The Epidemiology of Silent and Overt Strokes in Adults With Sickle Cell Disease: a Prospective Cohort Study
Conditions: Anemia, Sickle Cell, Sickle Cell Disease, Stroke, Sickle Cell Thalassemia, Sickle Cell-Beta0-Thalassemia
Sponsor: Vanderbilt University Medical Center

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
1 
 
Table of Contents 
 
 
 
 
Page 
Number 
1.0 
Abstract 
2 
2.0 
Introduction 
3 
3.0 
Specific aims 
5 
4.0 
Rationale for surveillance MRI and MRA in individuals with pre-
existing silent cerebral infarcts or overt strokes  
5 
5.0 
Background and significance 
6 
6.0 
Methods 
9 
7.0 
Statistical analysis 
17 
8.0 
Study organization structure 
17 
9.0 
Stroke adjudications 
18 
10.0 
Definitions 
18 
11.0 
Study responsibilities 
20 
12.0 
Privacy and confidentiality issues 
21 
13.0 
Human subjects research 
21 
14.0 
Adverse Event Reporting 
24 
15.0 
Study withdrawal and discontinuation 
25 
16.0 
Follow-up and retention record 
25 
17.0 
Long term objectives 
25 
17.0 
References 
26 
 
 
 
 
 
 
 
 
 
 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
2 
 
 
The natural history of overt or silent strokes in individuals with sickle cell disease treated 
with blood transfusion or hydroxyurea therapy  
1.0 ABSTRACT 
Overt strokes and silent cerebral infarcts (SCI), also referred to as silent strokes, contribute 
significantly to morbidity in children and adults with sickle cell disease (SCD), resulting in 
functional impairment, challenges with school and job performance, and premature death. While 
five NIH-funded randomized controlled trials have identified therapies for primary and secondary 
overt stroke or SCI prevention in children (≤17 years old) with SCD, no prospective multi-center 
cohort study has systematically evaluated therapeutic options for secondary cerebral infarct 
prevention in adults. The optimal evidence based therapeutic option for primary and secondary 
stroke prevention in children with SCD is burdensome monthly blood transfusion therapy, 
eventually requiring iron chelation therapy. The less burdensome treatment is hydroxyurea 
therapy, though overall efficacy of this approach is still being evaluated, and some families 
choose observation alone. These competing choices between optimal blood transfusion 
therapy, hydroxyurea therapy or observation highlight the challenges of developing acceptable 
therapeutic strategies, other than blood transfusion, for preventing cerebral infarct recurrence in 
individuals with SCD. Preliminary data from our cohort suggest that among children with SCI, 
despite treatment with regular blood transfusions, the presence of magnetic resonance 
angiogram (MRA) defined cerebral vasculopathy at baseline is a significant risk factor for 
cerebral infarct recurrence (defined as overt stroke, new or enlarging SCI) compared to those 
without vasculopathy, 31% (5 of 16) versus 12% (11 of 89), respectively (p= 0.053). In seven 
sickle cell disease (SCD) centers, we propose a retrospective and prospective cohort study 
describing the natural history of overt strokes and SCI in individuals ≥ 1 year old with SCD 
receiving uninterrupted disease modifying therapy (regular blood transfusions, hydroxyurea or 
both) versus interrupted therapy or no treatment for secondary cerebral infarct prevention. 
Inclusion criteria will include: a history of prior cerebral infarct identified on entry magnetic 
resonance imaging (MRI)/MRA of the brain with a least a 1.5 tesla magnet strength. Exit 
MRI/MRA will be completed with a minimum and average of 3.5 and 4.5 years of follow-up 
between imaging studies, respectively. Specifically, based on evidence from the pediatric 
literature, we will test the hypotheses that, among individuals with SCD, those with baseline 
vasculopathy will have a higher incidence of stroke recurrence (overt or silent) compared with 
those without baseline vasculopathy, and those with progressive vasculopathy will have a 
higher incidence of overt stroke recurrence compared with those without progressive 
vasculopathy. The aims include: 1) compare the incidence of cerebral infarct recurrence (overt 
strokes, including intracranial hemorrhage, and SCI) in individuals (≥1 year of age) with SCD 
and evidence of prior overt stroke (n=44) or SCI (n=180) treated with disease-modifying therapy 
or interrupted or no therapy for prevention of overt stroke or SCI; 2) determine whether baseline 
MRA vasculopathy predicts cerebral infarct recurrence in individuals with prior overt stroke or 
SCI. Critical data obtained from this first multi-center natural history cohort study of individuals 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
3 
 
with SCD will facilitate the development of new therapies, other than arduous regular blood 
transfusions, for secondary prevention of overt stroke and SCI in SCD.  
 
2.0 INTRODUCTION 
Untreated silent cerebral infarcts are common and progressive in young adults with 
sickle cell disease 
The natural history of overt strokes (also referred to as strokes with associated neurological 
symptoms) or silent cerebral infarcts (SCI, also referred to as silent strokes without associated 
neurological symptoms), in adults (≥ 18 years of age) with sickle cell disease (SCD), receiving 
disease-modifying therapy for secondary stroke prevention (regular blood transfusions, 
hydroxyurea or both) is unknown. All five NIH-sponsored randomized controlled trials (RCT) for 
primary and secondary stroke prevention in SCD included only children ≤ 18 years of age(1–5). 
Untreated SCIs are progressive, with a prevalence of ~37% and ~50% in children and adults, 
respectively, Figure 1 (6–13). Hence, both children and adults with untreated SCI or strokes 
have a high risk of infarct recurrence, although only in children has a RCT been completed 
demonstrating that regular blood transfusion therapy for SCI is associated with a relative risk 
reduction of infarct recurrence of 58% when compared to observation alone (8), Figure 2. Some 
individuals elect to take the less burdensome, yet less effective hydroxyurea therapy (3), Figure 
3, instead of regular blood transfusion therapy, while others elect for no treatment. The absolute 
numbers of adults with cerebral infarcts will continue to increase because 98% of adolescents 
with SCD are surviving to adulthood(9). 
 
 
 
 
 
 
Figure 1. Prevalence of SCI in children and young adults with SCD. The figure displays the cumulative 
prevalence of SCI in children and young adults based on 5 cross-sectional studies and 1 longitudinal study. The 
cumulative prevalence of SCI suggests that the incidence of SCI does not plateau in young adulthood to ≥30 
years of age.  [Kassim et al. Blood. 2016 Apr 21;127(16):2038-40]. 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
4 
 
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Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
5 
 
the brain at study entry and exit. Participants will include those with uninterrupted disease-
modifying therapy (regular blood transfusion, hydroxyurea, or both) versus no therapy or 
interrupted therapy (defined as disease-modifying therapy which is not given continuously over 
the study period). Among the 420 potential participants that we have identified, we will enroll a 
total of 224 individuals in this study with overt strokes (n=44, including those with intracranial 
hemorrhage) or silent cerebral infarcts (n=180) who have already been receiving either 
uninterrupted disease-modifying therapy, interrupted therapy or no therapy for secondary stroke 
prevention prior to enrollment in the study. For each group (overt strokes and silent cerebral 
infarcts), we anticipate that approximately half of the patients will already be receiving 
uninterrupted disease-modifying therapy for secondary stroke prevention, and the other half will 
be receiving either no therapy or interrupted therapy prior to enrollment. The decision to receive 
or not to receive any disease modifying therapy for secondary stroke prevention will be left at 
the discretion of both the study participants and their treating hematologists. As this is a natural 
history study, there will be no randomization to any interventions, and all treatment received will 
be per standard care. We anticipate the following four groups of participants: 
a. At least 90 individuals with SCI treated with uninterrupted disease-modifying 
therapy  
b. At least 90 individuals with SCI receiving either no treatment or interrupted 
disease-modifying therapy 
c. At least 22 individuals with overt strokes treated with uninterrupted disease-
modifying therapy  
d. At least 22 individuals with strokes receiving either no treatment or interrupted 
disease-modifying therapy 
 
3.0 SPECIFIC AIMS 
Aim 1: Determine the incidence of infarct recurrence in individuals (≥ 1 year of age) with 
SCD and prior overt stroke or SCI treated with disease-modifying therapy. Entry criterion 
will include: a history of prior cerebral infarct and evidence on prior or entry MRI/MRA of the 
brain with a least a 1.5 tesla magnet strength. Exit MRI/MRA will be completed with a minimum 
and average of 3.5 and 4.5 years of follow-up between imaging studies, respectively, for 112 
individuals with either overt stroke or SCI receiving uninterrupted disease-modifying therapy 
(regular blood transfusions, hydroxyurea or both) and 112 individuals receiving either no 
treatment or interrupted disease-modifying therapy. Primary outcome measures are adjudicated 
stroke, SCI recurrence or transient ischemic attack (TIA). Imaging and clinical data collected as 
part of standard care will be used for secondary analysis. Combining retrospective and 
prospective cohorts will enable us to potentially recruit a higher proportion of subjects as it 
includes those already in treatment. We are able to include combined retrospective and 
prospective cohort data for the participants because all of the participating sites have already 
been practicing the standard care for secondary stroke prevention.  

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
6 
 
Aim 2: Determine whether baseline MRA-defined cerebral arterial vasculopathy 
(vasculopathy) predicts cerebral infarct recurrence in individuals with overt stroke or SCI 
(n=224). Vasculopathy as visualized on entry MRA will predict cerebral infarct recurrence (overt 
strokes, including intracranial hemorrhage, and SCI) and progression of vasculopathy between 
entry and follow up MRI/MRA of the brain.  
Critical data obtained from this first international, multi-center natural history cohort study of 
individuals with SCD will facilitate the development of new therapies, other than arduous blood 
transfusion therapy, for primary and secondary prevention of overt strokes and SCI in 
individuals with SCD. 
4.0 RATIONALE FOR SURVEILLANCE MRI AND MRA IN INDIVIDUALS WITH PRE-
EXISTING SILENT CEREBRAL INFARCTS OR OVERT STROKES 
In children and adults with pre-existing strokes or SCI, routine MRI/MRA of the brain are 
required to determine if new or progressive cerebral infarcts or cerebral vasculopathy have 
occurred. The presence of progressive or new central nervous system disease informs the 
patient, family and health care provider as to whether more intensive interventions should be 
considered to prevent further progression. More aggressive care could include: increasing the 
frequency of blood transfusion therapy, optimizing hydroxyurea therapy to maximum tolerated 
dose (MTD), adding hydroxyurea therapy to regular blood transfusion, indirect re-vascularization 
procedures or hematopoietic stem cell transplant (HSCT). 
 
5.0 BACKGROUND AND SIGNIFICANCE  
For Aim 1: 
SCI is defined and can only be detected with MRI of the brain. The Cooperative Study for 
Sickle Cell Disease (CSSCD) included an observational study of children with sickle cell disease 
who were identified as infants and followed closely over the subsequent decade. This study 
used surveillance MRI of the brain to detect abnormally increased T2-weighted signal intensity 
on multiple MRI views in children without corresponding focal neurological deficits as assessed 
by a hematologist. Subsequently our group, The SIT (Silent Cerebral Infarct Transfusion Multi-
Center Clinical Trial) team, expanded on the previously validated definition of SCI to include an 
MRI lesion measuring at least 3 mm in greatest linear dimension, and visible on at least two 
planes of T2-weighted images(10–12).  
There is a paucity of data regarding optimal strategies for secondary stroke prevention in 
individuals with sickle cell disease 
Central nervous system complications contribute significantly to morbidity and mortality in adults 
with sickle cell disease (SCD). The most complete definition of a silent or overt cerebral infarct 
includes neuroimaging and a thorough examination by a neurologist(4). Since the 1990’s, five 
NIH-funded randomized clinical trials (RCT) have been completed, providing evidence-based 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
7 
 
guidelines for the primary and secondary prevention of strokes in children with SCD living in 
high income countries(2–6). However, there are unfortunately no evidence-based guidelines for 
primary and secondary stroke prevention for adults with SCD. Multiple risk factors have been 
identified for the development of cerebral ischemic infarcts in SCD, including low cerebral 
oxygen content(7–9), cerebral vasculopathy(13), fever(14), presence of a prior cerebral 
infarct(8,15), and rapid increases in hemoglobin levels(16,17). Additionally, acute stroke in SCD 
has been associated with a number of comorbid conditions that are known risk factors for stroke 
within the general population, such as hypertension in children, and hypertension, diabetes 
mellitus, hyperlipidemia, atrial fibrillation and renal disease in adults. Most acute strokes (75%) 
and in-hospital deaths from stroke (91%) occur in adults(18). The rate of stroke in SCD has 
been shown to peak in older adults and found to be three-fold higher than rates previously 
reported in African-Americans of similar age range (35-64 years) without SCD(15). Silent and 
overt cerebral infarcts in patients with SCD are associated with increased risk of future infarcts 
and a drop of five IQ points on average(19). Hence, secondary prevention of strokes in this high 
risk population is essential for reduction of morbidity and mortality. Studies for the primary and 
secondary prevention of strokes in individuals with SCD should include investigation for 
traditional risk factors for strokes in adults and are urgently needed.  
 
Despite compliance with regular blood transfusion therapy, some patients continue to 
have progressive cerebral infarcts 
The progressive nature of strokes despite regular blood transfusion was also demonstrated in a 
retrospective cohort study of 137 children with SCD and strokes who were followed for a median 
of 10 years. Over the course of a decade, approximately 22% had a second stroke despite 
regular blood transfusion therapy, and among those with a second stroke, approximately 30% 
had a third stroke(20). In both the retrospective (10) and prospective cohorts (18), participants 
had strokes when their hemoglobin S levels were 1%, 9%, 22%, 25%, and 26%. The 
hemoglobin S levels at the time of stroke recurrence were all < 30%, the maximum target 
percent for hemoglobin S concentration. Based on the strong results from these studies and 
other large retrospective observational studies(8,20), regular blood transfusion therapy has 
limited effectiveness for secondary prevention of cerebral infarcts (overt strokes and SCI) over 
the long term and has to be continued indefinitely in order to reduce the lifetime risk of recurrent 
cerebral infarcts. 
Hydroxyurea, while less effective than regular blood transfusion therapy, is better than 
observation alone for secondary stroke prevention in individuals with SCD 
 
The SWiTCH trial was designed to test the hypothesis that hydroxyurea therapy with 
phlebotomy (alternative therapy) was non-inferior to regular blood transfusion therapy and 
chelation (standard therapy) for secondary stroke prevention(3). As part of the interim analysis, 
the trial was stopped prematurely based on futility assessment for decreasing iron stores in the 
alternative therapy group. However, the rates of strokes in the alternative and standard therapy 
group were 10.5% (7 of 67) and 0% (0 of 66) respectively, p < .05, Figure 3, suggesting that 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
8 
 
blood transfusion therapy is more effective for secondary stroke prevention than hydroxyurea. 
While the SWiTCH trial did not demonstrate any advantage of hydroxyurea compared to regular 
blood transfusion therapy, in a cohort of children with SCD, hydroxyurea decreased the rate of 
stroke recurrence compared with no therapy(21). Additionally, three observational studies found 
decreased stroke rates in children treated with hydroxyurea compared to observation alone(21–
23), and children receiving hydroxyurea were shown to have reduced transcranial Doppler 
(TCD) velocities(23,24), suggesting a benefit of hydroxyurea for secondary stroke prevention 
compared to observation alone.  A recent published pooled analyses from multiple studies of 
secondary stroke prevention in individuals with SCD showed a significantly higher rate of stroke 
recurrence in individuals receiving no therapy compared to those receiving either regular blood 
transfusions or hydroxyurea(25), Figure 4. 
  
 
Figure 4. Incidence rates of recurrent stroke in individuals with SCD receiving either hydroxyurea therapy, 
blood transfusion therapy, or no therapy for secondary stroke prevention in published studies since 1995. 
The expected incidence rates of stroke recurrence while on regular blood transfusion therapy, hydroxyurea therapy, 
or no therapy were found to be 1.9 (95% CI, 1.0-2.9), 3.8 (95% CI, 1.9-5.7), and 29.1 (95% CI, 19.2-38.9) events per 
100 patient years, respectively.  
 
Understanding the natural history of progressive cerebral infarcts in individuals with 
SCD may help identify higher-risk subgroups of patients who would benefit from novel, 
risk-based secondary stroke preventions strategies, such as hematopoietic stem cell 
transplant and experimental disease modifying therapies 
In summary, all large multi-center cohort studies describing the natural history of strokes have 
been conducted in children.  However, given the increased lifespan of adolescents with SCD, 
with an expected 98% surviving into adulthood(29), understanding the natural history of pre-
existing strokes or SCI, or even new strokes in adults has taken on an urgent importance. For 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
9 
 
adults with SCD with either pre-existing or new cerebral infarcts, there is a paucity of age 
appropriate data to guide management and to develop new therapeutic pathways. The evidence 
provided from this natural history study is required to optimally select current therapeutic options 
(regular blood transfusion, hydroxyurea or both) for secondary stroke prevention.  Furthermore, 
the natural history data are required to establish the relative merits of new treatment options 
such as re-vascularization procedures(30), HSCT(31), or even new targeted agents that may 
decrease future CNS events such as GBT440, a novel small molecular hemoglobin modifier 
which increases hemoglobin oxygen affinity and decreases hemolysis(32).   
 
For Aim 2: Baseline vasculopathy in individuals with SCI may predict future SCI.  
MRA-defined cerebral arterial vasculopathy (vasculopathy) in children with sickle cell disease is 
associated with subclinical stroke(33) and recurrent stroke(34). Within the pediatric literature, 
there is evidence to suggest that the presence of cerebral vessel abnormalities may predict the 
risk for future SCI. Specifically, children with SCD and SCI are more likely to have vasculopathy 
compared to those without SCI; 15.9% and 6.3%, respectively (P < 0.001)(35). In children with 
SCD and overt strokes, progressive vasculopathy is associated with infarct recurrence in 
100%(13).  Our lab (DeBaun et al.) has preliminary data from a longitudinal secondary analysis 
of  participants from the Silent Cerebral Infarct Multi-Center Clinical Trial (SIT) trial with SCD 
and SCI who were followed for 3 years, showing an overall higher incidence of cerebral infarct 
recurrence in children with vasculopathy, regardless of transfusion (n=16). Those with 
vasculopathy had a higher rate of CNS events (stroke, new or enlarging SCI) compared to those 
without; 31% (95% CI, 8.34% to 53.66%) versus 12% (95% CI, 5.25% to 18.75%), respectively 
(95% CI, -0.0016 to 0.4381; p= 0.053). Additionally, we have preliminary data from a 
prospective study of children with SCD and prior SCI to suggest that the prevalence of cerebral 
blood vessel abnormalities detected on MRA is predictive of progressive cerebral infarcts in 
children with SCD, Figure 5. 
 
MRA-defined 
cerebral 
vasculopathy 
ABSENT 
MRA-defined 
cerebral 
vasculopathy 
PRESENT 
Total 
Percentage 
of patients 
with MRA-
defined 
cerebral 
vasculopathy 
ABSENCE of  
SCI progression on 
MRI brain 
43 
8 
51 
15.7% 
 
PRESENCE of SCI 
progression on MRI 
brain 
 
1 
4 
5 
80.0% 
 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
10 
 
While there is evidence to support the use of MRA as a predictor of future cerebral infarcts in 
children with SCD, the data in adults is lacking. This study will provide evidence of the 
usefulness of MRA to follow vasculopathy in adults with prior SCI or overt strokes and establish 
that vasculopathy predicts new stroke or TIA. MRA is already being used clinically for this 
purpose in the pediatric population, in light of the evidence that the presence of baseline 
vasculopathy detected on MRA is associated with an increased risk of silent cerebral infarcts in 
children with SCD (P = 0.0007, odds ratio (OR) 2.84; 95% CI = 1.55-5.21)(35). 
While regular blood transfusion therapy is accepted as standard care management for primary 
and secondary stroke prevention in children with SCD, Bishop et al. showed that children with a 
history of stroke experience progression of vasculopathy despite uninterrupted blood transfusion 
therapy(36), suggesting the need for augmented and alternative therapies for this high-risk 
population. To date, no prospective studies have longitudinally quantified the risk of stroke in 
adults with SCD and MRA-defined cerebral vasculopathy. This will be the first prospective 
and retrospective study of adults with SCD to validate the findings of pediatric studies 
that vasculopathy is a significant risk factor for cerebral infarct progression. Identification 
of a subset of high-risk adults with baseline vasculopathy could lead to earlier augmentation of 
current secondary stroke prevention strategies or alternatively, justify the use of novel 
therapeutic approaches for secondary stroke prevention, such as HSCT and experimental 
disease-modifying therapies. In addition, addressing other comorbid conditions that are known 
risk factors for stroke within the general population, such as hypertension, diabetes mellitus, 
hyperlipidemia, atrial fibrillation, and renal disease in adults early on may lead to further stroke 
risk reduction. 
6.0 METHODS 
6.1 Study Design: Multi-center, retrospective-prospective observational cohort study 
6.2 Study Outcomes 
Primary outcome: Radiographic and/or neurological evidence of recurrent cerebral infarction 
(overt strokes, including intracranial hemorrhage, and SCI) in individuals with SCD and prior 
overt stroke or SCI. 
Secondary outcome: Mortality 
6.3 Centers and settings 
This study is an observational, retrospective and prospective cohort study. Patients who meet 
eligibility criteria will be recruited from multiple participating sites that have tentatively agreed to 
participate. Site selection was based on the following criteria 1.) estimated number of patients at 
Figure 5. Preliminary data [DeBaun et al.] shows an increased rate of cerebral infarct progression among patients with SCD 
and prior SCI who have cerebral vasculopathy detected on MRA compared to those without cerebral vasculopathy.  

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
11 
 
each site with a history of SCD and prior overt stroke and SCI, 2.) presence of a seamless 
transition between pediatric and adult sickle cell disease clinics at each site, and 3.) prior 
experience of each site in conducting clinical trials involving patients with sickle cell anemia.  
The seven participating sites include:  
1. Vanderbilt University Medical Center (Nashville, TN)  
2. Washington University Medical Center (St. Louis, MO)  
3. Children’s Hospital Oakland (Oakland, CA)  
4. Medical University of South Carolina (Charleston, SC)  
5. London consortium consisting of 2 sites: Guy’s and St. Thomas’ (London, UK) and Royal 
London Hospital (London, UK) 
6. University of Alabama (Birmingham, AL)  
7. St. Jude-Methodist Sickle Cell Disease Transition Clinic (Memphis, TN)  
Informed consent will be obtained from all study participants. A log to identify all individuals who 
were approached will be kept as part of this study and will be maintained at each site. A one-
sentence reason will be recorded as to why eligible participants did not agree to participate. 
Every attempt will be made by the Study Team to contact participants regularly and keep them 
from becoming lost to follow-up. Identifying a stable contact person (such as a grandparent, 
neighbor, teacher, friend), not living with the study participants’ family but who will always know 
the family’s whereabouts, will assist in tracking participants and decrease the number lost to 
follow-up.   
A maximum of 224 participants ( 1 years of age) with sickle cell disease of any genotype and 
prior overt strokes (n=44) or silent cerebral infarcts (n=180) who have received either 
uninterrupted disease-modifying therapy (regular blood transfusion, hydroxyurea or both) or no 
treatment or intermittent therapy for secondary stroke prevention will be recruited and enrolled 
in the study. Among the 224 study participants, we will include the following 4 groups of patients 
who are receiving uninterrupted disease-modifying therapy for secondary stroke prevention 
versus interrupted therapy or observation alone prior to enrollment: 
a. At least 90 individuals with SCI receiving uninterrupted disease-modifying 
therapy  
b. At least 90 individuals with SCI receiving either no treatment or interrupted 
disease-modifying therapy 
c. At least 22 individuals with overt strokes receiving uninterrupted disease-
modifying therapy  
d. At least 22 individuals with strokes receiving either no treatment or interrupted 
disease-modifying therapy 
Each participant will be prospectively followed for a minimum of 3.5 years between the time of 
the entry and exit MRI/MRA of the brain. We will collect and record all acute neurological events 
(stroke and TIA; see table 2 for definitions) requiring hospitalization or ED visits during the 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
12 
 
study. Imaging and clinical data collected as part of standard care will be used for secondary 
analysis.  
6.4 During the initial visit 
Informed consent, assent or both will be obtained if required. All information, including labs and 
neuroimaging studies, will be collected as standard care. Annual screening for snoring, and 
other sleep disordered breathing will be performed using the previously validated Pittsburgh 
Sleep Quality Index (49) given the impact on cognition as well as cerebral flow (38). A stroke 
free questionnaire will also be administered to children and adult participants at the initial visit 
and at 6 month intervals at routine follow-up visits to assess for any new focal neurological 
deficits which may have occurred, in addition to a standard care neurological examination at 
that follow-up visit.  
Minimum eligibility criterion includes an MR of the brain with at least a 1.5 tesla magnet 
describing the presence of the cerebral infarct. The image must be archived and available to be 
evaluated by the neuroradiology team for central adjudication.  
Follow-up visits: 
Any of the regular office visits will serve as research study follow-up visits. Follow-up visits will 
occur as per standard care. However, data entry will occur every 6 months. The following 
information will be collected during follow-up visits:  
• 
The results of any MRI or MRA of the head, performed routinely as a part of the office 
visit per standard care, will be collected. At minimum, MRI/MRA of the head will occur at the 
time of study entry and exit as part of standard care management for individuals with SCD and 
prior overt strokes or SCI. Crucial to the study is that this will be a retrospective and prospective 
cohort study. Thus, individuals who have had a stroke or SCI prior to 2017 (expected funding 
year), with at least a 1.5 tesla MRI and MRA, are eligible to participate. Thus, some participants 
may be eligible for the protocol who were detected to have a stroke or SCI as early as 2010.  
Inclusion of the past MRI of the brain prior to signing the informed consent, as part of the pre-
existing medical data allows us to capture the baseline events and increases the average 
number of patient years that each person contributes to the cohort. This strategy will prove most 
efficient in determining the natural history of strokes and SCIs in individuals with SCD. 
• 
Clinical information such as height, weight will be collected and vital signs including 
blood pressure, temperature and respiratory rate as part of the routine medical care. Data 
regarding other co-morbid medical conditions and treatment and results of diagnostic lab testing 
will also be collected. We will collect and record all acute neurological events (stroke and TIA) 
requiring hospitalization or ED visits during the study. Imaging and clinical data collected as part 
of standard care will be used for secondary analysis. Study participants receiving interrupted 
disease-modifying therapy and no therapy for secondary stroke prevention will be analyzed in 
the same group.  

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
13 
 
• 
The participant’s primary hematologist will have the option of consulting a neurologist to 
assess for any interval changes in neurological examination at his or her clinical discretion.   
Assuring uniformity of treatment guidelines and compliance with disease-modifying 
therapies:  
Regular blood transfusion therapy defined as scheduled will occur at least every 6 weeks, to 
maintain HbS concentrations less than 30% for at least the first 2 years after the initial overt 
stroke or SCI and less than 50% thereafter for asymptomatic patients. In the event of a 
recurrent stroke or TIA, the hematologist treating the participant will be permitted to individualize 
the transfusion regimen with the goal to lower HbS concentration. The specific type of regular 
blood transfusion therapy used for secondary stroke prevention (simple transfusion, exchange 
transfusion or pheresis) will be recorded in the case report forms (CRF’s) and compared for 
efficacy in the secondary analysis. For patients receiving hydroxyurea for secondary stroke 
prevention, all investigators will titrate the hydroxyurea to either a dose of 20 mg/kg/day or to the 
maximum tolerated dose (MTD). Adherence to hydroxyurea will be defined as an MCV ≥ 10 
ficoliters above baseline. Participants who are not receiving continuous disease-modifying 
therapy for secondary stroke prevention (hydroxyurea, regular blood transfusion, or both) over 
the study period will be included in the no treatment group.  
 
6.5 Chart Review and completion of case report forms  
Through chart review of standard of care procedures, the clinical information recorded will 
include but will not be limited to: 
 
First neuroimaging study demonstrating a cerebral infarct with a minimum 1.5 tesla MRI 
and MRA 
 
Completion of case report form (CRF) for hospitalization visit or emergency department 
visit associated with an acute neurological event, defined as overt stroke or TIA (see 
table 2 for definitions) 
 
Standard care CRFs will be completed every six months. The data will include: pre-
transfusion hemoglobin S levels and hemoglobin levels, neurological examination forms 
(optional and preferably to be completed by a neurologist) 
 
Baseline CRFs focused on co-morbidities and clinical demographic information to be 
completed annually 
 
CRFs related to SAE forms (limited to death, life threatening events, suspected and 
confirmed CNS events 
 
CRFs related to AEs to be completed annually, focused on reasons for hospitalizations 
only 
 
All of this information will be included in our secondary analysis to assess their impact on 
recurrent stroke and SCI. There will be no interventions in this study that are not part of 
standard care management for individuals with SCD who have had prior strokes or SCI. 
 
 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
14 
 
6.6 Schedule of study tests: The table below shows the tests that will be performed at 
baseline and at interval follow-up visits. All testing will be part of standard care management.  
 
 
Baseline 
Every 6 months 
data entry 
Every 12 months 
data entry 
Prior to 
study exit 
Physical 
examination and 
vitals 
X 
X 
 
 
Review of medical 
record, demographic 
factors + 
comorbidities, & 
hospitalizations  
X 
X 
 
 
Assessment for 
snoring & sleep 
disordered breathing 
X 
 
X 
 
Stroke free 
questionnaire 
X 
X 
 
 
MRI/MRA of the brain 
* 
X 
 
 
X 
Neurological 
examination ** 
X 
 
X 
 
Laboratory testing 
(CBC with 
differential, HbF, 
HbS, reticulocyte 
indices) 
X 
X 
 
 
* For the purpose of the primary analysis, only the first and last MRI in the cohort will be 
evaluated to determine progression based on imaging of the brain. 
** Neurological exams will be performed by the participant’s hematologist. Involvement 
of a neurologist to perform neurological exams will be optional.  
 
 
 
Table 1. Schedule of tests that will be performed at baseline and at interval follow-up visits per standard care management.  

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
15 
 
  
 
Figure 6. Flow diagram of study enrollment.  

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
16 
 
 
 
 
 
Inclusion Criteria 
1 
Participants with sickle cell disease of any genotype confirmed on hemoglobin analysis 
and/or other confirmatory documentation of phenotype 
2. 
Patients ≥ 1year of age  
3. 
Patients followed regularly (at least 3 visits per year) in the hematology clinics and who 
have demonstrated adherence with follow-up visits for ≥ 3 years 
4. 
Patients willing to be followed prospectively for a minimum of 3.5 years and agree to a 
standard care exit MRI/MRA of the brain. 
5.  
Evidence of a prior cerebral infarction based on MRI with ≥1.5 m Tesla strength 
6 
Willingness to comply with study protocol, routine clinic visits 
Exclusion Criteria 
Figure 7. Study timeline outlined above.   

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
17 
 
1) Participants judged not to be non-compliant by the hematologist based on previous 
experience in terms of clinic appointments and following advice   
2) Participants with any contraindications to MRI, including individuals with foreign metal objects 
and claustrophobia 
3) Participants with cirrhosis of the liver 
4) Participants with known seropositivity for HIV 
5) Participants with end-stage renal disease (ESRD) 
6) Participants with a baseline oxygen requirement to oxygen saturation < 90% due to chronic 
pulmonary disease 
7) Participants with any baseline rheumatologic disease associated with end-organ disease 
8) Participants who do not meet study criteria for having a history of overt strokes or SCI 
 
6.7 Study Procedures 
6.7.1 Patient Recruitment 
Patients will be recruited starting September 2017 through September 2018. A screening history 
and physical examination, MRI/MRA with ≥1.5 m Tesla strength and neurological examination 
will be completed prior to enrollment of participants to confirm their eligibility for the study. 
Additionally, individuals who have had a stroke or SCI prior to 2017 (expected funding year), 
and have had at least a 1.5 m Tesla MRI and MRA are also eligible to participate. After 
obtaining informed consent, patients will undergo MRI/MRA of the brain at minimum at the time 
of study entry and exit as per standard care management for individuals with SCD and a history 
of strokes or SCI. If participants have had a prior brain MRI and MRA of  1.5 m Tesla strength 
after 2010, this can be used as the initial entry MRA and MRI. Participants will not be 
randomized to any specific interventions for secondary stroke prevention during the course of 
this trial. Standard of care labs, including CBC with differential, reticulocyte count, HbS and HbF 
levels will be ordered at a minimum of every 6 months. Patients will be followed retrospectively 
and prospectively for a minimum of 3.5 years (refer to section 6.6 schedule of study testing). 
6.7.2 Data Collection 
Research information, including consent forms and questionnaires will be maintained in a 
secure fashion in research charts in a locked file room or locked file cabinet.  Only the study 
personnel will have access to the research charts, the database, and to any identifying 
information on participants. All medical information collected prior to signing the informed may 
be reviewed and included in the data set. 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
18 
 
To maintain the confidentiality of research participants, each participant will be identified with a 
unique identifier number in REDCap. Any data shared with our collaborators will be provided 
without participant identifiers (i.e. in coded form only). A de-identified study code for each 
participant will be generated by an independent statistician.  
Annual screening for snoring, and other sleep disordered breathing will be performed using the 
previously validated Pittsburgh Sleep Quality Index (49) given the impact on cognition as well as 
cerebral flow (38). At baseline and every 6 months, a stroke free questionnaire will be 
administered to assess for the possibility of any new focal neurological deficits which may have 
developed in the interim. We will also record specific type of transfusion strategy employed 
(simple transfusion, exchange transfusion or pheresis) and any medications received during the 
course of the study that could have an impact on stroke or SCI recurrence (such as aspirin, 
warfarin and other anticoagulants). We will record any encephaloduroarteriosynangiosis (EDAS) 
procedures, which are performed to improve cerebral blood flow by transposing scalp arteries 
onto the surface of the brain, or administration of tPA into RedCap that occur during the course 
of this study. All of this information will be included in our secondary analysis to assess their 
impact on recurrent stroke and SCI. 
This information will be recorded for all participants prior to study entry, in additional to their 
other medical comorbidities, and updated every 12 months. One investigator will review all 
participants' inpatient and outpatient records to capture critical clinical data. Information 
extracted will include clinical events at the time of the first stroke, initial stroke symptoms and 
treatment, occurrence of second or subsequent strokes, HbS concentration at the time of the 
second stroke, and other neurologic events, such as TIAs or cerebrovascular surgery. As part of 
standard care, the MRI and MRA scans of the brain will be sent to the imaging reading center. 
Typically the provider will order the scan at least once every year to once every two years.  We 
will also review medical records to identify prior or concurrent risk factors for strokes or SCI. The 
information may be included in the prior medical records. Regardless of funding, 6 of 7 sites 
have agreed to follow the cohort for at least 5 years and will plan to follow the participants 
through 2027. Results of annual neurological examinations and neuroimaging (MRI/MRA of the 
brain) will also be entered into RedCap. Data entry will be double-checked by a second 
investigator to maximize accuracy of data entry.   
6.7.3 Data Management 
The data gathered will be entered into the REDCap data system with each ID having a unique 
identifier for each participant. REDCap is a secure, web-based application for building and 
managing online databases. All data will be de-identified and samples will only have a unique 
identifier that will be linked with the patient’s name.  
The study team members will be trained in HIPAA privacy regulations and other applicable site 
privacy policies. No information will be released, nor will participation in the research be 
acknowledged, to any party except where compulsory according to law or intuitional policy.  

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
19 
 
The results of the research study may be published, but participants’ names or identities will not 
be revealed. Records will remain confidential. In order that confidentiality be maintained, the 
principal investigators will keep records in locked cabinets and results of tests will be coded to 
prevent association with participants’ names. Participants’ records will be available to the study 
staff and to each site’s IRB.  
6.7.4 Sample Size 
Sample Size and Power for Aim 1:  
Potential participants will be recruited from seven sites and may include patients followed prior 
to opening the study. We will aim to include a minimum of 224 patients, of which we expect 
approximately 44 to have overt stroke and 180 to have SCI. Based on prior results in the 
children with SCD, the primary hypothesis tested is that the incidence of cerebral infarct 
recurrence in adults with overt strokes or SCI receiving disease-modifying therapy (transfusion 
or hydroxyurea) will be lower than the incidence in adults not receiving disease-modifying 
therapy (or receiving interrupted treatment). 
The primary hypotheses will be tested independently within the overt stroke and SCI groups. 
Based on data from children with SCD, we anticipate that the incidence of stroke recurrence will 
be at least 8 events per 100 patient-years in treated patients with prior overt stroke and at least 
29 events per 100 patient-years in untreated patients with prior over stroke. For patients with 
prior SCI, we anticipate rates of at least 2 events per 100 patient-years for treated patients and 
5 events per 100 patient-years for untreated patients. We calculated statistical power for 
detecting the difference between the treated and untreated groups assuming a constant hazard 
rate; an alpha of .05; a two-sided test; an average loss to follow-up of 10% per year over the 
course of 3 years in all groups, and an average follow-up time of 4.5 years. For SCI, 90 patients 
receiving treatment and 90 receiving standard care results in a power of 81.0% to detect a 
difference in recurrence rates of 2 per 100 patient years versus 5 per 100 patient years. For 
overt stroke, 22 patients receiving treatment and 22 receiving standard care results in a power 
of 88.4% to detect a difference in recurrence rates of 8 per 100 patient years versus 29 per 100 
patient years. 
Sample size calculations for aim 2:  
We calculated our sample size needs based on aim 1, given that aim 1 is the primary aim. 
Nevertheless, power calculations for aim 2 are informative. Based on prior literature, we 
anticipate that among the group with overt strokes, approximately 38% will experience 
progressive vasculopathy, with a relative risk of 12.0 of recurrence for those with progressive 
vasculopathy compared to those without progressive vasculopathy, over 3.5 years of follow-up. 
A sample size of 44 individuals with overt strokes gives us a power of 80.9% to detect a 
difference this large in relative risk between the two groups, assuming an alpha level of 0.05 
and a two-sided Mantel-Haenzel test.  

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
20 
 
For those with SCI, we base the analysis on baseline cerebral vasculopathy since progression 
of vasculopathy occurred in only a very small proportion in prior pediatric studies. Based on data 
from children with SCD, we anticipate a baseline prevalence of vasculopathy of 16% of those 
with SCI, an odds ratio for stroke recurrence of 6.26 comparing those with baseline 
vasculopathy to those without, and a cumulative incidence of stroke recurrence of 10.8% in 
those without baseline cerebral vasculopathy over 3.5 years of follow-up. A sample size of 180 
individuals with SCI gives us a power of 93.9% to detect a difference this large in relative odds 
between the two groups, assuming an alpha level of 0.05 and a two-sided Mantel-Haenzel test. 
6.8 Value of natural history data for stroke recurrence for designing a phase III trial to 
prevent cerebral infarct recurrence.  
We have included sample size calculations for the purpose of illustrating how data collected in 
this proposal will be useful for a planned phase III randomized controlled trial (or non-
randomized parallel comparison trial) comparing standard medical care (regular blood 
transfusion and/or hydroxyurea therapy) to hematopoietic stem cell transplant (HSCT). In an 
entirely separate study protocol from this, participants will be randomly allocated to either 
receive blood transfusion therapy or HSCT (experimental therapy). Participants will be followed 
for a minimum of 3.5 years with the following assumptions: an alpha of 0.05, two-sided is used, 
lost to-follow up rate of 3% per year, and hazard rates with an exponential distribution. The 
standard care arm (blood transfusion and/or hydroxyurea) will have 100 participants and the 
experimental arm (HSCT) will have 50 participants with an expected infarct recurrence 
incidence of 8.1(39) and 1.0 events(6,40), respectively. Based on these assumptions with an 
alpha level of 0.05, the power will be 80%. Clearly without the ability to collect the natural history 
data for stroke recurrence in individuals with SCD, we will not have sufficient information to 
design a phase III trial for HSCT or for that matter other promising therapies that may prevent 
infarct recurrence in this population. 
7.0 
STATISTICAL ANALYSIS  
Statistical analysis for aim 1 and 2: The same descriptive statistics will be used for both aims 
1 and 2. Differences in the distribution of relevant demographic and clinical factors will be 
compared between groups using two-tailed chi-square tests for categorical variables, t-tests for 
continuous variables, and negative binomial regression for count variables. For inferential aims, 
we will use multivariate Cox proportional hazards regression. The first multivariate model will 
use all covariates potentially associated with stroke recurrence, TIA, or death (primary 
endpoints) in one block (forced). A subsequent model will include only those covariates that 
were nominally significant predictors (p<0.20) from the first model. These covariates will include, 
but will not be limited to, age, gender, treatment, length of treatment, baseline MRA-defined 
vasculopathy, and progressive vasculopathy.  
 
As a secondary analysis, we will also explore the relationship between the presence of standard 
risk factors for strokes (hypertension, diabetes mellitus, hyperlipidemia, atrial fibrillation, renal 
disease) and primary endpoints. We will have a sub-analysis focused on: 1) the efficacy of the 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
21 
 
specific blood transfusion therapy method (simple blood transfusion, exchange blood 
transfusion or pheresis) for secondary stroke prevention, 2) hemoglobin S and hemoglobin 
levels at the time of stroke as well as the mean hemoglobin S and hemoglobin level while being 
transfused or receiving hydroxyurea therapy; and 3) the role of excessive iron storage at the 
time of stroke recurrence based on the most recent liver iron content assessment via MRI of the 
liver. Given that the associated morbidity of 5 mm cerebral infarcts has not been uniformly 
assessed, this will be included as part of our secondary analysis. We will record any 
medications taken during the course of the study (such as aspirin, warfarin and other 
anticoagulants) or any interventional procedures (such as EDAS) that could potentially have an 
impact on stroke or silent strokes recurrence, as well any tPA administration during this study. 
This information will be included as part of a post-hoc analysis to estimate their impact on 
recurrent overt strokes and SCI. Statistical analysis will be completed by Dr. Mark Rogeghier, 
Ph.D., an independent statistician based in Chicago, IL. For the past 15 years, Dr. Rodghier has 
been the dedicated statistician for Dr. DeBaun’s research team and he currently has 33 peer-
reviewed publications. 
 
8.0 STUDY ORGANIZATION STRUCTURE  
To accomplish the aims of the study, we will employ the same successful methods of operation 
and procedures from previous stroke prevention trials for children with SCD.  
Executive Committee: The executive committee will review enrollment, protocol conduct, 
scientific merit of proposed ancillary studies or publications, and issues raised by the clinical 
site. Dr. DeBaun will chair the committee, and at least one member from each participating site 
will serve as a member of the executive committee. The committee will meet via 1-hour weekly 
Skype conference calls. Other investigators on the committee will include multi-PI Lori Jordan, 
MD, PhD (Vanderbilt), Bob Mckinstry MD, PhD (Washington University), Adetola Kassim, MD, 
MS (Vanderbilt); and Fenella Kirkham, MD (University College London), an international expert 
in pediatric strokes, with >10 years of experience working with Dr. DeBaun on NIH-funded 
studies and > 20 years of experience in training teams in the UK and Africa on the use of TCD. 
Other investigators on the committee will include multi-PIs: Lori Jordan, MD, PhD (Vascular 
Neurologist), 
Robert 
Mckinstry 
MD, 
PhD 
(Neuroradiologist), 
Adetola 
Kassim, 
MD, 
MS,(Hematologist); and Andria Ford, MD (Vascular Neurologist).  
At least one member of the executive team will visit each site at least once to review research 
governance integrity and to inspect the quality of the source document that was entered into 
REDCap. 
 Neurology Committee: The role of the committee will be to assess and adjudicate patient 
eligibility and endpoint determinations (overt stroke, SCI, TIA) through central review of clinical 
history, neurological examinations and neuroimaging studies when available.  
Neuroradiology Committee: The Washington University neuroradiology committee, led by 
Robert Mckinstry, will form the adjudication committee and will review initial and exit MRI/MRAs 
of the brain. The MRIs/MRAs will be used to assess interval changes when compared to 
baseline neuroimaging studies.  

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
22 
 
There will be no funding for standard care neuroimaging studies (MRI/MRA) and neurology 
examinations. We will have phone conferences monthly to discuss progress and will meet 
annually as well. 
 
9.0 STROKE ADJUDICATIONS 
The stroke adjudication process will provide inclusive and systematic evaluation of all new acute 
neurologic events by treatment-masked neurologists or hematologists and neuroradiologists. 
Neurological examinations can be performed either by the participant’s hematologist or a 
neurologist.  
10.0 DEFINITIONS 
Table 2.  Definition of Neurological Events 
 
Diagnosis = 
Transient 
ischemic attack 
(TIA) 
 
Diagnosis = 
Clinical Stroke 
 
Diagnosis = Clinical 
Stroke 
 
Diagnosis = 
Neurological 
Deficit Due to 
Other Cause 
 
< 24 hours 
Neurological 
Deficit* 
 
 
< 24 hours  
Neurological Deficit* 
 
> 24 hours 
Neurological Deficit* 
 
> 24 hours 
Neurological 
Deficit* 
Negative Head CT 
or MRI 
or no Head CT or 
MRI obtained 
Positive Head CT or 
MRI if obtained for 
clinical indications  
(abnormality on 
brain imaging which 
could explain deficit) 
Positive Head CT or 
MRI if obtained for 
clinical indications  
(abnormality on brain 
imaging which could 
explain deficit) 
Based on 
investigations, 
physicians believe 
deficits are 
consistent with 
complex migraine, 
seizure or other 
non-stroke etiology 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
23 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
10.1 Definitions of acute neurological events 
Overt Strokes 
First and subsequent overt strokes will be defined as new focal neurologic deficits attributable to 
stroke lasting greater than 24 hours. If symptoms resolved within 24 hours, the event will be 
considered a stroke if the patient has had an imaging study demonstrating acute cerebral 
ischemia, based on restricted diffusion on diffusion-weighted MRI, in a region of the brain 
*Neurological deficit in this setting means an abnormality consistent with a stroke. 
Usually, a stroke is associated with weakness of face, arm, and/or leg.   
**The influence of blood transfusion on acute symptoms of stroke has not been well 
defined in SCD; some neurologic symptoms may last less than 24 hours after the 
treatment with blood transfusion therapy obscuring the diagnosis of stroke.  Transfusion 
therapy may decrease the persistence of neurological findings.  Neurologic deficit in 
this setting means an abnormality consistent with a stroke.  

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
24 
 
corresponding with the symptoms if the MRI is done within 7 days of symptom onset. If done 
more than 7 days after symptom onset, a cerebral infarction may only show increased signal 
intensity on T2-weighted MRI as defined below. Intracranial hemorrhage will be counted as an 
overt stroke and will be recorded as a severe adverse event. 
Transient ischemic attacks 
Transient (lasting < 24 hours) weakness or sensory deficits without acute cerebral infarcts on 
MRI of the brain will be considered TIA’s. 
New cerebral infarctions 
New cerebral infarctions, defined as new 3 mm or greater lesions of increased intensity on MRI 
sequences visible in at least 2 views, will be denoted as present or absent.  
Enlarging cerebral infarctions 
Enlarging infarctions will be defined as an increase in a previously existing area of signal 
hyperintensity of at least 3 mm in one dimension.   
Silent cerebral infarctions 
New or enlarging cerebral infarctions on imaging without co-localizing clinical signs or 
symptoms will be defined as silent cerebral infarctions. A silent cerebral infarction could be 
either an enlargement of an existing infarct lesion or a new cerebral infarct, as long as there is 
no new focal neurologic deficit that was called an overt stroke by the treating hematologist. 
Time to first recurrent cerebral infarction 
Time to the first recurrent overt stroke or silent cerebral infarct will be noted as the time to the 
progression of the cerebral infarct after the first stroke. The date for progression of a silent 
infarction will be defined as the date of the MRI on which the new lesion was noted, whereas the 
date for second overt strokes will be defined as the date of symptom onset or recognition.  
10.2 Radiographic definitions of cerebral infarct progression and arterial stenosis 
MRI classification 
Each MRI will be independently reviewed by the study neuroradiologists and subsequently, 
consensus will be achieved. The scan will be judged as 1) no infarct or 2) infarct if there is an 
ischemic lesion ≥ 3 mm in diameter visible in two planes (axial and coronal) using established 
criteria(10). The radiologists will record the total infarct count. The MRI scan will last 
approximately 30 minutes.  
 
Arterial stenosis 
The study radiologists will also grade the MRA brain for vasculopathy. In the major intracranial 
vessels, the first segments of the anterior, middle, and posterior cerebral arteries bilaterally (M1, 
A1, P1) and the intracranial vertebral arteries will be examined. Brain MRA examinations will be 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
25 
 
reviewed for vessel stenoses in both the anterior and posterior circulations. A standardized 
scale for vessel stenosis will provide a means of accurately assessing the location, extent, and 
severity of vascular disease (50). Four segments of each internal carotid artery (ICA), and 3 
segments of each anterior cerebral artery (ACA) and middle cerebral artery (MCA) will be 
examined, with results recorded for a total of 20 vessel segments per subject (10 per cerebral 
hemisphere). Vessel stenosis will be recorded by vascular location, frequency (number of 
stenoses), length (millimeters), and severity (percentage vessel occlusion). Severity of vessel 
segment occlusion will be graded as mild (25-49%), moderate (50-74%), severe (75-99%), or 
occlusion (>99%). These definitions are per prior pediatric SCD and vasculopathy literature 
where stenosis >50% was considered significant cerebral vasculopathy(35,39).The MRA scan 
will last approximately 20 minutes. 
 
10.3 Rationale for Utilization of National Institutes of Health Stroke Scale: 
The National Institutes of Health Stroke Scale (NIHSS) is a validated, standardized neurological 
examination that will be used for this study to perform neurological examinations. The 
neurological examination has been standardized in order to minimize variability that could be 
created by having multiple examiners at various sites. The rapid, standardized NIHSS 
examination is efficiently performed and has been validated for use (44-48). The NIHSS is a 
quantitative neurological examination developed for use in following strokes of various 
etiologies. Specifically, it is easily performed by non-neurologists, detects changes in 
neurological status well, and has excellent inter rater reliability, which can be further improved 
through the use of standardized video training. The scale consists of 11 items. Normal 
performance in any area receives a score of zero; hence a completely normal patient will have a 
cumulative score of 0. The involvement of a neurologist to complete neurological examinations 
for the purposes of this study will remain optional and left to the discretion of the treating 
hematologist. All study personnel responsible for neurological evaluations will be required to 
provide proof of certification on the NIHSS to the Coordinating Center (CC). For subjects under 
18 years of age, the previously validated pediatric NIH Stroke Scale and Modified Rankin Scale 
for Children will be used to record the presence of any focal neurological deficits on examination 
(51,52).  
 
11.0 STUDY RESPONSIBILITIES 
11.1 Coordinating Center  
The Coordinating Center (CC) is directly responsible for the monitoring and oversight of all 
fiscal, regulatory, clinical and other general administration needed to centralize and coordinate 
the research endeavors of all sites participating in the trial.  The CCC will be located at 
Vanderbilt University Medical Center and will be led by the Principal Investigator, Michael R. 
DeBaun, MD, MPH.  Two project managers, with overlapping responsibilities, will individually be 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
26 
 
responsible for the clinical and fiscal duties. The project manager will also plan and coordinate 
all meetings and travel related to the study. 
The responsibilities of the CCC will include:   
• 
protocol development and amendments 
• 
facilitating and monitoring protocol conduct at the participating sites 
• 
overseeing regulatory compliance  
• 
monitoring of adverse effects and events 
• 
assuring quality control via site audits 
• 
providing annual and quarterly reports on the progress of the study 
• 
participating in the analysis and interpretation of data 
• 
manuscript preparation and prioritization  
• 
maintaining ongoing communication with all participating study sites 
• 
plan and organize investigator meetings 
• 
coordinate all NIH requirements related to the grant 
 
12.0 PRIVACY AND CONFIDENTIALITY ISSUES 
All key personnel involved in the design or conduct of research involving the human subjects will 
receive the required education on the protection of human research participants prior to funding 
of this project. The patients will not be individually identifiable.  Confidential data, including 
name, address, employer, and name and address of relatives, are collected to aid in 
maintaining patient follow-up. However such information is kept separate from study forms in a 
secure file.  
Patients’ risks of participating in research are kept to a minimum with measures to protect 
confidentiality and planned interim analyses for safety monitoring and early termination.  Steps 
to protect privacy will include assignment of study codes to all records, without patient names, 
with the key for connecting patient names and study codes kept in a secure site separate from 
the study records and at the local site only.  All study records will be kept in a secure location 
that will be locked to prevent unauthorized access. 
13.0 HUMAN SUBJECTS RESEARCH 
13.1 IRB Review 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
27 
 
The protocol, informed consent documents, and all types of participant educational information 
must be submitted to the IRB for review and must be approved before the study is initiated.  Any 
amendments to the protocol must also be approved by the IRB prior to implementing any 
changes in the study.   
The investigator is responsible for keeping the IRB updated of the progress of the study and of 
any changes made to the protocol as deemed appropriate, but in any case at least once a year. 
The investigator must also keep the IRB informed of any significant adverse events.  
The study may be discontinued for administrative reasons and/or IRB.   
If for any reason the study is prematurely terminated, the Investigator should promptly inform 
the study participant and the IRB.  
All key study staff regularly undergo training in human subjects issues. 
13.2 Consent Process 
Written informed consent will be obtained from all study participants, without exception, before 
their enrollment into this study.  A waiver of consent will not be sought for this study. The study 
investigator or coordinator will review all portions of the consent in detail.  Any questions will be 
addressed prior to asking for a signature.  Study staff will emphasize that participants may 
continue to ask questions at any time during the study and they may withdraw consent at any 
time.  
A copy of the consent will be given to the participant before participation in the study begins.  
The original signed consent and will be available for review at any time. 
Patients with a history of stroke and significant associated cognitive deficit(s) precluding their 
ability to provide informed consent will not be excluded if they have a family member or 
significant other with Power of Attorney to consent on their behalf.  
Members of the Data Safety Monitoring Committee (DSMC) will be selected to reflect a mix of 
appropriate clinical expertise in sickle cell disease and knowledge of the design, monitoring, 
analysis and ethical issues of clinical research to protect the participants’ safety.  The DSMC will 
monitor study quality, safety of participants, and efficacy.  
 
The DSMC will monitor the study performance by reviewing: 
 
participant recruitment 
 
 
flow of forms  
 
 
quality control of the data 
 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
28 
 
 
adequacy of medical monitoring  
 
 
adverse event reporting  
 
 
adherence to protocol  
 
 
appropriateness of protocol changes with regard to scientific integrity 
 
 
The DSMB will monitor the safety of participants by reviewing: 
 
risk of harm inherent in participating in the study  
 
 
adverse events (type, incidence, and severity) 
 
 
 
effect of protocol changes on risk 
The DSMC will monitor the efficacy of the study by reviewing: 
 
data  
 
 
planned and/or unplanned interim analyses  
 
 
stopping rules, their implementation, and resulting decisions 
 
 
results and conclusions 
13.3  Statistical Coordinating Center  
The Statistical Coordinating Center (SCC) will reside at Vanderbilt University Medical Center.  
The data will be developed, managed and entered using REDCap, a secure, web-based 
application designed exclusively to support data capture for research studies. The SCC will 
coordinate the collection and analysis of data generated for the study. 
13.4 
Preparation of study documents 
SCC staff will collaborate with staff from the CCC and other investigators to develop:  protocol, 
Manual of Operating Procedures (MOP), case report forms (CRFs), and Question by Question 
guidelines (QxQs), which give the details on answering the questions on every CRF. 
13.5 
Training site personnel 
SCC staff will train site personnel in all aspects of the study related to: details of the study 
protocol, data collection procedures, screening procedures, submitting forms to the SCC and 
responding to queries from the SCC. SCC staff will also monitor each site to assure that the site 
coordinators have completed the required training. 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
29 
 
13.6 
Receipt and processing of study forms at the SCC 
SCC staff will design and implement the data management system for the study, including: 
electronic data entry, comprehensive editing of the data and preparation of protocol adherence 
aids, and data extraction for analyses related to research objectives. Data are submitted to the 
SCC through REDCap, a web-based data entry system. Data are comprehensively edited once 
received at the SCC, prior to insertion into the main study database. Edit queries will be 
resolved within a few days with the site coordinators. Protocol adherence aids, such as 
delinquent forms listings and patient schedules, are produced by the data management system 
to help the site coordinators keep the patient visits on schedule with the appropriate forms 
submitted. SCC staff will also enforce patient confidentiality and privacy rules so that patient 
health information (PHI) data are not shared inappropriately and so that data are protected by 
multiple layers of security. 
13.7 
Monitoring study progress and data quality 
SCC staff will ensure that clinic site staff is properly trained, that performance of the required 
procedures is monitored, and that deviations from the protocol or from study norms are 
investigated. Routine site monitoring reports, including patient recruitment, performance of 
follow-up visits, and protocol compliance, will be produced at regular intervals. 
SCC and CCC staff will develop and implement quality assurance procedures and programs will 
be developed and implemented for central reading centers, data collection, and data entry.   
13.8  Quality Assurance Procedures 
The study investigators and study personnel will conduct quality assurance site visits throughout 
the study as a means of building rapport. In addition, quality assurance procedures and 
programs will be developed and implemented for data collection and for data entry by the 
Statistical Coordinating Center and Coordinating Center.  At least one site visit will occur by 
members of the executive team to each site to review the research governance documents and 
quality of the collected data, including sources documents. 
13.9 
Collaboration in Study Publications 
SCC staff will collaborate in all study publications, providing statistical analysis, preparing the 
statistical methods and results sections of manuscripts. SCC staff will also participate in the 
development and review of abstracts for presentations and in the review of the statistical 
analysis for ancillary publications. 
14.0 ADVERSE EVENT REPORTING 
For the purposes of this study an adverse event (AE) will be defined as any unfavorable and 
unintended sign, symptom or disease associated with a subject’s participation in this study.  As 
this is an observational study, reportable adverse events are only those directly related to an 
individual’s participation in this study 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
30 
 
Although adverse events are not anticipated from the nature of this study, all participants will be 
under the care of experienced study staff who will be available to provide necessary care to 
participants who may experience an adverse event during the study.   
The investigator or designate is responsible for the detection and documentation of AE and 
serious adverse events (SAE) in persons participating in this study.  At each clinical evaluation 
during the study, the investigator or site personnel should document any AEs or SAEs, as 
detailed in this protocol.  AEs and SAEs should be reported to the IRBs. 
All adverse events (AE), regardless of causal relationship, will be monitored and reported on a 
form that includes the description of the event, onset date, stop date, and outcome. The 
Investigator will determine the intensity of any AE according to the NCI Common Terminology 
Criteria for Adverse Events (CTCAE) Version 4.03 (see http://ctep.info.nih.gov) and their causal 
relationship. Adverse events will be reported to the institutional review board at the time of study 
renewal or final report submission. All SAE will be reported to the DSMC and IRB within 24 
hours of awareness of the SAE. The DMSC will review the progress and safety of the study 
every 3 months. Recruitment, follow-up rates, compliance with the study protocol and safety will 
be reviewed.  
This study is currently not federally funded. However, if this research study becomes federally 
funded in the future, any action resulting in a temporary or permanent suspension of this project 
will be reported to the grant program director within 48 hours in writing.  
 
Each individual participating site will be responsible for completing case report forms (CRFs) 
related to serious adverse events (SAE), limited to death, life threatening events, suspected and 
confirmed CNS events. Each participating site will also be responsible for completing CRFs 
related to adverse events (AEs) annually, focused on reasons for hospitalizations only. Each 
participating site will be responsible for submitting CRFs for SAE and AE to the coordinating 
center's project manager. The coordinating center's project manager will be responsible for 
disseminating the AE and SAE case report forms to other participating centers, the participating 
site IRBs, data safety monitoring boards and applicable regulatory agencies. 
 
14.1 Potential risks to the study participant  
MRI/MRA of the brain: During the MRI scan, the patient lies in a closed area inside the magnetic 
tube. Some patients may experience claustrophobia during the procedure. Therefore, 
participants with any history of claustrophobia will be asked to report this to their hematologist 
so that he or she can make appropriate accommodations for the participant’s claustrophobia if 
necessary. Patients with metal devices in their body or any other contraindications to a 
magnetic resonance study will be excluded from this study for their own safety (see Exclusion 
Criteria in Section 6.6 for a full list of contraindications). 
Phlebotomy: Phlebotomy is often associated with mild pain and occasionally with a small bruise 
at the site. Rarely, patients feel lightheaded or have syncope. There is also a very small risk of 
infection at the site of the needle puncture. 

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
31 
 
Questionnaires:  The participant may experience emotional discomfort when answering some 
questions. If a particular question makes the parent or participant uncomfortable, the parent or 
participant can discuss its importance and the need to answer it with the reviewer. Completing 
the questionnaires may be inconvenient because of the time it takes to complete them. 
Disclosure of confidential information:  There is a small risk of breaches in confidentiality.  This 
may affect the participant’s future ability to obtain employment or medical, disability, or life 
insurance. 
15.0 STUDY WITHDRAWAL AND DISCONTINUATION 
The Principal Investigator (PI) may withdraw the patient from the study if considered 
appropriate. It may be in the best interest of the patient to allow follow-up outside the study.   
The PI will share any new information that could change how patients feel about continuing in 
the study. The study may be discontinued by the Investigator, and/or IRB.  If the study is 
prematurely terminated or suspended, the Investigator shall promptly inform the IRB and 
enrolled patients. 
16.0 FOLLOW-UP AND RECORD RETENTION 
Patients will be followed for 2 years after the completion of the trial. Records will be kept for 2 
years after the completion of the study.  
17.0 LONG TERM OBJECTIVES 
This will be the first observational cohort study to prospectively and retrospectively describe the 
natural history of individuals with SCD and prior SCI or overt strokes receiving standard care 
management (regular blood transfusion therapy and/or hydroxyurea) to attenuate further 
neurological injury. The discovery of progressive or new central nervous system disease will 
inform the patient, family and health care provider as to whether more intensive interventions 
should be considered to prevent further progression of cerebral infarcts. Descriptive outcomes 
from this study could help guide the development of hypotheses-driven randomized clinical trials 
to optimize the management of patients with SCD and prior cerebral infarcts. 
 
18.0 REFERENCES: 
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Investigators. Discontinuing prophylactic transfusions used to prevent stroke in sickle cell disease. N 
Engl J Med. 2005 Dec 29;353(26):2769–78.  

Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
32 
 
3.  Ware RE, Helms RW, SWiTCH Investigators. Stroke With Transfusions Changing to Hydroxyurea 
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Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
33 
 
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Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
34 
 
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Principal Investigator:  Dr. Michael R. DeBaun 
 
Revision Date:  11-21-2016 
Study Title:  The natural history of overt or silent strokes in individuals with sickle cell disease treated with blood transfusion or hydroxyurea 
therapy 
Institution/Hospital: Vanderbilt University Medical Center 
 
35 
 
42.  Prasad K, Dash D, Kumar A. Validation of the Hindi version of National Institute of Health Stroke 
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