afterwards, VIOLET-BUD’s data collection, including the long-term outcomes, will be 
performed. Patients will be provided financial compensation once the long term follow-up 
visit has been completed. 
 
5.0 
Study Procedures 
5.1 Data to be obtained from the VIOLET parent study 
The VIOLET parent study has collected patient demographics, home medications, 
including vitamin D and calcium supplementations, Sequential Organ Failure 
Assessment (SOFA) Score to day 7, Lung Injury Prediction Score (LIPS), calcium levels, 
Charlson Comorbidity Index, duration of mechanical ventilation, and ARDS status. 
These data were obtained through patient/caregiver interview and medical record 
review. The SOFA score is a marker of illness severity and estimates risk of mortality.30 
It quantifies CNS, respiratory, cardiovascular, hepatic, renal, and coagulation 
dysfunction/ failure and ranges from 0 to 24 (>90% risk of ICU mortality).31 ARDS was 
determined by local review of daily ventilator status/settings, PaO2 and SpO2 values, 

Jin Han, MD, MSc 
 
 
VIOLET-BUD  
 
 
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Protocol Date:  04/12/2019 
 
and chest x-ray. These data will be obtained from the PETAL Clinical Coordinating 
Center at Massachusetts General Hospital.  
 
 
5.2. Data to be collected specifically for VIOLET-BUD (Table 1) 
The primary outcomes for VIOLET-BUD are long-term global cognition and executive 
function. Only patients who have LC-MS confirmed Vitamin D deficiency (plasma 25-
OHD < 20 mg/dL) at enrollment will have their outcomes determined. All outcome 
assessments will be blinded to treatment group. We will use comprehensive 
neuropsychological assessment which allows for detailed evaluation of specific cognitive 
domains. Prior work suggests Vitamin D deficiency predominantly affects short-term and 
working memory,44,45 attention,45,46 and executive function.47  
 
Long-term global cognition will be assessed for using the Repeatable Battery for the 
Assessment of Neuropsychological Status (RBANS, 30 min, Appendix C),  
which we have extensively used in our prior work.3 The RBANS is a comprehensive 
neuropsychological battery for the evaluation of global cognition and has been validated 
in subjects with mild cognitive impairment, moderate to severe traumatic brain injuries, 
vascular dementias, and Alzheimer’s disease.34-38 In addition to providing a score for 
global cognition, it also provides individual scores for immediate and delayed memory, 
attention, visuospatial construction, and language.  
 
To quantify executive function, we will use tests from the Delis–Kaplan Executive 
Function System (D-KEFS) which is the most definitive executive functioning battery in 
existence.39  Executive functioning refers to a wide array of “higher order” abilities that 
exist across a spectrum of cognitive sub-domains. Recent factor analyses have 
suggested that the majority of executive sub-domains fall broadly under conceptual 
flexibility, inhibition, and monitoring.39  As such, we have selected tests of executive 
functioning from the D-KEFS to assess these three broad abilities – the DKEFS 
Proverbs subscale (conceptual flexibility, 5 min, Appendix D), the DKEFS Trail 
Making-Number/Letter Switching subscale (inhibition, 2 min, Appendix E), and 
DKEFS Verbal Fluency Category Switching subscale (monitoring, 3 min, Appendix 
F).  An executive function composite score will be calculated from these three DKEFS 
subscales.  
 
The RBANS and executive function tests will be conducted by experienced 
neuropsychologists from the Vanderbilt ICU Delirium and Cognitive Impairment Study 
Group via videophone-assisted neuropsychological testing. Eight to 26 months post-
randomization, study participants will return to their local study site where they were 
originally enrolled. At the local study site, there will be a trained proctor (research nurse 
or research assistant) with the patient to provide on-site support and assist the 
neuropsychologist rater (e.g., hand the appropriate instrument to the patient). To 
minimize the risk of poor connectivity, computers will connect to the internet using a local 
area network (LAN) connection rather than a wireless device.  In the rare event of a poor 
internet connection, we have developed specific contingency plans and protocols (e.g., 
training the proctors to administer portions of the neuropsychological tests) to overcome 
any connectivity issues.  
 
 

Jin Han, MD, MSc 
 
 
VIOLET-BUD  
 
 
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Protocol Date:  04/12/2019 
 
Study Assessment 
Description 
Source 
Length  
Past history of dementia or is on 
an acetylcholinesterase inhibitor 
medication (galantamine 
[Reminyl], Donepezil [Aricept] 
and rivastigmine [Exelon]) prior 
to randomization 
This will be used to characterize 
the presence of pre-existing 
cognitive impairment. 
Medical 
record 
5 mins 
Age, race, ethnicity sex, level of 
education, region of residence, 
and highest occupation32,33 
These data will be used to estimate 
premorbid intelligence using an 
actuarial formula. 
Informant or 
patient 
5 mins 
Repeatable Battery for the 
Assessment of 
Neuropsychological Status 
(RBANS, Appendix C).34-38 
This neuropsychological battery will 
quantify long-term global cognition 
which is our primary outcome. It 
assesses immediate and delayed 
memory, attention, visuospatial 
construction, and language. 
Patient 
30 mins 
Delis–Kaplan Executive 
Function System (D-KEFS) 
Proverbs subscale (conceptual 
flexibility, Appendix D), the 
DKEFS Trail Making-
Number/Letter Switching 
subscale (inhibition, Appendix 
E), and DKEFS Verbal Fluency 
Category Switching subscale 
(monitoring, Appendix F).39     
This neuropsychological battery will 
be used to quantify long-term 
executive function which is our 
primary outcome. It assesses  
conceptual flexibility, inhibition, and 
monitoring which are the 
cornerstones of executive 
function.39  An executive function 
composite score will be calculated 
from these three DKEFS 
subscales. 
Patient 
10 mins 
Katz Activities of Daily Living 
(ADL, Appendix G)40 
Quantifies basic ADLs -- bathing, 
dressing, toileting, transferring, 
continence, and feeding. 
Informant or 
patient 
5 mins 
Lawton Instrumental  Activities 
of Daily Living Scale (IADL, 
Appendix H)41 
Quantifies the patient’s IADLs such 
as his/her ability to use the 
telephone, shopping, food 
preparation, housekeeping, 
laundry, transportation, medication 
management, and finances. 
Informant or 
patient 
10 mins 
Outcomes After Critical Illness 
and Surgery (OACIS) 
Employment Status 
Questionnaire (Appendix I) 42,43 
9-item survey that characterizes 
the patient’s baseline (prior to the 
critical illness) and current level of 
employment (full, partial, or not 
employed).  
Informant or 
patient 
10 mins 
Death 
While attempting patient contact, 
we will determine death status. If 
the patient died before the long-
term follow-up, we will record date 
of death. 
 
Medical 
record, 
informant, 
obituary web 
searches, or 
National 
Death Index  
5 mins 
Nursing Home Placement 
During patient contact, we will 
determine if the patient placed in n 
nursing home. The date of 
placement will be recorded. 
Medical 
record, 
patient, 
informant 
5 mins 
Table 1. VIOLET-BUD data to be collected by the local study team. The gray cells indicate data 
collected during the long term (8 to 26 month) follow-up visit and should take approximately 70 
minutes. 

Jin Han, MD, MSc 
 
 
VIOLET-BUD  
 
 
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Protocol Date:  04/12/2019 
 
 
Several studies have found results for neurocognitive testing to be very similar when 
assessments are completed by videoconferencing and by face-to-face evaluations.48-50 
Galusha-Glasscock et al. observed that the videoconference-assisted RBANS, our 
primary cognitive measure, was highly correlated with the in-person RBANS with a 
correlation coefficient of 0.88 (p-value < 0.0001) in patients with and without dementia.48   
 
The local study team will also collect other long-term secondary outcome measures: 
functional status as measured by the Katz activities of daily living (ADL) scale and 
Lawton instrumental activities of daily living (IADL) scale, employment as measured by 
the Outcomes After Critical Illness and Surgery (OACIS) Employment Status 
Questionnaire, nursing home placement, and death. 
 
For subjects who are unable to come to the local site for the long-term follow-up visit 
(e.g., due to mobility issues), the local site local study team will have the option to 
conduct the visit at their place of residence. To perform the videophone-assisted 
neuropsychological testing at the subject’s resident, the study team will connect their 
laptop to the subject’s local area network via wireless or ethernet connection.  The local 
site study team may also use a mobile hotspot with a 4G LTE signal to perform the 
videophone-assisted neuropsychological testing. 
 
VIOLET-BUD will also collect whether or not a patient has a past history of dementia or 
is on an acetylcholinesterase inhibitor medication (galantamine [Reminyl], Donepezil 
[Aricept] and rivastigmine [Exelon]) from the medical record.  This will help determine if 
the patient has a history of pre-existing cognitive impairment prior to randomization. The 
determination of pre-existing cognitive impairment via medical record review will also be 
conducted in VIOLET subjects who were enrolled at the local site but died prior to the 
long-term outcome assessment or refused follow-up visit. These data will allow us to 
conduct survivor average causal effect (SACE), which will help determine the extent in 
which the competing of risk of death biased our findings 
 
To facilitate merging of data sets, VIOLET-BUD participants will be assigned the same 
unique identification number as assigned in VIOLET.  All VIOLET-BUD data will be 
entered into a secure REDCap database.  Because the written and drawing portions of 
RBANS and executive function assessments need to be scored by the Vanderbilt ICU 
Delirium and Cognitive Impairment neuropsychologists, the local study team will upload 
these forms to the REDCap database. 
 
6.0 
Risks 
This is a minimum risk study since there is no intervention. The Vitamin D intervention 
and safety monitoring has already been conducted as part of the VIOLET parent study.  
The VIOLET-BUD ancillary study will obtain long-term data using neuropsychological 
testing and survey instruments. For VIOLET-BUD specifically, subjects will undergo a 
neuropsychological examination at 8 to 26-months that can take approximately 40 
minutes to perform. There is a small risk that a patient enrolled in VIOLET-BUD may 
become fatigued or distressed during the study neuropsychological assessments. In 
these cases, we will immediately stop the assessment and give the patient an 
opportunity to rest. Afterwards, we will ask the patient if we can continue with the 
neurocognitive assessments or if we should reschedule the long-term (8 to 26) follow-up 
appointment.  Occasionally, patients and their families may get visibly frustrated and 

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VIOLET-BUD  
 
 
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Protocol Date:  04/12/2019 
 
distressed if the patient is unable to perform the cognitive tasks. We will reassure them 
that these tasks can be difficult to perform, and that such difficulties are common. If the 
patient or his or her family continue to be distressed after this reassurance, we will skip 
that particular task or stop the assessment altogether.  
 
After the neuropsychological assessments, we administer surveys to the patient or 
authorized surrogate to collect additional data about the patient’s functional status, 
baseline and current employment, education, region of residence, and highest 
occupation. These surveys will take an approximately 30 minutes to collect. If the patient 
is completing the surveys and becomes fatigued, then we will give the patient an 
opportunity to rest. If the patient can’t continue with the survey, then we will attempt to 
get the survey data by phone at a later date. 
 
Because patient identifiers are accessed throughout all phases of the study, there is a 
small risk of loss of patient confidentiality. Federal regulations at 45 CFR 46 111 (a) (7) 
requires that when appropriate, there are adequate provisions to protect the privacy of 
subjects and to maintain the confidentiality of data. To maintain confidentiality, all 
evaluation forms and reports will be identified only by a coded number. The same coded 
number will be used for subjects enrolled in both VIOLET and VIOLET-BUD. All related 
participant study records will be kept in a locked, password protected computer. The 
coded number will be generated by a computer at the PETAL CCC, and only the study 
team will have access to the codes. All records will be kept in a locked, password 
protected computer. All computer entry and networking programs will be done with 
coded numbers only. All paper case report forms will be maintained inside a locked 
office. Study information will not be released without the written permission of the 
patient, except as necessary for monitoring by the Vanderbilt Clinical Coordinating 
Center. 
 
7.0 
Reporting of Adverse Events or Unanticipated Problems involving Risk to 
Participants or Others 
As this is a non-intervention study, we do not anticipate having any adverse events.  
However, should any occur, they will be reported to the IRB per institution policies and 
procedures.  
 
8.0 
Study Withdrawal/Discontinuation 
If patient or caregiver declines participation at the outset or does not meet inclusion 
criteria, they will not be included in the study. If patients wish to discontinue taking part in 
the study, they will be instructed to contact local site principal investigator or the 
research staff and let them know that they wish to withdraw.  At that time, no further data 
will be collected on the patient. All health data previously collected before they withdraw 
their consent will still be used for reporting and research quality.  
 
9.0 
Statistical Considerations 
We will determine if single, high-dose enteral Vitamin D treatment improves long-term 
global cognition and executive function compared with placebo using multiple linear 
regression.  One model will be constructed for long-term RBANS and another for long-
term executive function. We will use multivariable linear regression over unadjusted 
analyses because it is hypothesized that there will be a 5% absolute risk reduction in 
death for those who receive enteral Vitamin D. This may imbalance potential 
confounders between the treatment and placebo groups. Additional imbalances may 

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VIOLET-BUD  
 
 
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Protocol Date:  04/12/2019 
 
occur because of random chance. Multivariable regression will minimize this bias and 
lead to moderate gains in power.51  
 
The models will be adjusted for the covariates 
listed in Table 2. Nonlinear predictor effects for 
continuous variables will be incorporated into 
the models. If necessary, non-normally 
distributed continuous variables will be 
transformed. To account for correlation within a 
center, we will use robust standard errors.52   
 
In the event of significant missing covariate 
data, multiple imputation will be considered.53-55 
Multiple imputation will also be performed if portions of the cognitive assessments are 
missing;3,56,57  patients with completely missing cognitive data will be excluded from the 
study. Because several of our covariates (e.g., premorbid cognition and intelligence) 
may be highly correlated, we will assess for collinearity using variance inflation factors 
and use only one of those variables if a high degree of collinearity is observed. The final 
models will be validated using bootstrap internal validation and cross validation 
approaches.58 After the initial analysis has been completed, we will also evaluate if there 
is any effect modification of age, dementia status (medical record), baseline severity of 
illness (SOFA), mechanical ventilation status, and ARDS status. We will also perform a 
sensitivity analysis where death within the follow-up window will be considered 
equivalent to the worst cognitive outcome. This will be performed to further determine 
the effect of death on our findings. 
 
Secondary analyses will be performed where we will determine how Vitamin D treatment 
affects the individual RBANS cognitive domains (e.g., immediate and delayed memory, 
attention, visuospatial construction, and language). We will also determine if treatment 
significantly improves long term functional status, employment as measured by the ICAP 
survey, nursing home placement, and death. 
 
To estimate our sample size requirements, we used half a standard deviation (SD) of 
that particular test to define the minimum clinically important difference (MCID) 
threshold.59-62 Based upon the preliminary studies, the SD for the RBANS was 12.3 As a 
result, we considered a 6-point difference to be the MCID threshold for the RBANS. 
Based upon a two-sided alpha of 0.05 and 81% power, we will need outcome data in a 
total of 140 patients (~70 patients in each group) to detect a 6-point difference in RBANS 
between treatment groups. This sample size is feasible to obtain over 10 months. From 
September 2017 to July 2018, the six VIOLET-BUD sites enrolled 339 patients in 
VIOLET.  Based upon the BRAIN-ICU study,3  we anticipate that 99% will not be deaf or 
blind, 62% will survive and 75% will follow-up at 8 to 26 months.  We have the potential 
to enroll 155 patients making our required sample size of 140 patients very achievable.  
For our executive function composite score, the SD was 2.34 (Section C6), so the MCID 
threshold would be a 1.17 point difference. If we complete long-term executive function 
assessments on 140 patients as planned, we will have 81% power to detect this 1.17-
point difference in the executive function composite score assuming a two-sided alpha of 
0.05.  
 
10.0 
Privacy/Confidentiality Issues 
Covariates 
df 
Vitamin D vs placebo 
Age 
1 
2 
Severity of illness (SOFA) 
2 
Comorbidity (Charlson) 
2 
Pre-existing cognitive impairment 
1 
Time to long-term cognitive 
assessment 
2 
Premorbid intelligence 
ARDS development 
1 
1 
Table 2. Covariates for the 
multivariable linear regression models. 

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VIOLET-BUD  
 
 
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Protocol Date:  04/12/2019 
 
The patient’s information, without identifiers, may be shared with other institutions or 
universities. Dr. Han, his co-investigators, and their staff will comply with any and all 
laws regarding the privacy of such information.  There are no plans to pay the patient for 
the use or transfer of their de-identified information or specimens.   
 
 

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Protocol Date:  04/12/2019 
 
11.0 
Follow-up and Record Retention 
The duration of this study is approximately one year.  We will try to enroll 140 patients 
during this time.  Data collected will be retained indefinitely.    
 
12.0 
References 
1. 
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Mortality for Patients With Acute Respiratory Distress Syndrome in Intensive 
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Hopkins RO, Weaver LK, Pope D, Orme JF, Bigler ED, Larson LV. 
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care medicine. 1999;160(1):50-56. 
3. 
Pandharipande PP, Girard TD, Jackson JC, et al. Long-term cognitive impairment 
after critical illness. The New England journal of medicine. 2013;369(14):1306-
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4. 
Rothenhausler HB, Ehrentraut S, Stoll C, Schelling G, Kapfhammer HP. The 
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Norman BC, Jackson JC, Graves JA, et al. Employment Outcomes After Critical 
Illness: An Analysis of the Bringing to Light the Risk Factors and Incidence of 
Neuropsychological Dysfunction in ICU Survivors Cohort. Critical care 
medicine. 2016. 
6. 
McCann JC, Ames BN. Is there convincing biological or behavioral evidence 
linking vitamin D deficiency to brain dysfunction? FASEB J. 2008;22(4):982-
1001. 
7. 
D'Ambrosio D, Cippitelli M, Cocciolo MG, et al. Inhibition of IL-12 production 
by 1,25-dihydroxyvitamin D3. Involvement of NF-kappaB downregulation in 
transcriptional repression of the p40 gene. J Clin Invest. 1998;101(1):252-262. 
8. 
Muller K, Haahr PM, Diamant M, Rieneck K, Kharazmi A, Bendtzen K. 1,25-
Dihydroxyvitamin D3 inhibits cytokine production by human blood monocytes at 
the post-transcriptional level. Cytokine. 1992;4(6):506-512. 
9. 
Schleithoff SS, Zittermann A, Tenderich G, Berthold HK, Stehle P, Koerfer R. 
Vitamin D supplementation improves cytokine profiles in patients with 
congestive heart failure: a double-blind, randomized, placebo-controlled trial. Am 
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