ECMO Hemostatic Transfusions in Children
Phase: PHASE4
Conditions: Extracorporeal Membrane Oxygenation Complication, Hemorrhage, Thromboembolism, Transfusion Adverse Reaction
Sponsor: Yale University

Study Protocol
Version 1.4 – April 22, 2024
IRB APPROVED
05/15/2024
NCT05796557

ECMO Hemostatic Transfusions in Children (ECSTATIC)
Study Protocol – version 1.4 – April 22, 2024
2
1.
Table of Contents
1.
TABLE OF CONTENTS ....................................................................................................................................2
2.
ABSTRACT ....................................................................................................................................................5
Background...........................................................................................................................................................5
Design ...................................................................................................................................................................5
Population.............................................................................................................................................................5
Interventions.........................................................................................................................................................6
Allocation..............................................................................................................................................................6
Outcome Measures...............................................................................................................................................6
Statistical Design and Power ................................................................................................................................6
Subject Participation Duration..............................................................................................................................6
3.
STUDY TEAMS ..............................................................................................................................................7
A.
CO-PRINCIPAL INVESTIGATORS.................................................................................................................................7
B.
EXECUTIVE COMMITTEE..........................................................................................................................................7
C.
STATISTICIANS AND DATA COORDINATING CENTER.......................................................................................................7
D.
PARTICIPATING INSTITUTIONS AND PRINCIPAL INVESTIGATORS........................................................................................8
4.
ABBREVIATIONS...........................................................................................................................................9
5.
INTRODUCTION..........................................................................................................................................10
A.
BACKGROUND ....................................................................................................................................................10
Pediatric ECMO...................................................................................................................................................10
Epidemiology of Platelet Transfusions................................................................................................................11
Outcomes Associated with Platelet Transfusions ...............................................................................................12
Mechanisms of Poor Outcomes ..........................................................................................................................12
Summary.............................................................................................................................................................12
B.
PRELIMINARY STUDIES..........................................................................................................................................13
C.
JUSTIFICATION FOR THE STUDY ...............................................................................................................................13
D.
HYPOTHESES AND AIMS........................................................................................................................................14
E.
INNOVATION ......................................................................................................................................................14
6.
PATIENT ELIGIBILITY ...................................................................................................................................15
A.
ANTICIPATED COHORT..........................................................................................................................................15
B.
CRITERIA TO IDENTIFY PARTICIPATING SITES..............................................................................................................15
C.
ELIGIBILITY.........................................................................................................................................................16
D.
INCLUSION CRITERIA ............................................................................................................................................16
E.
EXCLUSION CRITERIA............................................................................................................................................16
7.
CONSENT....................................................................................................................................................17
A.
INFORMED CONSENT............................................................................................................................................17
8.
STUDY DESIGN ...........................................................................................................................................18
A.
INTERVENTION....................................................................................................................................................18
B.
SAFETY..............................................................................................................................................................20
C.
CO-INTERVENTIONS.............................................................................................................................................21
D.
COMPLIANCE MEASURES ......................................................................................................................................22
Measures to Ensure Compliance.........................................................................................................................22
 
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ECMO Hemostatic Transfusions in Children (ECSTATIC)
Study Protocol – version 1.4 – April 22, 2024
3
E.
RANDOMIZATION ................................................................................................................................................22
F.
BLINDING...........................................................................................................................................................23
G.
TREATMENT PERIOD AND FOLLOW UP.....................................................................................................................23
H.
TEMPORARY SUSPENSION OF INDIVIDUAL PATIENTS....................................................................................................24
9.
OUTCOMES ................................................................................................................................................24
A.
PRIMARY OUTCOME ............................................................................................................................................24
B.
SECONDARY OUTCOMES.......................................................................................................................................25
C.
TERTIARY OUTCOMES...........................................................................................................................................26
Bleeding and/or Thrombotic Outcome Adjudication ..........................................................................................27
D.
OTHER OUTCOMES..............................................................................................................................................28
E.
APPLICATION OF RESULTS TO THE IMPLEMENTATION OF THE FULL TRIAL.........................................................................29
10.
SAMPLE SIZE AND STATISTICAL ANALYSIS...................................................................................................29
A.
SAMPLE SIZE ......................................................................................................................................................29
B.
PLANNED RECRUITMENT RATE...............................................................................................................................30
C.
STATISTICAL ANALYSIS..........................................................................................................................................30
Statistical Support...............................................................................................................................................32
Interim Analysis ..................................................................................................................................................32
D.
LIMITATIONS WITH STRATEGIES TO OVERCOME .........................................................................................................33
11.
STUDY MONITORING AND STUDY ORGANIZATION.....................................................................................34
A.
STUDY MONITORING ...........................................................................................................................................34
B.
DATA SAFETY MONITORING BOARD........................................................................................................................35
DSMB ..................................................................................................................................................................35
Monitoring..........................................................................................................................................................36
DCC Features and Safeguards.............................................................................................................................37
Adverse Events....................................................................................................................................................38
Adverse event reporting......................................................................................................................................40
C.
STUDY ORGANIZATION .........................................................................................................................................40
Study primary investigators................................................................................................................................40
Executive Committee ..........................................................................................................................................40
Steering Committee ............................................................................................................................................41
Writing Committee .............................................................................................................................................41
Data Coordinating Center...................................................................................................................................42
D.
STUDY TIMELINE .................................................................................................................................................43
12.
REFERENCES...............................................................................................................................................43
13.
APPENDICES...............................................................................................................................................48
A.
PROPOSAL FOR FUTURE LARGE RCT ........................................................................................................................48
Aims ....................................................................................................................................................................48
Anticipated Cohort..............................................................................................................................................48
Design .................................................................................................................................................................48
Site Eligibility.......................................................................................................................................................48
Patient Eligibility.................................................................................................................................................48
Exclusion Criteria ................................................................................................................................................49
Intervention ........................................................................................................................................................49
Co-interventions..................................................................................................................................................49
Randomization....................................................................................................................................................50
Concealment of randomization ..........................................................................................................................50
 
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ECMO Hemostatic Transfusions in Children (ECSTATIC)
Study Protocol – version 1.4 – April 22, 2024
4
Blinding...............................................................................................................................................................50
Temporary suspension........................................................................................................................................50
Primary Outcome................................................................................................................................................50
Secondary Outcomes ..........................................................................................................................................51
Sample Size .........................................................................................................................................................51
Statistical Analyses .............................................................................................................................................52
B.
BLEEDING ASSESSMENT SCALE IN CRITICALLY ILL CHILDREN (BASIC) SCORE ...................................................................53
C.
NHLBI DEFINITION OF SEVERE THROMBOTIC COMPLICATIONS.....................................................................................54
D.
PCPC AND POPC SCORES.....................................................................................................................................54
E.
WORKSHEET FOR SITE ASSESSMENT OF SEVERE BLEEDING/CLOTTING EVENTS.................................................................56
F.
PLATELET TRANSFUSION STRATEGY ADHERENCE (COMPLIANCE)...................................................................................58
 
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ECMO Hemostatic Transfusions in Children (ECSTATIC)
Study Protocol – version 1.4 – April 22, 2024
5
2.
Abstract
Background
Due to coagulopathy and thrombocytopenia induced by hemodilution and 
the extracorporeal circuit itself, children supported by extracorporeal 
membrane oxygenation (ECMO) are at significant risk of bleeding. To 
prevent bleeding, pediatric intensivists often prescribe prophylactic 
platelet transfusions. However, in observational studies, prophylactic 
platelet transfusions to children on ECMO have been independently 
associated with increased thrombosis, mortality, and paradoxically, 
increased bleeding. Guidelines to direct platelet transfusions in this 
patient population are limited by the lack of evidence and therefore based 
on expert opinion alone. Given the significant associated risks, it is crucial 
to provide evidence to guide clinicians.
ECSTATIC is designed to address the lack of evidence on platelet 
transfusion strategies in pediatric patients on ECMO, through a 
randomized controlled trial (RCT), comparing two prophylactic transfusion 
approaches in non-bleeding children.
The proposed pilot trial is innovative in that it is focused on children 
supported by ECMO, a population in whom transfusion strategies have 
never been tested previously; it involves the largest separation between 
the two arms of any platelet transfusion trial conducted in the past; and it 
involves two newly developed definitions of bleeding and thrombosis 
particularly applicable to children supported by ECMO.
The R34 pilot trial will provide necessary and sufficient information to 
proceed with the definitive ECSTATIC RCT to evaluate the impact of a 
low platelet transfusion threshold on the clinical outcomes in children on 
ECMO. ECSTATIC has the potential to optimize efficacy, to reduce 
platelet transfusion exposure and to decrease mortality and morbidity of 
these extremely ill neonates and children.
Design
Pilot randomized controlled trial
Population
We will include critically ill children (0 to <18 years of age), admitted to a 
participating PICU/NICU/CICU, on ECMO, and who have either no 
bleeding or minimal bleeding within 24 hours of cannulation. 
We will exclude children with 1) post conception age < 37 weeks at time 
of screening; 2) underlying oncologic diagnosis or recipient of bone 
marrow transplant in the last year; 3) congenital bleeding disorder or 
thrombocytopenia; 4) pregnant or admitted post-partum (within 6 weeks 
after giving birth); 5) decision to withdraw or withhold some critical care or 
interventions; and 6) known objection to blood transfusions; or 7) on 
ECMO for > 24 hours at time of enrollment. 
Fifty patients will be enrolled in the pilot trial in ten sites (9 in the United 
States and 1 in Israel). Enrolled means consented and randomized.
 
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ECMO Hemostatic Transfusions in Children (ECSTATIC)
Study Protocol – version 1.4 – April 22, 2024
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Interventions 
Comparator: Higher prophylactic platelet threshold strategy: Patients 
randomized to this arm will be transfused as soon as the platelet count is 
< 90 x 10^9 cells/L.
Intervention: Lower prophylactic platelet threshold strategy: Patients 
randomized to this arm will be transfused as soon as the platelet count is 
< 50 x 10^9 cells/L.
Allocation 
Subjects will be randomized in a 1:1 ratio to either arm. Subjects will be 
stratified by age (≤28 days vs >28 days).
Outcome 
Measures 
Primary Study Outcome: Pre-transfusion platelet count (supplemented 
with total volume of platelet transfusions, in mL/kg/day and in mL/kg/run); 
primary safety outcome: progression to severe bleeding, severe 
thrombosis, and/or all-cause mortality.
Secondary Endpoints: Observed values of estimated separation in mean 
pre-transfusion platelet counts; eligibility rate relative to predicted rate; 
enrollment of eligible patients percentage; adjudication rate of 
progression to severe bleeding/thrombotic outcomes; approach for 
consent percentage; consent percentage; transfusion compliance rate; 
transfusion strategy suspension percentage; withdrawal and or lost-to-
follow-up percentage; violation of local non-study-specific circulatory 
support protocol percentage.
Tertiary Outcome: A novel composite outcome of severe bleeding and/or 
severe thrombosis (supplemented with component outcomes of 
progression to severe bleeding, progression to severe thrombosis, and 
progression to both severe bleeding and severe thrombotic thrombosis.
Statistical 
Design and
Power 
For this pilot trial, enrolling 50 patients (25 per arm) would lead to 
approximately 81% power to detect a difference in mean platelet levels at 
the time of transfusion, assuming an autocorrelation between platelet 
levels at the time of transfusion of 0.3, a difference between the pre-
transfusion platelet counts of 30, and a standard deviation of the pre-
transfusion platelet count of 40.
Subject 
Participation 
Duration 
Subjects will be enrolled over a 24-month period (quarter 3 to 10 of the 
study). Each patient will be followed daily until 24 hours past the treatment 
strategy’s expiration (i.e., until at most day 22 after randomization). 
Mortality will be measured at day 28 (+/- 4 day window) and day 90 (+/- 
10 day window).
 
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05/15/2024

ECMO Hemostatic Transfusions in Children (ECSTATIC)
Study Protocol – version 1.4 – April 22, 2024
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3.
Study Teams
A. Co-Principal Investigators
Marianne Nellis, MD, MS
Division of Pediatric Critical Care Medicine
Weill Cornell Medicine
525 East 68th St, M-512
New York, NY 10065
man9026@med.cornell.edu
Oliver Karam, MD, PhD
Section of Pediatric Critical Care Medicine
Yale School of Medicine
333 Cedar Street
New Haven, CT 06520-8064
oliver.karam@yale.edu
B. Executive Committee
Jennifer Muszynski, MD
Nationwide Children’s Hospital
Columbus, OH
Jennifer.Muszynski@nationwidechildrens.org
Philip Spinella, MD
University of Pittsburgh
Pittsburgh, PA
spinella@pitt.edu
Marisa Tucci, MD
Sainte Justine Hospital
Montreal, QC, Canada
marisa.tucci@gmail.com
Jacques Lacroix, MD
Sainte Justine Hospital
Montreal, QC, Canada
jlacroix052@gmail.com
Simon Stanworth, MD
John Radcliffe Hospital
Oxford, UK
simon.stanworth@nhsbt.nhs.uk
Paul Clarke, MD
Norfolk and Norwich University Hospital
Norwich, UK
paul.clarke@nnuh.nhs.uk
Scott Weiss, MD, MSCE
Nemours Children’s Hospital 
Wilmington, DE
scott.weiss@nemours.org
C. Statisticians and Data Coordinating Center
Bradley Barney, PhD
Statistician
University of Utah
Salt Lake City, UT
bradley.barney@hsc.utah.edu
Chella Palmer
Statistician
University of Utah
Salt Lake City, UT
Chella.palmer@hsc.utah.edu
Eun Hea Unsicker
Coordinator
Trials Innovation Network
University of Utah
Salt Lake City, UT
eunhea.unsicker@hsc.utah.edu
 
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ECMO Hemostatic Transfusions in Children (ECSTATIC)
Study Protocol – version 1.4 – April 22, 2024
8
D. Participating Institutions and Principal Investigators
Jesse Bain, DO
Children’s Hospital of Richmond
Richmond, VA
jesse.bain@vcuhealth.org
Jamie Furlong-Dillard, MD
Norton Children’s Hospital 
Louisville, KY
jamie.furlong-
dillard@louisville.edu
Umesh Joashi, MBBS
Komansky Children’s Hospital of NY Presbyterian
New York, NY
ucj4001@med.cornell.edu
Eva Cheung, MD
Morgan Stanley Children’s 
Hospital of NY Presbyterian
New York, NY
ec2335@cumc.columbia.edu
Garrett Coles, MD
Golisano Children's Hospital
Rochester, NY
garrett_coles@urmc.rochester.edu
Aditya Badheka, MBBS, MS
University of Iowa Health Care
Iowa City, IA
aditya-badheka@uiowa.edu
Robert Niebler, MD
Children’s Hospital of Wisconsin
Milwaukee, WI
rniebler@mcw.edu
Matthew Paden, MD
Children’s Healthcare of Atlanta – 
Emory
Atlanta, GA
matthew.paden@choa.org
Caroline Ozment, MD
Duke University School of Medicine
Durham, NC
caroline.ozment@duke.edu
Ofer Schiller, MD
Schneider Children’s Medical 
Center
Petach Tikvah, Israel
schillero@clalit.org.il
 
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ECMO Hemostatic Transfusions in Children (ECSTATIC)
Study Protocol – version 1.4 – April 22, 2024
9
4.
Abbreviations
BASIC
Bleeding Assessment Scale in Critically Ill Children
CICU
Cardiac intensive care unit
DCC
Data Coordinating Center
DSMB
Data Safety Monitoring Board
ECMO
Extracorporeal membrane oxygenation
ECPR
Extracorporeal cardiopulmonary resuscitation
ECSTATIC
ECMO hemostatic transfusions in children
EFIC
Exception from informed consent
IRB
Institutional Review Board
NHLBI
National Heart, Lung, and Blood Institute
NICU
Neonatal intensive care unit
PICU
Pediatric intensive care unit
RCT
Randomized controlled trial
TEG
Thromboelastometry
V-A
Veno-arterial
V-V
Veno-venous
 
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ECMO Hemostatic Transfusions in Children (ECSTATIC)
Study Protocol – version 1.4 – April 22, 2024
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5.
Introduction
A. Background
The ultimate goal of our program is to improve outcomes in pediatric patients on extracorporeal 
membrane oxygenation (ECMO). We propose a randomized controlled trial (RCT), comparing 
two prophylactic platelet transfusion approaches in non-bleeding children, to identify if a low 
platelet threshold transfusion strategy is non-inferior to a high platelet threshold transfusion 
strategy. Prior to conducting the definitive trial, we propose to undertake a pilot trial to establish 
the degree of separation between intervention arms, the consenting process, and the adjudication 
of outcomes.
Pediatric ECMO
Over 3,600 neonates and children are supported worldwide each year by ECMO, with an increase 
of 5-10% in the number of cases each year (1,2). ECMO serves as cardiopulmonary support in 
critically ill patients with severe, refractory respiratory or heart failure. In addition, extracorporeal 
cardiopulmonary resuscitation (ECPR) can be employed during cardiac arrest. The system 
includes an extracorporeal circuit (tubing) with large cannula draining blood from the body, an 
external pump draining systemic venous blood towards a membrane allowing gas exchange 
(oxygenation and CO2 removal) and recirculating warmed blood into a main vessel. In veno-
arterial (V-A) ECMO, blood is drained from a large vein and returned to an artery, supporting both 
the heart and lungs, whereas in veno-venous (V-V) ECMO, the blood is drained from a vein and 
returned to a vein, thereby providing primarily pulmonary support. This implies that central vessels 
(internal jugular vein or femoral vein, and aorta or femoral artery) are cannulated, either 
percutaneously or by surgical cut-down. Cannulation sites depend on the patient age, weight and 
clinical context. Central ECMO via median sternotomy and exposure of cardiac structures is a 
preferred option when used immediately after cardiopulmonary bypass during cardiac surgery. 
The ECMO circuit is typically primed with red blood cells and plasma or whole blood in infants 
and small children and may be primed with isotonic fluid in older children and adults.
While ECMO may be lifesaving, bleeding is a common complication due to extracorporeal circuit-
induced coagulopathy and platelet dysfunction due to exposure of blood to the ECMO circuit (3–
5). Given the risk of thrombosis, children are also placed on anticoagulation which may contribute 
to the risk of bleeding. In a cohort of over 500 infants and children supported by ECMO, 70% had 
significant bleeding (as defined by the Extracorporeal Life Support Organization) by day 10 and 
bleeding events were independently associated with time to death (OR 1.75, 95% CI 1.20-2.55) 
(6). Furthermore, the amount of quantifiable bleeding (as measured from chest tube output) was 
associated with mortality in a dose-dependent fashion (7). 
Patients can develop either thrombocytopenia and/or platelet dysfunction while supported by 
ECMO. Studies of neonates have shown drops in platelet counts ranging from 25-60% shortly 
after cannulation (3,8), whereas pediatric patients, with a larger baseline circulating blood volume 
and less hemodilution, have been reported to drop their platelet count by approximately 30% (9). 
Few studies have examined platelet dysfunction in children on ECMO. In a case series of 10 
neonates on ECMO for respiratory failure, platelet aggregation studies 15 minutes after the 
initiation of ECMO demonstrated a 46% mean decrease in the response to collagen from 
baseline, and a significantly reduced response to ristocetin and to adenosine 5'-diphosphate (8). 
 
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ECMO Hemostatic Transfusions in Children (ECSTATIC)
Study Protocol – version 1.4 – April 22, 2024
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In a retrospective study of 24 children who had thromboelastography (TEG) testing performed 
during ECMO support (10), severe qualitative platelet dysfunction was more commonly seen for 
adenosine diphosphate (ADP)-mediated aggregation (92%) compared to arachidonic acid (AA)-
mediated aggregation (75%). In addition, ADP-mediated percent of platelet aggregation was 
significantly lower than AA-mediated platelet aggregation (15% vs. 49%, p<0.001). However, no 
significant changes in aggregation were seen for bleeding versus non-bleeding patients. In 
general, platelet function testing is limited in neonates and children due to the blood volume 
required, as well as lack of standardization for pediatric norms (11).
Epidemiology of Platelet Transfusions
To 
prevent 
bleeding 
and 
its 
associated 
outcomes, 
clinicians 
frequently 
prescribe 
platelet transfusions, to enhance hemostasis. In 
the same cohort of 514 children on ECMO, 
children received platelet transfusions on over 
two-thirds of all ECMO days (Figure 1) (12). 
As compared to other critically ill children, 
pediatric patients on ECMO receive a large 
proportion of platelet transfusions. In an 
epidemiologic study of critically ill children 
receiving platelet transfusions on ECMO, each 
child received an average of 92 mL/kg of platelet 
transfusions over the course of their ECMO 
support (13). Ninety-seven percent of patients received at least one platelet transfusion during 
their ECMO course (6,9). The transfusion of platelets, as well as other blood components, can 
lead to microthrombi in the ECMO circuit (14), as well as clinically relevant thrombotic 
complications in the patient. In the cohort of 514 children on ECMO, 33% had a significant 
thrombotic event by day 10 (Figure 2) (6).
Clinicians have very little evidence-based guidance 
as to the indications for the prescription of 
prophylactic platelet transfusions for children on 
ECMO. ECSTATIC seeks to provide evidence to 
guide prophylactic platelet transfusion strategies for 
children requiring ECMO support. Currently, both 
ELSO (15) and the Association for the Advancement 
of Blood & Biotherapies (AABB) (16) have 
recommended platelet transfusion thresholds of 80-
100x109/L, but both groups acknowledge the 
recommendations are based on expert opinion and 
little 
evidence. 
Observational 
data 
show 
no 
association between platelet count and subsequent 
bleeding, both in adults and children (17). 
Transfusion protocols are largely based on anecdotal 
institutional experience (18). There is significant 
heterogeneity in transfusion practices on ECMO 
(19). In a recent international survey of pediatric 
Figure 1: Venn diagram of the ECMO days with plasma, platelet, and 
cryoprecipitate transfusions. [Karam et al. Epidemiology of Hemostatic 
Transfusions in Children Supported by Extracorporeal Membrane 
Oxygenation. Crit Care Med 2020]
Figure 2: Cumulative percent of patients who have suffered 
mortality, bleeding events, or thrombotic events by day of 
extracorporeal membrane oxygenation support. Over the first 10 
days, 30% of patients who are not bleeding on day 1 will have a 
bleeding event, and 33% of the patients will have a thrombotic 
events. [Dalton et al. AMJCCM 2017]
 
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ECMO Hemostatic Transfusions in Children (ECSTATIC)
Study Protocol – version 1.4 – April 22, 2024
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ECMO centers, platelet transfusions were prescribed at a median (IQR) total platelet count 
threshold 72 (54-88) x109/L for non-bleeding patients (13).
Outcomes Associated with Platelet Transfusions
The decision to use prophylactic platelet transfusions as a hemostatic agent must be balanced 
with the fact that platelet transfusions are associated with significant morbidity and mortality, 
including bleeding and thrombosis. In critically ill children receiving platelet transfusions, each 
additional dose (10mL/kg) was independently associated with a 2% increase in mortality (20). The 
data from the only available randomized controlled trial (RCT) focused on pediatric platelet 
transfusion strategies, conducted in preterm infants, showed that a high platelet threshold for 
transfusion led to a 1.5-fold increase in the odds of major bleeding or death (95%CI 1.06-2.32) 
(21). 
In observational studies, adjusting for minimum lactate within 48 hours of ECMO initiation, ECPR, 
renal failure, new neurologic event on ECMO, preterm neonatal age, and a diagnosis of congenital 
diaphragmatic hernia, platelet transfusions have been independently associated with increased 
bleeding, thrombosis and mortality in children supported by ECMO (9). The association between 
platelet transfusion and both 90-day and 1-year mortality has also been confirmed in adults on 
VA ECMO (22).
Mechanisms of Poor Outcomes
The mechanisms of the observed morbidities and mortalities associated with platelet transfusions 
are poorly understood. Animal models of thrombocytopenia have suggested that it is not the low 
platelet count itself that increases the risk of bleeding, but rather the low platelet count in an 
inflamed environment (23,24). Platelet transfusions have been noted to be pro-inflammatory 
through a variety of mechanisms. Platelets themselves can synthesize inflammatory mediators 
and release them through interactions with endothelial cells and immune cells (25). Platelets 
release biological response modifiers during storage that include oxygenated moieties of 
membrane lipids (26), microparticles (27), serotonin, histamine and ADP/ATP (28), all of which 
can induce further inflammation upon transfusion. It is therefore plausible, in an attempt to correct 
thrombocytopenia, the platelet transfusion itself actually worsens inflammation and can lead, 
counterintuitively, to increased bleeding and thrombosis. 
Summary
There clearly remain knowledge gaps in both the optimal strategy to transfuse platelets to prevent 
bleeding in children supported by ECMO, as well as mechanisms to target to prevent the 
associated morbidities. Studies investigating these questions are urgently needed (29,30) to test 
the efficacy of such strategies. If the proposed aims of this pilot study are achieved, we will be 
able to undertake a full trial which will provide a better understanding of how to transfuse platelets 
prophylactically to children supported by ECMO. This contribution will be significant because it 
ultimately will influence transfusion medicine guidelines and reduce the morbidity associated with 
these therapies.
 
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ECMO Hemostatic Transfusions in Children (ECSTATIC)
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B. Preliminary Studies
We conducted a large, point prevalence study of platelet transfusions in critically ill children (20). 
Upon adjusted analysis, total administered platelet dose was independently associated with 
increased ICU mortality (odds ratio for each additional 1 mL/kg platelets transfused 1.002; 95% 
CI 1.001-1.003; p=0.005). We examined only those children within the cohort who were supported 
by ECMO (13); seventy-nine percent (71/90) of the platelet transfusions were given for 
prophylaxis in non-bleeding patients. The median (IQR) total platelet count prior to transfusion 
was 72 (54-88) x109/L and did not vary based on bleeding versus non-bleeding indications 
(according to local assessment). Through the course of their admission, children supported by 
ECMO received a total median (IQR) dose of 92 (42-239) mL/kg of platelet transfusions. 
Institutional protocols varied.
We also examined the use of platelet transfusions in children supported by ECMO in the Bleeding 
and Thrombosis on ECMO (BATE) dataset (12), and showed that platelets were transfused on 
68% of ECMO days. Platelet transfusion dose was independently associated with chest-tube 
output (p<0.001), other bleeding requiring red blood cell transfusion as defined by the authors 
(p=0.03), and daily set platelet goal (p=0.009), but not with total platelet count (p=0.75). These 
results mirrored other published associations for children supported by ECMO (9) and further 
justified our proposed RCT.
C. Justification for the study
Justifications to undertake a randomized controlled trial (RCT) to address platelet transfusion in 
children on ECMO include the following:
•
Many potential confounders that preclude from inferring causation from observational studies, 
thus only an RCT can evaluate the outcomes associated with platelet transfusion strategies.
•
Pediatric ECMO is associated with an in-hospital mortality rate of approximately 50% - the 
highest mortality associated with any standard of care pediatric treatment (1). Since platelet 
transfusions are independently associated with an increased morbidity and mortality, in both 
critically ill adults (31) and children (13), a low platelet threshold transfusion strategy could 
improve outcomes. 
•
In addition, the currently available observational data suggest that platelet transfusions are 
associated with an increased risk of both bleeding and thrombosis (9) which may be related 
to the inflammation induced by platelet transfusions. 
•
Despite the large volume of platelet transfusions that they receive, the current platelet 
transfusion recommendations for children supported by ECMO are based solely on expert 
opinion (1). 
•
There is a huge knowledge gap (29,30) and no high-quality evidence to guide platelet 
transfusion practice in these children supported by ECMO (15).
•
There is clearly equipoise with respect to platelet transfusion in pediatric ECMO patients (29).
In summary, platelet transfusions are widely used in children on ECMO (12) with little evidence 
to guide practitioners. There is no evidence that platelet transfusions are associated with 
improved outcomes; some data even suggest that platelet transfusion may be harmful (9). There 
is a need for a greater understanding of appropriate platelet transfusion strategies in these 
vulnerable children.
 
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ECMO Hemostatic Transfusions in Children (ECSTATIC)
Study Protocol – version 1.4 – April 22, 2024
14
D. Hypotheses and Aims
This protocol pertains to the pilot clinical trial, which will evaluate the feasibility of randomizing a 
LOW platelet threshold transfusion strategy, in which a platelet count of < 50x109/L will trigger a 
platelet transfusion, compared to a HIGH platelet threshold transfusion strategy, in which a 
platelet count of < 90x109/L will trigger a platelet transfusion (Figure 3). We hypothesize that, in 
non-bleeding children on ECMO, a low prophylactic platelet threshold transfusion strategy is non-
inferior to a high platelet threshold transfusion strategy, specifically related to bleeding or 
thrombotic complications, and this hypothesis is intended as a candidate primary hypothesis of 
the full RCT. 
The objective of this pilot trial is to determine the 
feasibility of the full RCT through the following:
•
Aim 1 is to demonstrate a separation between the 
LOWER 
and 
HIGHER 
platelet 
threshold 
transfusion arms. Specifically, we will demonstrate 
a difference in the average pre-transfusion platelet 
count between the two arms from randomization to 
when the treatment strategy was permanently 
discontinued. In addition, we will estimate the 
difference in average daily platelet dose during the 
same period, which we anticipate will be a difference 
of at least 20% between the groups randomized to 
each threshold.
•
Aim 2 is to demonstrate the ability for observed 
endpoints 
to 
satisfy 
various 
prespecified 
feasibility thresholds. Specifically, there are 10 
different criteria, and we will show that at least 5 of 
the 10 thresholds are satisfied by the estimated 
endpoints. 
•
Aim 3 is to provide additional information on the epidemiology of a novel composite outcome 
of progression to severe bleeding and/or severe thrombosis, to incorporate in the design of 
the full RCT. Specifically, we will demonstrate that the estimated rate of adjudication by an 
independent panel is more than 90% of the enrolled subjects for whom the outcome was 
reported by the site as having occurred.
We anticipate that if this pilot study should demonstrate feasibility of the larger study, the 
information gathered in the larger trial will provide the rationale for an evidence-based and rational 
strategy for platelet transfusions in pediatric ECMO patients, with the hope of reducing 
transfusions, as well as preventing additional complications. 
E. Innovation 
Our proposal is innovative in numerous ways:
•
Focus on children supported by ECMO. Children supported by ECMO receive a 
disproportionately large number of transfusions and these cumulative transfusions have been 
associated with mortality, bleeding and thrombosis. The R34 pilot trial will be the first to assess 
Figure 3: Design of the pilot RCT. In addition to the 
listed outcomes, we will also collect epidemiological 
information on progression to severe bleeding and/or 
clotting, and we will further assess safety by considering
mortality and adverse events.
 
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feasibility for consenting and enrolling subjects into a multicenter RCT of children on ECMO. 
The ability to enroll after regular informed consent will be relevant for any future ECMO trial.
•
The pragmatic design will allow the results to be applicable across a variety of ECMO centers 
and relevant for any ECMO trial.
•
Large separation in transfusion strategies. To ensure the outcomes can be caused by the 
intervention, it is important that the two interventions are separated. To date, limited trials of 
platelet transfusion threshold strategies in pediatrics have compared 50 to 25 x109 cell/L (21) 
in premature neonates and in adolescents and adults receiving chemotherapy have compared 
20 to 10 x109 cell/L (32). The sites participating in this pilot have all agreed to randomize to a 
much wider separation in transfusion strategies between arms (90 vs 50 x 109 cell/L). These 
thresholds are based on the 25th and 75th percentile of real-life transfusion thresholds (13), to 
ensure equipoise similar to other transfusion studies (33).
•
Novel approach to associated outcomes. Platelet transfusions are routinely prescribed to 
prevent bleeding. However, platelet transfusions, due to their inflammatory nature, may 
actually worsen both bleeding and thrombosis, based on the results of the RCT in neonates 
and the observations in children on ECMO. We propose a novel composite outcome that will 
address both bleeding and thrombosis.
6.
Patient Eligibility
A. Anticipated Cohort
Our aim is to enroll consecutive non-bleeding or minimally bleeding critically ill children on ECMO, 
who are at risk of progression to severe bleeding and/or to severe thrombosis, and at risk of 
receiving platelet transfusions. 
Among the children on ECMO who are not bleeding on Day 1, 30% will be bleeding by Day 10 
(Figure 2, page 11) (6). Furthermore, by day 10, 33% of patients will have experienced a 
thrombotic complication. Finally, 66% of those patients will receive at least one platelet transfusion 
on any given day on ECMO. 
B. Criteria to Identify Participating Sites
The following criteria were used to identify participating sites in the pilot study: 
1) Offering a pediatric ECMO program
2) Unanimous agreement to participate from surgical and medical practitioners (NICU, 
PICU, and/or CICU) and ECMO program leadership
3) Unanimous agreement to follow the PEACE anticoagulation, transfusion and 
hemostatic agent replacement guidelines
4) Well-established track record of excellence in clinical trials participation. 
For this pilot study, there is no need for our results to be generalizable to non-study PICU groups, 
in terms of clinical trial experience and geographic and demographic scope. However, we will cap 
the number of enrolled patients per center to 10, to avoid over-representation of large centers. 
 
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In addition, as the future large RCT may require participation of international sites, we will 
include one international site in the pilot to provide some evidence on the feasibility of the trial 
outside of the US. 
The ten participating sites are (alphabetical order): Children’s Healthcare of Atlanta – Emory, 
Atlanta, GA; Children’s Hospital of Richmond, Richmond, VA; Children’s Hospital of Wisconsin, 
Milwaukee, WI; Duke University School of Medicine, Durham, NC; Golisano Children's Hospital, 
Rochester, NY; Norton Children’s Hospital, Louisville, KY; Komansky Children’s Hospital of New 
York Presbyterian, New York, NY; Morgan Stanley Children’s Hospital of New York Presbyterian, 
New York, NY; Schneider Children’s Medical Center, Petach Tikvah, Israel; and University of Iowa 
Health Care, Iowa City, IA.
C. Eligibility
Children will be eligible if admitted to a participating tertiary care PICU, NICU or CICU for whom 
the decision to cannulate for ECMO support has been made.
D. Inclusion Criteria
Critically ill children (0 to <18 years of age), admitted to a participating PICU/NICU/CICU, on 
ECMO, and who have either no bleeding or minimal bleeding, according to the BASIC definition 
(appendix B) (34) within 24 hours of cannulation (any of these criteria define minimal bleeding):
•
streaks of blood in endotracheal tube or during suctioning only
•
streaks of blood in nasogastric tube
•
macroscopic hematuria
•
subcutaneous bleeding (including hematoma and petechiae) < 5 cm in diameter
•
quantifiable bleeding < 1 mL/kg/hr (e.g., chest tube). Since the objective is to capture 
bleeding, chest tube output can be higher than 1 mL/kg/hr, provided the output is 
serosanguinous and the site investigator judges the bleeding portion of the chest tube 
output is < 1 mL/kg/hr.
•
bloody dressings required to be changed no more often than every 6hr, or weighing no 
more than 1 mL/kg/hr if weighed, due to slow saturation
This definition, which was developed by a large consortium of experts, including ECMO 
specialists, has been validated with substantial inter-rater reliability: the free marginal kappa 
between two observers is 0.74 (95% CI, 0.57–0.91) (34).
If a patient experiences more than minimal bleeding after ECMO cannulation, they can be 
enrolled once bleeding meets criteria for minimal bleeding, with the condition that the bleeding 
resolves within the first 24 hours of ECMO initiation. 
E. Exclusion Criteria
To ensure our trial is as pragmatic as possible, we have as few exclusion criteria as possible: 
1) post conception age < 37 weeks at time of screening
2) underlying oncologic diagnosis (defined as receipt of chemotherapy or radiation in the last 
six months) or recipient of bone marrow transplant in the last year
 
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3) congenital bleeding disorder or congenital thrombocytopenia
4) pregnant or admitted post-partum (within 6 weeks of giving birth)
5) decision to withdraw or withhold some critical care or interventions
6) known objection to blood transfusions
7) on ECMO for > 24 hours at time of enrollment
Inclusion criteria and exclusion criteria will be ascertained by research staff in conjunction with 
the pediatric critical care clinical team. While we recognize that neonates (< 28 days of life) have 
distinctly different coagulation systems as compared to older children (35), they make up at least 
half of all children supported by ECMO (6) and therefore, strategies for prophylactic platelet 
transfusion must be understood in this group as well. Randomization will be stratified by age (see 
below).
7.
Consent
A. Informed Consent
All critically ill children cannulated for 
ECMO in a participating center will be 
screened on a 24 hour and 7 days a 
week basis. The ECMO team will be 
educated on this trial and will call the 
study team for each patient who is 
being approached for consent for 
ECMO or who is on ECMO at the time 
of PICU admission. To detect any 
potential 
biases, 
a 
de-identified 
screening log will be maintained at 
each site to record the number of 
eligible 
patients, 
the 
number 
of 
patients eligible for consent not 
randomized, the number of platelet 
transfusions between cannulation and 
consenting process (and if those 
transfusions were prophylactic or 
therapeutic), if the patient experienced 
bleeding between cannulation and 
enrollment, and the reason for their 
exclusion. This should allow detection 
of any selection bias. 
When the patient is expected to 
require ECMO within the next few 
hours, we will attempt to obtain consent 
prior to ECMO cannulation. However, 
we recognize that some parents might not able to make an informed choice in a situation where 
Figure 4: Inclusion, exclusion, and consenting process.
 
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they fear for their child’s life and where they might feel coerced (36–39).Therefore, we will 
approach the parents for consent as soon as possible, but at least within 24 hours of cannulation. 
The screening, inclusion and exclusion, and consent process is shown in Figure 4. 
Of note, for patients who will turn 18 years of age before the 28- and/or 90-day follow up call, we 
will provide the parents with a separate consent form for follow-up phone calls, at the time of ICU 
discharge, as well as a pre-stamped envelope. Once the patient turns 18 years of age, we will 
ask him to sign the consent form to allow us to be recontacted and mail it back to the site PI. The 
site PI or coordinator will call the patient before the 28- or 90-day evaluation, and remind them to 
mail the consent form. Once the research team has received the consent form, they will set up 
the 28- and/or 90-day follow-up evaluation call.
8.
Study Design
A. Intervention
Subjects will be randomized in a 1:1 ratio to either a lower or a higher platelet prophylactic 
transfusion strategy (Figure 3, page 14). All platelet units will be prepared in accordance to 
existing national standards (40). 
We chose platelet count as the basis for transfusion since 100% of sites in our study rely on this 
number, rather than platelet function or viscoelastic testing, to make transfusion decisions [9]. 
Platelet counts will be assayed at least twice daily.
To ensure clinical equipoise, we chose platelet transfusion thresholds that are based on 
observational data, similar to the thresholds chosen for red blood cell transfusion studies (33). 
The 25th and 75th percentile of real-life transfusion thresholds were used as a basis for the trial's 
thresholds to avoid extremes and promote equipoise.
In our original epidemiologic study from 2017-2018, among non-bleeding children on ECMO, the 
25th percentile of platelet count before transfusion was 54 x109 cell/L, whereas the 75th percentile 
was 88 x109 cell/L (13). 
In addition, we performed two additional studies to assess clinical equipoise: an analysis of 
contemporary data from the Recipient Epidemiology and Donor Evaluation Study (REDS) IV-P 
and a survey of ECMO providers at the participating sites. 
 
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We conducted a retrospective multicenter study from four academic children's hospitals, 
participating in the REDS-IV-P study between April 2019 and July 2022. All children (0 to 18 years 
old) supported with ECMO and transfused platelets 
were included. Transfusion doses were categorized 
as lower (≤ 75th percentile, i.e. ≤ 24 mL/kg) and 
higher doses (> 75th percentile, i.e. > 24 mL/kg), as 
a surrogate marker for bleeding status. Three 
hundred ten children, median age 6.4 months (IQR 
25 days-5.4 years) and median body weight 6.6 kg 
(IQR 3.7-18.0) on ECMO received 3437 platelet 
transfusions. Of these, 2578 were lower doses and 
859 were higher doses. Among the former, the 
median pre-transfusion platelet count was 72 x 109/L 
(IQR 56-94) in neonates (<28 days) and 71 x 109/L 
(IQR 48-102) in older infants and children. Among 
the latter, the median pre-transfusion platelet 
count was 113 x 109/L (IQR 72-166) in neonates 
and 112 x 109/L (IQR 66-177) in older infants and 
children (Figure 5). The median time between the platelet count and the platelet transfusion was 
2.6 hours (IQR 1.3-4.7). There was no association between the patient’s age and pre-transfusion 
platelet count (p=0.86), but there was a significant difference between the four sites (p=0.01).
We 
surveyed 
ECMO 
providers 
at 
all 
the 
participating 
sites. 
The 
overall response rate was 
56% 
(114/204). 
66% 
(68/103) of the responders 
were pediatric intensivists 
while 37% (38/103) were 
pediatric 
cardiac 
intensivists. 26% (27/103) 
had 10 to 14 years of 
experience, 
while 
45% 
(46/103) had more than 15 
years of experience.
 
When looking at the overall 
platelet 
transfusion 
thresholds, the median was 
70 (IQR 50-99). There were 
differences 
between 
the 
various 
scenarios. 
The 
median platelet threshold 
was 50 for non-bleeding 
children, 
70 
for 
non-
bleeding neonates, 75 for 
minimally bleeding children, 
Figure 5. Pretransfusion platelet thresholds from REDS-IVP
Figure 6. Platelet transfusion thresholds from survey of participating centers. There are 
six different scenarios, and each site’s responses are displayed, with the median (circle) 
and the interquartile range (line). The averaged responses per scenario are summarized 
by the diamonds and bold lines. The pink zone indicates the ECSTASTIC transfusion 
thresholds.
 
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75 for minimally bleeding neonates, 75 for resolved bleeding in children, and 80 for resolved 
bleeding in neonates. As shown in Figure 6, there was a wide heterogeneity among sites as well 
as within sites: each line representing the median (circle) and IQR (line) of one site. 59% (61/103) 
of the responders were uncertain or very uncertain about the level of evidence, 33% (34/103) 
were neutral, and 8% (8/103) were certain or very certain.
Given the evidence presented above from three studies, we will use transfusion thresholds of 90 
(higher platelet threshold arm) vs 50 x109/L (lower platelet threshold arm).
Patients should not be transfused above the upper transfusion thresholds, i.e., patients should 
not be transfused for a platelet count ≥ 90 x109 cell/L in the higher platelet threshold arm and 
≥ 50 x109 cell/L in the lower platelet threshold arm. Patients must be transfused below the 
transfusion thresholds, i.e., patients should be transfused for a platelet count < 90 x109 cell/L in 
the higher platelet threshold arm and < 50 x109 cell/L in the lower platelet threshold arm. The 
transfusion should occur within 12 hours of the sample collection time for the platelet count 
laboratory test. If a platelet count in either direction of the threshold has a platelet count within the 
next 12 hours in the opposite direction of the threshold (e.g., went from below threshold to not 
below threshold) without an intermediate platelet transfusion, the earlier platelet count’s implied 
action is superseded by the most recent platelet count to allow sites the flexibility to obtain a 
confirmatory platelet count in instances where the earlier count is considered unreliable. Specific 
criteria for temporary suspensions are detailed below. 
The transfusion dose is intended to be 10-20 mL/kg of apheresis platelets (whole blood-
derived platelets may be used if apheresis platelets are not available), which is the median platelet 
dose reported in two large observational studies (6,20), though adjustments to the dosing to be 
in line with local implementations are permitted so long as dosing is not differentially applied 
between assigned transfusion strategies. Though a range of dosing is provided, all doses given 
at each site will be the same across both arms of the study and will be in line with local guidelines. 
For patients more than 20-30 kg (per site guidelines), one unit of platelets will be transfused. 
For the sites in which the information is available (currently 5 out of the 10 participating sites), the 
total platelet count (per mL of transfusate) that each patient receives from each transfusion will 
be recorded. For sites in whom this exact information is not known, we will record if they receive 
“low-yield” platelet transfusions in which the platelet count is < 3 x 1011 platelets. In addition, we 
will collect information about irradiation, pathogen reduction and ABO compatibility of the platelet 
product.
A platelet count must be drawn two to four hours after the end of the platelet transfusion. 
Another platelet transfusion should be given if the platelet count drops again under the 
threshold of the RCT arm to which the participant was allocated.
B. Safety
Although ELSO guidelines suggest keeping the platelet count > 100 x109 cell/L in neonates (vs 
80 in older children) (41), there is currently no evidence to suggest neonates are at higher risk of 
bleeding than older children, at a similar platelet count. 
 
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Additional analyses from 514 pediatric ECMO 
runs, comparing bleeding in neonates versus 
infants at particular platelet counts, as shown in 
Figure 7, are reassuring (unpublished analysis, 
from the BATE dataset (6)). The proportion of 
children experiencing bleeding is stable 
throughout the range of platelet counts, 
which suggests that lower platelet counts are not 
associated 
with 
increased 
proportion 
of 
bleeding. Furthermore, neonates seem to have 
an overall lower proportion of bleeding than 
older children.
In addition, thrombocytopenia was not a risk factor for bleeding on ECMO, either from chest tubes 
or other sites (p=0.82) (7). 
C. Co-Interventions
As this is an open-label trial, we need to ensure co-interventions that may be associated to the 
primary clinical outcome in the larger trial (progression to severe bleeding, severe thrombosis or 
both) are managed identically in both arms, to avoid potential interactions and biases (e.g., 
providers managing patients in the low platelet threshold arm might be more prone to prescribe a 
lower dose of heparin). However, as the pragmatic design is important for the generalizability 
of the future results, independently of co-interventions, and as there is variability in the 
anticoagulation (42) and plasma transfusion strategies on ECMO (13), Therefore, we will be 
imposing the adherence to the new international guidelines (Pediatric ECMO Anticoagulation 
Initiative, to be published in 2024).
In addition, all participating sites will have agreed to follow their own local guidelines:
1. Plasma transfusions (which coagulation test is to be measured, how frequently is it 
measured, what is the threshold for plasma transfusion, what is the dose of plasma) 
2. Anticoagulation (what is the heparin bolus and starting dose, what is the primary test 
used to adapt heparin, what values will prompt a change in heparin and by how much, 
when to recheck the anticoagulation test)
3. Cryoprecipitate or fibrinogen concentrate (when to measure fibrinogen level or 
perform viscoelastic testing, what level triggers administration of cryoprecipitate or 
fibrinogen concentrate, what is the dose, and when to recheck the fibrinogen level)
4. Antithrombin concentrate (when to measure antithrombin level, what level triggers 
administration of antithrombin concentrate or plasma transfusion, what is the dose, and 
when to recheck the antithrombin level)
5. Priming of circuit (with plasma, saline, platelets, standard RBC units and/or whole 
blood)
Data on these co-interventions will be collected daily and compliance with the local 
guidelines will be measured, so that the effect of any imbalances on the primary outcome 
can be examined after the trial is complete.
Other co-interventions, such as ECMO flow management, vasopressors, ventilation, red blood 
cell transfusion, sedation, and the use of hypothermia, will not be protocolized, due to the 
Figure 7: Proportion of bleeding according to morning platelet count, 
by age (neonates versus older infants and children) [unpublished 
analysis from Dalton et al, AJRCCM 2017]
 
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pragmatic nature of the trial. In addition, the use of other hemostatic agents, such as tranexamic 
acid, aminocaproic acid, DDAVP, and von Willebrand factor will not be protocolized as these are 
typically used in patients with active bleeding and not to prevent bleeding (7). However, all these 
co-interventions will be recorded.
D. Compliance Measures
Patients in the lower platelet threshold arm of the study will be considered adherent to protocol if 
90% or more of pre-transfusion platelet counts are < 50 x109 cell/L, and likewise patients in the 
higher platelet threshold arm of the study will be considered adherent to protocol if 90% or more 
of pre-transfusion platelet counts are < 90 x109 cell/L. 
Should a center have more than 10% of their patients with non-compliant transfusions, we will 
work with the primary investigator to understand the barriers to following the trial’s protocol, and 
may provide additional measures, such as team education. 
In addition to the aforementioned compliance measures, the sixth feasibility criterion pertains to 
a combined metric of transfusion protocol compliance that considers platelet counts and 
transfusions. Details of this combined metric are provided in Appendix F.
Measures to Ensure Compliance
Trial conduct depends upon administration of platelet transfusions according to the appropriate 
transfusion threshold to patients in the two study arms. Study procedures will ensure timeliness 
and accuracy for platelet transfusion prescriptions. Such measures include: 
1) recruitment and engagement of ECMO directors as co-site-investigators, as well as 
inclusion of ECMO expertise on all important trial committees
2) development of institute-specific protocols
3) in-person and/or virtual meetings prior to initiation of the trial at every participating site
4) giving lectures and/or grand rounds to all participating centers
5) twice daily monitoring of the platelet levels of all patients on ECMO
6) Poster at the patient’s headboard with the platelet transfusion threshold
In similar trials, the adherence was 97% in the PlaNet-2 trial (21), whereas it was 92% in a low 
platelet threshold transfusion trial in oncology patients (PLADO trial) (32). Whereas neither of 
these studies were in ECMO patients, they occurred in the critically ill who were at high risk for 
bleeding. As such, they demonstrated clinical equipoise for randomization to two separate platelet 
transfusion thresholds. 
Given that ECMO programs protocolize nearly every aspect of a patient’s care, we believe ECMO 
centers are the ideal environment to conduct the study. 
E. Randomization
An independent biostatistician will generate the randomization scheme based upon instructions 
from the study statistician. The randomization process will consist of a computer-generated 
random listing of treatment allocation using a pre-established algorithm. 
 
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Local study teams will contact the REDCap randomization module when patients are eligible for 
enrollment. Patient characteristics that establish eligibility will be confirmed. 
Time zero will be the time that the patient is randomized to one arm of the study within the web 
randomization system. A site-specific back-up randomization envelope will be available for one 
use per site in the event that one is emergently needed. Only the study statisticians and designate 
at the Data Coordinating Center (DCC) will have knowledge of the randomization codes.
Allocation will be a 1:1 ratio. 
Subjects will be stratified by age (≤28 days vs >28 days) because it is infeasible to stratify by 
more than one variable given the sample size. We prioritized this variable since most of the safety 
concerns are around lower transfusion strategies in infants. Other variables considered for 
stratification in the future large trial will be type of ECMO support (V-A vs V-V) and site. 
Patients will not be stratified based on the severity of the disease, as all scores predicting outcome 
on ECMO use data from the first few days of the ECMO run (43–45). In addition, scores that 
evaluate severity at admission (PRISM III and PIM2) are not validated nor calibrated in ECMO 
patients (46,47). In the pilot, we will neither stratify according to the indication (medical vs post-
surgical), as this would significantly increase the sample size. However, this may be explored in 
regression models and sensitivity analyses. 
Research personnel at each site and PICU caregivers will not have access to the randomization 
schedule. Moreover, in order to conceal randomization, 3 sizes of blocks permutation will be 
used on a random sequence: 2, 4 and 6 subjects per block.
F. Blinding
This is an open-label trial, since the transfusion strategy cannot be blinded from the clinical 
team, as they must order the transfusions based on the platelet count. The research team 
(Marianne Nellis, Oliver Karam, and members of the steering committee) will be blinded to 
treatment allocation until the end of the analysis. Since the statisticians will need to report to the 
Data Safety Monitoring Board, they will not be blinded.
G. Treatment Period and Follow Up
The intervention will be implemented immediately upon randomization and maintained in effect 
until four possible events (whichever occurs first): 
1) progression to severe bleeding, according to the BASIC definition, and/or severe 
clotting, according to the NHLBI consensus definition, outside of temporary 
suspensions
2) decannulation from ECMO (or ECMO flow stopped, even if cannulas left in place)
3) twenty-one days post-randomization (as the progression to new bleeding and new 
thrombosis has plateaued by day 21 and 95% of the patients will be off ECMO by 
then (6))
4) or temporary suspension lasting more than 24 hours. 
5) Withdrawal from the study, either by the attending physician or the parents.
 
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The mortality follow-up will be at 28 (+/- 4 days) and 90 (+/- 10) days after randomization.
H. Temporary suspension of Individual Patients
The protocol will allow for temporary suspensions, i.e., transfusion at a higher platelet count 
threshold than allowed for the randomization arm, if the clinical situation warrants immediate 
transfusion for certain conditions including: 1) chest-tube insertion; 2) surgical intervention while 
on ECMO (such as tracheostomy or repair of congenital diaphragmatic hernia); 3) during ECMO 
circuit change; or 4) in preparation for decannulation. In addition, while patients will be monitored 
for severe bleeding and/or severe clotting events during this time, they will not have the 
transfusion strategy permanently discontinued if they develop such an event during the temporary 
suspension, provided that the severe bleeding/clotting has resolved within the 24 hours of the 
temporary suspension. If a patient continues to have severe bleeding or severe clotting past the 
24 hours of the temporary suspension, the intervention is stopped.
Adherence to the trial protocol must be resumed as soon as possible once these events are 
controlled or completed. Data monitoring and collection will be unchanged during suspension 
(including pre-transfusion platelet counts and platelet transfusion volumes), but data on length 
and justification of suspension will also be collected. Suspensions less than 24 hours will not be 
considered a breach of adherence to the protocol. After 24 hours of temporary suspension, the 
patient will have all assigned treatment strategy implications lifted (i.e., no restrictions on the 
range at which any subsequent platelet transfusions may occur) but will continue to be followed 
for ascertainment of safety and study outcomes. Temporary suspensions will only necessitate 
exclusion from analyses for one of the secondary endpoints (#6: proportion of compliant platelet 
transfusion actions), as explicitly noted in Section 9.B. The safety outcomes will still be followed 
but will be analyzed as a sub-group (e.g., severe bleeding during temporary suspensions).
9.
Outcomes
The objective for the pilot study will be to assess the feasibility of the full RCT. 
A. Primary Outcome
The primary objective is to assess the separation between the higher and lower platelet threshold 
arms. Indeed, for a transfusion trial to be successful, there has to be a clinically meaningful 
separation between the transfusion strategies, to be able to imply causality for the outcome. Since 
the higher platelet threshold is <90 x109/L and the lower platelet threshold is < 50x109/L, we 
expect the separation of the pre-transfusion platelet count (primary outcome) to be at least 
30 x109/L. To assess the separation between both arms, we will collect the pre-transfusion 
platelet count before each transfusion event. 
In addition, we will collect the total daily platelet transfusion dose for a patient while the patient is 
on ECMO. The average daily dose (in ml/kg/day) will be computed by the research team, by 
dividing the total platelet transfusion volume by the patient’s weight at admission and the number 
of days on ECMO. 
 
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These outcomes will be measured outside of temporary suspensions; that is, transfusions 
occurring during a temporary suspension are ignored from these analyses.
The primary safety outcome, which is assessed in conjunction with the early trial stopping rule, is 
occurrence of any of the following from the time of randomization (enrollment) to 24 hours after 
the transfusion strategy threshold was permanently discontinued: progression from no bleeding 
or minimal bleeding to severe bleeding, according to the BASIC definition (34); severe thrombotic 
event, according to the NHLBI Hemostasis Clinical Trial Outcomes Working Group (need for 
circuit change, need for thrombectomy, ischemic stroke, or distal thromboembolism, as diagnosed 
clinically) (48); death (i.e., all-cause mortality). The primary safety outcome ignores severe 
bleeding/severe clotting that begins during a period of temporary suspension, but does not ignore 
deaths that occur during a period of temporary suspension.
The adjudication committee will evaluate patients who are deemed by site investigators to meet 
this endpoint (severe bleeding or clotting) in batches of 5 patients. The adjudicators will rotate, 
which will help prevent bias and ensure that all adjudicators have an equal opportunity to 
contribute to the study. We will report the inter-rater reliability (Kappa) at the end of the study. The 
committee will make determinations for whether bleeding was the primary cause of the worsening 
clinical outcome or if clotting was severe. Specifically, the sites will complete the severe 
bleeding/clotting daily worksheet (see Appendix E) on a daily basis. If a severe bleeding event, 
or clotting event, the event will be recorded in the database (and the intervention will stop for that 
participant since this safety outcome has been reached). Once five new patients have any of 
these events reported, the adjudication committee will review the events and report if they agree 
with the determination of the outcome within 72 hours of their review or if they need additional 
information from the site PI. Patient enrollment will continue during their reviews.
B. Secondary Outcomes
Our second objective is to demonstrate the ability for observed endpoints to satisfy various 
prespecified feasibility thresholds.   The information the pilot trial will provide is both necessary 
and sufficient for the implementation of the full clinical trial. Based on PALISI and BloodNet’s 
suggestions, we will seek further funding and progress to the full trial if at least 5 of the following 
10 criteria are met:
1.
Estimated separation in mean pre-transfusion platelet count is > 30 x109 cell/L;
2.
Eligibility rate is > 50% of the predicted eligibility rate of 7.9 patients per site per year;
3.
Proportion of patients whose parents were approached for consent within 24 hours of 
cannulation (number of patients approached / number of patients eligible for consent) 
> 90%;
4.
Proportion of patients for whom we were able to obtain informed consent (number of 
patients consented / number of patients approached) > 50%;
5.
Randomization of consented patients is > 66%;
6.
Proportion of compliant platelet transfusion actions (examples of noncompliance are 
transfusion given at or above threshold or without any platelet count in the previous 
12 hours, excluding transfusions given during a temporary suspension, or transfusion 
not given within 12 hours of a platelet count below threshold) > 90% (see Appendix F 
for details);
7.
Proportion of patients with one or more temporary suspensions not due to preparation 
for decannulation < 10%;
 
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8.
Proportion of patients who withdraw from the study and/or are lost to follow-up (i.e., 
withdraw and/or are missing 90-day mortality assessment) < 6%;
9.
Proportion of patients with non-platelet transfusion protocol violations (violation of site-
specific management protocol for any aspect besides platelet transfusion threshold 
compliance) < 5%;
10. First instance of progression to severe bleeding or thrombotic outcomes can be 
adjudicated in > 90% of the patients for whom such an outcome was reported by the 
site.
Similar to pre-transfusion platelet counts, overall platelet transfusion doses, and progression to 
severe bleeding and/or clotting, feasibility outcomes 1, 6, and 9, are to be analyzed outside of 
temporary suspensions.
C. Tertiary Outcomes
Our tertiary outcome is to demonstrate the ability for an adjudication committee to determine the 
severity of bleeding and/or thrombotic outcomes. 
This is similar to the primary safety outcome, except that it does not include mortality in the 
definition. That is, progression to severe bleeding or a severe clotting event must occur, whether 
or not there is aa all-cause death. We will demonstrate that the recorded data is sufficient for the 
adjudication of severe bleeding, severe thrombus or both in 90% of enrolled subjects with this 
outcome reported by the site.
Justification of BASIC and NHLBI definitions of bleeding and thrombosis: 
We believe that the BASIC (34) (Table 1) and NHLBI definitions (48) of bleeding and thrombosis 
provide the most relevant descriptions as they relate to children supported by ECMO. Importantly, 
central nervous system involvement, a frequent site of bleeding on ECMO, may be captured by a 
change in the neurologic variables of the PELOD-2 
score (both the Glasgow Coma Scale and the 
pupillary reactions are in the PELOD-2 score) 
which is incorporated in the BASIC definition. In 
order to ensure that neurologic variables are not 
missed (since the majority of patients will be deeply 
sedated and paralyzed), we will also consider any 
new and/or expanding intracranial hemorrhage as 
severe bleeding and collect information regarding 
the location of the bleeding.
Importantly, several of the organ dysfunction 
criteria of the PELOD-2 score may be normalized 
by the ECMO circuit (blood pressure, PaO2/FiO2, 
pCO2), which might lead to an underestimation of 
the organ failure. Severe bleeding is likely to cause 
a change in these variables (such as hypotension 
or worsening respiratory function), which might be 
mitigated by changes in ECMO support. Therefore, 
we will not only record the PELOD-2 variables but 
also the potential increased ECMO support. Although we recognize some limitations to the 
PELOD-2 score on ECMO, similar scores, such as the adult SOFA score, have been used to 
Table 1: Severe Bleeding Definition [27] (at least 
one of the following criteria)
1) bleeding that leads to at least one organ 
dysfunction, using PELOD-2 score criteria of 
organ dysfunction
2) bleeding that leads to hemodynamic instability, 
defined as an increase in heart rate by > 20% from 
baseline or a decrease in mean arterial pressure 
by > 20% from baseline (i.e., prior to bleeding 
event)
3) bleeding leading to a drop in hemoglobin > 20% 
within 24 hr
4) quantifiable bleeding ≥ 5 mL/kg/hr for ≥ 1hr 
(e.g., chest tube)
5) intraspinal bleeding leading to loss of neurologic 
function below the lesion, nontraumatic intra-
articular bleeding leading to decreased range of 
movement, or intraocular bleeding leading to 
impaired vision
6) new and/or expanding intracranial hemorrhage
 
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report organ failure on ECMO (49,50). Therefore, in the absence of a validated score to evaluate 
organ failure on ECMO, it seems justified to use the validated PELOD-2 score contained within 
the BASIC definition. 
Of note, we will not use the ELSO definition for bleeding as the primary clinical outcome of the full 
trial, as it considers bleeding only if the patient requires RBC transfusions or interventions 
(“hemorrhagic complications requiring packed red blood cell (PRBC) or whole blood transfusion 
(>20 ml/kg/calendar day of PRBCs or >3 U PRBCs/calendar day in neonates and pediatrics and 
> 3U PRBCs/calendar day in adults) or other intervention such as surgical or endoscopic 
intervention”) (51). Indeed, this definition will lead to potential biases and increased heterogeneity, 
because of the high variability of the indications to transfuse and/or to intervene surgically (52). 
Similarly, the recently developed NHLBI bleeding definition is also tied to interventions (including 
transfusions and changes in anticoagulation) (48). However, we will measure the ELSO definition 
as a secondary outcome. 
Severe thrombotic events will be defined according to the NHLBI Consensus Conference (48) as 
circuit clot leading to circuit change, need for thrombectomy, ischemic stroke, or distal 
thromboembolism.
Bleeding and/or Thrombotic Outcome Adjudication
ECSTATIC is an open-label trial, but the bleeding and/or thrombosis outcome will be adjudicated 
by an independent central committee, blinded to the allocation group. This committee will be 
composed of three members, a pediatric cardiothoracic surgeon, a pediatric cardiac intensivist 
and a neonatologist, all of whom have significant experience caring for children on ECMO. The 
members of the adjudication committee will not be associated with any of the enrolling sites, nor 
members of the research consortiums supporting this trial. The adjudication committee will 
determine if the bleeding was the primary cause of organ dysfunction, hemodynamic instability, 
acute anemia, loss of spinal function, articular range of motion, or impaired vision, as per the 
definition of severe bleeding (34), or if clotting was severe. The members of the adjudicating 
committee will not receive any information on allocation and on the platelet counts of participants. 
Two members of the adjudicating committee will review the same information that the Data 
Coordinating Center will provide (please see Appendix E). They will determine independently if 
there is a cause-effect relationship between the bleeding episode and one or many of the 
outcomes listed above. In case of discordance between the two members, a third reviewer will be 
asked to state what they think, using the same information. 
The information that will be given to the adjudicators include: description of the bleeding or clotting 
episode (when it starts, its duration, the volume of blood loss, etc.), description of intervention 
(allocation will be kept blinded as much as possible) and of co-interventions (including procedural 
sedation, boluses of medications with hemodynamic or vasodilatory effects, initiation of CRRT, 
etc.), and description of outcomes (when the adverse event appears, what adverse events, etc.). 
The adjudication will be done in batches of 5 subjects at a time with the primary safety outcome 
(per site report), as described above. The adjudications will be made within 72 hours of the start 
of the review. Patient enrollment will not be paused during the time of their review. In addition, the 
adjudication committee will report their assessment of the outcomes immediately prior to the 
 
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DSMB meetings that are during the enrollment phase so that the most up to date safety data can 
be provided to the DSMB.
D. Other Outcomes
Although many adult studies evaluate mortality, which is a more objective outcome, there are 
many other variables that will directly influence mortality on ECMO, in addition to platelet 
transfusions (6,9). However, mortality will be one of the secondary outcomes of the full trial.
Mortality at 28 and 90 days will be assessed by phone calls to the families in the case that patients 
have been discharged at those times. 
In addition, we will collect the proportion of serious adverse events, which will include transfusion-
related reactions (including hyperkalemia, allergic reactions, transfusion-related acute lung injury, 
transfusion-associated circulatory overload and nosocomial infections), and ECMO-related 
complications not already captured in the outcomes (including seizures, need for neurosurgical 
intervention, cardiac arrhythmias, tamponade, pneumothorax, compartment syndrome, 
fasciotomy and amputation). Please see 11C for details on expected vs unexpected serious 
adverse events.
We will also collect the platelet processing of each unit received (i.e., apheresed versus whole 
blood derived, platelet number per mL of transfusate, ABO compatibility, irradiation, volume 
reduction, pathogen reduction, and storage age). For the sites in which the information is available 
(currently 5 out of the 10 participating sites), the total platelet count (per mL of transfusate) that 
each patient receives from each transfusion will be recorded. For sites in whom this exact 
information is not known, we will record if they receive “low-yield” platelet transfusions in which 
the platelet count is < 3 x 1011 platelets.
Additional data to be collected are:
•
Hemorrhagic complications according to the ELSO definition (51)
•
Thrombotic complications according to the ELSO definition (51)
•
Blood product exposure, defined as the total number of transfusions (red blood cell, plasma, 
platelets, cryoprecipitate, fibrinogen concentrate) and other hemostatic therapies (activated 
Factor VII, tranexamic acid, aminocaproic acid, DDAVP, and von Willebrand factor, etc.)
•
Re-explorations and other surgical procedures to treat bleeding
•
Duration of ECMO, defined as 90-day ECMO-free days (e.g., if a patient is decannulated after 
7 days, the 90-day ECMO-free days is 83 days; whereas if a patient dies on ECMO after 7 
days, the 90-day ECMO-free days is 0 days)
•
Length of PICU stay, defined as the 90-day PICU-free days
•
Functional outcome, defined as the discharge Pediatric Overall Performance Category 
(POPC) and Pediatric Cerebral Performance Category (PCPC) (53) which have been 
validated in infants and older children and have been used to measure outcomes in infants, 
neonates and children on ECMO (54) (Appendix C).
•
Mortality on ECMO, within 24 hours of the treatment strategy’s expiration, at 28 days, and at 
90 days.
 
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We will therefore be collecting six of the eight core outcome measures for ECMO research 
recommended by Hodgson et al (we will not be collecting the health-related quality of life and 
ability to return to work in the pilot trial) (55). Of note, neurodevelopment at one year is beyond 
the scope of the pilot trial, but will be assessed as an ancillary study of the full RCT.
E. Application of Results to the Implementation of the Full Trial
•
The information gained from the pilot will help to identify revisions to the plans for the full RCT.
•
We will optimize and document the ability to adjudicate a bleeding and/or thrombotic outcomes 
using both the BASIC and NHLBI developed definitions.
•
Eligibility numbers and enrollment rates will inform the number of centers, size of centers (high 
volume versus low volume), and location of centers (US-only versus international) required to 
accrue the required sample size for the subsequent ECSTATIC RCT.
•
We will document the ability to enroll patients through standard consent, which will allow us 
to determine if an Exception From Informed Consent (EFIC) will be needed for the larger RCT. 
This finding will also contribute to the selection of sites for the larger RCT as some countries 
have no provision for EFIC. 
•
Protocol deviations, should they occur in the pilot trial, will provide opportunities to refine key 
protocol elements, including reasons for protocol suspension, in order to maximize protocol 
adherence and minimize risk of bias in the subsequent ECSTATIC RCT.
•
We will document and classify temporary suspensions to allow for refinement in the 
subsequent RCT.
10. Sample Size and Statistical Analysis
A. Sample Size
Salient assumptions to permit power estimation include the number of observed transfusion days 
per subject, the within-subject dependence structure, the within-treatment variability, and the 
difference in means between treatments. Previous data, albeit not with an enforced low platelet 
threshold transfusion strategy, were used to inform the first three assumptions. It was assumed 
the distribution of the number of transfusion days per subject will mimic historical data (e.g., 
approximately 10%-20% will have no transfusions). It was assumed a compound symmetry 
covariance structure will adequately account for within-subject dependence; such a structure is 
induced by a mixed effects model with subject-specific random intercepts. Fitting a model with 
such a covariance structure to historical data [6] yielded an estimated autocorrelation slightly 
above 0.2; to be conservative the power estimation assumed ρ=0.3 (6). The assumed within-
treatment standard deviation, 40x109/L, resembles historical data but is anticipated to be a 
conservative estimate for the trial. 
Although the difference in platelet strategy thresholds is 40x109/L (i.e., 90 x109/L vs 50 x109/L), 
this full effect will not be realized for the mean difference because the strategies depend on 
platelet counts which are not continuously observed, and as such, by the time of measurement a 
patient may have a platelet count considerably below the threshold. Patients with a platelet count 
 
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under 50 x109/L would be transfused under either strategy. Therefore, a more realistic separation 
for the difference in means between strategies is 30 x109/L.
Assuming pre-transfusion platelet levels will be approximately normally distributed with a 
difference in means of 30 x109/L, then with 50 patients (25 per arm) enrolled there would be 
approximately 81% power to detect a difference in mean platelet levels at the time of transfusion. 
Power may be higher due to conservative assumptions.
The design of the full RCT, which will test the non-inferiority of a low platelet threshold strategy, 
will be informed by the separation in pre-transfusion platelet counts in both arms, median number 
of transfusions in each group, proportion of patients who develop the primary safety outcome 
(progression to severe bleeding and/or severe thrombotic event and/or study-related death) or 
tertiary outcome (progression to severe bleeding and/or severe thrombotic event), proportion of 
patients who require temporary suspension, proportion of patients with non-platelet transfusion 
protocol violations, and the consent rate.
B. Planned Recruitment Rate
There is a wide heterogeneity in the number of ECMO runs per center and per year. The sum of 
the eligible patients from the ten centers who have committed to participate in the pilot trial is 79 
patients per year. We therefore would require an enrollment rate of 32% to recruit all of the 
patients within 24 months [50 patients / (2 years x 79 patients per year) = 0.32]. As a result, we 
are very confident we will complete the pilot trial within 3 years.
All the participating sites have at least 10 ECMO runs per year (the larger sites have up to 50 a 
year). Therefore, we anticipate that even the smaller sites should be able to enroll 5 patients over 
18 months. However, should the enrollment be slower than expected, we will lift the cap on the 
larger sites (currently set at 10 subjects). 
C. Statistical Analysis
For the pilot study, the analysis of the primary outcome measure will be assessing the pre-
transfusion platelet count to describe the separation between the two platelet transfusion 
strategies, while accounting for intra-subject correlation. For each arm, we will estimate the mean 
pre-transfusion platelet count throughout the ECMO runs. This primary outcome will be measured 
for each transfusion and represent the latest available collected platelet count that is before the 
transfusion began. In the event that there was no preceding platelet count since a previous 
transfusion, the outcome for that transfusion will be regarded as missing. Participants who do not 
have a non-missing value for this outcome (e.g., they had no transfusions) will not be included in 
the analysis of the primary outcome. Because we anticipate intra-subject correlation when there 
are multiple transfusions for a patient, we will fit a linear model to pre-transfusion platelet counts 
with a fixed effect for assigned treatment (higher threshold vs. lower threshold). We will account 
for repeated measures by considering three within-subject covariance structures with constant 
variances: compound symmetry; the continuous analog of an autoregressive order 1 process, 
also known as CAR(1); and a hybrid of compound symmetry and CAR(1) achieved by including 
random subject effects and CAR(1) correlations in the within-subject errors. We will use restricted 
maximum likelihood (REML) for model estimation and use the model with the lowest Akaike 
Information Criterion (AIC) to select the best covariance structure. Once the covariance structure 
 
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is determined, the Kenward-Roger calculation for denominator degrees of freedom (df) will be 
applied in the F-test (equivalently, for the df in the t-test) of the treatment strategy's model 
coefficient. The test of the null hypothesis that the mean pre-transfusion platelet count is the same 
for each treatment arm vs the alternative that the higher threshold has a higher mean will be 
considered statistically significant if and only if the one-sided p-value is less than 0.025. In 
addition, a two-sided 95% confidence interval for the difference in means will be reported along 
with estimates of each treatment’s mean and the difference in means, with all inferences coming 
from the repeated measures model described above.
In addition, we will measure the effect of the days on ECMO, by providing summaries of the daily 
platelet counts for each specific ECMO day with ≥ 5 patients still on ECMO, independent of 
receiving a platelet transfusion. This will allow evaluating trends over time and a possible 
attenuation of the separation over time. 
Furthermore, we will also estimate the difference in the median platelet transfusion dose (in 
mL/kg/run) between the lower and higher platelet threshold strategy arms, and the median daily 
platelet transfusion dose (in mL/kg/day) to account for different lengths of ECMO runs. These 
outcomes will use two-sided nonparametric percentile bootstrap 95% confidence intervals, based 
on a minimum of 10,000 bootstrap replicates, for the difference in medians because by 
construction for each of these outcomes, there is only one outcome per person. As such, repeated 
measures modeling is not warranted.
Finally, we will also report the screening and inclusion rates, the proportion of patients who were 
approached within 24 hours of cannulation, the proportion of patients for whom we will be able to 
obtain informed consent, the number of platelet transfusions prior to randomization, the proportion 
of compliance with transfusion thresholds, number of temporary suspensions, and the ability to 
record data and adjudicate bleeding and/or clotting outcomes. In short, we will provide summaries 
of outcomes, with special emphasis for these summaries on the ten feasibility criteria of Aim 2.
Unless stated otherwise, the outcomes mentioned throughout the protocol are not intended to be 
tested for significance in this trial, but rather will be summarized descriptively (e.g., frequency and 
percentage for categorical variables, means and standard deviations or medians and interquartile 
ranges for continuous variables).
Subgroup analyses will be considered exploratory and will be interpreted with care given the
expected small sample sizes. For all Aim 1 outcomes, we will perform two subgroup analyses, 
one to evaluate the effect of V-V vs V-A ECMO, the other to evaluate neonates vs infants and 
older children (≤ vs. > 28 days of age). For the Aim 1 model, the subgroup analyses are intended 
to be based on reapplying the model with the same covariance structure already identified in the
overall analysis to the subset of participants from a particular subgroup (where subgroups
are defined based on ECMO type or age). However, if estimation is deemed too unwieldy
for some subsets, an alternative approach will be to use all participants in fitting one model
but include ECMO type or age as covariates in the model. For all secondary outcomes, we will 
perform two subgroup summaries, one to evaluate the effect of V-V vs V-A ECMO, the other to 
evaluate neonates vs infants and older children (≤ vs. > 28 days of age).
While the primary safety outcome will be tested for a difference in the proportion of patients in the 
safety population (i.e., those randomized, even in the very unlikely event that an unconsented 
 
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patient was randomized), other safety outcomes, including all-cause mortality and adverse event 
occurrences, will be summarized, overall and by treatment assignment, for the safety population.
Statistical Support
This proposal, and the study’s statistical analyses, were supported by the Trial Innovation 
Network. The statistician involved in the design and sample size calculation is Bradley Barney, 
PhD, School of Medicine, University of Utah.
Interim Analysis
There are two classes of interim analyses: a set of inferential statistical analyses for a formal 
interim stopping rule; and largely descriptive summaries focusing on data and safety monitoring 
used in reports prepared for the DSMB. The statistical testing underlying the formal stopping rule 
will be conducted when every new batch of five site-reported primary safety outcomes has been 
adjudicated. After every 5 patients who are thought to have the primary safety outcome of severe 
bleeding or thrombotic event or all-cause mortality, we will conduct a two-sided Fisher’s exact test 
using an alpha threshold of 0.0333. This p-value threshold was chosen so that, accounting for the 
varying number of interim tests that may be conducted, the overall type I error rate is close to 
0.05. Each test will include those with a known outcome at the time the adjudication process is 
completed (even if for a participant enrolled after the fifth new reported event), with participants 
for whom the outcome has not yet been determined being excluded until the outcome has been 
determined. We will test the null hypothesis that the two treatment strategies have the same 
expected proportion of patients who will experience the primary safety outcome. If a test is 
significant, we will pause the trial for safety reasons. The DSMB will evaluate the risk and make 
a recommendation to stop or continue the study to the NIH.
A DSMB report with safety and data completion metrics will be prepared and distributed to the 
DSMB for every DSMB meeting during the enrollment phase. Scheduled DSMB meetings are 
intended to be held twice during the study’s enrollment period: approximately 8 weeks after the 
tenth participant randomization, and approximately 8 weeks after the 25th participant 
randomization, to allow time for outcome assessment and report preparation.
Since the adjudication committee will be reviewing the major safety concerns (severe bleeding, 
and/or clotting) for every patient with site-reported major safety concerns and because there are 
formal tests when a set of five additional patients has a primary safety outcome per site report, 
there will be no need to pause enrollment during the interim analysisThe study will be paused if 
there is evidence of harm, as noted earlier, i.e., a statistically significant difference in the 
proportion of patients progressing to severe bleeding, severe thrombosis or all-cause mortality 
using p≤0.0333. If such a pause occurs, the study might then be stopped after the NHLBI, DSMB, 
and PIs have the opportunity to discuss findings that led to the automatic pause. In addition, 
serious adverse events related to the intervention will be evaluated and reported in real time 
(within 48 hours of occurrence) by the Medical Monitor and reported to the DSMB.
Note that the primary safety outcome will also be tested for a difference in arms after all 
participants have been enrolled, again using the 0.0333 threshold to be declared statistically 
significant, unless a previous test was already significant at this threshold. However, this is not 
considered an interim analysis because it is after all participants were enrolled.
 
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Many factors may influence exactly what the stopping rule thresholds are in the trial, such as 
primary safety outcomes that are overruled by the adjudication panel (expected to be 
uncommon), the underlying event rate, and the balance of randomization to treatment arms 
throughout the trial.
An example of how the rule translates to go/no-go thresholds under one particular set of 
assumptions is provided. Assuming that at every interim look, there have been equal numbers 
randomized to each treatment, and that at every interim look, half of the total number of enrolled 
patients have experienced the primary safety outcome (though the numbers can differ by arm), 
then there would be tests after 10, 20, 30, 40, and all 50 participants were enrolled, so long as 
there has not been a previous test that was significant. For any given test to have a two-sided p-
value ≤ 0.0333, the primary safety outcome rates would have to be as follows:
•
With n=10 participants, 0/5 (0%) or 5/5 (100%) in the lower threshold arm
•
With n=20 participants, either ≤2 (≤20%) or ≥8 (≥80%) out of 10 in the lower 
threshold arm
•
With n=30 participants, either ≤4 (≤26.7%) or ≥11 (≥73%) out of 15 in the lower 
threshold arm
•
With n=40 participants, either ≤6 (≤30%) or ≥14 (≥70%) out of 20 in the lower 
threshold arm
•
With n=50 participants, either ≤8 (≤32%) or ≥17 (≥68%) out of 25 in the lower 
threshold arm
D. Limitations with Strategies to Overcome
•
Sites may not be able to maintain clinical equipoise throughout the study period, as the study 
is not blinded. To compensate for this, prior to the study initiation, we will present the study 
protocol to leaders in the ECMO team at each site, including general surgeons, cardiothoracic 
surgeons, anesthesiologists, neonatologists, and pediatric intensivists to ensure compliance. 
We will review compliance for each site at monthly meetings and at the interim analysis and 
provide further education at each site as needed. Moreover, we will highlight the fact that the 
full ECSTATIC study will be a non-inferiority RCT, which underlines that we expect to find no 
statistically significant difference in the outcomes of the patients who will participate in the 
large study.
•
The proportion of platelets transfused during temporary suspensions may dilute the 
separation between platelet exposure between the two arms. For this reason, we have limited 
the number of reasons for temporary suspensions. We will also analyze the impact of 
suspensions on the platelet count separation.
•
Different methods can be used to obtain platelet products: 13% of platelet transfusions given 
to ECMO patients are whole blood derived - either by platelet-rich plasma (USA and UK) or 
the buffy-coat method (Europe and Canada), and 87% are obtained by apheresis (single-
donor) (13). The volume of a whole blood derived platelet unit is about 50 mL while that of an 
apheresis platelet unit ranges from 200 to 300 mL. We will not be able to control the methods 
of processing platelets at each site. However, processing details of each platelet transfusion 
will be recorded. At least five of the ten participating sites will be reporting the number of 
platelets per mL of transfusate received so that we can describe the dose of platelets 
 
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transfused and consider controlling for this in the larger trial. Though it is not feasible in the 
pilot due to cost, we will consider assaying the transfusates themselves in the larger trial to 
determine the actual platelet count transfused.
•
It will be possible to consider the pilot ECSTATIC trial as a vanguard phase of the full RCT 
and to merge their data to the data of the patients enrolled in the full trial if no significant 
changes are made to the design of the full RCT. On the other hand, merging patients of the 
pilot and the full trials would be inappropriate if significant changes are made (e.g., the 
threshold platelet count is changed). In that case, we will combine the data of the pilot and of 
the full RCT by performing a meta-analysis.
11. Study Monitoring and Study Organization
Approval for use of this protocol by the Human Studies Committee (HSC) must be obtained in 
accordance with the institutional assurance policies of the U.S. Department of Health and Human 
Services. Institutional Review Board (IRB) approval of the ECSTATIC protocol and consent forms 
will be required prior to patient participation on the trial. The single IRB at BRANY will coordinate 
IRB approvals at each site within the United States. All reasonable measures will be taken to 
protect the confidentiality and identity of the patient and patient’s records according to State and 
Federal laws. Patient identity will not be revealed in any publication.  
A. Study Monitoring
Data will be collected at PICU entry (e.g., PRISM and PELOD-2 scores, co-morbidity, coagulation 
evaluation, etc.), at ECMO cannulation, at randomization, daily while patient is on ECMO, at 
ECMO end, and at PICU discharge. Data will be collected at least on a daily basis from ECMO 
initiation to ECMO end. Mortality (28-day and 90-day) is the only data that would be collected 
after PICU discharge. The application of the RCT protocol (lower or higher platelet threshold 
strategy) will end at the earliest of any of the following five events: (1) progression to severe 
bleeding, according to the BASIC definition, and/or severe clotting, and/or death; (2) 
decannulation from ECMO; (3) Twenty-one days post-randomization (as the progression to new 
bleeding and new thrombosis has plateaued by day 21 and 95% of the patients will be off ECMO 
by then); 4) temporary suspension lasting more than 24 hours; or 5) study withdrawal.
Data management will include an audit trail, a security system, query functionality and quality 
control done according to US CFRs. Data management will be performed at the Trial Innovation 
Network at the University of Utah under the supervision of the study statistician.  
Data will be entered on site in the web-based eCRF. For validation purposes only, double data 
entry will be used for the first two patients enrolled at each site. During the validation phase, CRF 
and entries will be considered adequate if the frequency of discordance is lower than 2% in the 
eCRF. 
The data coordinating center (DCC, see below) will be responsible for data quality assurance 
done through eCRF (via regular data extraction) and queries.
 
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B. Data Safety Monitoring Board
There are two principal mechanisms to achieve the mandate of protecting participant safety and 
being proper stewards of study data for the ECSTATIC Trial. The first is the oversight of a Data 
Safety and Monitoring Board (DSMB) to assess participant safety and the integrity of the study. 
The second is the proven collection of processes employed by the University of Utah Data 
Coordinating Center (DCC) to collect, protect, and analyze study data. 
DSMB 
The purpose of the DSMB is to advise the Federal funding agency (NHLBI) and the ECSTATIC 
Principal Investigators (Drs. Karam and Nellis) regarding the continuing safety of study subjects 
and the continuing validity and scientific merit of the study. The DSMB will be a panel of specialists 
chosen by the NHLBI with input from the PIs. The panel will include a pediatric intensivist, two 
pediatric hematologists, a bioethicist, and a statistician. One member, directed by the NHLBI, will 
chair the group. Any conflicts of interest of the members of the DSMB will be declared and 
reviewed by the executive secretary and the NHLBI ethics officer prior to their first meeting.
The DSMB is responsible for monitoring accrual of study subjects, adherence to the protocol, 
assessments of data quality, performance of individual clinical sites, review of serious adverse 
events and other subject safety issues. The DCC will provide reports relating to these topics to 
DSMB members prior to the interim DSMB meeting(s). It is anticipated that the DSMB will meet 
four times: (1) prior to patient enrollment, (2) approximately 8 weeks after 10 patients are enrolled, 
(3) approximately 8 weeks after 25 patients are enrolled and (4) at trial completion. The DSMB 
and/or Medical Monitor will have the final say in determining if there is a need for more frequent 
interim analyses and/or need for more frequent meetings.
As described above, the DSMB will meet prior to the start of the ECSTATIC Trial to review the 
protocol prior to implementation. The NHLBI executive secretary will draft the DSMB charter, 
which will be reviewed by the DSMB for approval. The charter will include responsibilities of the 
DSMB, definitions of a meeting quorum, information about meeting logistics and frequency, and 
an outline of report contents the DSMB will be given prior to the biannual meetings. In addition, 
they will define event triggers that would call for an unscheduled review, stopping procedures that 
are consistent with the protocol, unmasking (unblinding), and voting procedures. After the DSMB 
has approved its charter and the final protocol, the Data Coordinating Center will send this 
information to the single Institutional Review Board (sIRB) at BRANY.  If the sIRB has changes 
they would like made to the study protocol and/or consent, the changes will be sent to the NHLBI 
and DSMB for review and approval. Certification of IRB approvals will be sent to the NHLBI Grants 
Management Officer (GMO) before enrollment may begin at each site. In addition, after the start 
of the trial, any amendments to the protocol and/or consent will be submitted to the NHLBI and 
DSMB for approval prior to submission to the sIRB. Finally, should the sIRB request any additional 
study actions, such as holding enrollment, requiring additional monitoring, etc, the co-PIs (Drs. 
Nellis and Karam) will notify the DSMB and the NHLBI within 24 hours, by email.
 
 
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DSMB meetings to evaluate study protocols, prior to study implementation, may be open or closed 
according to the decision of the DSMB members. We suggest that pre-enrollment meetings be 
open to members of the ECSTATIC investigative team. After enrollment commences, we suggest 
DSMB meetings consist of both open and closed sessions. Open sessions would allow for 
inclusive attendance in order to facilitate the review and appropriate alterations of the protocol in 
response to DSMB concerns. Any especially sensitive information, such as interim review of trial 
data by treatment arm, will be limited to the closed session. Attendance at the closed session will 
be for DSMB membership, DCC biostatisticians, and possibly administrative personnel supporting 
DSMB members (e.g, the DSMB executive secretary and others) and/or Medical Monitor.  
 
The DSMB can recommend whether or not to terminate enrollment in ECSTATIC because of 
potential safety concerns or study feasibility issues. In addition, they can recommend 
modifications to the trial. We proposed two interim DSMB meetings: (1) approximately 8 weeks 
after 10 patients are enrolled, (2) approximately 8 weeks after 25 patients are enrolled. The focus 
of the interim analysis will be on patient safety and enrollment characteristics. DSMB members 
must be satisfied that the timeliness, completeness, and accuracy of the data submitted to them 
for review are sufficient for evaluation of the safety and welfare of study participants. The interim 
analyses may show overall rates of adverse events in the open session, but summaries by 
treatment arm would be restricted to the closed session. Formal interim analysis for efficacy or 
futility of the primary outcome, separation in pre-transfusion platelet count, is not recommended. 
Our preference for not conducting an efficacy analysis mid-trial is in part because of the small 
sample size, but also because the primary outcome is relevant to the design of the future study, 
but not to the safety of the pilot. Patient safety is of paramount importance.
 
As per NHLBI practices, the NHLBI ES is responsible for preparation and transmission of the 
formal DSMB minutes to the Director of the applicable Division within 14 calendar days of each 
meeting or call. The NHLBI program office will prepare a Summary Report of Board 
Recommendations and submit it to primary study investigators(s) and DCC within 30 calendar 
days of each meeting, if the DSMB does not identify any safety or other protocol-related 
concerns.  If the DSMB does identify concerns, the NHLBI staff will distribute, as soon as feasible, 
preferably within 7 calendar days of the DSMB meeting. Primary study investigators(s) or DCC 
will forward the Summary Report to each participating research site. In the unlikely event that the 
DSMB recommends emergent cessation of enrollment in ECSTATIC because of safety concerns, 
this communication will be made as soon as possible upon the conclusion of the closed session. 
If the NHLBI concur with this recommendation, the DCC will notify all ECSTATIC clinical sites to 
cease enrollment immediately. 
Monitoring
Adverse Events: Each site PI will be responsible, in coordination with their local site research 
coordinators, to monitor the enrolled patients for adverse events on a daily basis. The adverse 
events will be reported to the PIs and DSMB accordingly as described below and as follows. 
An Independent Medical Monitor will be appointed by the investigators to review and monitor all 
serious and unexpected adverse events in real time and perform a periodic review of patient 
safety. Any serious adverse events that are considered by the site to be possibly or probably 
related to the study intervention will be reported to the Medical Monitor within 24 hours of the 
 
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event. For each of these serious 
adverse 
events, 
the 
site 
investigator will provide sufficient 
medical history and clinical details 
for a safety assessment to be 
made with regard to continuation 
of the trial. The Medical Monitor 
will review the event with the site 
within 24 hours and determine if 
the event is related to the 
intervention and if it should 
immediately trigger DSMB review. 
The Independent Safety Monitor 
will be a physician with clinical 
expertise in ECMO with no real or 
perceived conflicts of interest. The 
Independent Safety Monitor will not be part of the DSMB, but the DSMB may review these reports 
or consult with the monitor as needed.
Data Quality: The DCC at the University of Utah will be responsible for monitoring the quality of 
data entry in real-time as described below.
DCC Features and Safeguards
The DCC in the Department of Pediatrics at the University of Utah School of Medicine provides 
data coordination and management services for a variety of national research networks.  
Anchoring these services is a state-of-the-art, energy efficient data center. The DCC currently 
manages over 125 terabytes (TB) of data and supports more than 3500 users around the country 
and provides a secure, reliable, enterprise-wide infrastructure for delivering mission critical DCC 
systems and services.
The DCC employs the following utilities and practices to ensure the safety and security of data 
stored at the facility.
•
The DCC’s electrical power system contains an uninterruptible power supply (UPS) with 
diesel backup generator.  
•
The DCC is protected with an FM-200 backed fire suppression system.  
•
Incremental backups of data occur Monday through Friday.  A full data backup occurs 
weekly. Full backups are sent off-site on a weekly basis to a secure secondary location. 
•
On premise security guards monitor and enforce access control 24 hours a day, 7 days a 
week, 365 days a year. 
•
The DCC and external building access points are monitored with video surveillance and 
entry is restricted by card access and layered security measures and controls.  
•
Direct access to data center machines is only available while physically on premise or via 
a VPN client. 
•
All network traffic is monitored for intrusion attempts.  
•
All DCC personnel have completed Human Subjects Protection and Health Information 
Portability and Accountability Act (HIPAA) education. 
Figure 8: Serious safety event and medical monitor’s reporting process
 
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•
The DCC requires all users to sign specific agreements concerning security, 
confidentiality, and use of our information systems before access is provided.
Data management efforts extend beyond collecting and securely storing data. To promote the 
collection of valid data, the DCC uses an in-house application named QueryManager to support 
a variety of studies. Rules are built to identify certain types of potentially problematic data. Clinical 
sites will be sent queries in real time when data for participants at their site violate these rules, 
and these queries are tracked until the problem is automatically resolved (e.g., by correcting 
aberrant data entries) or manually resolved (e.g., confirmation received from the site that the 
original data were correct). Critical derived variables and analyses reported in DSMB reports, 
abstracts or manuscripts are routinely dual-programmed or code-reviewed by a second 
statistician. These safeguards instill confidence that the collected data and reported results are 
accurate. 
Adverse Events
Definition of an Adverse Event
An adverse event (AE) is an unfavorable occurrence in a study subject that begins or worsens 
during the study (from the time of randomization through study completion). The reporting of an 
adverse event does not imply a causal relationship between the event and study participation. 
Determination of the relationship between the event and study drug or a study mandated 
procedure must be made by the Site Investigator. All adverse events will be categorized as being 
possibly, probably, or not related to study intervention. For any unexpected serious adverse 
events that are considered by the site to be possibly or probably related to the study intervention, 
a report will be made to the Medical Monitor within 24 hours of the event. The Medical Monitor 
will review the event with the site within 24 hours and determine if the event is related and if it 
should immediately trigger DSMB review (automatic emails send by REDCap).
Clinical Adverse Events
Children requiring support from extracorporeal membrane oxygenation (ECMO) are at the 
extreme of critical illness and therefore we expect a wide range of adverse events, including organ 
dysfunction, hospital-acquired infections, and death (which occurs in up to 50% of all children on 
ECMO), to happen as part of the routine clinical care. Adverse events to be collected will include, 
but not be limited to, transfusion-related reactions (including hyperkalemia, allergic reactions, 
transfusion-related acute lung injury, transfusion-associated circulatory overload and nosocomial 
infections), and ECMO-related complications not already captured in the outcomes (including 
seizures, need for neurosurgical intervention, cardiac arrhythmias, tamponade, pneumothorax, 
compartment syndrome, fasciotomy and amputation). The occurrence of adverse events will be 
monitored by the site PI or the site coordinator on a daily basis from the time of enrollment until 
24 hours after when the transfusion strategy was permanently discontinued; strategy 
discontinuation is initiated by five possible events (whichever occurs first): 1) progression to 
severe bleeding, according to the BASIC definition, and/or severe clotting, and/or death; 2) 
decannulation from ECMO; 3) Twenty-one days post-randomization (as the progression to new 
bleeding and new thrombosis has plateaued by day 21 and 95% of the patients will be off ECMO 
by then); 4) temporary suspension lasting more than 24 hours; or 5) study withdrawal. If AEs are 
identified, the AEs will be reported as they arise as described below and then will be followed 
through to resolution or hospital discharge, whichever comes first.  
 
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Severity of AEs will be defined as follows:
•
No severity: mild event that does not impact the expected clinical course.
•
Moderate: moderate event that leads to prolonged time on ECMO, prolonged length of 
stay or additional procedures.
•
Severe: severe event that leads to long-term sequelae or death.
A distinction will be drawn between serious and severe AEs. A severe AE is a major event of its 
type.  A severe AE does not necessarily need to be considered serious.  For example, nausea 
which persists for several hours may be considered severe nausea, but would not be an SAE.  
On the other hand, a stroke that results in only a limited degree of disability may be considered a 
mild stroke, but would be an SAE.
Relatedness
The site investigator must determine if an AE is “probably related”, “possibly related”, or “not 
related” to the platelet transfusion strategy. Determinations will be based on the biologic 
plausibility, the temporal relationship, and the presence or absence of an alternative explanation 
for an AE.
•
Probably Related: The event follows a reasonable temporal sequence from the time of 
beginning the assigned study intervention, and cannot be reasonably explained by other 
factors such as the subject’s clinical state, therapeutic interventions, or concomitant drugs 
administered to the subject.
•
Possibly Related: The event follows compatible temporal sequence from the time of 
beginning the assigned study intervention, but could have been produced by other factors 
such as the subject’s clinical state, therapeutic interventions, or concomitant drugs 
administered to the subject.
•
Not Related: The event is clearly related to other factors, such as the subject’s clinical state, 
therapeutic interventions, or concomitant drugs administered to the subject.
Expectedness
All adverse events, including serious adverse events, will be evaluated as to whether their 
occurrence was expected or unexpected. An adverse event is considered expected if it is known 
to be associated with ECMO, underlying medical conditions of the subject, is directly related to 
study outcome, or is otherwise mentioned in the protocol, informed consent, investigator 
brochure, or other study documents.
 
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Adverse event reporting
The period for collecting adverse 
events 
is 
from 
the 
time 
of 
randomization through 24 hours past 
the 
time 
when 
the 
transfusion 
strategy was last applicable. AEs that 
are not SAEs will only be reported if 
judged by the investigators to be at 
least 
possibly 
related 
to 
the 
transfusion strategy.  SAEs will be 
reported regardless of whether these 
are judged to be related to the 
transfusion strategy, with unexpected 
(unanticipated), serious (including all 
deaths), and related SAEs reported 
promptly as noted in Figure 9. The Medical Monitor will notify by email the DSMB,  NHLBI, and 
PIs within 24 hours of being notified by the site investigator (using REDCap’s automatically-
generated emails).
Of note, deaths occurring after ICU transfer or discharge will be reported within 24 hours of it 
coming to the attention of either of the co-primary investigators (e.g. a patient might die 38 days 
after the end of the ECMO run, in another institution; in which case, the co-PIs will only be made 
aware at the 90-day follow-up call).
C. Study Organization
Study primary investigators 
Drs. Karam and Nellis (study primary investigators) will have overall responsibility for the project. 
They will ensure the smooth operation of all committees and facilitate communication between 
committee members. They will have the final approval of all reports and scientific publications 
emanating from the study. The study primary investigators will preside over all Executive and 
Steering Committee meetings. They will also appoint a study Vice-Chair at its first meeting who 
will assume all responsibilities for the Chairs in their absence.
Executive Committee
The voting members of the Executive Committee will consist of Oliver Karam, Marianne Nellis, 
Marisa Tucci, Jacques Lacroix, Philip Spinella, Jennifer Muszynski, Scott Weiss, Simon 
Stanworth and Paul Clarke. 
The Executive Committee will oversee all aspects of the ECSTATIC trial including but not limited 
to:
1.
Will ensure that the study is conducted in accordance to the protocol.
2.
Will closely monitor the progress of the study.
3.
Will monitor study expenses.
4.
Will develop or modify policies and instructions regarding daily operations of the trial.
5.
Will oversee analysis of the results.
6.
Will be responsible for the writing of all publications pertaining to the trial.
Figure 9:  Reporting of Adverse Events
 
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7.
May develop modifications to the design, execution and analysis of the trial, if applicable.
8.
Will have ultimate responsibility for data management and quality assurance, with the help 
of the staff of the Data Coordinating Center
9.
Will receive reports from the DSMB.
10. Will report all safety concerns encountered to the Steering Committee and the Data and 
Safety Monitoring Board.
11. Will approve all ancillary research and/or sub-group analyses involving study participants 
proposed by principal investigators or by outsider researchers.
It is expected that one member of the Executive Committee will present a study progress report 
at all official meetings of the PALISI Network, which holds 2 meetings per year.
The Executive Committee may wish to delegate some responsibilities to other subcommittees 
such as a writing committee, a quality assurance committee, etc. The Executive Committee will 
convene regularly (on Zoom) according to a pre-defined schedule and according to the need to 
discuss any significant issues that may arise. A quorum will require at least three members.
Steering Committee
The voting members of the Steering Committee will consist of the members of the Executive 
Committee, and each of the site primary investigators from the participating sites.  Non-voting 
members of the Steering Committee will make use of expertise in the fields of pediatric critical 
care medicine, transfusion medicine, data management, statistics, and finances/administration. 
The Steering Committee will convene at least once a year once the trial has begun.
The Steering Committee will have the overall responsibility for the design, execution, analysis and 
publication of results of the ECSTATIC Trial including but not limited to:
1.
implementation of all major policy changes made by the Executive Committee
2.
reporting all potential safety problems encountered by the Executive Committee
3.
review of accrual patterns
4.
reporting randomization and data collection procedures problems as required
5.
discussion of all other concerns of any member of the Committee.
Writing Committee
The Writing Committee will reflect trial participation, preserving the writing/authorship roles of the 
primary study leaders, if they so desire. Broad distribution of authorship among Study 
Investigators is encouraged. The Writing Committee will be chaired by the study primary 
investigators. All other principal investigators and co-investigators may be members of the Writing 
Committee, as determined by the Executive Committee.
The Writing Committee is responsible for the following:
•
Review of proposed abstracts, conference presentations and publications, and any 
discussion necessary to clarify issues concerned with the preparation and publication of 
manuscripts, such as overlap and data availability.
 
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•
Forwarding the detailed plan for the proposed publication to the Executive Committee for 
review (to ensure accurate representation of trial results and adherence to trial policies) 
and final approval within 4 weeks.
•
All publications must be approved by the Scientific Committee of BloodNet as well as the 
PALISI Network before it is submitted to a scientific journal; they must also be published 
on behalf of BloodNet and the PALISI Network.
•
Ensure that publication, authorship criteria and rules with respect to conflict of interest are 
adhered to by the authors.
•
Proper acknowledgment of the funding agency is required in all publications 
•
Periodically provide a listing of the status of submitted manuscripts to the Executive 
Committee.
The Writing Committee will convene by teleconference as necessary to address and prepare 
proposed publications.
Data Coordinating Center
The Data Coordinating Center (DCC) in the Department of Pediatrics at the University of Utah 
School of Medicine will provide data coordination and management services. Anchoring these 
services is a state-of-the-art, energy efficient data center. The DCC currently manages over 125 
terabytes (TB) of data and supports more than 3500 users around the country and provides a 
secure, reliable, enterprise-wide infrastructure for delivering mission critical DCC systems and 
services.
The DCC employs the following utilities and practices to ensure the safety and security of data 
stored at the facility.
•
The DCC’s electrical power system contains an uninterruptible power supply (UPS) with 
diesel backup generator.  
•
The DCC is protected with an FM-200 backed fire suppression system.  
•
Incremental backups of data occur Monday through Friday.  A full data backup occurs 
weekly. Full backups are sent off-site on a weekly basis to a secure secondary location.
•
On premise security guards monitor and enforce access control 24 hours a day, 7 days a 
week, 365 days a year. 
•
The DCC and external building access points are monitored with video surveillance and 
entry is restricted by card access and layered security measures and controls.  
•
Direct access to data center machines is only available while physically on premise or via 
a VPN client. 
•
All network traffic is monitored for intrusion attempts.  
•
All DCC personnel have completed Human Subjects Protection and Health Information 
Portability and Accountability Act (HIPAA) education. 
•
The DCC requires all users to sign specific agreements concerning security, 
confidentiality, and use of our information systems before access is provided.
Data management efforts extend beyond collecting and securely storing data. To promote the 
collection of valid data, the DCC uses an in-house application named Query Manager to support 
a variety of studies. Rules are built to identify certain types of potentially problematic data. Clinical 
sites are sent queries when data for participants at their site violate these rules, and these queries 
 
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are tracked until the problem is automatically resolved (e.g., by correcting aberrant data entries) 
or manually resolved (e.g., confirmation received from the site that the original data were correct). 
Critical derived variables and analyses reported in DSMB reports, abstracts or manuscripts are 
routinely dual-programmed or code-reviewed by a second statistician. These safeguards instill 
confidence that the collected data and reported results are accurate.
D. Study Timeline
The study will be conducted over three years (Table 2). 
The first six months will be primarily spent on the IRB submission, database development, and 
site on-boarding and training. 
The following two years will be spent on patient enrollment. Due to the support of the Data 
Coordinating Center at the University of Utah, data entry will be monitored in real-time and time 
for data cleaning will not be required at the end of the trial. 
The second half of the third year will be spent on data analysis and manuscript preparation. 
Table 2: Timeline
Year 1
Year 2
Year 3
Q1
Q2
Q3
Q4
Q1
Q2
Q3
Q4
Q1
Q2
Q3
Q4
IRB submission
Database development
Site onboarding
Patient enrollment
Data validation
Analysis
Manuscript preparation
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biogenesis. Platelets. 2017 Feb 17;28(2):138–46. 
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MacLaren G, Monagle P. Platelet Transfusion During Extracorporeal Membrane 
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30.
Biagas KV, Muratore CS. Plasma and Platelet Utilization in Children on Extracorporeal 
Membrane Oxygenation: A Call for Targeted Anticoagulation*. Pediatr Crit Care Med. 2020 
Mar;21(3):296–7. 
31.
Ming Y, Liu J, Zhang F, Chen C, Zhou L, Du L, et al. Transfusion of Red Blood Cells, 
Fresh Frozen Plasma, or Platelets Is Associated With Mortality and Infection After Cardiac 
Surgery in a Dose-Dependent Manner: Anesth Analg. 2020 Feb;130(2):488–97. 
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Josephson CD, Granger S, Assmann SF, Castillejo MI, Strauss RG, Slichter SJ, et al. 
Bleeding risks are higher in children versus adults given prophylactic platelet transfusions for 
treatment-induced hypoproliferative thrombocytopenia. Blood. 2012 Jul 26;120(4):748–60. 
33.
Lacroix J, Hébert PC, Hutchison JS, Hume HA, Tucci M, Ducruet T, et al. Transfusion 
strategies for patients in pediatric intensive care units. N Engl J Med. 2007 Apr 
19;356(16):1609–19. 
34.
Nellis ME, Tucci M, Lacroix J, Spinella PC, Haque KD, Stock A, et al. Bleeding 
Assessment Scale in Critically Ill Children (BASIC): Physician-Driven Diagnostic Criteria for 
Bleeding Severity. Crit Care Med. 2019 Dec;47(12):1766–72. 
35.
Pal S, Curley A, Stanworth SJ. Interpretation of clotting tests in the neonate. Arch Dis 
Child - Fetal Neonatal Ed. 2015 May;100(3):F270–4. 
36.
van Stuijvenberg M, Suur MH, de Vos S, Tjiang GCH, Steyerberg EW, Derksen-Lubsen 
G, et al. Informed consent, parental awareness, and reasons for participating in a randomised 
controlled study. Arch Dis Child. 1998 Aug 1;79(2):120–5. 
37.
Tait AR, Voepel-Lewis T, Siewert M, Malviya S. Factors that influence parents’ decisions 
to consent to their child’s participation in clinical anesthesia research. Anesth Analg. 1998 
Jan;86(1):50–3. 
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Murtha TD, Faustino EVS. Is “Informed Consent” Truly Informed?*. Pediatr Crit Care 
Med. 2020 Jun;21(6):589–90. 
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Gertsman S, O’Hearn K, Gibson J, Menon K. Parental Understanding of Research 
Consent Forms in the PICU: A Pilot Study*. Pediatr Crit Care Med. 2020 Jun;21(6):526–34. 
 
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40.
FDA Standards for Human Blood and Blood Products - Platelets [Internet]. Electronic 
Code of Federal Regulations. Available from: https://www.ecfr.gov/cgi-bin/text-
idx?SID=5d10701242f6aa41e680efeaef1d712c&mc=true&node=sp21.7.640.c&rgn=div6
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Extracorporeal Life Support Organization (ELSO). Guidelines for Neonatal Respiratory 
Failure [Internet]. 2017. Available from: 
https://www.elso.org/Portals/0/ELSOGuidelinesNeonatalRespiratoryFailurev1_4_1.pdf
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Bembea MM, Schwartz JM, Shah N, Colantuoni E, Lehmann CU, Kickler T, et al. 
Anticoagulation Monitoring during Pediatric Extracorporeal Membrane Oxygenation. ASAIO J. 
2013 Jan;59(1):63–8. 
43.
Barbaro RP, Boonstra PS, Paden ML, Roberts LA, Annich GM, Bartlett RH, et al. 
Development and validation of the pediatric risk estimate score for children using extracorporeal 
respiratory support (Ped-RESCUERS). Intensive Care Med. 2016 May;42(5):879–88. 
44.
Şık G, Demirbuğa A, Annayev A, Temur B, Aydın S, Demir Hİ, et al. Clinical and 
laboratory predictors of survival for pediatric patients on non-postcardiotomy extracorporeal 
membrane oxygenation (ecmo). Turk J Pediatr. 2020;62(1):24. 
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Bokman CL, Tashiro J, Perez EA, Lasko DS, Sola JE. Determinants of survival and 
resource utilization for pediatric extracorporeal membrane oxygenation in the United States 
1997–2009. J Pediatr Surg. 2015 May;50(5):809–14. 
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Pollack MM, Patel KM, Ruttimann UE. PRISM III: An updated Pediatric Risk of Mortality 
score. Crit Care Med. 1996 May;24(5):743–52. 
47.
Slater A, Shann F, Pearson G, for the PIM Study Group. PIM2: a revised version of the 
Paediatric Index of Mortality. Intensive Care Med. 2003 Feb;29(2):278–85. 
48.
Spinella PC, El Kassar N, Cap AP, Kindzelski AL, Almond CS, Barkun A, et al. 
Recommended primary outcomes for clinical trials evaluating hemostatic blood products and 
agents in patients with bleeding: Proceedings of a National Heart Lung and Blood Institute and 
US Department of Defense Consensus Conference. J Trauma Acute Care Surg. 2021 Aug 
1;91(2S Suppl 2):S19–25. 
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Akin S, Caliskan K, Soliman O, Muslem R, Guven G, van Thiel RJ, et al. A novel 
mortality risk score predicting intensive care mortality in cardiogenic shock patients treated with 
veno-arterial extracorporeal membrane oxygenation. J Crit Care. 2020 Feb;55:35–41. 
50.
Muller G, Flecher E, Lebreton G, Luyt CE, Trouillet JL, Bréchot N, et al. The 
ENCOURAGE mortality risk score and analysis of long-term outcomes after VA-ECMO for acute 
myocardial infarction with cardiogenic shock. Intensive Care Med. 2016 Mar;42(3):370–8. 
51.
ELSO Database Definitions [Internet]. [cited 2020 Apr 27]. Available from: 
https://www.elso.org/Portals/0/Files/PDF/ELSO%20Database%20Definitions%202018-2-1.pdf
52.
Karam O, Nellis ME, Zantek ND, Lacroix J, Faustino EVS, BloodNet and the Pediatric 
Acute Lung Injury and Sepsis Investigators (PALISI) Network. Criteria for Clinically Relevant 
Bleeding in Critically Ill Children: An International Survey. Pediatr Crit Care Med J Soc Crit Care 
Med World Fed Pediatr Intensive Crit Care Soc. 2019;20(3):e137–44. 
53.
Fiser DH. Assessing the outcome of pediatric intensive care. J Pediatr. 1992 
Jul;121(1):68–74. 
54.
Pollack MM, Holubkov R, Funai T, Clark A, Moler F, Shanley T, et al. Relationship 
Between the Functional Status Scale and the Pediatric Overall Performance Category and 
Pediatric Cerebral Performance Category Scales. JAMA Pediatr. 2014 Jul 1;168(7):671. 
55.
Hodgson CL, Burrell AJC, Engeler DM, Pellegrino VA, Brodie D, Fan E. Core Outcome 
Measures for Research in Critically Ill Patients Receiving Extracorporeal Membrane 
Oxygenation for Acute Respiratory or Cardiac Failure: An International, Multidisciplinary, 
Modified Delphi Consensus Study*. Crit Care Med. 2019 Nov;47(11):1557–63. 
 
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56.
Faes C, Molenberghs G, Aerts M, Verbeke G, Kenward MG. The Effective Sample Size 
and an Alternative Small-Sample Degrees-of-Freedom Method. Am Stat. 2009 Nov;63(4):389–
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Chan AW, Tetzlaff JM, Gotzsche PC, Altman DG, Mann H, Berlin JA, et al. SPIRIT 2013 
explanation and elaboration: guidance for protocols of clinical trials. BMJ. 2013 Jan 9;346(jan08 
15):e7586–e7586. 
58.
Loudon K, Treweek S, Sullivan F, Donnan P, Thorpe KE, Zwarenstein M. The PRECIS-2 
tool: designing trials that are fit for purpose. BMJ. 2015 May 8;350(may08 1):h2147–h2147. 
 
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13. Appendices
A. Proposal for future large RCT
Aims
The primary research aim for the future RCT will be to demonstrate the non-inferiority of a lower 
platelet threshold transfusion strategy, as compared to a higher platelet threshold strategy, in non-
bleeding children on ECMO. 
Anticipated Cohort
Our aim is to enroll non-bleeding or minimally bleeding 
children on ECMO, at risk of receiving platelet 
transfusions. 
Design
This will be a multicenter, non-inferiority, randomized, 
controlled, investigator-initiated, open-label, trial with 
blinded adjudication of the outcomes (Figure 10). We will 
take into account the recommendations of SPIRIT to 
elaborate our final version of the protocol for the full 
ECSTATIC RCT (57). We will use the PRECIS-2 tool to 
estimate the pragmatic or explanatory nature of the trial 
(58).
Site Eligibility
PICUs will be considered eligible to participate in the full 
ECSTATIC RCT only if there is an ECMO program in the 
site. 
Site Inclusion Criteria: Pediatric and neonatal ICUs, 
including cardiac PICUs, with at least a mean of 5 ECMO 
runs per year. 
Site Exclusion Criteria: PICUs will be excluded if 1) a 
consensus from all physicians working in the PICU to 
apply the research protocol is not reached, and/or 2) if 
there is no local scientific infrastructure that can support 
the participation in a large multicenter RCT.
Patient Eligibility
Children will be eligible if admitted to a participating tertiary 
care PICU, NICU or CICU for whom the decision to 
cannulate for ECMO support has been made. 
Inclusion Criteria: Critically ill children (0 to <18 years of age), admitted to a participating 
PICU/NICU/CICU, on ECMO for less than 24 hours, and who have either no bleeding or 
minimal bleeding. If a patient experiences more than minimal bleeding after ECMO 
Figure 10: Overall proposal of the future RCT
 
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cannulation, they can be enrolled once bleeding meets criteria for no/minimal bleeding, with the 
condition that the moderate or severe bleeding resolves within the first 24 hours of ECMO 
initiation. 
Exclusion Criteria
To ensure our trial is as pragmatic as possible, we have as few exclusion criteria as possible: 
1) post conception age < 37 weeks at time of screening
2) underlying oncologic diagnosis (defined as receipt of chemotherapy or radiation in the last 
six months) or recipient of bone marrow transplant in the last year
3) congenital bleeding disorder or congenital thrombocytopenia
4) pregnant or admitted post-partum (within 6 weeks of giving birth)
5) decision to withdraw or withhold some critical care or interventions
6) known objection to blood transfusions
7) on ECMO for > 24 hours at time of enrollment
 Inclusion criteria and exclusion criteria will be ascertained by pediatric critical care clinical or 
research staff.
Intervention
Subjects will be randomized in a 1:1 ratio to either a lower platelet threshold or a higher platelet 
threshold prophylactic transfusion strategy. 
Subjects in the lower platelet threshold arm can be transfused only if their platelet count is below 
50 x109 cell/L, whereas subjects in the higher platelet threshold arm should be transfused if 
their platelet count is below 90 x109 cell/L. 
The transfusion dose is intended to be 10-20 ml/kg but with some allowance based on local 
policies as long as dosing is not differentially applied by treatment arm. For patients > 30 kg, the 
dose will be one unit.
This intervention will be maintained in effect until five possible events that will signal permanent 
discontinuation, whichever occurs first: 1) death, or progression to moderate or severe bleeding 
or severe thrombosis, as diagnosed by the attending physician (i.e., primary safety outcome); 2) 
decannulation from ECMO; 3) 21 days elapse post-randomization, 4) temporary suspension 
lasting more than 24 hours, or 5) study withdrawal. 
Co-interventions
As this is an open-label trial, we need to ensure co-interventions that may be associated to the 
primary clinical outcome are managed identically in both arms, to avoid potential interactions and 
biases. However, as the pragmatic design is important for the generalizability of the future 
results, we will be imposing the adherence to the Pediatric ECMO Anti-Coagulation 
CollaborativE (PEACE) recommendations. Of note, none of the centers have anticoagulation 
protocols that are dependent on the platelet count. Data on co-interventions (plasma transfusion 
strategy, anticoagulation strategy, cryoprecipitate or fibrinogen concentrate, antithrombin 
concentrate, and priming of circuit) will be collected daily and compliance with the guidelines will 
be measured, so that the effect of any imbalances on the primary outcome can be examined after 
the trial is complete. Other co-interventions, such as ECMO flow management, vasopressors, 
ventilation, red blood cell transfusion, sedation, and the use of hypothermia, will not be 
protocolized, due to the pragmatic nature of the trial. In addition, the use of other hemostatic 
agents, such as tranexamic acid, aminocaproic acid, DDAVP, and von Willebrand factor will not 
be protocolized as these are typically used in patients with active bleeding and not to prevent 
bleeding. However, all these co-interventions will be recorded.
 
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Randomization
Allocation will be a 1:1 ratio. Only the study statistician and designate at the DCC will have 
knowledge of the randomization codes. Subjects will be stratified by site (as there might be a 
site effect, due to the co-interventions mentioned above) and by age (≤28 days vs >28 days), 
which is of principal interest to keep balanced between arms. Patients will not be stratified based 
on the severity of the disease or indication (medical vs post-surgical), as this would significantly 
increase the sample size. Moreover, in order to conceal randomization, 3 sizes of blocks 
permutation will be used on a random sequence: 2, 4 and 6 patients per block.
Concealment of randomization
Research personnel at each site and PICU caregivers will not have access to the randomization 
schedule. 
Blinding
This is an open-label trial, since the transfusion strategy cannot be blinded from the clinical 
team, as they must order the transfusions based on the platelet count. The research team, 
including the adjudicators, will be blinded to treatment allocation, until the end of the analysis. 
Temporary suspension
The protocol will allow for temporary suspensions, i.e., transfusion at a higher platelet count 
threshold than allowed for the randomization arm, if the clinical situation warrants immediate 
transfusion for certain conditions including: 1) chest-tube insertion; 2) surgical intervention while 
on ECMO (such as tracheostomy or repair of congenital diaphragmatic hernia); 3) during ECMO 
circuit change; or 4) in preparation for decannulation. Adherence to the trial protocol must be 
resumed as soon as possible once these events are controlled or completed. Data monitoring 
and collection will be unchanged during suspension, but data on length and justification of 
suspension will also be collected. Suspensions less than 24 hours will not be considered a breach 
of adherence to the protocol. After 24 hours of temporary suspension, the patient will have all 
assigned treatment strategy implications lifted (i.e., no restrictions on the range at which any 
subsequent platelet transfusions may occur) but will continue to be followed for ascertainment of 
safety and study outcomes. Suspended patients will be included in the intention-to-treat analysis.
Primary Outcome
The primary outcome will be the proportion of patients who progress from no bleeding or minimal 
bleeding to severe bleeding and/or severe thrombosis, ignoring events that begin during a period 
of temporary suspension.
Using the BASIC definition plus one additional element, severe bleeding will be defined as: 
1) Bleeding that leads to at least one organ dysfunction, using PELOD-2 score criteria of 
organ dysfunction
2) Bleeding that leads to hemodynamic instability, defined as an increase in HR by > 20% 
from baseline or a decrease in BP by > 20% from baseline (i.e., prior to the bleeding event)
3) Bleeding leading to a drop in hemoglobin > 20% within 24hr
4) Quantifiable bleeding ≥ 5 mL/kg/hr for ≥ 1hr (e.g., chest tube)
5) New and/or expanding intracranial hemorrhage on CT scan or head ultrasound
6) Intraspinal bleeding leading to loss of neurologic function below the lesion, nontraumatic 
intra-articular bleeding leading to decreased range of movement, or intraocular bleeding 
leading to impaired vision. 
 
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Severe thrombotic events will be defined according to the NHLBI Consensus Conference as 
circuit clot leading to circuit change, need for thrombectomy, ischemic stroke, or distal 
thromboembolism.
As this is an open-label trial, the primary outcome will be adjudicated by an independent central 
committee, blinded to the allocation group. This committee, composed of three members, will 
determine if the bleeding was the primary cause of organ dysfunction, hemodynamic instability, 
acute anemia, loss of spinal function, articular range of motion, or impaired vision.
Secondary Outcomes
The secondary outcomes will be: 
1) hemorrhagic complications according to the ELSO definition
2) thrombotic complications according to the ELSO definition
3) blood product exposure, defined as the total number of transfusions (red blood cell, 
plasma, platelets, cryoprecipitate)
4) duration of ECMO, defined as 90-day ECMO-free days
5) length of stay, defined as the 90-day PICU-free days
6) functional outcome at hospital discharge, defined as the discharge Pediatric Overall 
Performance Category (POPC) and Pediatric Cerebral Performance Category (PCPC)
7) all-cause mortality on ECMO, at 28 days, and at 90 days.
Sample Size
Preliminary data from the BATE study suggests about 65% of the non-bleeding patients will 
experience severe bleeding, severe clotting, and/or death (6). Each ml/kg of platelet transfusion 
was independently associated with 1% increased odds of bleeding and 5% increased odds of 
mortality. Considering the median dose is 92 ml/kg per ECMO run and a low platelet threshold 
transfusion strategy is expected to decrease blood exposure by at least 20%, the effect of the low 
platelet threshold strategy may be a 17% relative decrease in the odds of bleeding [(1/1.01) odds 
ratio for each 1 ml/kg reduction and median reduction of 20% x 92 ml/kg), i.e., an absolute risk 
reduction in percentages from 65% to approximately 60.7%. These results, which are 
extrapolated from observational data where no transfusion strategies were enforced, need to be 
confirmed in the pilot trial. 
Non-inferiority margins of safety of 5% and 10% (33) have previously been used in transfusion 
medicine. With a margin of 5% and based on these very uncertain data, if there is a true relative 
difference in favor of the low platelet threshold strategy of 6.6% (i.e., 65% vs 60.7% progression 
to severe bleeding and/or severe clotting), and we anticipate a 5% drop-out rate (similar to what 
was observed in previous studies), using an alpha of 0.025 and a 90% power, the sample size 
to enroll for the test of non-inferiority would be 1238 patients (619 per arm), The primary 
outcome would be ascertained for an expected 588 per arm. We will calculate the 95% confidence 
intervals (95% CI) around the absolute risk reduction in the proportion of patients progressing to 
severe bleeding and conclude non-inferiority if the upper limit of the 95% CI is lower than 5%. For 
example, if the proportion of patients who progress to the primary outcome is 376/588 (64%) in 
the higher platelet threshold group and 359/588 (61%) in the lower platelet threshold group, the 
absolute risk reduction would be 3%, but the 95% CI would be -9% to 3%. In this example, 
although superiority would not have been demonstrated (p=0.34), we could conclude to a non-
 
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inferiority as the upper limit of the absolute risk reductions (+3%) is below the +5% pre-defined 
limit of the margin of safety.
If non-inferiority were not demonstrated, experts believe such trials still provide important 
information, since we will be able to report multiple secondary and tertiary outcomes, such as 
blood product exposure and lengths of ECMO or stay. Such results would still be impactful, as a 
potential decrease in blood product exposure or in lengths of care would be clinically meaningful. 
In terms of feasibility, enrolling 1238 patients in 30 centers who would each enroll 10 
subjects by year, would require just over 4 years.
Statistical Analyses
The analysis of the primary outcome measure will be conducted on an intention to treat (ITT) 
basis for all patients randomized in ECSTATIC. The principal analysis, i.e., the influence of 
platelet transfusion strategy (“higher platelet threshold” vs. “lower platelet threshold”) on the 
primary outcome (progression to severe bleeding and/or severe clotting, ignoring such events 
that begin during a period of temporary suspension), will be done using an unadjusted z-test (with 
continuity correction) of non-inferiority, comparing the proportion of patients who progress to the 
primary outcome after randomization in both groups. The principal effect measure will be an 
unadjusted absolute risk reduction (ARR) with a 95% CI for the primary comparison. 
Secondary analyses of the primary composite outcome will include logistic regression 
modeling to further elucidate the measure of effect while adjusting for known prognostic risk 
factors. For adjusted models, risk factors such as site, age, co-morbid illnesses, type of ECMO, 
indication to ECMO, and severity of illness scores will be added based on clinical (not statistical) 
rationale. Continuous risk factors (e.g., PELOD-2, number of transfusions) will be entered into the 
models as a continuous measure to improve statistical efficiency. Regression diagnostics will be 
performed on all models. We will also compare Kaplan-Meier curves using a log rank test followed 
by proportional hazards modeling for liberation from ECMO alive, 28- and 90-day mortality, and 
time to the composite outcome: these analyses will compare the length of time between 
randomization and appearance of those outcomes.
We will also do a per-protocol analysis using an unadjusted absolute risk reduction with 95% CI 
on the effect of “higher platelet threshold” vs. “lower platelet threshold” transfusion strategies, but 
such per-protocol analysis will be done only for the primary outcome. We will perform the following 
pre-specified subgroup analyses: types of ECMO (V-V vs V-A), indication to ECMO (medical 
vs surgical), age (< 7 days, 7 to 28 days, 29 to 365 days, > 365 days), location of the patients 
(NICU, PICU, CICU), and indication for ECMO (cardiac, respiratory, and ECMO cardiopulmonary 
resuscitation).
We will also perform some sensitivity analysis pertaining to the primary outcome. For example, 
we will repeat statistical analyses, adding lost to follow-up patients, using best-case and worst-
case scenario.
 
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B. Bleeding Assessment Scale in Critically Ill Children (BASIC) Score
Any of the following criteria define severe bleeding:
•
Bleeding that leads to at least one organ dysfunction, using PELOD-2 score criteria, within 
24 hours of the previous assessment (if there is no previous assessment, the results are 
presumed to be normal). The organ dysfunction should be associated with the bleeding, 
in absence of other causes.
•
Bleeding that leads to hemodynamic instability, defined as an increase in heart rate (HR) 
by > 20% from baseline or a decrease in blood pressure (BP) by > 20% from baseline 
(i.e., prior to bleeding event). The hemodynamic instability should be associated with the 
bleeding, in absence of other causes.
•
Bleeding leading to a drop in Hb >20% within 24 hours. The drop in Hb should be 
associated with the bleeding, in absence of other causes.
•
Quantifiable bleeding ≥ 5 ml/kg/hr for ≥ 1 hour (eg. chest tube, drain).
•
Intraspinal bleeding leading to loss of neurologic function below the lesion, non-traumatic 
intra-articular bleeding leading to decreased range of movement, or intraocular bleeding 
leading to impaired vision.
•
New and/or expanding intracranial hemorrhage (not in the original BASIC definition, but 
added as explained earlier)
All of the following criteria must be present to define moderate bleeding: 
•
Bleeding more than minimal bleeding but without criteria for severe bleeding.
•
Bleeding not leading to organ dysfunction, as measured by the PELOD-2 score. 
•
Bleeding not leading to hemodynamic instability, i.e. change in HR > 20% or BP < 20% 
from baseline (i.e., prior to bleeding event). 
•
Bleeding leading to a drop in Hb ≤ 20%. 
•
Quantifiable bleeding ≥ 1 ml/kg/hr but < 5 ml/kg/hr for ≥ 1 hour (e.g. chest tube, drain). 
Any of the following criteria define minimal bleeding: 
•
Streaks of blood in endotracheal tube (ETT) or during suctioning only. 
•
Streaks of blood in nasogastric (NG) tube.
•
Macroscopic hematuria, or less than or equal to 1+ RBCs present on urine dipstick if 
available.
•
Subcutaneous bleeding (including hematoma and petechiae) < 5 cm (2 in) in diameter.
•
Quantifiable bleeding < 1 ml/kg/hr (e.g. chest tube, drain). Since the objective is to capture 
bleeding, chest tube output can be higher than 1 mL/kg/hr provided the output is 
serosanguinous and the site investigator judges the bleeding portion of the chest tube 
output is < 1mL/kg/hr.
•
Bloody dressings required to be changed not more often than each 6 hours, or weighing 
no more than 1mL/kg/hr if weighed, due to slow saturation. 
Progressive bleeding is bleeding that either progresses to a higher severity category (e.g. from 
minimal to moderate bleeding, or from moderate to severe bleeding) or to a higher number of 
criteria within the same category (e.g. hemodynamic instability progressing to organ failure, or 
streaks of blood in the ETT and subsequent slightly blood-tinged urine). 
Fatal Bleeding is bleeding that is the direct cause leading to death.
 
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C. NHLBI Definition of Severe Thrombotic Complications
Circuit clot leading to circuit change
Need for thrombectomy
Ischemic stroke
Distal thromboembolism
D. PCPC and POPC scores
Pediatric Cerebral Performance Category (PCPC) Scale (53)
Score 
Category 
Description 
1 
Normal 
At age-appropriate level; school-age child attends 
regular school 
2 
Mild disability 
Conscious, alert, able to interact at age-appropriate 
level; regular school, but grades perhaps not age-
appropriate, possibility of mild neurologic deficit 
3 
Moderate disability 
Conscious, age-appropriate independent activities 
of daily life; special education classroom and/or 
learning deficit present 
4 
Severe disability 
Conscious, dependent in others for daily support 
because of impaired brain function 
5 
Coma or vegetative state 
Any degree of coma, unaware, even if awake in 
appearance, 
without 
interaction 
with 
the 
environment; no evidence of cortex function; 
possibility 
for 
some 
reflexive 
response, 
spontaneous eye-opening, sleep-wake cycles 
6 
Brain death/death 
Brain death, death 
 
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Pediatric Overall Performance Category (POPC) Scale (53)
Score 
Category 
Description 
1 
Good overall performance 
PCPC 1; healthy, alert, and capable of normal 
activities of daily life 
2 
Mild overall disability 
PCPC 2; possibility of minor physical problem that 
is still compatible with normal life 
3 
Moderate overall disability 
PCPC 3; possibility of moderate disability from non 
cerebral systems dysfunction alone or with cerebral 
dysfunction; performs independent activities of daily 
life but disabled for competitive performance at 
school 
4 
Severe overall disability 
PCPC 4; possibility of severe disability from non 
cerebral systems dysfunction alone or with cerebral 
dysfunction; conscious but dependent on others for 
activities of daily living support 
5 
Coma or vegetative state 
Any degree of coma, unaware, even if awake in 
appearance, 
without 
interaction 
with 
the 
environment; no evidence of cortex function; 
possibility 
for 
some 
reflexive 
response, 
spontaneous eye-opening, sleep-wake cycles 
6 
Brain death/death 
Brain death, death 
 
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E. Worksheet for Site Assessment of Severe Bleeding/Clotting Events 
Severe Bleeding and/or Severe Clotting Evaluation Form
Severe clotting:
 
Description of the clotting event: 
_____________________________________________________________________________________
_____________________________________________________________________________________
 
Circuit clot leading to circuit change: 
yes
no
Clot that required thrombectomy: 
yes
no
Ischemic stroke: 
yes
no
Clot that caused distal thromboembolism: 
yes
no
 
Severe bleeding:
 
Description of the bleeding event: 
_____________________________________________________________________________________
_____________________________________________________________________________________
1) Bleeding that leads to at least one organ dysfunction, using PELOD-2 score criteria of organ 
dysfunction 
•
Previous day’s PELOD-2 score: ___ (bit.ly/pelod2)
•
Bleeding day’s PELOD-2 score: ___
•
Change in PELOD-2 score (Bleeding-Previous): ___
•
Change in PELOD-2 score associated with the bleeding, in absence of 
other causes (e.g. natural progression of the underlying condition, 
suboptimal ECMO support, etc.): 
yes
no
•
Was there a bleeding event that led to organ dysfunction? yes
no
 
2) Bleeding that leads to hemodynamic instability, defined as an increase in heart rate by > 20% from 
baseline or a decrease in mean arterial pressure by > 20% from baseline (i.e., prior to bleeding event) 
•
Baseline heart rate: ___/min
•
Heart rate during bleeding event: ___/min
•
Change in heart rate (100% x (bleeding-baseline)/baseline): ___% 
•
Change in heart rate associated with the bleeding, in absence of other causes (e.g. 
medication, CRRT, etc.): 
yes
no
•
Baseline mean arterial pressure: ___
•
Mean arterial pressure during bleeding event: ___
•
Change in mean arterial pressure (bleeding-baseline/baseline): ___% 
 
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ECMO Hemostatic Transfusions in Children (ECSTATIC)
Study Protocol – version 1.4 – April 22, 2024
57
•
Change in mean arterial pressure associated with the bleeding, in absence of other causes 
(e.g. medication, CRRT, etc.): 
yes
no
•
Was there a bleeding event that led to hemodynamic instability? yes
no
 
3) Bleeding leading to a drop in hemoglobin > 20% within 24 hr
•
Baseline hemoglobin: ___
•
Hemoglobin during bleeding event: ___
•
Change in hemoglobin (100% x (bleeding-baseline)/baseline): ___%
•
Change in hemoglobin associated with the bleeding, in absence of other causes (e.g. 
hemodilution due to boluses of saline, fluid overload, etc.): yes
no
•
Was there a bleeding event that led to a drop in hemoglobin by > 20%? 
yes
no
4) Quantifiable bleeding ≥ 5 mL/kg/hr for ≥ 1hr (e.g., chest tube) 
•
Quantifiable bleeding: ___ mL/kg/hr
•
Quantifiable volume from chest-tube or drain believed to be bleeding (i.e. not 
serosanguinous): 
yes
no
•
Was there a quantifiable bleeding ≥ 5 mL/kg/hr for ≥ 1hr? 
yes
no
 
5) Intraspinal bleeding leading to loss of neurologic function below the lesion, nontraumatic intra- 
articular bleeding leading to decreased range of movement, or intraocular bleeding leading to impaired 
vision 
•
Intraspinal bleeding leading to loss of neurologic function below the lesion: yes
no
•
Intra-articular bleeding leading to decreased range of movement: 
yes
no
•
Intraocular bleeding leading to impaired vision: 
yes
no
•
Was there a bleeding event that led to intraspinal, intra-articular, or intraocular bleeding? 
yes
no
6) New and/or expanding intracranial hemorrhage as evidenced by CT, head ultrasound or MRI
yes
no
Epidural
yes
no
Subdural 
yes
no
Intraventricular
yes
no
Intraparenchymal
yes
no
 
IRB APPROVED
05/15/2024

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Study Protocol – version 1.4 – April 22, 2024
58
F. Platelet Transfusion Strategy Adherence (Compliance)
The sixth feasibility criterion’s outcome is defined for a combination of platelet-count level and 
platelet-transfusion-level observations. Adherence is indicated by having an appropriately dosed 
platelet transfusion within 12 hours after a below-threshold platelet count, and by not having any 
platelet transfusion when there is no below-threshold platelet count. In general, there is one 
adherence/non-adherence determination per platelet count (although some are excluded, and 
some additional determinations can apply when a transfusion occurs without any platelet count 
in the previous 12 hours).
Platelet Transfusion
Yes
No
Excluded 
transfusion(s) 
was the only 
transfusion 
within 12 
hours after 
platelet counta
Yes
Adherenceb,c
Non-adherencec
Adherence
No
Non-adherenceb
Adherence
ignored
Excluded 
platelet countd
Non-adherenceb
ignored
Ignored
Platelet count 
below threshold
a Platelet transfusions are excluded if any of the following apply: transfusion start time before 
randomization or after the time the strategy was permanently terminated; transfusion start time 
during a period of temporary suspension
b The transfusion is considered non-adherent if an included (i.e., not excluded) platelet 
transfusion did not have a below-threshold platelet count as the most recent platelet count, or if 
there was an absence of a platelet count within 12 hours prior to the transfusion start time, or 
the most-recent platelet count was at or above the threshold.
c The platelet count is considered non-adherent if an included (i.e., not excluded) platelet count 
was below the threshold and there was no platelet transfusion (not known to be at an 
inappropriate dose) within 12 hours after the platelet count.
d Platelet counts are excluded if any of the following apply: was more than 12 hours pre-
randomization; was after the time the strategy was permanently terminated; was prior to 
randomization, and before randomization had a transfusion and/or another pre-randomization 
platelet count; was a below-threshold count but within 12 hours after the blood sample was 
collected, and prior to a subsequent platelet transfusion, there was either a platelet count at or 
above the threshold or the transfusion strategy was permanently terminated; was a platelet 
count at or above the threshold but within 12 hours and prior to a blood transfusion, there was 
either a below-threshold platelet count or the transfusion strategy was permanently terminated.
Calculation: Adherence % = 100% * (Adherences)/(Adherences + Non-adherences)
Feasibility Criterion 6 Threshold: Adherence > 90%
 
IRB APPROVED
05/15/2024

Statistical Analysis Plan
Protocol Title: ECMO Hemostatic Transfusions in Children
Protocol Version and Date: 1.3.1; July 14, 2023 (BRANY IRB approved July 19, 2023)
SAP Authors: Bradley J. Barney, PhD (with reuse of protocol and DCC template language)
SAP Version and Date: 1.0; November 9, 2023
CONFIDENTIAL
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ECSTATIC: Statistical Analysis Plan
Page ii
Approvals:
Approved By:
Name
Title
Date
Name
Title
Date
Name
Title
Date
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Sutdy PI
Study PI
11/10/2023
Co-I, 
Statistician
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Contents
Abbreviations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
v
1
PREFACE
1
1.1
Purpose of SAP . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
1
1.2
Auxiliary/Other Documents . . . . . . . . . . . . . . . . . . . . . . . . . . .
1
2
STUDY OBJECTIVES AND OUTCOMES
1
2.1
Study Objectives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
1
2.1.1
Primary Objective(s) . . . . . . . . . . . . . . . . . . . . . . . . . . .
1
2.2
Study Outcomes
. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
2
2.2.1
Primary Outcome(s) . . . . . . . . . . . . . . . . . . . . . . . . . . .
2
2.2.2
Secondary Outcome(s) . . . . . . . . . . . . . . . . . . . . . . . . . .
2
2.2.3
Tertiary Outcome(s)
. . . . . . . . . . . . . . . . . . . . . . . . . . .
6
2.2.4
Additional Outcome(s) . . . . . . . . . . . . . . . . . . . . . . . . . .
7
2.2.5
Safety Outcome(s)
. . . . . . . . . . . . . . . . . . . . . . . . . . . .
8
3
STUDY DESIGN AND METHODS
9
3.1
Overall Study Design . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
9
3.2
Method of Treatment Assignment and Randomization . . . . . . . . . . . . .
9
3.2.1
Delivery of Randomization and Emergency Backup
. . . . . . . . . .
10
3.2.2
Handling of Incorrect Randomization in Study Analyses and Reports
10
3.3
Treatment Masking (Blinding) . . . . . . . . . . . . . . . . . . . . . . . . . .
10
3.4
Study Intervention Compliance
. . . . . . . . . . . . . . . . . . . . . . . . .
10
4
STUDY SUBJECTS AND ANALYSIS POPULATIONS
11
4.1
Eligibility
. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
11
4.2
Populations for Analyses . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
11
4.2.1
Screening Population . . . . . . . . . . . . . . . . . . . . . . . . . . .
11
4.2.2
Inclusion Population . . . . . . . . . . . . . . . . . . . . . . . . . . .
11
4.2.3
Eligible Population . . . . . . . . . . . . . . . . . . . . . . . . . . . .
11
4.2.4
Approached Population . . . . . . . . . . . . . . . . . . . . . . . . . .
12
4.2.5
Consented Population
. . . . . . . . . . . . . . . . . . . . . . . . . .
12
4.2.6
Intention-to-Treat Population . . . . . . . . . . . . . . . . . . . . . .
12
4.2.7
Safety Population . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
12
5
GENERAL ISSUES FOR STATISTICAL ANALYSES
13
5.1
Analysis Software . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
13
5.2
Methods for Withdrawals, Missing Data, and Outliers . . . . . . . . . . . . .
13
5.3
Multiple Comparisons and Multiplicity . . . . . . . . . . . . . . . . . . . . .
13
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5.4
Planned Subgroups, Interactions, and Covariates . . . . . . . . . . . . . . . .
14
5.5
Derived and Computed Variables
. . . . . . . . . . . . . . . . . . . . . . . .
14
5.6
Independent Review
. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
14
6
INTERIM ANALYSES
15
6.1
Frequency of and Timepoints for Interim Analysis . . . . . . . . . . . . . . .
15
6.2
Stopping Rules for Interim Efficacy Analysis . . . . . . . . . . . . . . . . . .
15
6.3
Futility Monitoring in the Interim Analysis . . . . . . . . . . . . . . . . . . .
15
6.4
Early Trial Termination for Safety . . . . . . . . . . . . . . . . . . . . . . . .
15
6.5
Subgroups in the Interim Analysis . . . . . . . . . . . . . . . . . . . . . . . .
17
6.6
Blinding in the Interim Analysis . . . . . . . . . . . . . . . . . . . . . . . . .
17
7
PLANNED ANALYSES
17
7.1
Description of Participant Characteristics . . . . . . . . . . . . . . . . . . . .
17
7.2
Primary Outcome Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . .
17
7.2.1
Primary Analysis for Aim 1 . . . . . . . . . . . . . . . . . . . . . . .
18
7.2.2
Additional Analyses
. . . . . . . . . . . . . . . . . . . . . . . . . . .
18
7.3
Secondary Outcomes Analyses (Analyses for Aim 2) . . . . . . . . . . . . . .
19
7.3.1
Primary Analysis for Aim 2 . . . . . . . . . . . . . . . . . . . . . . .
19
7.4
Tertiary Outcome Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . .
19
7.5
Additional Outcome Analyses . . . . . . . . . . . . . . . . . . . . . . . . . .
19
7.6
Technical Approaches for Subgroup Analyses . . . . . . . . . . . . . . . . . .
20
7.7
Safety Analyses . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
20
8
SAMPLE SIZE DETERMINATION
21
9
References
22
Appendix: INTERIM STOPPING RULE OPERATING CHARACTER-
ISTICS
23
A Interim stopping rule considerations
23
B Determining the appropriate p-value threshold for each interim stopping
rule analysis
24
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Abbreviations
Abbreviation
Definition
AIC
Akaike Information Criterion
APPROACHED
Approached Population; see Section 4.2
BP
Blood Pressure
CAR(1)
Continuous analog of first-order AutoRegressive process
CONSENTED
Consented Population; see Section 4.2
CRF
Case Report Form
CS
Compound Symmetry
CT
Computerized Tomography
DCC
Data Coordinating Center
df
degrees of freedom
DSMB
Data and Safety Monitoring Board
ECMO
ExtraCorporeal Membrane Oxygenation
eCRF
Electronic Case Report Form
ECSTATIC
ECMO Hemostatic Transfusions in Chldren
ELIGIBLE
Eligible Population; see Section 4.2
ENROLLED
Enrolled Population; see Section 4.2
gm
gram(s)
hr
hour(s)
HR
Heart Rate
ITT
Intention-To-Treat
kg
kilogram(s)
L
liter
mL
milliliter
NHLBI
National Heart, Lung, and Blood Institute
PICU
Pediatric Intensive Care Unit
REDCap
Research Electronic Data Capture
REML
Restricted Maximum Likelihood (also known as Residual Maximum
Likelihood)
(S)AE
(Serious) Adverse Event
SAFETY
Safety Population
SAP
Statistical Analysis Plan
SCREENING
Screening Population; see Section 4.2
SE
Secondary Endpoint (SE1=Secondary Endpoint #1, etc.)
V-A
Veno-Arterial
vs
versus
V-V
Veno-Venous
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1
PREFACE
1.1
Purpose of SAP
This Statistical Analysis Plan (SAP) describes the planned analysis and reporting for the
protocol: ECMO Hemostatic Transfusions in Children.
The structure and content of this SAP provides sufficient detail to meet the requirements
and standards set by the Data Coordinating Center (DCC).
1.2
Auxiliary/Other Documents
The following documents were reviewed in preparation of this SAP:
 Protocol: ECMO Hemostatic Transfusions in Children.
 Case Report Forms (CRFs) for the ECSTATIC protocol
The reader of this SAP is encouraged to read the protocol for details on the conduct of this
study, and the operational aspects of clinical assessments.
The purpose of this SAP is to outline the planned analyses to be completed for the EC-
STATIC trial. The planned analyses identified in this SAP will be included in future study
abstracts and manuscripts. Also, exploratory analyses not necessarily identified in this SAP
may be performed. Any post hoc, or unplanned, analyses not explicitly identified in this
SAP will be clearly identified as such in any published reports from this study.
It is possible that, due to updates or identification of errors in specific statistical software
discussed in this SAP, the exact technical specifications for carrying out a given analysis may
be modified. This is considered acceptable as long as the original, prespecified statistical
analytic approach is completely followed in the revised technical specifications.
2
STUDY OBJECTIVES AND OUTCOMES
2.1
Study Objectives
2.1.1
Primary Objective(s)
The primary objectives of the ECSTATIC trial are:
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 Aim 1: to demonstrate a separation between the LOWER and HIGHER platelet
threshold transfusion arms. Specifically, we will demonstrate a difference in the average
pre-transfusion platelet count between the two arms from randomization to when the
treatment strategy was permanently discontinued. In addition, we will estimate the
difference in average daily platelet dose during the same period, which we anticipate
will be a difference of at least 20% between the groups randomized to each threshold.
 Aim 2: to demonstrate the ability for observed endpoints to satisfy various prespecified
feasibility thresholds. Specifically, there are 10 different criteria and we will show that
at least 5 of the 10 thresholds are satisfied by the estimated endpoints.
 Aim 3: to provide additional information on the epidemiology of a novel composite
outcome of progression to severe bleeding and/or severe thrombosis, to incorporate in
the design of the full RCT. Specifically, we will demonstrate that the estimated rate
of adjudication by an independent panel is more than 90% of the enrolled subjects for
whom the outcome was reported by the site as having occurred.
2.2
Study Outcomes
2.2.1
Primary Outcome(s)
The primary outcome for Aim 1, hereafter primary outcome, is the platelet count imme-
diately (or as recently as available) before a transfusion. Note that this outcome is defined
for each transfusion, and as such, participants may contribute multiple values.
Though not considered primary outcomes, there are two outcomes that are especially related
to the primary outcome which are of additional interest: the total platelet transfusion dose
while on ECMO in mL/kg/run, and the transfusion dose in mL/kg/hr (or its redundant
value, in mL/kg/day, obtained by multiplying the dose in mL/kg/hr by 24). Refer to Sec-
tion 5.5 for details of the mL/kg/run and mL/kg/hr calculations.
For safety monitoring, there is a primary safety outcome, (occurrence of progression to
severe bleeding, severe thrombosis, or death, in the acute follow-up period more precisely
defined in Section 2.2.5). To avoid confusion, any reference to the primary safety outcome
contains the qualifier “safety.”
2.2.2
Secondary Outcome(s)
The secondary endpoints (SEs) are: observed values of estimated separation in mean pre-
transfusion platelet counts; eligibility rate relative to predicted rate; approach for consent
percentage; consent percentage; randomization of consented patients percentage; transfu-
sion compliance rate; transfusion strategy suspension percentage; withdrawal and or lost-
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to-follow-up percentage; violation of local non-study-specific circulatory support protocol
percentage; and adjudication rate of progression to severe bleeding/thrombotic outcomes.
Each endpoint is based on an underlying outcome and has an associated feasibility threshold.
To support a larger trial, at least five out of ten feasibility criteria must be
met. The underlying outcomes and thresholds for these 10 secondary endpoints (SE1–SE10)
criteria are defined in the next paragraphs.
(SE1) Estimated separation in pre-transfusion platelet counts
 Outcome: Uses the primary outcome, pre-transfusion platelet count, defined in Sec-
tion 2.2.1
 Endpoint: Model-based estimated effect of transfusion strategy; see Section 7.2 for
modeling details
 Feasibility threshold: Model-based estimated difference > 30 × 109 cells/L
(SE2) Eligibility rate relative to predicted eligibility rate
 Outcome: For each patient screened (see definition in Section 4.2.1), it will be deter-
mined whether the patient was indicated in the study database as meeting all study
eligibility criteria, did not meet all study eligibility criteria, or whether screening was
indeterminate
 Endpoint: The observed eligibility rate as a percentage of the predicted eligibility rate.
That is,
100% × observed eligibility rate
predicted eligibility rate,
where the predicted eligibility rate is 7.9 patients per site per year, and the observed el-
igibility rate per site per year will be calculated as (# in ELIGIBLE population)/(total
# of site-years enrollment was open). The ELIGIBLE population is defined in Sec-
tion 4.2.3. For each site, the enrollment duration is the number of calendar days from
the site being activated for enrollment to the site being closed to enrollment, inclusive,
divided by 365.25 (approximate # of days per year)
 Feasibility threshold: > 50%
(SE3) Approach for consent percentage
 Outcome: For each patient in the ELIGIBLE population (see Section 4.2.3), whether
the patient was in the APPROACHED population (approached for study participation
within 24 hours of cannulation; see also Section 4.2.4)
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 Endpoint: 100% × (# in APPROACHED population) / (# in ELIGIBLE population)
 Feasibility threshold: > 90%
(SE4) Consent percentage
 Outcome: For each patient in the APPROACHED population (see Section 4.2.4),
whether the person was consented to study participation (see also Section 4.2.5 for a
description of the CONSENTED population)
 Endpoint: 100% × (# in CONSENTED population) / (# in APPROACHED popu-
lation)
 Feasibility threshold: > 50%
(SE5) Randomization of consented patients percentage
 Outcome: For each patient in the CONSENTED population, whether the person was in
the ITT population (i.e., consented and randomized; equivalently, in the ENROLLED
population; see Section 4.2.6)
 Endpoint: 100% × (# in ITT population) / (# in CONSENTED population)
 Feasibility threshold: > 66% (previous data indicate approximately one-third of pa-
tients bleed within 24 hours of cannulation)
(SE6) Transfusion compliance rate
 Outcome: This is a transfusion-level outcome for the ITT population, such that some
patients could have multiple outcomes and others could have no outcomes. Any trans-
fusions that were performed under a period of temporary suspension will be ignored.
Among the remaining transfusions performed between randomization and when the
treatment strategy was permanently discontinued, the transfusion will be considered
compliant if and only if (a) the most recent nonmissing pre-transfusion platelet count
(obtained no more than 12 hours before the transfusion, and not obtained prior to
an earlier transfusion) was below the assigned threshold AND (b) the site principal
investigators agree the transfusion dose was the proper dose in use at the site (e.g., 10
mL/kg if that is the local guideline and the same guideline applies to each arm of the
study); if the pre-transfusion platelet count is not available, it will be assumed that the
transfusion was not compliant. Also, if a transfusion was indicated per the threshold
(i.e., the platelet count was below the threshold) but no transfusion was given within
12 hours, this will also be considered as an outcome and automatically be regarded as
noncompliance
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 Endpoint: 100% × (# of compliant transfusions not during a temporary suspension)/
(# of transfusions not during a temporary suspension + number of transfusions that
should have been given but were not)
 Feasibility threshold: >90%
(SE7) Transfusion strategy suspension (not due to preparation for decannulation)
percentage
 Outcome: For each patient in the ITT population (see Section 4.2.6), whether the
patient had at least one temporary suspension of the transfusion threshold not due to
preparation for decannulation before the permanent discontinuation of the treatment
strategy. A temporary suspension of the transfusion threshold will not require a trans-
fusion in excess of the threshold to have been given during the suspension period for
it to be counted as a suspension
 Endpoint: 100% × (# in ITT population with at least one temporary suspension
of the transfusion threshold not due to preparation for decannulation) / (# in ITT
population)
 Feasibility threshold: < 10%
(SE8) Withdrawal and/or lost-to-follow-up percentage
 Outcome: For each patient in the ITT population, whether the patient was withdrawn
from study participation by 90 days post-randomization and/or had an unknown mor-
tality status for the timepoint of 90 days after randomization. The 90-day vital status
will be considered known if the 90-day communication for mortality assessment is com-
pleted within ±10 days, inclusive, of 90 days after randomization, or if the patient was
known to have already died (e.g., died while on ECMO) such that the 90-day follow-up
mortality assessment is unnecessary
 Endpoint: 100% × (# in ITT population who were withdrawn from the study by 90
days after randomization or had unknown 90-day mortality) / (# in ITT population)
 Feasibility threshold: < 6%
(SE9) Percentage with any violations of local non-study-specific circulatory sup-
port protocols
 Outcome: For each patient in the ITT population, whether the patient had any re-
ported instances of non-adherence to the site’s non-study-specific circulatory support
strategies. For this outcome, any instances of platelet transfusions above the random-
ized threshold are immaterial; on the other hand, for example, if the anticoagulation
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strategy in place at the site was not followed, this is considered a protocol violation for
this outcome. The site PIs will determine whether or not local protocols were followed
 Endpoint: 100% × (# of ITT patients with any reported violations of local non-study-
specific circulatory support protocols)/(# of ITT patients)
 Feasibility threshold: < 5%
(SE10) Adjudication percentage of progression to severe bleeding/thrombotic
outcomes
 Outcome: This is defined for each patient in the ITT population (Section 4.2.6) for
whom there was any site-reported instance of progression to severe bleeding and/or
thrombosis in the acute follow-up period (the acute follow-up period consists of
the time from randomization through 24 hours past when the transfusion strategy was
last applicable). Because any site-reported severe bleeding or severe prognosis is cause
to terminate use of the transfusion strategy for that patient, we focus on whether the
initial such event can be adjudicated, whether or not additional distinct events were
later observed. The patient’s outcome is defined as adjudicated if the adjudication
panel reached a decision for the initial site-reported event (confirming or overturning
the site-level decision for the initial event), and defined as not adjudicated if the panel
was unable to reach either a confirmation or overturning decision for the reported event
even after requesting additional information from the site
 Endpoint: 100% × (# of ITT patients with site-reported severe bleeding/thrombotic
outcomes for which the adjudication panel was able to adjudicate the patient’s first such
reported outcome)/(# of ITT patients with site-reported severe bleeding/thrombotic
outcomes)
 Feasibility threshold: > 90%
2.2.3
Tertiary Outcome(s)
The tertiary outcome is whether the adjudication committee was able to reach an adjudi-
cation decision for progression to severe bleeding and/or a severe thrombotic event. Note
that the outcome of progression to severe bleeding and/or severe thrombosis is similar to the
primary safety outcome, but the primary safety outcome also includes mortality while the
tertiary outcome disregards mortality as a sufficient condition (though it is anticipated that
mortality may have already involved a severe bleeding and/or thrombotic event, making the
distinction less impactful). This patient-level outcome will be defined for those who have a
site-reported progression to severe bleeding and/or severe thrombosis and will use each such
patient’s first reported instance of a progression to severe bleeding or severe thrombosis.
Note that this tertiary outcome is the same as defined in SE10.
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2.2.4
Additional Outcome(s)
Additional outcomes mentioned in the protocol include the following, which include most of
the measures recommended in [1].
 Hemorrhagic complications according to the ELSO definition
 Thrombotic complications according to the ELSO definition
 Blood product exposure, defined as the total number of transfusions (red blood cell,
plasma, platelets, cryoprecipitate, fibrinogen concentrate) and other hemostatic ther-
apies (activated Factor VII, tranexamic acid, aminocaproic acid, DDAVP, and von
Willebrand factor, etc.)
 Re-explorations and other surgical procedures to treat bleeding
 Duration of ECMO, defined as 90-day ECMO-free days (e.g., if a patient is decannu-
lated after 7 days, the 90-day ECMO-free days is 83 days; whereas if a patient dies on
ECMO after 7 days, the 90-day ECMO-free days is 0 days)
– This is defined based on calendar days for days 1–90, with day 1 being the day
of randomization and day 90 being 89 calendar days after randomization, and
because every randomized patient will be on ECMO on day 1, the range for this
outcome is 0–89
 Length of Pediatric Intensive Care Unit (PICU) stay, defined as the 90-day PICU-free
days (as with ECMO-free days, using calendar days and with possible range of 0–89)
 Discharge Pediatric Overall Performance Category (POPC)
 Discharge Pediatric Cerebral Performance Category (PCPC)
 Mortality on ECMO
 Mortality within 24 hours of the treatment strategy being permanently discontinued
 Mortality at 28 days (obtained on or after 28 days after randomization, unless known
death before)
 Mortality at 90 days (obtained on or after 80 days after randomization, unless known
death before)
 Number of temporary suspensions of the transfusion strategy
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2.2.5
Safety Outcome(s)
Many safety outcomes are obtained during the acute follow-up period, defined as the period
beginning with randomization and ending 24 hours after the permanent discontinuation of
the transfusion strategy. The transfusion strategy for a randomized patient is permanently
discontinued at the earliest of any of the following five events: (1) per site report, progression
to severe bleeding, according to the BASIC definition, and/or severe clotting, and/or death;
(2) decannulation from ECMO; (3) twenty-one days post-randomization (as the progression
to new bleeding and new thrombosis has plateaued by day 21 and ≈95% of the patients will
be off ECMO by then); 4) temporary suspension lasting more than 24 hours; or 5) study
withdrawal.
The primary safety outcome is defined at the patient level for all patients in the ITT popula-
tion. It is defined as the occurrence of any of the following during the acute follow-up period:
progression to severe bleeding (defined below); progression to severe thrombosis (defined be-
low); death. The outcome is initially site-reported but then is to be adjudicated by the
adjudication panel. If the panel determines the outcome criteria were not met, the outcome
will be considered not to have occurred. Otherwise, if the panel confirms the criteria were
met, or if the panel is unable to reach a determination, the primary safety outcome will be
considered to have occurred.
Using the BASIC definition [2], severe bleeding will be defined as:
1. Bleeding that leads to at least one organ dysfunction, using PELOD-2 score criteria of
organ dysfunction
2. Bleeding that leads to hemodynamic instability, defined as an increase in heart rate
(HR) by > 20% from baseline or a decrease in blood pressure (BP) by > 20% from
baseline (i.e., prior to the bleeding event)
3. Bleeding leading to a drop in hemoglobin > 20% within 24hr
4. Quantifiable bleeding ≥5 mL/kg/hr for ≥1hr (e.g., chest tube)
5. Intracranial hemorrhage on computerized tomography (CT) scan or head ultrasound
6. Intraspinal bleeding leading to loss of neurologic function below the lesion, nontrau-
matic intra-articular bleeding leading to decreased range of movement, or intraocular
bleeding leading to impaired vision.
Severe thrombotic events will be defined according to the NHLBI Hemostasis Clinical
Trial Outcomes Working Group as any of the following, as diagnosed clinically: circuit clot
requiring circuit change; need for thrombectomy; ischemic stroke; or distal thromboem-
bolism. [3]
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Other patient-level safety outcomes include:
 the occurrence of any adverse event (AE) that is non-serious but at least possibly
related to the platelet transfusion strategy
 the occurrence of any serious adverse event (SAE)
 the occurrence of any SAE, or of any non-serious AE at least possibly related to the
platelet transfusion strategy (i.e., either of the two outcomes mentioned immediately
above)
 the occurrence of any serious adverse event at least possibly related to the platelet
transfusion strategy.
As with the primary safety outcome, the period during which new adverse events are to
be identified for subsequent entry in the study database is the acute follow-up phase from
randomization through 24 hours after the transfusion strategy was permanently discontinued.
Note that adverse events are recorded if they are non-serious but deemed at least possibly
related to the study intervention (transfusion strategy), or if they are serious regardless of
study-relatedness.
3
STUDY DESIGN AND METHODS
3.1
Overall Study Design
The ECSTATIC trial is a two-arm, 1:1 randomized controlled trial, to compare lower (50 ×
109 cell/L) and higher (90 × 109 cell/L) platelet thresholds for transfusions in children on
ECMO. Eligible and consented participants are to have transfusions managed using the
randomly assigned strategy from the time of their randomization until one of five prespecified
criteria are met which permanently discontinues the strategy, with allowances for temporary
suspension of the assigned protocol as described in the study protocol.
Because this is
a feasibility trial, analyses focus on the ability to demonstrate adherence to the protocol
as captured by separation in the average pre-transfusion platelet counts corresponding to
transfusions (Aim 1) and on summaries of ten criteria taken as indicators of the feasibility to
conduct a larger trial designed to assess efficacy of the transfusion threshold strategy (Aim
2). The larger trial is proposed to use a novel composite outcome of progression to severe
bleeding or severe prognosis (see SE10 in Section 2.2.2 or Section 2.2.3), and added attention
is given to the adjudication of this outcome (Aim 3).
3.2
Method of Treatment Assignment and Randomization
This study’s randomization scheme is 1:1 allocation to the lower or higher transfusion thresh-
old. Randomization will use random block sizes of 2, 4, or 6, and be stratified by age (≤28
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days or > 28 days). The randomization sequences will be generated using R version 4.2.1
or later, and the randomization sequences will be loaded to the study’s Research Electronic
Data Capture (REDCap) database.
3.2.1
Delivery of Randomization and Emergency Backup
When a research coordinator or other designated personnel is enrolling a participant, the
study team member will use the randomization module in REDCap to make the treatment
assignment. In case of emergency because REDCap is not functioning, each site will be
provided one backup randomization envelope containing a treatment assignment. It is very
unlikely REDCap will not be properly functioning at the time a patient is enrolled, and
therefore one backup envelope per site should be more than sufficient.
3.2.2
Handling of Incorrect Randomization in Study Analyses and Reports
We assume that misrandomization will be rare for this trial. For intention-to-treat analyses,
the participants will be analyzed according to the assigned treatment arm, even if it was
later determined that the incorrect randomization stratum (defined by age) was used.
3.3
Treatment Masking (Blinding)
The clinical team will be unblinded because they need to know the strategy to which they
are to adhere. The Data Coordinating Center (DCC), including the statistical team, will
also be unblinded. Otherwise, the steering committee will remain blinded except as needed
at their own sites to provided clinical care for participants they are treating. The trial’s
Data and Safety Monitoring Board (DSMB) will not be blinded.
3.4
Study Intervention Compliance
At the transfusion level, a transfusion is considered to be compliant if it was below the as-
signed threshold (with the count obtained less than 12 hours previously and not obtained
before a prior transfusion) and at the dose level to be used at the local site; it is considered
to be non-compliant otherwise (see also SE6 in Section 2.2.2). As a supplementary mea-
sure of study compliance, we will distinguish between transfusions that occurred during a
period of temporary suspension and those that did not occur during such a period. At the
individual level, the person is considered as having been managed in compliance with the
assigned treatment if and only if there were no non-compliant transfusions given, ignoring
any transfusions given during a period of temporary suspension. Any missed transfusions
(i.e., platelet counts below the threshold for which there were no transfusions within 12 hours
of the platelet count) are considered as noncompliance.
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An additional measure of compliance is whether the patient was treated according to in-
stitutional protocols for aspects of circulatory support other than the platelet transfusion
strategy (e.g., per the site’s anticoagulation strategy). The determination of whether local
protocols were followed will be made by the study PIs (Drs. Nellis and Karam). See also
SE9 in Section 2.2.2.
4
STUDY SUBJECTS AND ANALYSIS POPULA-
TIONS
4.1
Eligibility
This study is among children on ECMO. The exact eligibility criteria are specified in the
study protocol.
4.2
Populations for Analyses
4.2.1
Screening Population
The SCREENING population is all patients on ECMO under age 18.
4.2.2
Inclusion Population
The INCLUSION population is all patients who meet all study inclusion criteria and were
entered into the study database.
4.2.3
Eligible Population
The ELIGIBLE population is the subset of the INCLUSION population who were deter-
mined to meet all eligibility criteria. Note that this will include participants who provided
consent but were later found to be ineligible. For the purposes of evaluating the exclusion
criterion “on ECMO for > 24 hours at time of enrollment,” an unconsented patient who
otherwise satisifies all eligibility criteria will be considered eligible if the person would have
been eligible to be randomized within 24 hours of cannulation, had the patient been con-
sented. Furthermore, the exclusion criterion “decision to withdraw or withhold some critical
care or interventions” will be assumed to not apply unless such a decision was known to be
in place (e.g., a participant cannot withdraw if the participant was never consented, but for
purposes of determining missed eligibles the person would be counted as eligibile if all other
criteria would indicate eligibility).
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4.2.4
Approached Population
The APPROACHED population is the subset of the ELIGIBLE population for whom the
parents were approached for consent within 24 hours of cannulation.
4.2.5
Consented Population
The CONSENTED population is the subset of the ELIGIBLE population for whom informed
consent for study participation was provided.
4.2.6
Intention-to-Treat Population
The Intention-to-Treat (ITT) population is the subset of the CONSENTED population who
were randomized to one of the study’s two transfusion thresholds. For the ITT population,
the treatment arm is the arm to which the participant was assigned, even if this strategy
was not adhered to or there was a randomization error because the wrong randomization
stratum was used. The ITT population will equivalently be regarded as the ENROLLED
population. Note that a participant who consented and was randomized but was later found
to be ineligible will still be considered part of the ITT population for analysis and safety
assessment purposes. Although we note that the primary analysis of the primary outcome
requires being in the ITT population as a necessary condition, not all ITT participants will
necessarily contribute to the primary analysis because the primary outcome is a transfusion-
level outcome and as such is not applicable if an ITT participant had no transfusions during
the acute follow-up period. (Additionally, note that missing primary outcomes from ITT
participantes with transfusions will not be imputed.)
4.2.7
Safety Population
The SAFETY population is comprised of all patients in the ITT population. In the unlikely
event that a participant is randomized but did not have informed consent provided, this
person will be added to the SAFETY population. All individuals in the SAFETY population
will be analyzed according to the assigned treatment arm. This population will be used for
adverse event summaries and for the primary safety outcome’s interim and end-of-study
analyses.
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5
GENERAL ISSUES FOR STATISTICAL ANALY-
SES
5.1
Analysis Software
Analysis will be performed using SAS® Software version 9.4 or later when feasible. Other
software packages, including R and StatXact®, may be used for particular specialized pro-
cedures.
5.2
Methods for Withdrawals, Missing Data, and Outliers
Participants who withdraw from the study intervention should continue to be followed for
assessment of safety outcomes, if allowed. However, outcomes such as the primary outcome
of pre-transfusion platelet counts and adherence to study intervention will be ignored from
analyses beginning at the time of the withdrawal.
Because the outcomes generally relate to adherence or outcomes that should have no miss-
ingness (e.g., whether or not a patient consented), it is not expected that missing data will
be problematic for the main analyses. There is naturally some protection against detrimen-
tal effects due to missing data for model-based analyses that incorporate random effects or
other forms of within-subject correlation, such as for the primary outcome.
Outliers may affect the primary analysis because the primary analysis of Aim 1 is mean-
based. Pre-transfusion platelet counts that appear to be outliers may be queried to ascertain
if the values were correctly recorded. If it is determined that the values were not correctly
recorded, they will be corrected if the correct values are available. If it is determined the
values were not correctly recorded but the correct values are unavailable, the platelet counts
will be regarded as missing for the primary Aim 1 analysis of separation in mean platelet
counts.
5.3
Multiple Comparisons and Multiplicity
The first objective has one primary analysis: separation in the average pre-transfusion
platelet counts between treatment strategies. There is an associated one-sided test to be
conducted at α = 0.025. This test provides the key p-value for purposes of the Aim 1 feasi-
bility assessment.
Other tests of the feasibility, including the one-sided test for separation in the average total
volumes of transfusions, are considered supplementary or exploratory and as such will not
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be adjusted for multiple comparisons.
The early stopping rule for safety allows multiple tests and as such will account for multi-
plicity as described in Section 6.
5.4
Planned Subgroups, Interactions, and Covariates
Subgroups defined by the randomization strata (≤28 days vs > 28 days) will be considered.
The intended sample size does not facilitate precise subgroup inference, so subgroup analyses
will be considered as supplementary and interpreted carefully. We do not plan to allow for
interactions in exploratory models given there is limited information for estimation of even
main effects. ECMO type (Veno-Venous [V-V] vs Veno-Arterial [V-A]) may be used as a
covariate in some exploratory analyses, and site may be included as a random effect in some
exploratory analyses.
5.5
Derived and Computed Variables
Derived variables needed for analysis will be defined in a corresponding data dictionary.
Most variables of particular importance were already defined. Select variables of particular
importance are defined in the remainder of this subsection.
Additional outcomes: transfusion volumes in mL/kg/run and mL/kg/day.
These
are the total platelet transfusion doses while on ECMO in mL/kg/run, and the transfusion
dose in mL/kg/hr (or its redundant value, in mL/kg/day, obtained by multiplying the dose in
mL/kg/hr by 24). When determining the total mL/kg of the platelet transfusions, all doses
given from randomization until the permanent discontinuation of the transfusion strategy
will be counted, whether or not the dose was transfused during a temporary suspension
of the assigned strategy. The total transfusion dose measured in mL/kg/run is simply the
mL/kg during this acute follow-up phase, whereas the dose in mL/kg/day is the total dose
in mL/kg divided by the number of hours from randomization until permanent strategy
discontinuation.
5.6
Independent Review
All statistical analyses for primary reporting of trial results will be independently verified
through dual programming and/or code review.
Two statisticians will each program all
datasets and trial results analyses for the DSMB reports and the primary manuscript and
the results will be compared.
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6
INTERIM ANALYSES
6.1
Frequency of and Timepoints for Interim Analysis
There are two classes of interim analyses: a set of inferential statistical analyses for a for-
mal interim stopping rule; and largely descriptive summaries focusing on data and safety
monitoring used in reports prepared for the DSMB. The statistical testing underlying the
formal stopping rule will be conducted when every new batch of five additional participants
for whom has the site reported the primary safety outcome occurred has undergone the ad-
judication process. Details of this testing are provided in Section 6.4.
A DSMB report with safety and data completion metrics will be prepared and distributed
to the DSMB for every DSMB meeting during enrollment. Scheduled DSMB meetings are
intended to be held twice during the study: approximately 8 weeks after the tenth participant
randomization, and approximately 8 weeks after the 25th participant randomization.
6.2
Stopping Rules for Interim Efficacy Analysis
This study is not intended to provide definitive efficacy assessments, even at study conclusion.
As such, early stopping for efficacy is not a consideration.
6.3
Futility Monitoring in the Interim Analysis
Because the study is not intended to provide definitive efficacy assessments, early stopping
for the futility of demonstrating efficacy is also not a consideration.
6.4
Early Trial Termination for Safety
Tests of the primary safety outcome will be conducted from the SAFETY population (see
Section 4.2.7). Per the DSMB guidance, we increased the planned frequency of comparing
the primary safety outcome between treatment arms for the purpose of applying early trial
termination criteria. Instead of one interim analysis after 25 enrollments, testing for a differ-
ence between treatment arms in the proportion of enrolled patients experiencing the primary
safety outcome, a group sequential approach will be taken.
After the adjudication of every 5 new patients who were reported by the site to have had
the primary safety outcome, we will conduct a two-sided Fisher’s exact test using an alpha
threshold of 0.0333.1 Each test will include those with a known outcome at the time the
1This p-value threshold was chosen so that, accounting for the varying number of interim tests that
may be conducted, the overall type I error rate is close to 0.05. The SAP’s Appendix contains a detailed
motivation for the 0.0333 threshold at each interim look.
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adjudication process is completed (even if for a participant enrolled after the fifth new re-
ported event), with participants for whom the outcome has not yet been determined being
excluded until the outcome has been determined. We will test the null hypothesis that the
two treatment strategies have the same expected proportion of patients who will experience
the primary safety outcome. If a test is significant, we will stop the trial for safety reasons.
Many factors may influence exactly what the stopping rule thresholds are in the trial, such
as primary safety outcomes that are overruled by the adjudication panel (expected to be
uncommon), the underlying event rate, and the balance of randomization to treatment arms
throughout the trial.
Figure 6.4 is an illustrative example of how the rule translates to go/no-go thresholds under
one particular set of assumptions:
 At every interim look, there have been equal numbers randomized to each treatment.
 At every interim look, half of the total number of enrolled patients have experienced
the primary safety outcome (though the numbers can differ by arm)
Note that the primary safety outcome will also be tested for a difference in arms after all
participants have been enrolled, again using the 0.0333 threshold to be declared statistically
significant, unless a previous test was already significant at this threshold. However, this is
not considered an interim analysis because it is after all participants were enrolled.
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6.5
Subgroups in the Interim Analysis
Interim DSMB reports will provide information overall (and by treatment group for the
closed session). In addition, select summaries (e.g., enrollment rates) may be provided by
site to possibly inform training or monitoring efforts.
6.6
Blinding in the Interim Analysis
Interim DSMB reports will only show information by treatment arm in the closed session
report. The closed session reports will be made available to unblinded DCC statisticians and
DSMB members but not to clinical staff or study investigators.
7
PLANNED ANALYSES
7.1
Description of Participant Characteristics
Summaries of enrolled participants will be provided for baseline characteristics such as, but
not necessarily limited to, the following:
 age
 sex
 race/ethnicity
 number of platelet transfusions between cannulation and consenting process and if
those transfusions were prophylactic or therapeutic (i.e., to treat active bleeding)
 bleeding between cannulation and enrollment
Summaries of nominal variables will be comprised of frequencies and percentages. Summaries
of (quasi-)continuous variables will be comprised of means and standard deviations, and/or
medians, first quartiles, and third quartiles, as deemed appropriate. Summaries of ordinal
variables may use frequencies with percentages, means with standard deviations, or medians
with first and third quartiles, as deemed most suitable.
7.2
Primary Outcome Analysis
For the pilot study, the analysis of the primary outcome measure will be assessing the pre-
transfusion platelet count to describe the separation between the two platelet transfusion
strategies, while accounting for intra-subject correlation. For each arm, we will estimate the
mean pre-transfusion platelet count from randomization to the permanent discontinuation
of the treatment strategy.
This primary outcome will be measured for each transfusion
and represent the latest available collected platelet count that is before the transfusion
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began and not before a prior transfusion. In the event that there was no preceding platelet
count since a previous transfusion, the outcome for that transfusion will be regarded as
missing. Participants who do not have a nonmissing value for this outcome (e.g., they had no
transfusions) will not be included in the analysis of the primary outcome. Thus, the primary
outcome uses all transfusions with non-missing pre-transfusion platelet counts during the
period between randomization and the final termination of the transfusion strategy being in
effect (whether or not during a temporary suspension) from all ITT patients with at least
one such transfusion having a non-missing pre-transfusion platelet count.
7.2.1
Primary Analysis for Aim 1
The first objective is based on the primary outcome of platelet counts before transfusions.
Because we anticipate intra-subject correlation when there are multiple transfusions for a
participant, we will fit a linear model to pre-transfusion platelet counts with a fixed effect
for assigned treatment (Higher threshold, with tmt=1, vs Lower threshold, with tmt=0 ),
such that the outcome’s expected value is represented by β0 + βtmt for the higher threshold
and β0 for the lower threshold. We will account for repeated measures by considering three
covariance structures: compound symmetry (CS), continuous analog of autoregressive order
1, also known as CAR(1), and a hybrid of CS and CAR(1) achieved by including random
subject effects and CAR(1) correlations in the within-subject errors. We will use restricted
maximum likelihood (REML) for model estimation and use the model with the lowest Akaike
Information Criterion (AIC) to select the best covariance structure. Once the covariance
structure is determined, the Kenward-Roger calculation for denominator degrees of freedom
(df) will be applied in the F-test (equivalently, for the df in the t-test) of the treatment
strategy’s model coefficient. The one-sided p-value of the null hypothesis βtmt = 0 vs βtmt > 0
will be considered statistically significant if and only if this p-value < 0.025. In addition,
a two-sided 95% confidence interval for βtmt will be reported. Likewise, estimated means
for the lower threshold and the higher threshold arms will be reported as ˆβ0 and ˆβ0 + ˆβtmt,
respectively.
7.2.2
Additional Analyses
We will measure the effect of the days on ECMO, by providing summaries of the daily
platelet counts for each specific ECMO day with ≥5 patients still on ECMO, independent
of receiving a platelet transfusion. This will allow evaluating trends over time and a possible
attenuation of the separation over time. The raw summaries for such days will be reported.
As another analysis we will fit a linear model with a main linear effect for study day, a main
effect for assigned treatment, a treatment by time interaction effect, and the best-fitting
covariance structure, per AIC, among compound symmetry, CAR(1), the CS/CAR(1) hy-
brid, and subject-specific random intercepts and random time slopes. A two-df Wald test
with Kenward-Roger denominator df and REML estimation will be conducted for the null
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hypothesis that both the treatment main effect and the treatment by time interaction effect
are 0. If this test is rejected at α = 0.05, the interaction effect will be separately tested with
a two-sided alternative at α = 0.05. Then, if the 2-df test was significant but the 1-df test of
the interaction effect was not significant, the main effect of treatment will be tested against
a two-sided alternative at α = 0.05.
Furthermore, we will also estimate the difference in the median platelet transfusion dose
(in mL/kg/run) between the lower and higher platelet threshold strategy arms, and the
median daily platelet transfusion dose (in mL/kg/day, i.e., 24 × mL/kg/hr) to account for
different durations of treatment strategies. These outcomes will use two-sided nonparametric
percentile bootstrap 95% confidence intervals, based on a minimum of 10,000 bootstrap
replicates, for the difference in medians because by construction for each of these outcomes,
there is only one outcome per person. As such, repeated measures modeling is not warranted.
7.3
Secondary Outcomes Analyses (Analyses for Aim 2)
7.3.1
Primary Analysis for Aim 2
The first Aim 2 feasibility criterion, SE1, will be determined by whether the estimated
treatment effect from the model in Section 7.2.1 is > 30 × 109cells/L. The other nine Aim
2 feasibility criteria, SE2–SE10, are simple yes/no criteria which do not require statistical
modeling: simply a comparison of the estimated endpoint with the prespecified threshold.
For each criterion, the associated trial quantity will be reported along with the indication of
whether the criterion was met. The number of criteria met will also be reported.
7.4
Tertiary Outcome Analysis
The tertiary outcome is whether the adjudication committee was able to reach an adjudi-
cation decision for the outcome of progression to severe bleeding or thrombotic event for
all ITT patients for whom such progression was reported by the site. This outcome will be
summarized by frequency and percentage, overall and by assigned treatment arm.
7.5
Additional Outcome Analyses
As noted in the protocol, additional analyses will be conducted to assist with planning a
future large trial. Though it is not intended that these planning analyses be definitive given
the modest sample size in ECSTATIC, descriptive summaries of the additional outcomes will
be calculated overall and by assigned treatment arm. If there are suggested differences in
safety-related outcomes between assigned arms, the study procedures and eligibility criteria
for the larger trial might be revised. Given the nature of these additional outcome analyses
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in the ECSTATIC trial itself, it is not necessary to formally test for differences in the
additional outcomes, as the precision of estimation would likely be insufficient to overcome
issues associated with the multiplicity of testing.
7.6
Technical Approaches for Subgroup Analyses
Subgroup analyses will be considered exploratory and will be interpreted with care given
the expected small sample sizes. For all Aim 1 outcomes, we will perform two subgroup
analyses, one to evaluate the effect of V-V vs V-A ECMO, the other to evaluate neonates
vs infants and older children (≤vs > 28 days of age). For the Aim 1 model, the subgroup
analyses are intended to be based on reapplying the model with the same covariance struc-
ture already identified in the overall analysis to the subset of participants from a particular
subgroup (where subgroups are defined based on ECMO type or age). However, if estima-
tion is deemed too unwieldy for some subsets, an alternative approach will be to use all
participants in fitting one model but include ECMO type or age as covariates in the model.
For the primary outcome, a subgroup analysis by site would be expected to have very small
sample sizes that would make statistical testing suspect, and so adjustment for site as a ran-
dom effect would be considered in a supplementary analysis in place of subgroup analyses.
Because of a heightened risk that the model may not converge when including subject effects
and site effects, adjustment for site is not incorporated in the primary analysis.
For all secondary outcomes, we will perform two subgroup summaries, one to evaluate the
effect of V-V vs V-A ECMO, the other to evaluate neonates vs infants and older children
(≤vs > 28 days of age).
For outcomes that do not use hypothesis testing, descriptive summaries by site might be
reported, though they would require cautious interpretation.
7.7
Safety Analyses
The number and percentage of individuals with any reported adverse events will be summa-
rized at the end of the trial, overall and by treatment arm, for the SAFETY population. In
addition to possible end-of-trial DSMB reporting, the interim DSMB reports will summarize
adverse event rates, overall for the open session DSMB reports and also by assigned treat-
ment arm for the closed DSMB session.
End-of-study safety analyses will summarize descriptively each safety outcome. The only
planned safety hypothesis is for the primary safety outcome, which will be tested throughout
the study using the group-sequential approach described in Section 6.4, and if the null
hypothesis of no difference in proportion of patients experiencing the primary safety outcome
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was not previously rejected, the test will be conducted after all patients have had their
outcome determined, again using a nominal alpha threshold of 0.0333 for the two-sided
Fisher’s exact test so that the overall type I error rate for all sequential tests of the primary
safety outcome is approximately 0.05. (See also the Appendix.)
8
SAMPLE SIZE DETERMINATION
The trial has two primary types of formal statistical inferences: testing for separation in
the primary outcome of Aim 1—pre-transfusion platelet counts—between transfusions with
non-missing pre-transfusion platelet counts from ITT participants in the two study arms
at the end of the study, and ongoing testing for differences in the proportions of SAFETY
participants with the primary safety outcome between arms for an interim safety stopping
rule and/or end-of-study evaluation. While the study is powered for the Aim 1 primary
outcome, it is not intended to be powered for a difference in the primary safety outcome in
the ECSTATIC trial.
Salient assumptions to permit power estimation include the number of observed transfusion
days per subject, the within-subject dependence structure, the within-treatment variability,
and the difference in means between treatments. Previous data, albeit not with an enforced
low platelet threshold transfusion strategy, were used to inform the first three assumptions.
It was assumed the distribution of the number of transfusion days per subject will mimic
historical data (e.g., approximately 10%-20% will have no transfusions). It was assumed a
compound symmetry covariance structure will adequately account for within-subject depen-
dence; such a structure is induced by a mixed effects model with subject-specific random
intercepts. Fitting a model with such a covariance structure to historical data [4] yielded
an estimated autocorrelation slightly above 0.2; to be conservative the power estimation
assumed ρ = 0.3. The assumed within-treatment standard deviation, 40 × 109/L, resembles
historical data but is anticipated to be a conservative estimate for the trial.
Although the difference in platelet strategy thresholds is 40 × 109/L (i.e., 90 × 109/L vs
50×109/L), this full effect will not be realized for the mean difference because the strategies
depend on platelet counts which are not continuously observed, and as such, by the time of
measurement a patient may have a platelet count considerably below the threshold. Patients
with a platelet count under 50×109/L would be transfused under either strategy. Therefore,
a more realistic separation for the difference in means between strategies is 30 × 109/L.
Assuming pre-transfusion platelet levels will be approximately normally distributed with a
difference in means of 30 × 109/L, then with 50 patients (25 per arm) enrolled (and thus
an estimated effective sample size of 30 statistically independent observations per arm when
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accounting for patients with no transfusions, patients with multiple transfusions, and the
assumed within-subject dependence in the outcome) there would be approximately 81%
power to detect a difference in mean platelet levels at the time of transfusion.Power may be
higher due to conservative assumptions.
9
References
References
[1] Hodgson CL, Burrell AJC, Engeler DM, Pellegrino VA, Brodie D, Fan E. Core Outcome
Measures for Research in Critically Ill Patients Receiving Extracorporeal Membrane Oxy-
genation for Acute Respiratory or Cardiac Failure: An International, Multidisciplinary,
Modified Delphi Consensus Study*, Crit Care Med, 2019 Nov; 47(11): 1557–63.
[2] Nellis ME, Tucci M, Lacroix J, Spinella PC, Haque KD, Stock A, et al. Bleeding Assess-
ment Scale in Critically Ill Children (BASIC): Physician-Driven Diagnostic Criteria for
Bleeding Severity. Crit Care Med, 2019 Dec; 47(12): 1766–72.
[3] Spinella PC, El Kassar N, Cap AP, Kindzelski AL, Almond CS, Barkun A, et al. Recom-
mended primary outcomes for clinical trials evaluating hemostatic blood products and
agents in patients with bleeding: Proceedings of a National Heart Lung and Blood Insti-
tute and US Department of Defense Consensus Conference. J Trauma Acute Care Surg,
2021 Aug 1; 91(2S Suppl 2): S19–25.
[4] Dalton HJ, Reeder R, Garcia-Filion P, Holubkov R, Berg RA, Zuppa A, et al. Factors As-
sociated with Bleeding and Thrombosis in Children Receiving Extracorporeal Membrane
Oxygenation. Am J Respir Crit Care Med, 2017 Sep 15; 196(6): 762–71.
November 9, 2023
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Page 23
Appendix: INTERIM STOPPING RULE OPERAT-
ING CHARACTERISTICS
There are two reasons the trial could be stopped early for safety reasons. One is based on
the prerogative of the DSMB, based on their review of accumulating trial data in DSMB
reports, to recommend the study be terminated; the NHLBI could then enforce such a rec-
ommendation, if they were to support the recommendation. Such an action is unpredictable
and is not accounted for in examining the operating characteristics of the second reason the
trial could be stopped early: a significant observed difference between assigned treatment
arms in the proportion of patients experiencing the primary safety outcome. It is this formal,
prespecified stopping rule that is examined in the remainder of the appendix.
A
Interim stopping rule considerations
The interim stopping rule was calibrated based on several considerations:
Frequency of inferential interim analyses.
Because it is feasible for the adjudication
panel to review site-reported primary safety outcomes in batches of 5 new patients with this
outcome being reported by the enrolling site, with adjudication decisions received within 48
hours of the batch being provided to the panel, the interim stopping rule analyses will be
conducted on the same schedule: as soon as the most recent batch of 5 new patients with
adjudication decisions are available from the panel.
Ability to detect a difference.
A variety of methods have been proposed to conduct
sequential tests while preserving a desired type I error rate. Two popular approaches are
the O’Brien-Fleming and Pocock approaches. The O’Brien-Fleming approach requires very
strong evidence against the null hypothesis initially, but reduces the strength of evidence
required with subsequent tests to preserve more alpha for the end-of-study comparison.
The Pocock approach uses the same test statistic thresholds (equivalently, the same p-value
thresholds) at each pre-specified look and therefore has greater ability to stop the trial earlier
when there are underlying differences in the outcome between treatment arms. The Pocock
approach will be used to give early analyses more statistical power to detect a difference.
Small-sample inference.
Rather than use the standard-normal approximation for the
distribution of the test statistic comparing proportions in the two arms, we will use Fisher’s
exact test because the normal approximation is not justified for dichotomous data with very
small sample sizes. Analyses could be as early as when there are 5 participants.
November 9, 2023
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ECSTATIC: Statistical Analysis Plan
Page 24
Randomization balance.
Although a priori randomization will have the same expected
number allocated to each treatment at any given instant, the observed balance at any given
point may differ. We propagate variability in the observed allocations via simulation to
achieve a more realistic assessment of the interim analysis’s operating characteristics, such
as power and type I error rate.
Desired type I error rate.
We aim to have a 5% type I error rate.
B
Determining the appropriate p-value threshold for
each interim stopping rule analysis
There are standard power calculations available for, say, use of group-sequential testing with
Pocock significance thresholds, a test statistic with a standard normal distribution, and a
prespecified and (often equally spaced) fixed number of interim analysis timepoints. How-
ever, to accommodate the use of Fisher’s exact test (relaxing the dubious assumption of
a standard normal test statistic), unknown number of interim analyses, and variability in
randomization balance, we applied a custom but straightforward Monte Carlo simulation to
determine what p-value threshold would be appropriate to yield a 5% type I error rate.
Because of the nature of patients on ECMO, the eligible population is at high risk of experi-
encing the primary safety outcome. However, a precise estimate of the percentage of enrolled
participants who will experience the primary safety outcome is unknown. We considered
rates from 20% to 80% to calibrate the threshold for having approximately 5% overall type
I error given the multiplicity of testing (and variability in how many tests may be performed).
Monte Carlo simulation with random numbers of events was used to estimate a suitable
nominal alpha for each test, and 0.0333 worked well: average overall type I error rate for
all tests—including a test at trial completion—when averaging Monte Carlo results over the
range pA = pB = p such that p ∈(0.20, 0.21, 0.22, . . . , 0.80) is approximately 5.0%, based
on 2000 simulations per value of p to have adequate precision. The procedure can be con-
servative if the proportion of individuals with the primary safety outcome is < 0.30, which
is not surprising because then fewer tests would be expected to be conducted. However,
because the type I error rate varies with the true p, it was deemed preferable to have the
test be conservative in situations where there are fewer events while allowing the type I error
rate to be slightly above 5% when events are more common, as the latter indicates more
participants would be expected to have the primary safety outcome.
R code for simulating this process is included below, as well as a plot of the estimated type
I error rates as a function of p (Figure 1).
November 9, 2023
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E C S T A TI C: St atistic al A n al ysis Pl a n
P a ge 2 5
0. 2 
0. 3 
0. 4 
0. 5 
0. 6 
0. 7 
0. 8
0. 0 0 
0. 0 2 
0. 0 4 
0. 0 6 
0. 0 8 
0. 1 0
p ≡p A = p B
E sti m at e d o v er all t y p e I err or r at e
S m o ot h e d M o nt e C arl o e sti m at e c o n diti o n al o n p
Tar g et of 0. 0 5
Fi g ure 1:
Esti m ate d o ver all t y p e I err or r ate
w he n usi n g 2, 0 0 0 si m ul ate d tri als f or e ac h
p ∈
0 .2 0 ,0 .2 1 ,...,0 .8 0 a n d c o n d ucti n g e ac h i nteri m test a n d e n d- of-st u d y test usi n g t he
t hres h ol d p −
v al u e ≤
0 .0 3 3 3.
Alt h o u g h t he esti m ate d t y p e I err or r ate varies o ver p ,
w he n
m ar gi n alize d o ver t hese val ues of p , t he t y p e I err or r ate w as esti m ate d t o b e 0. 0 5 0 2,
e xtre mel y cl ose t o t he desire d 0. 0 5 le vel.
N o ve m b er 9, 2 0 2 3
S A P Versi o n 1. 0
C O N FI D E N TI A L

ECSTATIC: Statistical Analysis Plan
Page 26
simPocock <- function(p1,p2,totalN,alpha, nsims=1e3){
whenSig <- rep(NA, nsims)
nTests <- rep(NA, nsims)
tmtvec <- 1+rbinom(totalN*nsims, 1, 0.5)
tmt <- matrix(tmtvec, byrow=FALSE, nrow=totalN, ncol=nsims)
yvec <- rbinom(nsims*totalN, 1, c(p1,p2)[tmtvec])
y <- matrix(yvec, byrow=FALSE, nrow=totalN, ncol=nsims)
for (k in 1:nsims){
# find where we reach 5 more events, but also guarantee we look at study end
templooks <- (1:totalN)[diff(cumsum(c(0,y[,k])))==1 & cumsum(y[,k])%%5 == 0]
Ns.for.looks <- unique(c(templooks[templooks>=5], totalN))
#print(Ns.for.looks)
j <- 0
while (j < length(Ns.for.looks) & is.na(whenSig[k])){
j <- j+1
if (fisher.test(matrix(
c(sum(y[1:(Ns.for.looks[j]),k]==0 & tmt[1:(Ns.for.looks[j]), k]==1),
sum(y[1:(Ns.for.looks[j]),k]==1 & tmt[1:(Ns.for.looks[j]), k]==1),
sum(y[1:(Ns.for.looks[j]),k]==0 & tmt[1:(Ns.for.looks[j]), k]==2),
sum(y[1:(Ns.for.looks[j]),k]==1 & tmt[1:(Ns.for.looks[j]), k]==2)
),
nrow=2, ncol=2))$p.value <= alpha) {
whenSig[k] <- Ns.for.looks[j]
}
}
nTests[k] <- j
}
return(list(nTests=nTests, whenSig=whenSig, estPower = mean(!is.na(whenSig))))
}
pseq <- seq(0.2, 0.8, by=0.01)
powseq <- rep(NA, length(pseq))
set.seed(13250604)
for (j in 1:length(pseq)){
print(pseq[j])
powseq[j] <- simPocock( p1=pseq[j], p2=pseq[j], totalN=50, nsims=2e3,
alpha=0.0333)$estPower
}
library(mgcv)
November 9, 2023
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ECSTATIC: Statistical Analysis Plan
Page 27
print(paste0("Estimated overall type I error rate when using",
" alpha=0.0333 threshold for each test, marginalized",
" average over p in {0.20, 0.21, ",
"..., 0.80}, is ", round(mean(powseq),4)))
plot(gam(powseq~s(pseq)), shift=mean(powseq), se=FALSE, ylim=c(0,0.10),
xlab=expression(p %==% p[A]==p[B]),
ylab="Estimated overall type I error rate",
main="")
abline(h=0.05, lty=2, col="red")
points(pseq, powseq)
legend("topleft", c("Smoothed Monte Carlo estimate conditional on p",
"Target of 0.05"),
lty=c(1,2), col=c("black", "red"), bty="n")
November 9, 2023
SAP Version 1.0
CONFIDENTIAL

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