Ganciclovir to Prevent Reactivation of Cytomegalovirus in Patients With Acute Respiratory Failure and Sepsis
Phase: PHASE3
Conditions: Acute Respiratory Failure
Sponsor: Fred Hutchinson Cancer Center

 
 
Official Title 
Ganciclovir to Prevent Reactivation of Cytomegalovirus in 
Patients With Acute Respiratory Failure and Sepsis 
NCT Number 
NCT04706507 
Document Type 
Protocol and Statistical Analysis Plan 
Document Date 
7/23/2025 
 
 

 
 
 
 
PROTOCOL 
 
 
 
Ganciclovir to Prevent Reactivation of Cytomegalovirus in Patients                              
with Acute Respiratory Failure and Sepsis 
 
 
 
Clinical Trial Funded by the 
NHLBI 
Bethesda, Maryland, USA 
 
 
IND Exempt 152419 
 
Date 07/02/2025 
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ABBREVIATIONS 
 
ANC 
Absolute Neutrophil Count 
ARDS 
Acute Respiratory Distress Syndrome 
CMV 
Cytomegalovirus 
COMS 
Core Outcome Measurement Set 
ELISA 
Enzyme-Linked Immunosorbent Assay 
ETA 
Endotracheal Aspirate 
GCSF 
Granulocyte-Colony Stimulating Factor 
GRAIL 
Ganciclovir/Valganciclovir for Prevention of CMV Reactivation in Acute Injury of 
the Lung And Respiratory Failure 
HCT 
Hematopoietic Cell Transplant 
ICU 
Intensive Care Unit 
IG 
Immunoglobulin 
IL 
Interleukin 
ITT 
Intent-To-Treat 
LFA 
Lateral Flow Assay 
NPV 
Negative Predictive Value 
PEEP 
Positive End Expiratory Pressure  
PPV 
Positive Predictive Value 
RCT 
Randomized Controlled Trial 
RSFD 
Respiratory-Support-Free Days 
SBT 
Spontaneous Breathing Trial 
SOFA 
Sequential Organ Failure Assessment  
SOT 
Solid Organ Transplant 
VFD 
Ventilator-Free Days 
WBC 
White Blood Cell 
 
 
 
 

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TABLE OF CONTENTS 
1 
PROTOCOL SUMMARY ......................................................................................................... 6 
2 
BACKGROUND ......................................................................................................................... 8 
 Critical illness due to sepsis 
8 
 Acute respiratory failure 
8 
 Cytomegalovirus (CMV) overview 
8 
 CMV in immunocompetent ICU patients 
9 
 Use of ganciclovir in the context of respiratory failure 
12 
 A rapid, point-of-care, lateral flow assay (LFA) for CMV IgG serostatus detects CMV 
seropositivity with high sensitivity. 
14 
3 
GANCICLOVIR ....................................................................................................................... 17 
 Mode of action 
17 
 Clinical use 
17 
 Forms of ganciclovir 
17 
 Standard dosing regimens 
17 
 Safety profile 
18 
 Potential toxicities of ganciclovir 
18 
4 
RATIONALE ............................................................................................................................ 23 
 Rationale for study population 
24 
 Rationale for study intervention 
25 
 Rationale for the choice of drug, dose & regimen 
25 
 Rationale for timing of enrollment 
26 
 Rationale for choice of endpoints 
27 
5 
STUDY HYPOTHESES, OBJECTIVES AND ENDPOINTS ............................................. 29 
 Primary Hypotheses 
29 
 Secondary Objectives 
29 
 Exploratory Objectives 
30 
 Post Hoc Analyses 
32 
6 
STATISTICAL CONSIDERATIONS .................................................................................... 34 
 Power Calculations for Primary and Secondary Hypotheses 
34 
 Statistical Analyses for Endpoints. 
37 
 Randomization scheme 
41 
 Blinding 
41 
 Planned analyses prior to end of study 
42 
7 
SELECTION AND WITHDRAWAL OF SUBJECTS ......................................................... 46 
 Study population 
46 
 Randomization 
46 
 Inclusion criteria 
46 
 Exclusion criteria 
46 
 Subject withdrawal 
48 

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 Subject replacement 
49 
8 
STUDY DRUG ACQUISITION, PREPARATION, & ADMINISTRATION .................... 50 
 Study drug & placebo formulation 
50 
 Acquisition of study drugs & placebos 
50 
 Storage of study drugs & placebos 
50 
 Administration of study drugs & placebos 
50 
 Pharmacy Records 
50 
9 
CLINICAL PROCEDURES .................................................................................................... 51 
 Patient identification & recruitment 
51 
 Informed Consent 
51 
 Screening procedures 
52 
 Patient Registration 
54 
 Randomization procedure 
54 
 First dose of study drug 
55 
 Intervention (Study drug administration) 
55 
 Co-interventions 
56 
 Specimen collection 
56 
 
Patient-Centered Outcomes Survey (COMS) 
56 
 
Post-Enrollment Procedures 
57 
 
Monitoring of renal function 
58 
 
Monitoring for and managing neutropenia 
58 
 
Pregnancy 
59 
 
Unblinding 
59 
10 
LABORATORY PROCEDURES ........................................................................................... 60 
 
Laboratory procedures 
60 
 
Future use of stored specimens 
60 
 
Biohazard containment 
60 
11 
ADVERSE EVENT REPORTING ......................................................................................... 61 
 
Reportable Adverse Events 
61 
 
Reportable Serious Adverse Events 
62 
 
Reporting Adverse Events 
63 
 
Relationship to study drug 
67 
 
Pregnancy 
67 
 
Breaking the blind 
67 
 
Stopping rules 
68 
12 
DATA MANAGEMENT CONSIDERATIONS .................................................................... 69 
 
Overview 
69 
 
Data Collection 
69 
 
Data Management 
69 
 
Quality Control and Quality Assurance 
69 
 
Study monitoring 
70 

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13 
ETHICAL CONSIDERATIONS & HUMAN SUBJECTS PROTECTIONS .................... 71 
 
Ethical Review 
71 
 
Potential risks of study drugs and procedures 
71 
 
Risks of Endotracheal Aspirates 
71 
 
Risks of blood collection 
71 
 
Risk of loss of confidentiality 
72 
 
Potential benefit of enrollment 
72 
14 
PROTOCOL OVERSIGHT AND GOVERNANCE ............................................................. 73 
 
Principal investigator 
73 
 
Protocol Leadership Team 
73 
 
Safety and protocol adherence review team 
73 
 
Data Safety and Monitoring Plan (outlined briefly below and in more detail in 
Appendix F) 
73 
 
Data and Safety Monitoring Board 
73 
 
Study termination 
74 
15 
REFERENCES ......................................................................................................................... 75 
16 
INVESTIGATORS STATEMENT/PROTOCOL SIGNATURE PAGE ............................ 80 
APPENDIX A: TIME AND EVENTS SCHEDULE ....................................................................... 81 
APPENDIX B: NCI COMMON TOXICITY CRITERIA (CTC) ................................................. 83 
APPENDIX C:  COMMONLY PRESCRIBED IMMUNOSUPPRESSIVE AGENTS WITH 
KNOWN EFFECT ON CMV REACTIVATION ........................................................................... 84 
APPENDIX D: LUNG PROTECTIVE  VENTILATION PROTOCOL 
RECOMMENDATIONS ................................................................................................................... 85 
APPENDIX E: CONSERVATIVE FLUID MANAGEMENT ....................................................... 89 
APPENDIX F: DATA AND SAFETY MONITORING PLAN ...................................................... 90 
APPENDIX G:  SEPSIS CRITERIA ................................................................................................ 93 
APPENDIX H:  LIST OF INITIALLY SELECTED SITES .......................................................... 94 
APPENDIX I: RECOMMENDATIONS FOR HFNC AND NIV .................................................. 95 
 

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1 
PROTOCOL SUMMARY 
 
Title  
Ganciclovir to Prevent Reactivation of Cytomegalovirus in Patients 
with Acute Respiratory Failure and Sepsis 
 
Study drugs 
Ganciclovir sodium: 2-amino-9-76,9-dihydro-3H-purin-6-one.  
Placebo for ganciclovir: [normal saline] 
 
Patients 
Immunocompetent, CMV seropositive adults hospitalized with 
sepsis and acute respiratory failure requiring respiratory support  
Protocol Schema 
 
 
 
 
Schedule of administration* 
 
 
Day 1 through Day 5 
Day 6 through Day 28 or hospital discharge, 
whichever occurs earlier 
Arm 
N 
Twice daily 
Once daily 
1 
250 
Ganciclovir 5 mg/kg intravenously 
 
Ganciclovir 5 mg/kg intravenously  
 
2 
250 
Normal saline intravenously 
 
Normal saline intravenously 
 
Total 500 
 
 
 
* “Day” on this table refers to study day.  Day 1 is the first day of study drug administration. 
 
Primary Objective 
To evaluate whether administration of ganciclovir increases 
respiratory-support-free days in immunocompetent patients with 
sepsis-associated acute respiratory failure. 
Primary Hypotheses 
We hypothesize that IV ganciclovir administered early in critical 
illness will effectively suppress CMV reactivation in CMV 
seropositive adults with sepsis-associated acute respiratory failure, 
thereby reducing lung damage and accelerating recovery from 
respiratory failure by direct and indirect mechanisms, and leading to 
improved clinical outcomes. 

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Study Design 
Multicenter randomized placebo-controlled double-blind trial, 
[randomized in blocks for balance across study sites, with interim 
analyses of safety]. 
Study Duration 
180 days per patient 
Trial Safety Monitoring 
Safety and Protocol Adherence Review Team (see Section 14.3) 
Data Safety Monitoring Board (see Section 14.5 ) 
Study drug provider 
Commercially acquired 
Funding Agency 
U.S. National Institutes of Health (NIH) National Heart, Lung, & 
Blood Institute (NHLBI) 
Coordinating Center 
 Fred Hutchinson Cancer Center (Fred Hutch)/Vaccine & Infectious 
Disease Division (VIDD) 
Statistical and Data Management 
Fred Hutchinson Cancer Center/Vaccine & Infectious Disease Division 
(VIDD), Statistical Center for HIV/AIDS Research & Prevention 
(SCHARP) 
Endpoint Laboratory(ies) 
Boeckh Lab, Fred Hutch 
Protocol Leadership Team  
Renee Stapleton, MD, PhD, Division of Pulmonary and Critical Care 
Medicine, University of Vermont, Burlington, VT 
Michael Boeckh, MD, Vaccine and Infectious Disease Division, Fred 
Hutchinson Cancer Center, Seattle, WA 
Ajit Limaye, MD, Division of Infectious Diseases , Univ. of California 
San Francisco, San Francisco, CA 
Gordon Rubenfeld, MD, MSc Sunnybrook Medical Centre, Univ. of 
Toronto, Toronto, Canada 
 
Protocol Writing Committee 
 
Ajit Limaye, MD 
Michael Boeckh, MD 
Gordon Rubenfeld, MD, MSc 
Renee Stapleton, MD, PhD 
Louise Kimball, PhD, RN 
Ashley Sherrid, PhD 
 
Principal Investigator of the DCC/Protocol Statisticians:  
Ying Huang, PhD, Professor, Fred Hutchinson Cancer Center   
 
NIH Medical Officer:  
Matthew Craig, PhD 

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2 
BACKGROUND  
 Critical illness due to sepsis  
One million people per year in the US are hospitalized with sepsis and, as the population ages, its 
incidence is expected to increase [1]. Sepsis is considered the most expensive condition treated in 
US hospitals, with an overall cost to society of ~$38 billion in 2013, and the subset with acute 
respiratory failure  accounts for 10% of all intensive care unit (ICU) admissions [1, 2].  
 
 Acute respiratory failure 
Acute respiratory failure is a common and serious complication in hospitalized patients. Sepsis, 
and particularly sepsis from pneumonia, accounts for 73% of patients with acute respiratory 
failure [3]. Among patients with sepsis, 40% develop acute respiratory failure, with mortality 
rates of >30% [4, 5]. Despite this burden of illness for sepsis-associated acute respiratory failure, 
no pharmacologic treatments have been proven effective, and care is generally supportive [6] . 
The proposed intervention (IV ganciclovir to prevent cytomegalovirus [CMV]-mediated 
exacerbation of respiratory failure), if proven effective, would be applicable to all CMV 
seropositive adults, who account for at least 60% of all patients with sepsis, representing a 
significant improvement in care for a common, deadly, and expensive health condition.  
For this study, acute respiratory failure will be defined as in Section 4.1.1 of this protocol.  
 
 Cytomegalovirus (CMV) overview 
CMV is a human herpesvirus known to infect more than 50-90% of US adults, and prevalence of 
CMV infection increases with age [7]. CMV has generally been considered a pathogen only in 
severely immunocompromised patients (transplant, HIV), and CMV reactivation is a well-
established cause of morbidity and mortality in these populations. There are now multiple studies 
and meta-analyses demonstrating that CMV reactivation is also common in critically ill, 
otherwise immunocompetent patients with sepsis, pneumonia, and trauma and is associated with 
worse clinical outcomes [8-15] in this setting.  
CMV infection can be acquired through multiple means: mother-to-child (in utero, breast milk), 
infected body fluids (saliva, genital secretions), blood transfusion, or organ transplant. In 
immunocompetent persons, following primary infection by any of these routes, CMV is 
controlled by the immune system and establishes latency in multiple organs/cell-types for the life 
of the host. Importantly, the lung represents one of the largest reservoirs of latent CMV in 
seropositive hosts, which may explain the propensity for CMV-associated pulmonary disease in 
predisposed hosts [16]. During periods of immunosuppression (or as a result of specific stimuli), 
CMV can reactivate from latency (preferentially in the lung) and replicate, producing active 
infection. In persons with impaired cellular immunity, reactivation can progress to high-grade 
CMV replication and commonly leads to clinically evident disease such as CMV pneumonia. 
Lower-grade CMV reactivation that is otherwise clinically silent (“subclinical”) can also be 
detected in apparently immunocompetent persons with critical illness using sensitive techniques 
such as PCR [8].  
Even low-level, subclinical CMV reactivation can produce significant biologic effects such as 
inflammation, fibrosis and immunosuppression. Each of these effects of subclinical CMV 
infection has either previously been demonstrated or could theoretically be important in sepsis-
associated acute respiratory failure and its complications. These biological effects of CMV have 
been shown to occur through various mediators and other indirect means (reviewed in [17, 18]). 

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Importantly, several important CMV-associated adverse clinical outcomes in transplant 
populations [allograft rejection, secondary infections] are not necessarily accompanied by overt 
CMV disease and can only be detected by sensitive means of virus detection such as PCR [19-
21]. In the context of critical illness caused by sepsis [8, 12, 14, 22, 23], it has been postulated 
that the underlying mechanism behind CMV reactivation is the immune perturbations that occur 
during sepsis: intense and dysregulated inflammation, and a compensatory anti-inflammatory 
response [24-26]. To date, specific mediators have not been defined. 
 
 CMV in immunocompetent ICU patients 
 
2.4.1 
CMV reactivation is common in immunocompetent ICU patients 
CMV viremia occurs in ≥30% of CMV seropositive ICU patients (Figure 2-1) [12, 22, 27]. 
Importantly, CMV also preferentially reactivates in the lung compartment [28], which results in 
an overall reactivation rate of ~40% [23, 28]. However, predicting reactivation in individuals 
remains elusive, as specific risk factors for CMV reactivation are inconsistent across existing 
studies. In a prospective observational study performed by our group and published in JAMA [8], 
severity of illness at baseline was not independently associated with subsequent CMV 
reactivation, diminishing the possibility that reactivation is simply a marker for worse outcomes 
[8]. 
2.4.2 
CMV reactivation is associated with adverse clinical outcomes in immunocompetent ICU 
patients with sepsis. 
A compelling body of evidence implicates CMV reactivation as a causal contributor to morbidity 
and mortality in critically ill adults with sepsis-associated acute respiratory failure, based on 
observational studies, animal models, and data from our NHLBI-funded multicenter randomized 
controlled trial (RCT) of ganciclovir (Ganciclovir/Valganciclovir for Prevention of 
Cytomegalovirus Reactivation in Acute Injury of the Lung [GRAIL]) in 160 immunocompetent 
CMV seropositive adults, recently published in JAMA [8-10, 12, 15, 23]. In a recent meta-
analysis, CMV reactivation (compared to no reactivation) was associated with a 2-fold increased 
Figure 2-1: CMV reactivation in the ICU setting [22]. 
 

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odds of mortality in ICU patients (Figure 2-2) [29]. In addition to mortality, recent studies have 
demonstrated a strong and independent association between CMV reactivation and increased 
hospital and ICU length of stay [8] and duration of mechanical ventilation [10]. 
 
We performed a secondary pooled analysis of two prospective cohorts with sepsis: an 
observational cohort of ICU patients (n=40) and the placebo cohort from our phase 2 GRAIL 
RCT (n=66). In a multivariate model, CMV reactivation (assessed in multiple ways) was 
independently associated with worse clinical outcomes: fewer ventilator-free days (VFDs) and 
ICU-free days (Figure 2-3) [30]. 
Figure 2-3. Quantitative Association of CMV Reactivation with Clinical Outcomes in 
Critically Ill Patients with Sepsis, in the Absence of Ganciclovir Treatment [30]. a Estimates 
represent the difference in mean days and associated 95% CIs.     
 
a After adjustment for age, race, gender, baseline transfusion status, study cohort, and a standardized score 
created from APACHE II and III scores used in the individual studies. 
 
2.4.3 
Putative mechanisms linking CMV reactivation with sepsis-associated acute respiratory 
failure. 
Based on a large body of evidence (natural history data, animal data, evidence that CMV can 
cause tissue invasive lung disease in critically ill immunocompetent patients, laboratory studies of 
CMV immunomodulatory capacity, and our phase 2 GRAIL RCT), it appears less likely that 
CMV initiates acute respiratory failure in patients with sepsis, as CMV reactivation tends to occur 
after the development of respiratory failure [8, 22]. Rather, despite adequate antimicrobial and 
supportive care for sepsis, CMV reactivation occurs frequently and secondarily contributes to the 
Figure 2-2: Meta-analysis of mortality of in patients with CMV reactivation [12]. 
 

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severity of injury to the lung, worsening clinical outcomes. Results from our phase 2 GRAIL 
RCT support our hypothesis that preventing CMV reactivation interrupts this cascade, thereby 
improving clinical outcomes. The putative mechanisms by which CMV increases severity of 
respiratory failure are likely to involve several previously well-characterized biological effects of 
CMV (Figure 2-4).  
2.4.3.1 
CMV causes direct tissue injury and is a potent immune modulator.  
Cell lysis caused by rapid viral replication during CMV reactivation is directly destructive to 
cells, and this process is thought to exacerbate damage to tissues in which CMV reactivates, such 
as the lungs [10, 14, 31]. Remarkably, although CMV is typically considered to cause tissue-
invasive disease only in severely immunocompromised patients, histologic evidence of CMV 
pneumonitis was reported in 25-50% of selected previously immunocompetent critically ill 
patients with persistent unexplained respiratory failure and lung infiltrates who underwent lung 
biopsy or autopsy [31-33]. 
 
Numerous studies in both immunocompetent and immunocompromised patients show the 
profound effect of CMV on the human immune system [34-36]. While the single inflammatory 
pathway (interleukin [IL]-6) we studied in our phase 2 trial was not influenced by CMV 
prophylaxis, CMV interacts with the immune system and inflammatory pathways in many other 
ways, all of which could affect patients with sepsis. For example, CMV influences a broad range 
of immune cells, including effector CD8+ and gamma-delta T cells, and macrophage and 
cytokine response including IL-10 production [34, 37]. CMV also promotes immune activation 
[38] and upregulates adhesion molecules necessary for recruitment of inflammatory cells into the 
lung leading to tissue damage [37, 39, 40]. These immune activating and immunosuppressive 
properties of CMV may directly interact with or enhance key aspects of sepsis pathogenesis and 
profoundly interfere with compensatory mechanisms associated with recovery, especially because 
it is widely accepted that sepsis pathogenesis involves early aberrant activation of innate immune 
cells and is associated with a profound immunosuppression [41, 42]. Finally, active CMV 
Figure 2-4. Proposed Mechanism. Effects of CMV and ganciclovir on sepsis-associated 
respiratory failure (solid lines) and clinical sequelae.  
 

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infection is known to increase risk of invasive bacterial and fungal infections [43, 44], which 
could also profoundly affect the course of critical illness in sepsis. 
  
 Use of ganciclovir in the context of respiratory failure 
2.5.1 
Ganciclovir antiviral therapy decreases CMV-associated lung injury in a murine model.  
Studies in a murine CMV sepsis model (cecal ligation and perforation) demonstrate that in mice 
with latent CMV infection, CMV reactivation during sepsis causes lung injury (fibrosis) [45, 46]. 
Ganciclovir prophylaxis prevents the development of lung fibrosis after lung injury in this model 
by blocking murine CMV reactivation [45]. This effect is dependent on both the dose and timing 
of antiviral therapy, providing strong evidence of a mechanistic link between CMV reactivation 
and lung injury (Figure 2-5), consistent with improvement of lung-related outcomes with 
ganciclovir prophylaxis in our phase 2 GRAIL RCT.  
 
2.5.2 
Phase 2 RCT: Efficacy of ganciclovir for preventing CMV reactivation in patients with 
sepsis-associated acute respiratory failure.  
Our phase 2 GRAIL RCT showed that ganciclovir was effective in suppressing CMV reactivation 
(Figure 2-6). Specifically, CMV was suppressed in plasma and endotracheal aspirate (ETA) and 
bronchoalveolar lavage fluid (BAL). Safety data for this trial are detailed in Section 3.6.1.7. 
Similar success in preventing CMV reactivation was observed in a single-center, open-label RCT 
of valganciclovir for CMV seropositive, critically ill patients [47]. 
Figure 2-5. Ganciclovir Decreases Sepsis-associated Pulmonary Fibrosis (Represented 
by Percent of Pixels) in Mice: Effect of Dose and Duration [45]  
 

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2.5.3 
Phase 2 trial results demonstrate that ganciclovir prophylaxis improves several respiratory 
clinical outcomes in critically ill patients with sepsis-associated acute respiratory failure.  
The phase 2 GRAIL RCT did not demonstrate a difference in the primary outcome of serum IL-6 
levels between groups [23]. For that trial, IL-6 was selected as the primary endpoint because the 
trial was intentionally designed to measure a candidate biomarker with potential for predicting 
clinical outcomes of sepsis-associated acute respiratory failure. IL-6 was associated with 
mortality in prior ICU studies [48], and preliminary data linked increased IL-6 levels with CMV 
reactivation [49]. However, as described above (Section 2.4.3.1), CMV interacts with the immune 
system through multiple mechanisms, collectively resulting in profound impacts. It is likely that 
the failure to detect changes in IL-6 indicates that the observed clinical effects were mediated 
through pathways other than IL-6.  
In contrast to the negative findings for the primary endpoint, several clinical respiratory outcomes 
(all secondary outcomes) were improved among patients who received CMV prophylaxis, as 
compared to placebo [23]. In the pre-specified sepsis subgroup, the ganciclovir group had a 
statistically significant increase of 3 VFDs compared to placebo, corresponding to a reduction in 
duration of mechanical ventilation of 2.5 days (p=0.006) among survivors at day 28 and of 1 day 
overall (p=0.06) (Table 2-1), and no statistically significant difference in mortality. Patients 
Figure 2-6. Cumulative Incidence of any CMV Reactivation and High-Grade CMV 
Reactivation in Plasma through Day 28 [23].  
 
 
Table 2-1. Key Secondary Outcomes Among Patients with Critical Illness Receiving 
Ganciclovir vs Placebo in the Sepsis Subgroup [23]. 
Outcome Day 28 
Placebo 
Group, N=66 
Ganciclovir 
Group, N=71 
Absolute Difference 
(95% CI) 
P-value 
VFDs median (IQR), 
days 
20 (9-24) 
23 (16-25) 
3 (0-4) 
0.03 
Duration of 
Mechanical 
ventilation, median 
(IQR), days 
6 (3-11) 
5 (3-8) 
-1 (0-4) 
0.06 
Duration of 
mechanical ventilation 
in survivors, median 
(IQR), days 
6.5 (3-12.25) 
4 (2.25-7) 
-3 (-4,0) 
0.006 
 

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receiving ganciclovir also had a trend toward improved oxygenation as measured by PaO2/FiO2 
ratio (Figure 2-7).  
Figure 2-7. Median PaO2 to FiO2 Ratio over First 7 Days of Mechanical Ventilation 
(Intent-to-Treat [ITT] population, Sepsis Subset). Error bars indicate first and third quartiles. 
Blue = placebo, red = ganciclovir [23]. 
 
A recent separate single-center RCT of antiviral prophylaxis did not identify differences in the 
few clinical outcomes that were measured [47]. However, a careful assessment of this negative 
trial provides useful insights that have guided our study. First, it was a small (N=124), complex, 
3-arm trial primarily designed to investigate viral suppression efficacy for both herpes simplex 
virus and CMV using different drugs. Second, they used oral valganciclovir at half the standard 
dose used for prevention in other populations [23, 44, 50]. Third, only 20% of the subjects in this 
trial had sepsis/infection as their reason for ICU admission; the remainder were a heterogeneous 
population of trauma, cardiac arrest, and post-operative diagnoses. Finally, while the study did 
show an antiviral effect of valganciclovir, it did not examine clinical outcomes that are likely to 
be the main effect of CMV reactivation in sepsis (i.e. the exacerbation of existing respiratory 
failure).  
 
2.5.4 
An assessment of risk factors for CMV reactivation to define a target population for the 
phase 3 RCT (Figure 2-8).  
One key question is whether CMV prophylaxis can be targeted to a high-risk group, thereby 
enriching the study population for the phase 3 trial. The valuable data obtained from our phase 2 
GRAIL RCT and our initial observational study allowed us to examine this question by 
performing analyses of risk factors for CMV reactivation. However, we were unable to identify 
any such baseline patient variables that were predictive for CMV reactivation, and that would 
therefore allow us to enrich the study population for the proposed phase 3 trial. Specifically, the 
association of the APACHE score with CMV reactivation measures was found to be inconsistent 
and with small effect size. We also did not identify other patient variables (e.g. age, race, gender 
and baseline transfusion status) associated with subsequent CMV reactivation. No protective 
threshold of lymphocyte counts was identified. These findings provide the rationale to conduct 
the phase 3 trial in all CMV seropositive patients.  
 
 A rapid, point-of-care, lateral flow assay (LFA) for CMV IgG serostatus detects CMV 
seropositivity with high sensitivity.  
Prompt identification and initiation of prophylactic drug treatment in CMV seropositive patients 
is crucial both for this trial and for ultimate effective implementation of this strategy if the trial is  

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Figure 2-8. Baseline patient variables are not predictive of CMV reactivation in 
critically ill patients with sepsis, in the absence of ganciclovir treatment.a Data represent 
odds ratios for binary CMV responses (cross-sectional). Transfusion status was also not 
associated with CMV reactivation (data not shown). 
 
a After adjustment for age, race, gender, baseline transfusion status, study cohort, and a standardized score 
created from APACHE II and III scores used in the individual studies. 
positive. Standard enzyme-linked immunosorbent assays (ELISA) are routinely available, but the 
turnaround time varies by laboratory, and can require up to 48-72 hours. Delays in receiving 
results increase the likelihood that CMV reactivation has already occurred. However, empirically 
treating everyone would lead to unnecessary exposure to ganciclovir in the ~40% of patients who 
are CMV seronegative (and thus not at risk for CMV reactivation and associated adverse effects). 
To address this challenge, we will utilize a new, investigational, easy to perform, rapid, lateral 
flow assay (LFA) for CMV IgG antibodies (QooLabs, Inc). This technology is similar to that 
used for home pregnancy tests, and is easy to perform and interpret. The FDA has determined this 
to be a nonsignificant risk device study [Reference #Q202303]). All rapid testing results will be 
confirmed with a standard assay performed in a CLIA-certified lab. This approach will allow 
physicians to rapidly identify CMV seropositive patients in the ICU, so as to promptly initiate 
therapy.  
We directly compared a novel, NIH SBIR-supported, point-of-care LFA platform (QooLabs, Inc., 
N43AI170072), versus the clinically validated ELISA assay performed in the CLIA-certified UW 
laboratory, in 200 samples. Our results indicate that the assay from QooLabs, Inc, had excellent 
performance characteristics for serum and plasma. For serum, the LFA had 97% sensitivity and 
97% specificity when read by QNow UV Flashlight (the method that will be used for this trial) 
and 99% sensitivity and 93% specificity when read by QNow Automated reader (Table 2-2). For 
plasma, the LFA had 70% sensitivity and 98% specificity by UV Flashlight. Assuming a 
seroprevalence of ~60% as observed in our phase 2 RCT, the positive (PPV) and negative (NPV) 
predictive values for the test were 98% and 96% for serum by QNow UV Flashlight, 96% and 
98% for serum by QNow Automated reader, and 97% and 77% for plasma by UV Flashlight, 
respectively. Serum will be the preferred sample for this study, but plasma will be acceptable if 
serum is not readily available. Thus, the assay will identify ~98% of CMV seropositive patients 
within an hour, facilitating rapid trial initiation in the majority of patients. The false negative rate 

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will be minimal (~3%), but these will be identified by ELISA and remain eligible as long as they 
are within the specified enrollment window of 120 hours.  
 
 
 
 
 
Table 2-2. LFA assay performance characteristics. Results of the novel, point-of-care, rapid 
CMV IgG LFA test (Qoolabs) compared with the clinically validated CMV IgG assay (Zeus 
Scientific, Branchburg, NJ) in 200 consecutive serum samples from adults, performed at the 
CLIA-approved UW Clinical Virology Laboratory. PPV and NPV are calculated based on 
60% seroprevalence. 
Lateral Flow Assays 
Diagnostic Performance (%) 
Sensitivity 
Specificity 
PPV 
NPV 
QNow Automated 
Reader 
99 
93 
96 
98 
QNow UV Flashlight1 
97 
97 
98 
96 
1 only 100 samples were used for the QNow UV Flashlight reader 
 
 

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3 
GANCICLOVIR 
Ganciclovir [DHPG] is an FDA-approved antiviral agent with potent in vitro and in vivo activity 
against human cytomegalovirus and has been in widespread use in the United States and 
worldwide since it was approved in ~1988. More detailed information is contained within the 
package insert. 
 
 Mode of action 
The primary mechanism of action is inhibition of viral DNA polymerase in virally-infected cells. 
More detailed information is contained within the package insert. 
 
 Clinical use 
Ganciclovir is indicated for:  
o Sight-threatening CMV retinitis in severely immunocompromised people  
o CMV pneumonitis in bone marrow transplant recipients  
o Prevention of CMV disease in bone marrow and solid organ transplant recipients  
o Confirmed CMV retinitis in people with AIDS (intravitreal implant)  
It is also used for acute CMV colitis in HIV/AIDS and CMV pneumonitis in immunosuppressed 
patients. See the package insert for more information. 
 
 Forms of ganciclovir 
Ganciclovir is available in both intravenous (ganciclovir) and oral formulations (valganciclovir) 
and is proven efficacious for both prevention and treatment of CMV infection and disease in 
immunocompromised patients (transplant, HIV) and in neonates with congenital CMV infection 
[51, 52].  
3.3.1 
Ganciclovir (intravenous formulation) 
Ganciclovir is an FDA-approved, commercially-available antiviral medication used to treat or 
prevent cytomegalovirus (CMV) infections. Generic versions of ganciclovir sodium are marketed 
by different drug suppliers.  
Ganciclovir is a synthetic analogue of 2'-deoxy-guanosine. It is first phosphorylated to a 
deoxyguanosine triphosphate (dGTP) analogue. This competitively inhibits the incorporation of 
dGTP by viral DNA polymerase, resulting in the termination of elongation of viral DNA. See the 
package insert for more information. 
 
 Standard dosing regimens 
1. Treatment of active CMV infection (i.e. presence of CMV by culture, PCR, or antigen 
detection). 
Dosing of intravenous ganciclovir is 10 mg/kg daily, given as 5 mg/kg every 
12 hours (adjusted for renal function).  A minimum interval of 6 hours is 
required between the first and second dose. 

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2. Prevention of CMV reactivation (in CMV seropositive patients with latent CMV 
infection but without evidence of active CMV infection) 
Dosing of intravenous ganciclovir is 5 mg/kg once daily (adjusted for renal 
function). 
In this protocol we will use an initial 5-day regimen of twice daily dosing intravenous ganciclovir 
for hospitalized patients, followed by a daily dosing regimen of intravenous ganciclovir. All 
patients will receive a maximum of 28 days of study drug, provided that they have intravenous 
access. For patients discharged from the hospital prior to day 28, study drug will be discontinued 
at the time of hospital discharge or removal of intravenous access, whichever occurs earlier. For 
patients who remain hospitalized beyond day 28, study drug will be discontinued after day 28. 
Dose adjustments for reduced renal function will be done according to the package insert.  
 
 Safety profile 
It is estimated that tens of thousands of persons have received either intravenous or oral 
formulation ganciclovir over the last ~30 years since its initial approval. Based on its efficacy and 
general tolerability, ganciclovir is currently recommended as a first-line agent for prevention & 
treatment of CMV infection and disease in HIV, solid-organ transplant, and stem cell transplant 
populations [53, 54]. See the package insert for more information (Appendix I). 
 
 Potential toxicities of ganciclovir 
Ganciclovir is generally well-tolerated, with low rates of toxicity when given for less than 28 
days (the maximum possible duration of study drug in the present study). The most common 
adverse effects, which appear to be related to longer duration of exposure and use of concomitant 
drugs with similar toxicities, are various hematological adverse effects, most commonly 
leukopenia, neutropenia, and thrombocytopenia, all of which are considered reversible after drug 
discontinuation. The potential toxicities of ganciclovir have been extensively studied in vitro, in 
vivo and in placebo-controlled studies in humans. Based on animal and cell culture data 
ganciclovir is considered a potential human carcinogen, teratogen, and mutagen. It is also 
considered likely to cause inhibition of spermatogenesis. No human data exist that estimate the 
actual risk of these effects. Thus, it is used judiciously and handled as a cytotoxic drug in the 
clinical setting.  
3.6.1 
Human toxicity data relevant to the proposed trial 
In human studies (mostly involving immunocompromised solid-organ or stem-cell transplant 
recipients), the primary toxicity has been reversible leukopenia or neutropenia and has generally 
occurred after months of drug exposure and in patients receiving other marrow toxic agents. 
Baseline leukopenia/neutropenia is an uncommon finding in critically-ill patients with sepsis and 
acute respiratory failure and is thus not anticipated to be a significant issue but will be closely 
monitored. For all patients receiving study drug (ganciclovir), routine weekly monitoring (with 
absolute neutrophil and platelets counts) is recommended and will be performed in the present 
study. Monitoring will be performed by the research team at each site for all participants, with 
any reportable AE reporting per the protocol.  These research personnel will be blinded. Other 
potential side effects have generally been similar between ganciclovir and placebo groups in 
randomized trials. 

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3.6.1.1 
Hematotoxicity 
3.6.1.1.1 Platelets 
Most placebo-controlled randomized studies, including those in stem cell transplant patients, do 
not show a difference in the incidence of thrombocytopenia and platelet transfusion requirements 
[44, 55-59]. However, there are rare anecdotal reports of ganciclovir-related pancytopenia. One 
study of ganciclovir prophylaxis in hematopoietic cell transplant (HCT) recipients reported 
delayed platelet engraftment [60]. Overall, the potential to cause thrombocytopenia is considered 
low.  
3.6.1.1.2 Neutropenia  
Neutropenia is the principal toxicity of ganciclovir and valganciclovir. The incidence is highest in 
HCT recipients and HIV-infected individuals, followed by pediatric patients with congenital 
CMV disease and solid organ transplant (SOT) recipients. Many studies have demonstrated the 
effect occurs late after drug administration [56, 61, 62]. In fact, several studies in HCT recipients, 
the most susceptible population for this complication, show that the median time of onset is 5 
weeks after start of drug administration. The most relevant data for the proposed study come from 
a recent randomized trial of valganciclovir prophylaxis in kidney transplant recipients [56]. In 
that study, the incidence of neutropenia within 28 days (the duration of treatment proposed in the 
present study) was only 2%. Another recent randomized trial of valganciclovir vs. ganciclovir at 
treatment doses (900 mg twice daily and 5 mg/kg twice daily, respectively) for CMV disease in 
SOT recipients showed a neutropenia rate of 1.2% and 0%, respectively, at 21 days of treatment 
[57].  
Ganciclovir-related neutropenia is reversible [56, 57, 61]. The time to recovery can be hastened 
by administration of G-CSF [54].  
3.6.1.2 
HIV & hematotoxicity  
A trend towards anemia has been shown to occur in HIV-infected patients treated with 
valganciclovir. However, no strong evidence exists in transplant recipients and other patient 
populations, suggesting that the effect may be related to concomitant medications specific to the 
HIV setting. One phase III randomized trial of prolonged valganciclovir prophylaxis in HCT 
recipients, a population that would be considered at particularly high risk for this complication, 
did not show an increased rate of anemia or red blood cell transfusion requirements [63]. Other 
randomized trials also did not show an increased risk of anemia [50, 56, 64].  
3.6.1.3 
Renal toxicity 
Results from randomized trials do not support a role for ganciclovir or valganciclovir as causes of 
renal toxicity. None of the recently conducted randomized trials shows an increased risk or renal 
toxicity [50, 56]; however, two earlier trials, one in heart transplant recipients with IV ganciclovir 
[65, 66] showed increased rates of renal insufficiency. While the potential to cause direct toxicity 
appears to be low, we will monitor renal function closely and adjust doses according to the 
creatinine clearance according to the package insert.  
3.6.1.4 
Neurotoxicity 
Rarely observed. Not statistically significant between study arms of most randomized trials 
except one study in HCT recipients [50]. This effect probably occurs only in a setting of 
concomitant drugs with neurotoxic potential and high blood levels in the setting of subclinical 
renal insufficiency.  
3.6.1.5 
Carcinogenicity 
Ganciclovir is considered a potential human carcinogens (see package insert). No studies have 
been performed to systematically assess this potential in humans. Although tens of thousands of 

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transplant and HIV infected patients have been treated with these compounds over the past ~20 
years, no reports of an increased risk of cancer have been published. However, this does not rule 
out possible carcinogenic effect.  
3.6.1.6 
Teratogenicity 
There are reports of ganciclovir-associated teratogenicity in humans, and this drug is 
contraindicated in patients who are or are planning to become pregnant. For the purposes of this 
study, all patients will be screened and excluded for pregnancy/possible pregnancy. Participants 
who are women of childbearing potential will be advised to use effective contraception during 
treatment and for at least 30 days following treatment with study drug. Similarly, participants 
who are men will be advised to practice barrier contraception during and for at least 90 days 
following treatment with study drug. 
 
3.6.1.7 
Phase 2 RCT results for safety of ganciclovir in critically ill patients.  
Ganciclovir was well tolerated in critically ill patients with acute respiratory failure associated 
with sepsis or trauma in our NHLBI-funded phase 2 double-blind, multicenter GRAIL RCT of 
160 patients, of whom 84 received ganciclovir (Table 3-1) [23]. Despite concerns about 
hematologic toxicity of ganciclovir in other populations (HIV, transplant), no patients in the 
ganciclovir group developed neutropenia that was felt to be related to study drug. There were also 
no significant differences in transfusion requirements and other pre-specified adverse effects (use 
of hematopoietic growth factors, renal insufficiency) associated with study medication. Similarly, 
valganciclovir was well tolerated in a single-center, open-label RCT in CMV seropositive, 
critically ill patients [47]. In this trial, oral valganciclovir was used at half the standard dose used 
for prevention in other populations. No cases of neutropenia were observed in the valganciclovir 
group, and there were no significant differences were observed in renal insufficiency, platelet 
transfusions, or mortality between the valganciclovir group and the control group. A recent, 
separate RCT evaluated pre-emptive ganciclovir for CMV reactivation in 76 immunocompetent 
ICU patients requiring invasive mechanical ventilation (39 of whom received ganciclovir) [67]. 
Adverse event rates were comparable for for ganciclovir-treated patients versus placebo 
recipients, no adverse events related to hematological toxicity were observed, and no leucopenia 
or thrombocytopenia were reported. Furthermore, creatinine levels, white blood cell and platelet 
counts, and the percentage of patients requiring renal replacement therapy from randomization to 
the end of treatment were similar.  
Table 3-1. Safety Assessments Among Patients with Critical Illness Receiving Ganciclovir vs 
Placebo [23]. 

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3.6.1.8 
Summary of human toxicity data  
Ganciclovir is currently recommended and widely used as a first-line agent for prevention and 
treatment of CMV infection/disease in patients with HIV and solid-organ or hematopoietic cell 
transplant [54, 68]. Potential toxicities of ganciclovir are well known, with the primary toxicity 
being reversible leukopenia/neutropenia [43, 56] that can occur after several weeks of drug 
exposure, usually in patients receiving other marrow toxic agents. Other important toxicities, per 
the package insert, include neurotoxicity and thrombocytopenia. As stated above, data from our 
phase 2 RCT [23] and another trial [47] demonstrate that ganciclovir is well tolerated in 
immunocompetent patients with critical illness due to sepsis or trauma and has a favorable safety 
profile. The observed incidence of neutropenia and neurotoxicity in our phase 2 trial was 0%, and 
there was no significant increase in transfusion requirements and other cytopenias, including 
thrombocytopenia, in patients receiving ganciclovir versus placebo (Table 3-1). These data 
suggest that severe side effects of ganciclovir are rare in critically ill patients and ganciclovir-
related neutropenia occurs very uncommonly in persons without underlying bone marrow 
dysfunction and generally occurs at a median of 5 weeks after drug exposure (longer than the 
maximum 28 days in the proposed study). 
Anemia has been observed in HIV-infected subjects, but there is no evidence that it is a problem 
in transplant patients or ICU patients with sepsis, or in the treatment of congenital disease. 
There may be some risk of renal toxicity; however, this was not consistently observed across 
randomized trials. Data from our phase 2 RCT, in otherwise immunocompetent patients with 
critical illness due to sepsis or trauma, demonstrated no significant increase in renal insufficiency 
in the ganciclovir group compared with the placebo group [23]. Data from a second RCT had 
similar findings [47]. 
Other potential safety issues include teratogenicity and carcinogenicity.  
 
3.6.2 
Other recent investigational applications of ganciclovir 
A randomized clinical pilot trial of valganciclovir for chronic fatigue syndrome (NCT00478465) 
has been completed [69]. Valganciclovir was well‐tolerated and was not discontinued due to 
hematologic or hepatic adverse events. 
 

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A randomized placebo-controlled pilot trial of valganciclovir has also been completed in patients 
with glioblastoma multiforme (NCT00400322) [70]. Valganciclovir was safe and well tolerated. 
Treatment‐related adverse events were generally mild to moderate; the most common 
hematological events were thrombocytopenia and leucopenia. 

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4 
RATIONALE  
The study is a phase 3 randomized, double-blind, placebo-controlled multicenter (see Appendix H 
for a list of the 19 initial clinical sites) clinical trial with respiratory-support-free days (RSFD) as 
the primary outcome measure. The rationale for key aspects of the study design are summarized 
in Table 4-1 and described below.  
Table 4-1. Rationale for Key Aspects of the Study Design. 
Study Population 
Sepsis, Acute Respiratory Failure, and Respiratory Support are defined as in Section 4.1.1 below.    
Prophylaxis vs. Treatment 
 
PROS 
CONS 
Prophylaxis 
• 
Conceptually more attractive; prevents all 
CMV reactivation at any site (including 
lung) before CMV-associated effects begin 
• 
Standard of care for other populations 
where CMV is a clinical problem 
(transplantation) 
• 
Best experimental/clinical data for 
preventing CMV indirect effects 
• 
Logistically easier (would also be easier to 
implement clinically)  
• 
Effect “diluted” by high proportion of non-
reactivators  
• 
Relative “over-treatment” with risk for 
drug toxicity 
Treatment 
(Preemptive 
Therapy) 
• 
More targeted approach (“treating a 
known/diagnosed infection” vs. preventing 
a theoretical possibility) 
• 
Minimizes drug exposure and toxicity by 
targeting only infected patients 
• 
Logistically complicated 
• 
Likely too late to see any benefit of 
intervention (CMV-mediated effect cascade 
already initiated) [10] 
• 
Less effective in animal models  
• 
Plasma CMV PCR is an insensitive marker 
of CMV reactivation (preferentially local 
reactivation in lung)  
Choice of Drug 
Ganciclovir 
• 
FDA-approved for prophylaxis and treatment of CMV infection and disease 
• 
Favorable safety profile in ICU patients 
• 
Extensive experience in patients with reduced renal function 
• 
Insufficient data for alternative drugs (letermovir, maribavir) 
• 
Broad-spectrum antiviral activity (including HHV-6 [71]) 
Study drug duration 
Until death, 
hospital 
discharge, 
or 28 days 
(whichever 
occurs 
earliest) 
• 
Incident CMV reactivation occurs in critically ill patients up to 28 days after admission [7, 
12, 16];  therefore, discontinuing ganciclovir at ICU discharge would leave CMV 
seropositive patients at risk of CMV reactivation; 
• 
Time-dependent analyses have shown a delay between CMV reactivation and subsequent 
poor clinical outcomes; thus, early discontinuation of ganciclovir could have a substantial 
impact on subsequent patient outcomes; 
• 
In our phase 2 study, ~8% of days on invasive mechanical ventilation occurred after initial 
ICU discharge; this figure is likely similar for days on respiratory support. 
Timing of Enrollment 

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 Rationale for study population 
4.1.1 
Definitions of sepsis, acute respiratory failure, and respiratory support 
The study population is sepsis-associated acute respiratory failure.  
Sepsis will be defined according to the recent Sepsis-3 consensus definition (outlined briefly 
below and in more detail in Appendix G) [72]:  
• 
Infection suspected or confirmed by the treating physician and  
• 
Organ dysfunction measured as an acute change in total SOFA score ≥2 points after the 
infection (baseline SOFA score assumed 0 unless known pre-existing organ dysfunction).  
Acute respiratory failure will be defined as the concurrent presence of :  
• 
PaO2/FiO2 < 300mmHg if PaO2 is available. If PaO2 is not available, the SpO2 must be ≤ 
96% on any FiO2 setting ≥ 40%,* 
AND 
• 
New, or if no prior imaging, presumed new unilateral or bilateral infiltrates on chest 
imaging occurring in the setting of sepsis from any source, 
AND 
• 
Requiring respiratory support for respiratory failure. 
For this study, respiratory support includes high flow nasal cannula oxygen with a flow rate ≥ 
30 L/min, non-invasive ventilation by mask or helmet with any mode including pressure support, 
CPAP, or bi-level CPAP (BiPAP) with the equivalent of positive end-expiratory pressure (PEEP) 
≥ 5 cmH2O, or invasive mechanical ventilation via endotracheal tube or tracheostomy with PEEP 
≥ 5 cmH2O.  It does not include respiratory support for sleep apnea or conditions other than acute 
respiratory failure.  
* Although FiO2 is difficult to estimate at lower flow HFNC oxygen, for purposes of this 
study, the set FiO2 during HFNC oxygen therapy will be used as the study FiO2 
Although 67% of these patients meet criteria for the acute respiratory distress syndrome (ARDS), 
the main difference between the sepsis-associated acute respiratory failure population and ARDS 
is the chest radiograph. We chose to remove “bilateral radiographic opacities” from our inclusion 
criteria because: (1) There is no reason to suspect that the deleterious effects of future CMV 
reactivation differ by the chest radiograph on presentation; and (2) We have recently shown that 
training clinicians to apply this criterion accurately is ineffective; therefore we do not believe the 
populations can practically be distinguished [73]. Note that meeting these criteria for acute 
respiratory failure confers 2 points in SOFA score and qualifies all patients for sepsis if they have 
clinically suspected/documented infection as indicated by the treating physician and use of 
antibiotics.  
5-day 
window 
from 
hospital 
admission 
• 
Early initiation of antiviral therapy (before or soon after CMV reactivation) is important for 
efficacy 
• 
Use of CMV serologic test (FDA has determined that this is a nonsignificant risk device 
study) allows determination of eligibility within this window 
• 
Provides sufficient time to confirm serologic test  
• 
Competes less for participants with other RCTs that have very short enrollment windows  
 

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4.1.2 
Rationale for focusing on sepsis 
The rationale for focusing on sepsis relates to epidemiology and the fundamental biology of CMV 
reactivation. Sepsis, and particularly sepsis from pneumonia, accounts for 73% of patients with 
acute respiratory failure [3]. The other 27% of patients include trauma, burn, transfusion, COPD, 
and acute congestive heart failure. Since cytokine profiles, patient characteristics and outcomes 
vary considerably by these risk factors, restricting the study to patients with sepsis yields a more 
homogeneous study population. CMV reactivation also varies by risk factor, with septic patients 
at highest risk, likely due to sepsis mediated immune dysfunction [8]. Additionally, our phase 2 
trial found the greatest signal for prevention in the large subset of patients with sepsis- (rather 
than trauma-) associated respiratory failure [23].  
4.1.3 
Rationale for age of study participants 
We will include all patients ≥18 years of age who meet all other inclusion criteria. Studies have 
shown no association between age and CMV reactivation risk in critically ill patients [8, 23, 30]. 
 
 Rationale for study intervention 
We selected a “prophylactic” approach (in which antiviral therapy will be initiated prior to CMV 
reactivation in all eligible CMV seropositive patients) over a “treatment” approach (in which 
antiviral therapy would be started only after CMV reactivation was documented) for this trial (see 
Table 4-1). Despite potential limitations, use of a prophylactic strategy offers the best opportunity 
to assess for an effect of ganciclovir with an acceptable likelihood of toxicity. The major 
weaknesses of a treatment approach are that local CMV reactivation in the lung can occur even in 
the absence of reactivation in blood [74, 75] and that current methods of CMV measurement in 
blood (i.e., PCR) are not sensitive enough for detection of all CMV reactivation [76]. Indeed, a 
recent study showed that patients with sepsis had a much higher proportion of reactive CMV-
specific immune response than what would have been expected based on viral load monitoring in 
the blood [76]; thus reactivation at sites other than the blood (e.g. the lung, salivary gland) is 
probably more common than viremia. Also, since the kinetics of CMV replication in critically ill 
patients is so rapid, significant CMV replication and its negative consequences would likely occur 
before antiviral intervention would be possible. A recent multi-site RCT evaluated a treatment 
(preemptive therapy) approach, with ganciclovir or placebo administered to mechanically 
ventilated ICU patients in whom CMV reactivation was detected (by PCR) [67]. The trial was 
stopped for futility based on an interim analysis by the DSMB, and no significant difference was 
observed between ganciclovir vs. placebo recipients (n=39 vs. 37, respectively) in ventilator-free 
days (the primary endpoint), mortality, or duration of hospitalization or ICU stay. These findings 
lend support to the idea that with a treatment approach, ganciclovir may be administered too late 
to have clinical benefit. Thus, these data support the prophylactic approach of our study design. A 
non-controlled study using a test and treat approach (i.e. ganciclovir treatment instituted on the 
basis of a positive blood test for CMV) failed to demonstrate a clinical benefit [10], probably 
related to the issues discussed above. Finally, for a treatment strategy to be effective generally, 
hospitals would need to implement rapid CMV diagnostic techniques that are not available at all 
centers. 
 
 Rationale for the choice of drug, dose & regimen 
4.3.1 
Rationale for choice of drug 
Among clinically available medications, only ganciclovir and its oral analogue valganciclovir are 
FDA approved for both the treatment and prevention of CMV infection and disease. To date, 
insufficient data exists for use of alternative drugs (letermovir, maribavir) in this setting. 

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Furthermore, ganciclovir offers broad-spectrum activity against other viruses including HHV-6; 
this may offer added benefit since reactivation of both HHV-6 and CMV is associated with worse 
outcomes than reactivation of either virus alone in ICU patients [71]. There is extensive 
experience with ganciclovir due to its widespread clinical use for ~30 years in hundreds of 
thousands of patients. Its primary mechanism of action is inhibition of viral DNA polymerase and 
it does not appear to have other significant biologic effects (e.g. anti-inflammatory, antifibrotic). 
Information on clinical use of ganciclovir, available formulations, standard dosing regimens, 
safety and toxicity are detailed in Section 3.  
As stated above in the Section 2.5, our phase 2 RCT demonstrated that ganciclovir is well 
tolerated in immunocompetent patients with critical illness due to sepsis or trauma and has a 
favorable safety profile [23]. These data suggest that severe side effects of ganciclovir are rare in 
critically ill patients and are likely outweighed by the potential benefit in this high-risk target 
population. Other potential side effects (e.g., liver toxicity) have generally been similar between 
ganciclovir and placebo groups in randomized trials [44, 50, 55, 58, 60, 64, 68, 77]. Also of note, 
there is extensive experience with giving ganciclovir to patients with reduced renal function, 
including critically ill patients receiving renal replacement therapy.  
4.3.2 
Rationale for choice of formulation 
We chose IV ganciclovir only (rather than also using its oral analogue valganciclovir) due to 
unclear absorption of enteral medications in critically ill populations. Additionally, data from our 
phase 2 RCT suggest that oral therapy after hospital discharge occurred in <10% of subjects and 
(after having in-hospital IV therapy) added no apparent benefit. Participants will receive study 
drug twice daily for the first 5 days of therapy, then once daily until therapy ends; the initial twice 
daily dosing will be more effective in disrupting already ongoing reactivation in the small subset 
of patients that, despite several study design features to minimize this number, are expected to be 
PCR positive at baseline. Similar regimens have been used successfully in transplant recipients 
[50, 60].  
4.3.3 
Rationale for duration of drug 
We chose a study drug duration of until death, hospital discharge, or 28 days (whichever occurs 
earlier), for several reasons:  
• 
Incident CMV reactivation occurs in critically ill patients up to 28 days after admission [8, 
15, 23];  therefore, discontinuing ganciclovir at ICU discharge would leave CMV 
seropositive patients at risk of CMV reactivation; 
• 
Time-dependent analyses have shown a delay between CMV reactivation and subsequent 
poor clinical outcomes; thus, early discontinuation of ganciclovir could have a substantial 
impact on subsequent patient outcomes; 
• 
 In our phase 2 study, ~8% of days on invasive mechanical ventilation occurred after initial 
ICU discharge; this figure is likely similar for days on respiratory support.  
Administering ganciclovir was highly feasible across sites in our phase 2 study; 92% of patients 
received all post-ICU discharge doses, and 84% of post-ICU doses were administered 
successfully.  
 
 Rationale for timing of enrollment 
The 5-day enrollment window from the time of initial hospital admission reflects a tradeoff 
between trial efficacy and feasibility. Initiating prophylaxis before CMV reactivation is important 
as prophylaxis is less effective once reactivation has occurred. The shortest window would be 
ideal, and a 5-day window will minimize reactivation; fewer than 8% of patients reactivate CMV 

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in plasma before day 5 [8]. A shorter enrollment window would reduce the number of eligible 
patients and contribute to enrollment failure due to missed patients, weekend screening, and 
delays in consent, affecting feasibility. The 5-day window will allow the PETAL sites included in 
our trial to enroll patients who are beyond the 24–48 hour enrollment window of planned trials in 
that Network, and will minimize competition for patients. Finally, this replicates the enrollment 
window in our positive phase 2 RCT. Enrollment will be defined as occurring at randomization. 
 
 Rationale for choice of endpoints 
4.5.1 
Justification for Primary Endpoint.  
VFDs are a composite outcome of mortality and duration of mechanical ventilation in survivors 
that assigns zero points to all deaths within a 28-day window and 1 point to every day off the 
ventilator in 28-day survivors. It was designed, in part, to address the issue of competing 
mortality on duration of mechanical ventilation. More recently, changes in clinical practice have 
led to increased use of non-invasive ventilation (NIV) in patients with acute respiratory failure. 
This change is based on 20 years of accumulating evidence that NIV as initial management in 
patients with acute hypoxemic respiratory failure is associated with lower mortality and, in some 
patients, can completely prevent intubation [78]. The COVID-19 pandemic has accelerated the 
implementation of NIV in acute respiratory failure, and at present, most patients with sepsis 
associated respiratory failure are managed with NIV initially unless they have a contraindication 
to its use or need an airway to facilitate transport. While it is impossible to predict every aspect of 
clinical care 5-7 years in the future, the trend toward non-invasive management of sepsis 
associated respiratory failure is clear now. Thus, including participants receiving NIV is essential 
for sufficient enrollment in this trial and applicability of trial results to future clinical practice.  
Furthermore, while the mechanisms of sepsis associated respiratory failure are incompletely 
understood, the risk of CMV reactivation and its attendant harms are likely to be approximately 
the same whether the patient is managed with invasive versus non-invasive ventilation [8, 79, 80].  
In a study of risk factors for CMV reactivation in critically ill patients that included both 
intubated and non-intubated patients, baseline intubation and mechanical ventilation was not a 
statistically significant risk factor for CMV reactivation [8].  
As such, patients receiving NIV will be included in this trial and respiratory-support-free days 
(RSFDs) will be the primary endpoint for this trial. The following rationale supports use of RSFD 
as a primary endpoint, based in part on VFD data as well as available RSFD data.  
First, biologically, we believe that CMV prophylaxis is likely to exert its greatest effect on 
pulmonary related morbidity. Second, VFDs (and more recently, RSFDs or organ support-free 
days) are frequently used as outcomes in high profile RCTs and influence clinical practice. A 
recent review of 128 RCTs assessing duration of mechanical ventilation in 10 high-profile 
general medical and critical care journals found that 43% reported VFDs and 12% used it as the 
primary outcome variable [81]. The NHLBI ARDS Network FACTT Trial showed that a 
conservative approach to fluid management increased VFDs by 2.5 with no statistically 
significant effect on mortality at 60 days [82]. Based primarily on this trial evidence, the 
Surviving Sepsis Guidelines endorsed this approach with a strong recommendation. Finally, 
VFDs are the outcome that was significantly improved in our phase 2 GRAIL RCT.  Importantly, 
in recent trials enrolling NIV patients, there has been a change from VFDs to RSFDs and organ 
support free days in a 28 day window [83-85]. For example, the REMAP-CAP platform trials for 
COVID and community acquired pneumonia have adopted RSFDs and have determined that 1.5 
RSFDs are the minimal clinically important difference [84].   

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There are several possible limitations to using RSFDs at 28 days as our primary endpoint. As 
with all composites, patients and clinicians may not value these outcomes in the way they are 
weighted in the composite. RSFDs will not capture outcomes that go in different directions (e.g., 
treatments that increase mortality but also reduce duration of respiratory support in survivors). 
RSFDs can be difficult to interpret as they do not simply reflect either a change in mortality or a 
difference in duration of respiratory support. Generally, VFDs have a bi-modal distribution and 
must be analyzed appropriately [81]; we anticipate that the distribution of RSFDs will also have a 
bi-modal distribution. 
RSFDs were selected over several potential alternative approaches, which were deemed less 
suitable than using RSFDs to study duration of respiratory support with competing mortality. The 
preliminary data from our observational cohort and our phase 2 trial suggest that the biggest 
effect of CMV reactivation in the ICU was on duration of mechanical ventilation [8, 23]; in 
current practice, many of those who received invasive mechanical ventilation in these previous 
studies would now receive NIV and we anticipate that CMV reactivation would similarly affect 
duration of respiratory support.  Thus, mortality was not selected as the primary outcome for this 
trial. However, since high mortality is expected in this trial of septic patients requiring respiratory 
support or mechanical ventilation and there may in fact be an effect on mortality, we deemed it 
necessary to consider mortality in this trial’s outcome. Therefore, we did not choose respiratory 
support or mechanical ventilation alone as a primary endpoint. While advanced modeling 
approaches may allow the analysis of one outcome, we are not aware of any that have been used 
in the primary analysis of a clinical trial to address the effect of competing mortality on duration 
of mechanical ventilation. The Win-Ratio and the hierarchical outcome approach proposed by 
Finkelstein and Schoenfeld [86] have been used in trials [87], avoid the weighting issue inherent 
in VFDs, and are superior to binary composites as they allow time-to-event measures to be 
compared; however, these were deemed unlikely to offer additional clarity and have not been 
used extensively to study duration of respiratory support. RSFDs combine two outcomes 
(mortality and duration of respiratory support in survivors) considered of the same inseparable 
clinical importance as one endpoint, providing the reader with a single P value. Mortality, 
duration of respiratory support, and duration of mechanical ventilation will also be included as 
secondary analyses. 
4.5.2 
Secondary and Exploratory Endpoints.  
Secondary endpoints were selected either because of their known association with clinically 
significant outcomes in sepsis-associated respiratory failure or because they are clinically relevant 
themselves as outcomes or safety measures. VFDs will be a key secondary endpoint. Although 
the study is not specifically powered to detect significant differences in these secondary clinical 
endpoints, we have provided estimates of the differences that could be detected based on the 
sample size (see Statistical Considerations).  

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5 
STUDY HYPOTHESES, OBJECTIVES AND ENDPOINTS 
 Primary Hypotheses 
We hypothesize that IV ganciclovir administered early in critical illness will effectively suppress 
CMV reactivation in CMV seropositive adults with sepsis-associated acute respiratory failure, 
thereby reducing lung damage and accelerating recovery from respiratory failure by direct and 
indirect mechanisms, and lead to improved clinical outcomes.  
5.1.1 
Primary Objective 
To evaluate whether administration of ganciclovir increases respiratory-support-free days 
(RSFDs) in immunocompetent patients with sepsis-associated acute respiratory failure. 
5.1.2 
Primary Endpoint  
RSFDs will use a “last off” approach (detailed in Section 6.1.1), meaning that RSFDs will be 
counted when a participant gets off and stays off of respiratory support (as defined in Section 
4.1.1) to day 28. Participants who do not survive through day 28 are assigned zero RSFDs. 
 
 Secondary Objectives 
The secondary objectives of the study are: 
1. To evaluate whether administration of ganciclovir increases VFDs in immunocompetent 
patients with sepsis-associated acute respiratory failure. 
2. To evaluate whether administration of ganciclovir increases total RSFDs (all RSFDs, instead 
of last-off approach) in immunocompetent patients with sepsis-associated acute respiratory 
failure 
3. To evaluate whether mortality and time to death in the 28 and 180 days is different among 
ganciclovir recipients relative to placebo recipients, respectively. 
4. To evaluate whether duration of mechanical ventilation among survivors in the first 28 days 
is different among ganciclovir recipients relative to placebo recipients. 
5. To evaluate whether duration of respiratory support among survivors in the first 28 days is 
different among ganciclovir recipients relative to placebo recipients. 
6. To evaluate whether oxygenation is different among ganciclovir recipients relative to placebo 
recipients. 
7. To evaluate whether ICU-free days in the first 28 days are different among ganciclovir 
recipients relative to placebo recipients. 
8. To evaluate whether CMV DNA detection in plasma and endotracheal aspirate (ETA) by day 
28 is different among ganciclovir recipients relative to placebo recipients. 
9. To assess the number and severity of reportable adverse events and reportable serious adverse 
events in the first 28 days in both groups. 

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5.2.1 
Secondary Endpoints 
1. VFDs (defined to be 28 days minus the duration of mechanical ventilation through day 28 
since randomization [a key secondary endpoint]). Participants who do not survive through 
day 28 are assigned zero VFDs. 
2. Total RSFDs (calculated as 28 days minus each day of respiratory support through day 28 
since randomization). Participants who do not survive through day 28 are assigned zero 
respiratory-support-free ventilator-free days. 
3. Mortality by day 180 (day 28, day 180, time-to-event). 
4. Duration of mechanical ventilation (among survivors) by day 28. 
5. Duration of respiratory support (among survivors) by day 28. 
6. Oxygenation (PaO2/FiO2 ratio) daily on study days 1-7. 
7. ICU-free days by day 28. 
8. CMV DNA detection in plasma and endotracheal aspirate (ETA) by day 28 (>0 IU/mL, 
>1000 IU/mL). 
9. Number of patients with reportable adverse events of Grade 3 or higher by day 28. 
 
 Exploratory Objectives 
The exploratory objectives of the study are: 
1. 
To evaluate whether static respiratory system compliance at randomization, day 4 and 
day 7, is different among ganciclovir recipients relative to placebo recipients. 
2. 
To assess occurrence of invasive bacterial and fungal infections among ganciclovir 
recipients relative to placebo recipients. 
3. 
To evaluate if organ dysfunction scores (regular SOFA variables including respiratory, 
coagulation, liver, cardiovascular, CNS and renal; however, SpO2/FiO2 ratio will be used in 
place of P/F ratio used in regular SOFA)  are different among ganciclovir recipients relative to 
placebo recipients. 
4. 
To assess long-term life quality as measured by the Acute Respiratory Failure Core 
Outcome Measurement Set (COMS), which will include the Katz Index of Independence in 
Activities of Daily Living (ADL) and Lawton – Brody Instrumental Activities of Daily Living 
Scale (IADL) completed by legally authorized representatives (LARs) or participants at baseline; 
and the ADL, IADL, Hospital Anxiety and Depression Scale (HADS), EQ-5D-5L, and Impact of 
Events Scale – Revised (IES-R), completed by patients at day 180. 
5. 
To assess risk factors that may associate with CMV reactivation kinetics, including 
demographics, co-morbidity, severity of illness, organ dysfunction, lymphocyte count, time from 
hospital admission to enrollment, ventilation type, viral load prior to randomization, and duration 
of illness before hospital admission. 

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6. 
To characterize the relationship of CMV viral load kinetics in blood and lung 
compartments with RSFDs and VFDs and specific secondary clinical outcomes like oxygenation, 
static respiratory system compliance, mortality and duration of mechanical ventilation in 
survivors. 
7. 
To assess performance of rapid lateral flow CMV serostatus assay compared to clinically 
performed assays for IgG antibodies to CMV, as performed in CLIA-approved labs, throughout 
the study and at trial completion. 
8. 
To evaluate assays to characterize immunity to CMV (cellular immunity, neutralizing 
antibodies, antibody epitope expansion, and transcriptional signatures). 
9. 
To determine whether use and duration of ECMO differs among ganciclovir recipients 
relative to placebo recipients. 
10. 
To assess occurrence of neuromuscular blockade among ganciclovir recipients relative to 
placebo recipients. 
11. 
To assess the use of prone positioning among ganciclovir recipients relative to placebo 
recipients.  
5.3.1 
Exploratory Endpoints 
1. Static respiratory system compliance at randomization, day 4 and day 7. 
2. Invasive bacterial and fungal infections. 
3. Organ dysfunction scores (regular SOFA variables including respiratory, coagulation, liver, 
cardiovascular, CNS and renal; as well as SpO2/FiO2 ratio). 
4. The NHLBI-endorsed Acute Respiratory Failure Core Outcome Measurement Set (COMS) in 
survivors at day 180. 
5. Risk factors for CMV reactivation (>0 IU/mL, >1000 IU/mL) in plasma and lung 
a. Sex, age, race  
b. Co-morbidity 
c. APACHE III score at baseline 
d. SOFA score and individual components 
e. Lymphocyte count 
f. Time from hospital admission 
g. Viral load prior to randomization 
6. Relationship of CMV viral load with RSFDs and VFDs and secondary clinical outcomes: 
a. Viral load: initial, peak, slope, area under the curve (AUC) 
b. Association with RSFD and VFD, day-28 mortality, duration of mechanical ventilation in 
survivors, static compliance, and PaO2/FiO2. 
c. Baseline viral load in plasma and lung.  

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7. Viral load kinetics among survivors in day 7 and 14. 
8. Sensitivity, specificity, PPV and NPV of rapid lateral flow CMV serostatus assay. 
9. CMV cellular immunity, neutralizing antibodies, antibody epitope expansion, and 
transcriptional signatures. 
10. Use of ECMO at any time during the post-randomization period, and duration if used 
11. Occurrence of neuromuscular blockade. 
12. Prone positioning status. 
5.3.2 
Collection and banking of DNA and RNA, and study samples 
In order to perform future investigations into the causes of acute respiratory failure and any 
possible links between acute respiratory failure outcomes and with treatment with ganciclovir, we 
will collect DNA and RNA samples for gene association and gene expression studies. Other study 
samples (blood, endotracheal aspirates) as well as left-over material from clinical samples (e.g. 
endotracheal aspirates, biopsy, autopsy material) will be kept in a repository for future studies of 
other herpesviruses. IRB approval will be obtained for studies not related to herpesviruses.  
5.3.3 
Ancillary studies 
Cryopreserved samples may be used to perform additional assays to support standardization and 
validation of laboratory assays, and to evaluate additional endpoints and associations of interest. 
These assays may include, but are not limited to PCR testing for other pathogens, gene 
association studies, additional cytokines and chemokines, proteomics, microbiome, gene 
expression and immune function studies. We will identify specific sites to do certain ancillary 
pathogenesis studies.  
 
 Post hoc analyses 
We will conduct electronic medical record review to systematically capture potential additional 
adverse effects, that did not meet the protocol defined definitions of AEs and SAEs (e.g. 
neurologic, cardiac, pancreatic, hepatic, renal events). Additionally, we will extract parameters 
that are required to analyze population pharmacokinetics of the study product. A systematic 
review of the medical record according to previously published categorizations to determine the 
causes of death will be completed.  
 
We will also analyze additional parameters that are already captured in the database, for their 
association with study groups and outcomes, (e.g. lymphopenia, monocytopenia). Additional post 
hoc subgroup analyses will be performed to evaluate specific subgroups (e.g. COVID-19, 
ventilation type, sepsis phenotypes, cause of sepsis, CMV reactivation patterns). 
 
All details of the statistical analyses of these post hoc variables will be outlined in the statistical 
analysis plan.  
 
Using stored samples obtained during the study, we will conduct the following tests: 
- 
Drug levels (ganciclovir) 

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- 
Analysis of inflammatory patterns (e.g. to determine sepsis phenotypes) 
and pathways using system approaches (e.g. cytokine analyses and 
proteomic testing)  
Using stored study product, we will test for purity and contamination of the study product. 
 
Samples may be sent out to external institutions and testing facilities after removing identifiers. 
Additional tests may be conducted based on the results.  
 
 

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6 
STATISTICAL CONSIDERATIONS 
 Power Calculations for Primary and Secondary Hypotheses 
6.1.1 
Primary Endpoint 
The primary endpoint RSFD is defined as follows.  
Consistent with use of free-days in other studies, RSFDs will use a “last off” approach. This is the 
approach that has been used in PETAL Network studies (e.g., NCT02509078), is preferred by 
FDA as a more durable/sustained recovery landmark, and recommended by our External 
Advisory Board.  
“Last off” means that RSFDs will be counted when a participant gets off and stays off of 
respiratory support (as defined in Section 4.1.1) to day 28. Days off of respiratory support will be 
counted back from day 28 to the last day when respiratory support was received.  If a participant 
is still receiving respiratory support on day 28, no RSFDs will be counted, even if they were not 
receiving support for some number of days before day 28.  Following are 3 examples of this 
calculation: 
1. 
Participant receives respiratory support from days 1-10 and then receives no 
further respiratory support through day 28.  RSFDs=18. 
2. 
Participant receives respiratory support from days 1-10, then again from 16-20, 
then again from 24-26.  No respiratory support is received on days 27 or 28.  
RSFDs=2. 
3. 
Participant receives respiratory support from days 1-10, then again from 26-28.  
RSFDs=0. 
Participants who do not survive to day 28 are assigned zero respiratory-support-free days [23, 
88]. The 28-day landmark is chosen, because 
1. Interventional trials in acute respiratory failure typically involve a 28-day treatment 
or follow-up period after the patient enrolls in the trial; and 
2. Most patients with acute respiratory failure have either died or been successfully 
weaned from respiratory support by day 28 [23, 88]. 
To estimate the required sample size for the trial with adequate statistical power for the primary 
endpoint, we used the data of VFDs in the ganciclovir and placebo arms of the GRAIL Phase 2 
trial and available RSFD data in the literature [83-85]. The summary statistics of VFDs in the 
sepsis cohort of GRAIL Phase 2 trial are shown in the table below. Since standard deviation data 
for RSFD is sparse, we assume that standard deviation for RSFDs is similar to that of VFDs in 
our sample size calculations as informed by Table 6-1. 

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The sample size of the Phase 3 trial was determined for the primary endpoint, accounting for 
mortality, realistic and clinically relevant mean difference in RSFD between the arms, and 
feasibility. 
Assume that RSFDs follow a normal distribution within group 1 (ganciclovir) and group 2  
(placebo), respectively. 
!"#$!,# ~ '()!, +!
$),
-. -. /.,
- = 1, … , 3!  
!"#$$,% ~ '()$, +$
$),
-. -. /.,
4 = 1, … , 3$ 
 
The hypotheses are then: 
                                               5&: )! = )$ 
                                                    5!: )! = )$ + /,
where / is the difference between the means. 
 
Assume equal sample sizes, the sample size in each randomization group can be calculated using 
the following formula: 
3 = 2 GH!'!
",$('$ + H!'),$('$I
$
J
*#$$%&'
+
K
$
, where  
3 = estimated sample size for each randomization groupH!'!
",$('$ = J1 −
,
$K
-.
 quantile  MN H$('$ 
distribution (with 23 −2 degrees of freedom) 
H!'),$('$ = (1 −β)-. quantile MN H$('$ distribution (with 23 −2 degrees of freedom) 
+/0012+
$
 = pooled variance of both groups 
Note that 3 in the above equation does not have an explicit form, and thus, we use  
pwr.t.test in CRAN library pwr to derive sample size for the two-sample t-test here. 
Although distributional information for RSFD is limited in the literature, based on data that we do 
have available we anticipate the variance of RSFD to be similar to that of VFD. The pooled 
variance +/0012+
$
 was estimated from the GRAIL Phase 2 trial to be 9.42, which has been used as 
an estimate for +/0012+
$
 in our power calculations. A matrix is given below in Table 6-2. The 
numbers correspond to the estimated sample sizes required in each arm to achieve 80 - 90% 
power for various settings for difference in the means (minimum detectable difference in RSFDs 
between the two arms; 2.5 – 3 days) and the assumption that the pooled standard deviation of 
Table 6-1. Summary statistics of VFDs in the Sepsis Cohort of GRAIL Phase 2 Trial 
 

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RSFDs is 9.4 days, same as the estimate of +/0012+ for VFDs in GRAIL Phase 2, and at a type I 
error rate of 5%. 
Table 6-2. Single arm sample size required to detect a minimum difference of 2.5 – 3 days 
between the mean of RSFDs in the two arms, with different levels of power (80 – 90%), with 
the assumptions that pooled standard deviation of RSFDs in the two arms is 9.4 days (same 
as the estimate of σ_pooled for VFDs in GRAIL Phase 2), and at a type I error rate of 5%. 
 
Single arm Sample Size 
Power 
Detectable difference (P) 
2.5 days 
2.74 days 
3 days 
80% 
223 
186 
156 
85% 
255 
213 
178 
90% 
299 
249 
208 
 
Note: Although RSFDs (or VFDs) can be considered as the number of event days out of the total 
28 days, and conceptually a binomial assumption may be preferred, it is nevertheless not feasible 
for the RSFDs (or VFDs) given their complex definition involving both mortality and actual 
respiratory support (ventilator) usage. For the GRAIL Phase 2 Trial, the standard deviation (SD) 
of VFDs, assuming a binomial distribution, is estimated to be 6.67 days, smaller than the 
empirical estimate of 9.4 days, which suggests a substantial over-dispersion. Thus, normal 
assumption with empirical variances (as obtained from the GRAIL Phase 2 Trial and given that 
we anticipate variance for RSFDs to be similar to that of VFDs) instead appears to be justified for 
RSFDs.  Also note that considering that there is no substantial literature on the distribution of 
RSFDs, our trial design relies on GRAIL Phase 2 data on VFDs instead and our belief that 
variability in RSFDs will be similar to that of VFDs. However, to safeguard against the case if 
variance of RSFDs is substantially larger than that of VFDs, we have planned to conduct a 
sample size re-estimation analysis after ~50% of the endpoints have been observed (see Section 
6.5.3). 
In the GRAIL Phase 2 trial, VFDs difference was estimated to be 2.74 days. As shown in Table 
6-2, to detect a similar difference (of 2.74 days) in RSFDs between the two arms with 85% 
statistical power, a sample size of 426 subjects (213 × 2 = 426) total is required in this Phase 3 
trial’s primary analysis, and that is what we power for in this study. Findings from previous RCTs 
suggest that approximately 12% of subjects in this Phase 3 RCT will be CMV PCR positive at 
baseline when rapid lateral flow assay is used to assess serostatus. This 12% estimate is lower 
than the proportion reported in the GRAIL Phase 2 study due to the use of the rapid lateral flow 
assay, which we estimate will shorten the time to randomization from ICU admission by 
approximately 1 day and reduce the baseline PCR positivity rate. Furthermore, we anticipate that 
the percentage of subjects who are PCR positive at baseline will be lower among individuals on 
respiratory support (as defined in Section 4.1.1) compared with individuals on mechanical 
ventilation alone (as in the GRAIL Phase 2 trial) because we will be capturing some eligible 
participants earlier in the course of their respiratory failure when less time has elapsed to allow 
for CMV reactivation (both because we will allow enrollment of patients on NIV and HFNC 
oxygen and because time to obtain consent from participants able to consent for themselves may 
be shorter than consent through LARs. Since the population for primary analysis includes all 
patients without regard to PCR positivity at baseline, we inflate the sample size by a factor to 

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account for the power loss by including these cases in our primary analysis population.  We also 
assume a dropout rate of 1% lost to follow-up for the primary endpoint, based on findings from 
other large ICU trials, and a dropout rate of 2% for early discontinuation following a negative 
CLIA CMV ELISA test among randomized participants who have received at least one dose of 
study drug. Accounting for both of these effects, the sample size for GRAIL Phase 3 trial was 
increased to 500 subjects in total.   
6.1.2 
Power Calculations for Secondary Endpoints.  
We do not expect this intervention to lead to a statistically significant difference in mortality 
between treatment groups.  Nonetheless, mortality is a common and expected endpoint in critical 
care clinical trials, and it is a component of our primary outcome RSFDs.  As such, we will 
include mortality as a secondary endpoint in this RCT.  The risk of death is typically high 
(approximately 20%) in the patient population included in our trial. Table 6.3 shows the largest 
relative risk (for mortality) that we can detect in the treatment versus control group, with a sample 
size of 500 (including dropouts) and 80% power, and for different assumptions of overall 
mortality in the control arm (ranging from 10-35%). 
 
Table 6-3. Detectable Relative Risk (Hazard Ratios) under different scenarios of 
control arm mortality with a two-sided test, 80% power and type I error rate of 5%. 
Secondary outcome 
Control arm 
assumption (%) 
Hazard Ratio detectable* 
Mortality (at 28 days) 
35 
30 
25 
20 
15‡ 
10 
0.62 
0.60 
0.56 
0.52 
0.45 
0.35 
* Assumes control arm mortality rate in table, power = 80%, and 0.05 alpha 
‡ From GRAIL Phase 2 
 
Based on this table, with the current sample size of 500 and assuming a control arm mortality of 
20% (more realistic assumption of mortality for GRAIL Phase 3), the proposed trial will have 
80% power to detect a hazard ratio of 0.52 for mortality between the Ganciclovir arm and the 
Placebo arm.  
 
 Statistical Analyses for Endpoints.  
6.2.1 
Primary Endpoint. 
We will first compare baseline characteristics of the ganciclovir and placebo groups in 
assessment of the randomization process. Then to test the primary hypothesis that the mean 
difference in RSFDs between groups differs significantly from 0, we will use the 2-sample 
parametric t-test. The t-test is powerful and will be sufficient for our purpose, even if normality 
assumption for RSFDs fails to hold, as the t-test is robust to non-normality assumption for n>25. 
However, as a confirmatory test, we will also use the 2-sample permutation t-test. The 
permutation test does not require any distributional assumption on the data and is valid under the 
much weaker assumption of exchangeability (which will be satisfied under controlled settings of 
a trial). As the next step, we will use the semiparametric efficient and robust method of Davidian 

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et al. [89, 90] to estimate the mean difference in the primary endpoint (intervention vs. control) 
with a 95% confidence interval, and to test for whether the mean difference differs from 0. This 
method leverages information in baseline subject characteristics predictive of the primary 
endpoint to maximize power and precision and is more efficient than a t-test for comparing 
baseline subtracted levels or analysis of covariance. If subjects are missing a primary endpoint for 
reasons other than death, then the analysis method will accommodate for it by addressing the 
missing data mechanism and adjusting for it in the analysis plan (please see Section 6.4.1 for a 
discussion). Apart from the primary analysis, a classical intent-to-treat (cITT) and two modified 
intent-to-treat (mITT) analyses will be carried out for the primary endpoint. The primary analysis 
population will be all patients randomized who receive at least one dose of the assigned study 
product. The classical ITT population will consider all patients that are randomized, while the two 
modified intent-to-treat (mITT) populations will be: 1) excluding those patients who were 
withdrawn from the study for any reason (including initial positive CMV LFA test followed with 
confirmatory negative CMV ELISA test) after receiving at least one dose of study drug, and 2) 
only including patients who survived and were followed to day 28. 
In the primary analysis (as well as in the secondary and exploratory analyses), gender and 
race/ethnicity as biological variables will be included and examined. 
6.2.2 
Secondary endpoints 
For the quantitative secondary endpoints (including a key secondary endpoint of VFDs or 
ventilator-free days), the same method as described in the analysis plan for the primary endpoint 
will be used. For time to event endpoints including CMV reactivation at given thresholds, the 
Kaplan-Meier method will be used to estimate the probability of not experiencing CMV 
reactivation by Day 28 for each group. A 95% confidence interval about the group difference in 
event rates will be computed using the two Kaplan-Meier estimates and the two Greenwood 
variance estimates. Based on these estimates, a Z-statistic will be used for testing for difference in 
the group event rates. The multiple testing problem will be handled using a Bonferroni correction, 
whenever such situations arise. Additionally, multivariate models will be built for the primary 
endpoint (RSFD) as well as other secondary endpoints, after adjusting for baseline subject 
characteristics and risk factors for CMV reactivation, using generalized linear models or the 
semiparametric efficient and robust method of Davidian et al. [89, 90]. The list of baseline 
characteristics to be considered for adjustment are those listed in (1) under the Exploratory 
Objectives/Endpoints (Section 5.3). We will repeat the above analyses in several subgroups 
(separately) as well, including those who are PCR negative at baseline, as well as those that have 
septic shock, pneumonia, unilateral infiltrates, or lymphopenia at the time of randomization, and 
by age and sex. In addition, we will repeat the above analyses in the following subgroups as 
follows: (1) VFDs in those on invasive mechanical ventilation at randomization; (2) RSFDs in 
those on invasive mechanical ventilation at randomization; (3) RSFDs in those on NIV/high flow 
at randomization; (4) Intubation after enrollment in those on NIV/HFNC at randomization (i.e. 
does ganciclovir affect the need for intubation in this subgroup?); (5) RSFDs according to 
COVID-19 status at randomization; and (6) CMV detection in plasma by day 28 (28 (>0 IU/mL, 
>1000 IU/mL) in those with invasive mechanical ventilation vs. NIV/high flow at randomization.  
We will also compare the duration of mechanical ventilation and respiratory support by day 28 
among survivors between the two intervention arms.  If the rate of death by day 28 differs 
between the two groups, then the analysis in survivors may be biased. If there is evidence for a 
differential death rate, then a sensitivity analysis will be conducted to evaluate how the estimated 
mean difference changes with a range of assumptions about the degree of possible selection bias. 
The sensitivity analysis method of Shepherd will be used [91], which was designed to address 
“truncation by death”.  A mediation analysis will also be conducted to test if CMV viremia lies in 
the causal pathway linking the treatment (Ganciclovir) effect with the outcome (RSFDs), that is, 

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if CMV viremia, which is assumed to be affected by the treatment, itself affects the outcome or 
not. 
This analysis will be conducted in four steps: 
1. A regression analysis with Treatment predicting RSFDs to test for path c alone, 
2. A regression analysis with Treatment predicting CMV Viremia to test for path a, 
3. A regression analysis with CMV Viremia predicting RSFDs to test the significance 
of path b, 
4. A multiple regression analysis with Treatment and CMV Viremia predicting RSFDs. 
               
 
 
The purpose of Steps 1-3 is to establish that zero-order relationships among the variables exist. If 
one or more of these relationships are non-significant, it can be concluded that mediation is not 
likely. If there are significant relationships in Steps 1 through 3, Step 4 is conducted. In the Step 4 
model, some form of mediation is supported if the effect of CMV Viremia (path b) remains 
significant after controlling for Treatment. If Treatment is no longer significant when CMV 
Viremia is controlled, the finding supports full mediation. If effect of CMV Viremia is still 
significant, the finding supports partial mediation. Apart from this, static respiratory system 
compliance (Crs) will also be used in a separate mediation analysis to understand whether any 
effect of Ganciclovir is exerted via pulmonary mechanisms. 
Alternative approaches to Analyze RSFDs. Our secondary analysis plan also includes a pipeline 
for exploring alternative approaches to analyze RSFDs. Given that RSFDs are increasingly being 
used as an outcome in high profile RCTs (as mentioned in Section 4.5.1, RSFDs or organ 
support-free days have been used as the primary endpoint in several recent high-profile RCTs 
[83-85]), these proposed statistical considerations can serve as a reference point for RSFD 
analyses in future clinical trials. Thus, we believe this to be one of the statistical innovations 
proposed by the Data Coordinating Center (DCC) for this Phase 3 trial.  
One important limitation to using RSFDs as an endpoint arises, as previously stated, due to its 
composite nature. RSFD assigns points based on counting back from day 28 to the last day when 
respiratory support was received (see Section 6.1.1). The outcome also considers mortality and 
assigns an overall score of zero points to those who die within those first 28 days. Thus, the 
composite nature of this endpoint leads to it being a zero-inflated binomial random variable, 
which has a mixture distribution of a degenerate 0 and a binomial distribution. That is, we can 
write  
!"#$U ~ V × 0 + (1 −V) × X-(28, Z) 
 
where V is the probability that a death occurs, and X-(28, Z) is a binomial distribution such that Z 
is the probability of a subject experiencing a RSFD while being alive through the specified 
window, that is desired to be modeled. The zero inflated binomial response is a special instance 
of over-dispersed responses, and there is considerable literature on the numerous challenges and 

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statistical issues when modeling such data [92, 93]. Thus, modeling of RSFDs can be handled 
through a number of approaches: 
1. Account for over-dispersion through the zero-inflated binomial model: The zero-inflated 
binomial model is obtained by mixing a degenerate distribution at zero with a standard binomial 
distribution [94, 95], and has been subsequently used in epidemiology and health economics to 
model binomial counts with excessive zeros. The zero-inflated binomial model will be used as an 
alternative approach to model RSFDs in this analysis. 
2. Account for over-dispersion though the beta-binomial model: The beta-binomial distribution is 
a discrete probability distribution which arises when the probability of success in a fixed number 
of Bernoulli trials is either unknown or random. It has frequently been used in classical statistics 
to capture over-dispersion in binomial type distributed data [96, 97], and this will be used as 
another alternative approach. 
3. Separate modeling of composites: In addition to the above approaches, the composite outcome 
will also be modeled separately for the primary analysis population and considered for 
assessment. Mortality will be modeled as a time to event outcome, and the Cox model will be 
used to evaluate whether the treatment (ganciclovir) has any effect on time to death. On the other 
hand, duration of time off respiratory support (total time in days not spent receiving respiratory 
support within the first 28 days), regardless of the mortality status, will be analyzed separately in 
a Generalized Linear Model to evaluate if ganciclovir increases total time off respiratory support 
within the first 28 days. 
6.2.3 
Exploratory endpoints 
To identify risk factors for CMV reactivation, we will first build univariate generalized linear 
regression models to test the association between CMV reactivation and different potential risk 
factors identified in Section 1 C.2. We will also build multivariate Generalized Linear Models for 
association with CMV reactivation, by including (i) factors found significant at level 0.2 in the 
first step and (ii) all risk factors, and then using a step-wise backward selection using the Akaike 
Information Criterion (AIC) or the Bayesian Information Criterion (BIC) for model selection.  
We will use descriptive statistics and plots to examine the association of CMV viral load kinetics 
in blood and lung compartments with RSFDs and VFDs, mortality at day 28 and 180, duration of 
respiratory support in survivors, PaO2/FiO2, and static respiratory system compliance. We will 
also build regression models (generalized linear models) to examine these relationships more 
closely. We will also descriptively determine the sensitivity and specificity of the rapid lateral 
flow CMV serostatus assay, and assess its concordance with the standard clinical assays, using 
Cohen’s kappa statistic. 
6.2.4 
 Other pre-specified analyses  
Primary analysis, cITT, mITT, and survivors 
Ideally, any patient randomized would be included in the conventionally defined ITT analysis 
cohort. Nonetheless in an emergency care setting, such as this study, the following unusual but 
important scenario will need to be considered: a patient is eligible and randomized but by the time 
of drug delivery no longer meets eligibility criteria and thus, should have been excluded. 
Accordingly,it is intended that patients randomized but not having drug/placebo delivered will 
not be included in the primary analysis cohort. The integrity of randomization shall remain 
preserved, as the decision to deliver drug/placebo is determined a priori and completely 
independent of treatment assignment (14). If a participant is withdrawn from the study before a 
dose of study drug is given, no study procedures will be performed and no study data will be 
collected, and these participants will be excluded from the primary analysis. However, if a 
participant is withdrawn after receipt of study drug, safety procedures (e.g. monitoring of 

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creatinine and CBC with differential) will continue for another 48 hours, no study biosamples 
will be obtained, and study data in the medical record will continue to be collected per protocol 
(e.g. respiratory-support-free days), and they will be included in the primary analysis. In addition 
to the primary analysis, we will also conduct a randomized or classical ITT analysis, which will 
consider all patients randomized. This will be used as a conservative alternative to our primary 
comparison. Additionally, we will also perform 2 modified intent-to-treat (mITT) analyses: 1) 
excluding patients who were withdrawn from the study for any reason (including initial positive 
CMV LFA test followed with confirmatory negative CMV ELISA test) after receiving at least 
one dose of study drug, and 2) only including patients who survived and were followed to day 28. 
We will also test for an interaction of treatment period (pre and post drug use) and the primary 
treatment differences at the study conclusion. Finally, we will perform subgroup analyses 
evaluating treatment effect stratified on mode of respiratory support.  
 
6.2.5 
Post hoc analyses 
Statistical considerations for the post hoc analyses described in Section 5.4 are described in the 
statistical analysis plan.  
 
 Randomization scheme 
The randomization sequence will be uploaded by SCHARP into the Medidata’s Randomization 
and Trial Supply Management (RTSM) system and provided to each site through Medidata. The 
randomization will be block-randomized by site. At each institution, the pharmacist with primary 
responsibility for drug dispensing is charged with maintaining the security of the randomization 
list.  A research pharmacy designee may be provided access to the randomization assignment 
from Medidata RAVE. 
 
 Blinding 
Participants and site staff (except for site pharmacists) will be blinded as to treatment arm 
assignments (e.g., study drug or placebo). Study drug assignments are accessible to those site 
pharmacists, contract monitors (or the central site IDS pharmacist as backup), and unblinded 
statisticians who are required to know this information to ensure proper trial conduct. Access to 
randomization assignment is restricted to unblinded statisticians only. Emergency unblinding if 
ever needed is managed 24/7 through Medidata RTSM. Any discussion of study drug assignment 
between the site clinical and pharmacy staff is prohibited. The DSMB members also are 
unblinded to treatment assignment to conduct review of trial safety. Thus, closed reports for the 
DSMB will include the treatment indicators of the two participant groups. 
Unblinding procedures are discussed in Section 9.15.  
6.4.1 
Missing data 
Every effort will be dedicated to complete, comprehensive, and accurate data collection and data 
annotation by the DCC.  However, issues and challenges arise in every clinical trial. Due to the 
high proportion of deaths in this patient population, one major challenge in this RCT of which we 
must be aware and ready to correct is missing data. Missing data can occur to the primary 
endpoint and key secondary endpoints (e.g., CMV reactivation [missing viral load values] and 
organ dysfunction or the NHLBI-endorsed acute respiratory failure core outcome measurement 
set). We have carefully examined sources of missing data from the GRAIL phase 2 trial and 
adjusted the analysis plan accordingly. 

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6.4.1.1 
Missing Data in Primary Outcome.  
It is highly likely that missing data in RSFDs, the primary outcome in our RCT, will be MNAR 
(missing not at random), as most missingness will result from withdrawal of consent, a rare 
occurrence that can happen when patients and families are in crisis, and is more likely to occur in 
sicker patients. However, we only expect <3% missingness in our primary outcome (see Section 
2A), and thus this data can be ignored from the primary analysis. However, if for some 
unforeseen reasons, missingness in RSFDs does increase beyond a set threshold of 5%, we will 
have to adjust for it in our analysis plan. In that case, we will use baseline patient characteristics 
identified in (1) under the Exploratory Objectives/Endpoints (Section 5.3) to model the missing 
data mechanism in RSFDs, which will be used to estimate the treatment effect unbiasedly using 
maximum likelihood methods, provided the missing data model is correctly specified.   
6.4.1.2 
Missing Data in some secondary outcomes due to Study Design.  
The absence of data pertaining to some secondary endpoints (for example, static respiratory 
system compliance) may be problematic if participants are discharged from the ICU, are on 
pressure support after being extubated, or expire prior to our pre-specified time points for 
endotracheal aspirates and serum collection. For example, if the Ganciclovir intervention reduces 
duration of respiratory support, it may bias the results because participants cannot undergo 
endotracheal aspirate if they are extubated. Thus, to minimize missing data and to maximize the 
CMV detection rate, we have selected multiple time points for the endotracheal aspirate (Day 1 
and twice weekly until Day 28). If these issues still occur, we will address them in the analysis 
phase using data imputation methods, or by acknowledging it as a limitation of any analyses 
conducted with these variables. 
6.4.1.3 
Missing Data in covariates. 
Missing data can occur in covariates as well, due to factors such as death, withdrawal, or other 
reasons; this will need to be accounted for in our analyses, if overall missingness (in covariates) 
increases beyond 5%. Since we expect covariate missingness in this study to be MAR (missing at 
random), we will use weighting adjustments (for example, weighted generalized estimating 
equations) or multiple imputation methods [98, 99] to deal with covariate missingness. 
 
 Planned analyses prior to end of study 
6.5.1 
Safety 
The DSMB will have access to unblinded safety data at the time of planned reviews and upon 
request of the DSMB may review additional analyses. Operating details are specified in the 
DSMB charter. A scheduled interim safety analysis at midpoint will be performed.  
The safety and protocol adherence review team (SPART) will review all clinical and laboratory 
safety data during the course of the study, at least every 3 months. The site teams are responsible 
for regularly monitoring all Adverse Events and documenting reportable AEs to the DCC and 
CCC for review by the SPART.  The SPART will include CCC PIs (Drs. Boeckh, Stapleton, 
Rubenfeld, and Limaye), Clinical Site Manager Sara Ardren, and Clinical Trial Project 
Manager/Site Monitoring Manager Dr. Louise Kimball. Meetings of the SPART (by 
videoconference or teleconference) will occur regularly, and will not require attendance of all 
team members at each meeting. The SPART is responsible for the review of the clinical safety 
reports, discordant CMV serostatus test results (LFA versus CLIA test), and protocol violations, 
as well as communication with the IRB, NHLBI executive secretary, and DSMB as outlined in 
Figure 11.1. 
 

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6.5.1.1 
Interim safety analysis. 
It is expected in this trial that approximately 20% or approximately = 100 of the 500 participants 
will have death events relative to the safety endpoint. A two-sided interim safety analysis is 
planned to be performed at the midpoint, either at the 50th event or when 50% of participants (n = 
250) are randomized, whichever occurs earlier. 
Guidelines for early termination at the interim analysis due to concerns on the safety endpoint 
should (i) adjust for the nature of interim monitoring that involves repeated testing over time, (ii) 
reflect particular caution given the relative benefit-to-risk profile of the two arms. 
Specifically, a recommendation for stopping will be based on strong evidence for the hazard ratio 
(treatment/placebo, HR) of death to be less than 1 or greater than 1. The O’Brien-Fleming “upper 
boundary” will be used to establish if an elevated event rate in the intervention group preserves 
the (one-sided) 0.025 false positive error rate relative to the hypothesis: 
H0: the event rate for the intervention group relative to control ≤ 1.00, or HR ≤ 1.  
The O’Brien-Fleming “lower boundary” will be used to establish if an elevated event rate in the 
control group preserves the (one-sided) 0.025 false positive error rate relative to the hypothesis: 
H1: the event rate for the control group relative to intervention ≤ 1.00, or HR ≥ 1. 
For illustration, Table 6.5.1-1 below presents the O’Brien-Fleming boundaries for the hazard ratio 
(HR) estimates that would lead to rejection of H0 at the interim analysis performed when one has 
observed 50% and 100% of the trial’s expected total of 100 death events. 
 
 
 
 
 
 
 
 
Observe that, for the total of 50 events at the interim analysis, to reach the O’Brien-Fleming 
boundary for a lower death rate in the intervention group, the control group would need to have at 
least 20 excess events (15 in intervention group versus 35 in the control group) at the 50% 
information fraction. Similarly, to reach the O’Brien-Fleming boundary for a lower death rate in 
the control group for the total of 50 events at the interim analysis, the intervention group would 
need to have at least 20 excess events (15 in control group versus 35 in the intervention group) at 
the 50% information fraction. The Lan-DeMets implementation of the O’Brien-Fleming guideline 
will be used to provide flexibility in the timing and number (in the case of unplanned DSMB 
meetings) of interim analyses [100]. 
An efficacy analysis (of the primary endpoint) will also be conducted at the time of interim look. 
Although no formal stopping rule will be implemented for either efficacy or futility, number of 
participants (or outcome data) needed to change the direction of the observed treatment effect will 
be estimated at the time of the interim analysis, which is required for the DSMB’s consideration 
Table 6.5.1-1: Interim analysis assumptions 
 
Information Fraction 
(% of Total Events) 
Reject H0: 
HR ≤ 1.00 
Nominal one-sided p- 
values for rejection of H0 
Reject H1: 
HR ≥ 1.00 
50% (50 events) 
≥ 2.316 
P ≤ 0.0015; Z = 2.97 
≤ 0.432 
100% (100 events) 
≥ 1.480 
P ≤ 0.025; Z = 1.96 
≤ 0.676 

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of the data. Additionally, at the time of the interim analysis, variance of the actual data will be 
assessed in accordance with the assumptions made in the sample size calculations. 
6.5.2 
Other endpoint analyses 
Distribution will be limited to those with a need to know, for informing future trial-related 
decisions. To guarantee the unrestricted performance of its task, the DSMB may receive the 
individual study morbidity and mortality data from an unblinded statistician. The DSMB will 
have access to unblinded safety data at the time of planned reviews and upon request of the 
DSMB may review additional analyses. Any analyses conducted prior to the end of the study 
should not compromise the integrity of the trial in terms of participant retention or safety or 
immunogenicity endpoint assessments. 
6.5.3 
Sample size re-estimation plan 
Considering that there is no substantial literature on the distribution of RSFDs, our trial design 
relies on GRAIL Phase 2 data on VFDs instead and our belief that variability in RSFDs will be 
similar to that of VFDs. However, in case it happens that variance of RSFDs is substantially 
larger than that of VFDs (which can happen for a variety of reasons including the fact that RSFDs 
is expected to be measured on a wider population than those who typically only require 
ventilation, which may introduce additional sources of variability), the common SD used in the 
sample size determination (Section 6.1.1) for the study may not reflect the actual value. To ensure 
the study remains adequately powered, the assumption of variability will be checked when the 
study is halfway done, ie, when endpoints for '∗= 250 ~ 50% of total initially planned sample 
size subjects (500) have been measured. Specifically, total variance will be calculated in a blinded 
way using the following approach, similar to the one proposed by Kieser and Friede as well as 
Zucker et al [101, 102]. 
"]$ =
1
'∗−1 ^_!"#$% −!"#$
````````4∗a
$
4∗
%5!
−
'∗
4('∗−1) Δ$ 
Where '∗: total number of subjects at re-assessment; = '/2 where ' = 23 is total sample size 
in the study 
!"#$%: RSFDs in patient 4 in the reassessment. 
!"#$
````````4∗: average of !"#$%, 4 = 1, … , '∗ 
Δ: expected difference in RSFDs between the arms (based on GRAIL Phase 2 data on 
VFDs) 
Further investigation done by Keiser and Friede[101] indicates that this method has little impact 
on the type I error of the test for superiority. Therefore, no type I error adjustment needs to be 
made. The formula for blinded estimation of variance given above depends on the assumed 
treatment effects. It was noted that if the treatment effect is mis-specified, the estimated variance 
will be biased and may be under-estimated. However, as pointed by Kieser and Friede, such bias 
is generally negligible in most clinical trial situations. 
If the estimated pooled SD from all available subjects at sample size re-assessment is 
considerably larger than our assumption of 9.4 days, sample size will be recalculated based on the 
estimated SD from the re-assessment using a similar approach as described in Section 6.1.1. 
Table 6.5.3 below provides details on the sample size re-assessment under the same targeted 
power (85%), the same treatment differences (2.74 RSFDs), and numbers of additional subjects 
needed for the larger SD values. The blinded variance estimation will be performed by one of the 
blinded statisticians on the trial. In work by Laterre et al [83], ventilator and vasopressor free days 

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within 30 days had an estimated SD around 13 and we suspect SD of our RSFD primary endpoint 
would not go beyond that.  
 
 
 
Table 6.5.3: Sample size re-assessment to achieve 85% power 
 
Estimated 
SD 
Sample 
size per 
arm 
Adjusted 
Sample size 
per arm 
Total 
Sample 
Size  
Overall 
sample size 
increase 
≤ 9.4 days 
213 
250 
500 
0 
9.8 days 
231 
271 
542 
42 
10.2 days 
250 
293 
586 
86 
10.6 days 
270 
317 
634 
134 
11 days 
291 
341 
682 
182 
11.4 days 
312 
366 
732 
232 
11.8 days 
334 
392 
784 
284 
12.2 days 
357 
419 
838 
338 
12.6 days 
381 
447 
894 
394 
13 days 
406 
476 
952 
452 
 

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7 
SELECTION AND WITHDRAWAL OF SUBJECTS 
 Study population 
Five hundred adults will be randomized in a 1:1 ratio to receive either the study drug or placebo. 
All patients entered into this study will have established respiratory failure associated with sepsis. 
By virtue of their need for respiratory support (as defined in Section 4.1.1) within an intensive 
care unit (ICU), all patients will be considered critically ill.  
Final eligibility determination will depend on results of laboratory tests, medical history, and 
physical examinations. Those determined to be eligible, based on the inclusion and exclusion 
criteria, will be enrolled in the study. Investigators should always use good clinical judgment in 
considering a subject’s overall appropriateness for trial participation. Some subjects may not be 
appropriate for enrollment even if they meet all inclusion/exclusion criteria because medical, 
psychiatric, social, or logistic conditions may make evaluation of safety and/or efficacy difficult. 
Duration of participation in the study by individual patients will be 180 days. 
 
 Randomization 
Patients meeting inclusion and exclusion criteria will be randomized to standard ICU care 
(including standard lung protective ventilation and weaning protocols, if mechanically ventilated) 
+ intervention or placebo. 
  
 Inclusion criteria 
1. Subject/next of kin informed consent 
2. Age > 18 years 
3. CMV IgG seropositive by lateral flow assay (LFA) or standard serologic methods 
4. Receiving care from an ICU team/service 
5. Acute respiratory failure as defined in Section 4.1.1. 
6. Expected to require respiratory support for at least 2 more days after randomization 
7. Infection confirmed or suspected by the treating clinician and felt to be causing or contributing 
to acute respiratory failure (Respiratory failure associated with infection confers at least 2 
SOFA points above assumed baseline SOFA score of 0, thereby meeting Sepsis-3 definition).  
 
 Exclusion criteria 
1. Known or suspected immunosuppression, including:  
a. HIV+ (i.e. prior positive test or clinical signs of suspicion of HIV/AIDS; a negative HIV 
test is not required for enrollment) 
b. stem cell transplantation:  
i. 
within 6 months after autologous transplantation or 
ii. within 1 year after allogeneic transplantation (regardless of immunosuppression) 

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iii. greater than 1 year after allogeneic transplantation if still taking systemic 
immunosuppression or prophylactic antibiotics (e.g. for chronic graft versus host 
disease) 
Note: if details of stem cell transplantation are unknown, patients who do not take 
systemic immunosuppression and do not take anti-infective prophylaxis are 
acceptable for enrollment and randomization.  
c. solid organ transplantation with receipt of systemic immunosuppression (any time) 
d. cytotoxic anti-cancer chemotherapy within the past three months (Note: next-of-kin 
estimate is acceptable) 
e. congenital immunodeficiency requiring antimicrobial prophylaxis (e.g. TMP-SMX, 
dapsone, antifungal drugs, intravenous immunoglobulin) 
f. receipt of one or more of the following in the indicated time period (see Appendix C): 
i. within 6 months: prednisone, alemtuzumab, antithymocyte/antilymphocyte 
antibodies, or other immunosuppressive drugs associated with CMV reactivation  
Note:  
– 
If no information on these agents is available in the history and no direct or 
indirect evidence exists from the history that any condition exists that 
requires treatment with these agents (based on the investigator’s assessment), 
the subject may be enrolled. For all drug information, next-of-kin estimates 
are acceptable.  See Appendix C for commonly prescribed 
immunosuppressive agents.  Information on the use of biologics with 
moderate immunosuppressive effect but no known effect on CMV are 
permitted and will be recorded in the CRFs. 
– An average of >20mg/day of prednisone for the past 30 days prior to hospital 
admission. The total prednisone intake should be added and averaged over 30 
days.  For example, a patient taking 40mg/day for 14 days would not be 
excluded because (40x14)/30=18.7.  Other steroids should be converted into 
prednisone equivalents. Additionally, any dose of corticosteroids given for 
any reason on or after hospital admission is acceptable. 
2. Expected to survive < 72 hours (in the opinion of the investigator), or not committed to full 
intensive care support at the time of study enrollment (DNR but otherwise committed to full 
support is acceptable). 
3. Unable to start receiving first dose of study drug within 120 hours after hospitalization (as 
measured from admission or time of transfer; subjects who are transferred from a chronic 
care ward, such as a rehabilitation unit, with an acute event are acceptable). 
4. Pregnant or breastfeeding (either currently or expected within one month).  
Note: for women of childbearing age (18-60 years, unless documentation of surgical 
sterilization [hysterectomy, tubal ligation, oophorectomy]), if a pregnancy test has not been 
done as part of initial ICU admission work-up (within 120 hours before enrollment), it will 
be ordered stat and documented to be negative before randomization. Both urine and blood 
tests are acceptable.  
5. Absolute neutrophil count < 1,000/mm3 (if no ANC value is available, the WBC must be > 
2500/mm3) 

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6. Use of anti-CMV drugs (cidofovir, letermovir, foscarnet, valganciclovir, ganciclovir) within 
seven (7) days of patient randomization.  
7. Use of IVIG within four (4) weeks of patient randomization [103].  
8. Currently intubated for airway protection only.  
9. Currently enrolled in an interventional trial of an investigational therapeutic agent known or 
suspected to have anti-CMV activity or to be associated with significant known hematologic 
toxicity (prior approval required). 
10. At baseline patients who have a tracheostomy, and have been receiving any positive pressure 
ventilation through it during the 30-day period prior to ICU admission. 
11. Patients with Child Class C Cirrhosis.  
12. Patients with severe (requiring home oxygen) pre-existing interstitial lung disease. 
13. Allergy to ganciclovir 
14. Incarcerated 
15. Other, specify (e.g. clinician refusal) 
 
 Subject withdrawal 
Under certain circumstances, an individual patient must be terminated from participation in this 
study. Specific events that will result in early termination include: 
• 
Randomized after positive LFA results but prior to negative CLIA-approved ELISA test 
result, 
• 
 Need for respiratory support ends unexpectedly between time of randomization and first 
study product administration, 
• 
Unexpected death between time of randomization and first study product administration, 
• 
Subject has been inappropriately enrolled based on inclusion/exclusion criteria (e.g. when 
information through next of kin was inaccurate),  
• 
Site investigator decides to terminate participation for reasons of patient’s safety or to prevent 
compromising the scientific integrity of the study, 
• 
It is determined that side effects are severe,  
• 
New scientific developments indicate that the treatment is not in the patient’s best interest,  
• 
Patient or next of kin refuses further participation, 
• 
Study is terminated.  
Patients may be withdrawn at any time once the study team becomes aware that the patient meets 
one or more criteria for withdrawal. If the participant is withdrawn from the study before a dose 
of study product is given, no study procedures will be performed, no study data will be collected, 
and they will not be included in the primary analysis population.  If a participant is withdrawn 
after receipt of study product, safety procedures (e.g. monitoring of creatinine and CBC with 
differential) will continue for another 48 hours, no further study biosamples will be obtained, 
study data in the medical record will continue to be collected per protocol and as allowed in the 
signed consent (e.g. respiratory-support-free days), and they will be included in the primary 
analysis population. 

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In the event that a participant does not start receiving first dose of study product within 120 hours 
after hospitalization, and this is discovered before 5 doses of study drug are administered, they 
must be withdrawn. In this circumstance, no further study procedures will be performed, no 
further study data will be collected, and they will not be included in the primary analysis 
population. If initiation of study drug past the 120-hour window is discovered after 5 or more 
doses of study drug have been given, the participant may remain in the study. 
 
 Subject replacement 
A participant may be replaced in the study under certain circumstances of early withdrawal. The 
purpose of participant replacement is to compensate for potential data loss. The following 
circumstances may qualify a participant for replacement:  
• 
Randomized after positive LFA results with subsequent negative CLIA-approved ELISA test 
result and before administration of study drug,  
• 
Participant did not start receiving first dose of study product within 120 hours after 
randomization, and was withdrawn, 
• 
 Need for respiratory support ends unexpectedly between time of randomization and first 
study product administration, 
• 
Unexpected death between time of randomization and first study product administration, 
• 
Early termination between time of randomization and first study product administration (e.g., 
due to patient or next of kin refusing further participation, new scientific developments 
indicate that the treatment is not in the patient’s best interest, or subject has been 
inappropriately enrolled based on inclusion/exclusion criteria),  
• 
Participant who declines further participation and requests complete deletion of all data after 
regaining consciousness. 
 
 
 
 

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8 
STUDY DRUG ACQUISITION, PREPARATION, & ADMINISTRATION 
 Study drug & placebo formulation  
Intravenous ganciclovir and matching placebo. 
 
 Acquisition of study drugs & placebos 
Study drug will be purchased by UW Pharmacy and supplied to study sites. In the event that UW 
Pharmacy is unable to source sufficient supply or if the drug cannot be directly supplied (e.g. to 
international sites), study drug from a commercial supplier may be used or purchased directly by 
study sites (costs to be passed through to the CCC). This provides necessary flexibility to prevent 
disruption of per-protocol administration of study drug due to global supply chain challenges or 
drug shortages. The drug supply must be FDA approved commercial ganciclovir sourced from a 
licensed distributor/manufacturer. The site will be responsible to appropriately document all 
doses prepared from local drug supply. 
 
 Storage of study drugs & placebos 
Study drug will be stored as per manufacturer’s recommendations.  
 
 Administration of study drugs & placebos 
Ganciclovir (or IV placebo) will be administered via central or peripheral venous access.  
Ganciclovir doses must be adjusted according to renal function as per package insert.  A subject 
who is on hemodialysis should continue IV dosing according to the package insert.  
 
 Pharmacy Records 
The site pharmacist is required to maintain complete records of all study drugs received from the 
sponsor or purchased by the site and subsequently dispensed. When using site supplied drug, sites 
are to maintain a record of manufacturer/lot number used for dose preparations.  

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9 
CLINICAL PROCEDURES 
 Patient identification & recruitment 
Patients with sepsis-associated acute respiratory failure will be identified via prospective 
screening of all ICU patients. This process is done by trained and experienced research 
coordinators who review charts using a standardized screening tool. Additionally, patients may be 
identified by the attending physician based on eligibility criteria.  
 
 Informed Consent 
Informed consent is the essential processes of ensuring that study subjects or legal guardians fully 
understand what will and may happen to them while participating in a research study. Before any 
protocol-specific questions are asked or procedures to determine protocol eligibility performed, a 
screening consent form or protocol-specific consent form (described below) must be obtained. 
Patients or family members must be provided with a copy of all consent forms that they sign. 
Since all potential patients will be intubated and sedated or requiring high levels of respiratory 
support, initial consent for patients who cannot make decisions for themselves will be from the 
patients’ LAR. Subsequent consent from the patient will be obtained whenever possible. 
Interested surrogates will be given information about the study, explaining potential risks. They 
will then undergo informed consent. Consent forms will be approved by the Human Subjects 
Committee.  
Participation in this study is voluntary. The nature of the study will be fully explained to each 
patient during the informed consent process. If the patient is deemed unable to provide written 
informed consent, informed consent for the patient’s participation must be obtained from a LAR 
using practices and procedures that are acceptable as defined by local law and the Institutional 
Review Board. In this situation (the use of surrogate consent), subsequent in-person consent will 
be obtained from the patient when possible before hospital discharge.  In circumstances where the 
patient has regained the capacity to consent but is discharged before study personnel have an 
opportunity to schedule an in-person meeting, then a remote consent method of phone/video 
conference will be used instead. The patient (or authorized representative, when applicable) will 
have the opportunity to ask questions. The patient (or authorized representative, when applicable) 
and the individual who performs the consent discussion will sign an informed consent document. 
The investigator will retain the informed consent document according to Good Clinical Practice. 
HIPAA authorization will also take place during the informed consent process. 
The determination of appropriate “next-of-kin” will be made in accordance with the standard 
practices used in provision of medical care. Detailed documentation of all attempts to obtain 
consent from the patient and/or the patient’s next-or-kin will be kept. 
9.2.1 
Consenting process 
Informed consent is not limited to the signing of the consent form; it also includes all written or 
verbal study information site staff discuss with the patient, before and during the trial. Once  an 
eligible participant is identified, the study team will coordinate a consent conference with the 
patient or LAR. This can be done in person or via phone/video conference. The patient or LAR 
will have a copy of the consent form to review during the conference. The patient or LAR will be 
given ample time to review the consent and ask questions.  
When a previously incapacitated patient regains capacity to consent, the study team will schedule 
a follow up consent conference with the participant and document their willingness to continue 
participating in the study.  

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 The consenting process for each site will be documented in the individual “Participating Site 
Application”. Participating sites (initial sites listed in Appendix H) may use resources available to 
their site to conduct eConsent if it meets FDA 21 CFR Part 11 compliance.  A template 
Prescreening consent (for participants where leftover blood sample is not available) and Main 
consent template will be given to the participating sites to modify. 
9.2.2 
Consent form 
The informed consent form documents that a prospective patient or their agent (1) understands 
the potential risks, benefits, and alternatives to participation, and (2) is willing to participate in a 
study. 
This study will conduct multi-site research under the regulatory approval of a Single Institutional 
Review Board (sIRB) in accordance with the NIH Policy on the Use of a Single Institutional 
Review Board for Multi-Site Research. The Fred Hutchinson Cancer Center IRB 
(FWA00001920) will serve as the sIRB of record that is responsible for overseeing the conduct of 
this study. All collaborating sites (initial sites listed in Appendix H) have agreed to rely on the 
sIRB and any sites added after study initiation will also be required to rely on the sIRB. The 
Principal Investigator (PI) will disseminate the proposed informed consent to all participating 
sites and will submit these materials to the sIRB for review and approval. Any subsequent 
changes will be distributed to sites and submitted to the sIRB as a modification to be approved 
prior to implementation. The consent form(s) must be developed in accordance with local 
IRB/IEC requirements and the principles of informed consent as described in Title 45, Code of 
Federal Regulations (CFR) Part 46 and Title 21 CFR, Part 50, and in the International Conference 
on Harmonisation (ICH) E6: Good Clinical Practice: Consolidated Guidance 4.8. It must be 
approved by all responsible ethical review bodies before any subjects can be deemed to have 
consented for the study.  
 
 Screening procedures 
Screening procedures are done to determine eligibility and to provide a baseline for comparison 
of data. Baseline data are obtained during screening. All inclusion and exclusion criteria must be 
assessed within 120 hours before randomization. Importantly, the patient can only be randomized 
once these test results are available.  
Before randomization, the following procedures are performed: 
• 
Clinical laboratory tests as defined in the inclusion and exclusion criteria, including: 
o Serum or urine pregnancy test—the results of this must be negative before 
proceeding, since ganciclovir is suspected to be teratogenic. 
o CMV serology by lateral flow assay (LFA) or gold standard test. See Figure 9-1 and 
Section 9.3.1 for the specific algorithm by which the LFA and confirmatory gold 
standard test will be used for CMV serology.  
o Leftover serum (preferred) or plasma may be used if available.  
o If leftover material is not available, a small amount of extra blood (0.5-1.5 ml) 
should be obtained after prescreening consent is administered to permit such 
testing. Prescreening consent may be obtained verbally or as written consent, as 
permitted per IRB guidelines. 
o After lateral flow testing is obtained, a portion of the same sample should be sent 
to a CLIA-approved lab for testing. 
o Absolute neutrophil count/total white blood cell counts 

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• 
Collection of medical history 
• 
Collection of samples for CMV PCR in plasma and endotracheal aspirates as well as for 
storage for subsequent CMV immunity testing, genomic analysis, and ancillary studies 
• 
Assessment of select concomitant medications: steroids, immunosuppressive medications 
and antivirals  
• 
Net fluid balance at ICU admission 
• 
Obtaining of patient demographics in compliance with  the NIH Policy on Reporting 
Race and Ethnicity Data: Subjects in Clinical Research, Aug. 8, 2001. Available at 
http://grants.nih.gov/grants/guide/notice-files/NOT-OD-01-053.html 
 
9.3.1 
CMV Serology Screening Procedures 
 
CMV serology test results (for rapid CMV serology [LFA] and CLIA test) are reported as a 
qualitative “Positive” or “Negative”, and very rarely “Indeterminate.” 
 
Randomization can occur in the follow instances: 
• 
If the Rapid CMV Serology (LFA) comes back positive, randomization can occur prior to 
the CLIA test. 
• 
If the Rapid CMV Serology (LFA) comes back indeterminate or negative, and then the 
CLIA test comes back as positive, randomization can occur. 
• 
If sites do not perform a Rapid CMV Serology (LFA) test, and the CLIA test comes back 
as positive, randomization can occur. 
If Rapid CMV Serology (LFA) testing is done, a site progresses to the CLIA test as follows: 
• 
Regardless of whether the Rapid CMV Serology (LFA) comes back positive, 
indeterminate, or negative, progress to the CLIA test (using part of the same sample used 
for the rapid test, as described in section 9.3 above). 
• 
CLIA test result must be recorded as either positive or negative; an indeterminate CLIA 
test result should be recorded as negative.  
Rapid CMV Serology (LFA) is NOT required to progress to CLIA testing: 
• 
Sites can perform only CLIA testing and randomize with a single positive test result. 
 
Discordant results (between the LFA and CLIA test) that lead to administration of study drug will 
be tracked and reviewed regularly by the Safety and Protocol Adherence Review Team (Section 
14.3). 

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Figure 9-1. Flow diagram of serologic assessment strategy for inclusion using LFA with 
confirmation via standard serologic assay. PPV and NPV for LFA are calculated based on 
60% seroprevalence, using QNow UV Flashlight reader as is planned for this study. 
 
 
 Patient Registration 
Participating sites can verify a patient’s consent and eligibility and register the patient into the 
study. DCC will provide patient accrual reports. Randomization will be through MediData’s 
Balance system, 24/7, with a backup system in place. 
 
 Randomization procedure 
Randomization will occur after assessment of positive CMV serostatus as detailed in Section 
9.3.1, and negative pregnancy test. Randomization and first dose of study drug should occur as 
close to each other as possible. Randomization and first dose of drug must occur within 120 
hours of hospital admission and on the same calendar day. Patients who are randomized but in 
whom the need for respiratory support ends, who die, are outside of the 120 hour window, or 
have consent withdrawn before receiving first dose of study drug should not receive their first 
dose of drug and are withdrawn from the study (see Section 7.5).  
 The randomization sequence will be uploaded by SCHARP into the Medidata’s Randomization 
and Trial Supply Management (RTSM) system and provided to each site through Medidata. This 

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system automatically notifies the site pharmacist of the treatment assignment. The randomization 
will be block-randomized by site. At each institution, the pharmacist with primary responsibility 
for drug dispensing is charged with maintaining the security of the randomization list. Patients 
will be stratified at the time of randomization according to treatment center.  
When the patient is randomized, the following information is required by NIH reporting 
guidelines: date of birth, race/ethnicity, and gender. For the purpose of this study, each patient 
will be assigned a study number, which will be used for all communications with outside 
institutions to assure confidentiality. 
A CLIA-certified standard serologic assay will be used to confirm CMV positivity, but patients 
with a positive rapid test result may be randomized prior to the return of the CLIA test results (see 
Section 9.3.1).   
Randomization will continue until the target accrual number for participants in the primary 
analysis are met. The primary analysis population will be all patients randomized who receive at 
least one dose of the assigned study product. 
 
 First dose of study drug 
Study day 1 is defined as the first calendar day on which the patient is randomized and the first 
dose of study drug is administered. All subsequent study days will start accordingly.  At baseline, 
but before administration of study drug, the following procedures will need to be performed: 
• 
Blood: Creatinine, CMV cell-mediated immunity, platelets, CMV plasma PCR, CBC 
w/differential, research samples. 
• 
Endotracheal aspirate (ETA).  If patient is intubated, collect an ETA specimen at baseline 
(± 1 day) and twice weekly, at the time that this procedure is routinely performed by 
respiratory therapy. Specimens will be labeled and stored frozen for subsequent CMV 
PCR analysis at the coordinating lab at Fred Hutch.  
• 
Clinical Assessments: Apache III, SOFA, COMS survey (see Section 9.10), static 
respiratory compliance, ventilator parameters, assessment of select concomitant 
medications (steroids, immunosuppressive medications, neuromuscular blockers, and 
antivirals), use of ECMO and prone positioning, and bilateral vs. unilateral infiltrates on 
chest radiograph. See Section 9.11 for details on these assessments. 
 
 Intervention (Study drug administration) 
Patients will be randomized in a 1:1 ratio to receive either ganciclovir or placebo. Study drug 
delivery should begin as soon after randomization as possible; it must occur on the same day as 
randomization and within 120 hours of hospitalization. The first day of study drug is considered 
Day 1 of this study. 
• 
Study drug will be administered for a maximum of 28 days. For the initial 5 days of study 
treatment, the dose will be ganciclovir 5mg/kg or Placebo IV q 12hr. 
• 
If the patient is discharged from the hospital prior to day 28 or when intravenous access is 
removed, the patient will stop receiving study drug.   
• 
After 5 days, the dose will be reduced to ganciclovir 5mg/kg or placebo IV once daily. 
• 
Ganciclovir doses must be adjusted according to renal function as per package insert. 

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• 
Biopsy-proven CMV disease (eg colitis/esophagitis) can occur very infrequently in very 
sick but immunocompetent patients.  In such circumstances, one of the PIs should be 
contacted and it should be specified that study medication (which could be GCV or 
placebo) is to be held, and open-label GCV administered.  This would likely lead to 
unblinding since it would be presumed that breakthrough CMV disease is very unlikely 
to occur while receiving IV GCV at prophylactic doses (those used in this study after day 
5). See section 9.15 for more information on unblinding procedures. 
 
 Co-interventions 
All patients will receive standard intensive care unit care, which includes respiratory support and 
ventilator management (standardized lung protective ventilation [Appendix D] and fluid 
management [Appendix E] protocols will be used at all sites for mechanically ventilated patients, 
and Appendix I provides recommended approaches for high-flow nasal cannula oxygen and non-
invasive ventilation), antimicrobial therapy, blood glucose control, and ICU sedation. Many of 
these co-interventions occur under local protocols used as a part of routine clinical care at the 
clinical sites (initial sites listed in Appendix H).  
 
 Specimen collection 
Patients will undergo serial blood draws at study entry (+ 1 day of randomization) and plasma 
CMV PCR samples every seventh day while on study. Not more than 200 mL of blood will be 
collected over the initial 28 days of the study. A substudy of patients will have plasma 
collected every four days while hospitalized. 
Collect an ETA specimen at baseline (± 1 day), and also every fourth day (± 2 days) at 
intensified monitoring sites, while the patient is intubated, at the time this procedure is 
routinely performed by respiratory therapy. BAL fluid (from a standard or "mini" 
bronchoscopy) may be collected in the place of ETA. Specimen will be collected, labeled, and 
stored frozen for subsequent CMV PCR analysis at the coordinating center lab (Boeckh Lab at 
Fred Hutch).  
After hospital discharge, patients will not be followed daily, but they will be contacted at Day 
28 (±4 days) if not hospitalized, and Day 180 for a telephone follow-up to ascertain reportable 
adverse events, vital status, pregnancy status (at Day 28), and pregnancy within 30 days of 
study drug (at Day 180). 
 
 Patient-Centered Outcomes Survey (COMS) 
Patients will complete a survey at baseline and at 6 months (NHLBI COMS), by previously 
agreed-upon communication preference. The purpose of this survey is to compare functional 
assessment and well-being (patient-centered outcomes), at 6 months between ganciclovir and 
placebo recipients. 
At baseline, patients or LARs will complete the Katz Index of Independence in Activities of 
Daily Living (ADL) and Lawton – Brody Instrumental Activities of Daily Living Scale 
(IADL), which will take approximately 4 minutes to complete.  
At 6 months, patients will be asked to again complete the ADL, IADL, and to complete the 
Hospital Anxiety and Depression Scale (HADS), EQ-5D-5L, and Impact of Events Scale – 
Revised (IESR). The survey will take approximately 15 minutes to complete. 
 

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 Post-Enrollment Procedures 
See the schedule of procedures for specific time points (including permissible windows) in 
Appendix A. Duration of participation in the study by individual patients will be 180 days. 
• 
Blood:  
o Creatinine, Platelets, CBC w/differential - Days 4, 7, 11, 14, 18, 21, 25, 28 (all + 1 
day); can be obtained from clinical testing results if available 
o CMV PCR (all + 1 day): 
§ 
At intensified monitoring sites: Days 4, 7, 11, 14, 18, 21, 25, 28 
§ 
At all sites: 7, 14, 21, 28 at all sites 
• 
ETA: At intensified monitoring sites, CMV PCR, twice weekly (± 2 days) while intubated.  
• 
Clinical Assessments:  
o SOFA score daily on Days 1-7, then twice weekly (Day 11, 14, 18, 21, 25, 28) 
through day 28 if still in the ICU  
o Vital status (collected as ICU and hospital mortality at discharge, and 180-day 
mortality) 
o Assessment of concomitant antiviral medications daily on Days 1-7, then twice 
weekly through day 28  
o Ventilator parameters and static respiratory compliance (daily if mechanically 
ventilated), NIV parameters, and HFNC parameters daily. 
o Net fluid balance at ICU admission and net cumulative fluid balance until ICU 
discharge 
o Use at any point during study (and if used, duration/number/dose) of ECMO, prone 
positioning, steroids, neuromuscular blockers; collected at hospital discharge 
o Occurrence of bacteremia and fungemia; collected at discharge 
o Use of antibacterials or antifungals in the event of ventilator-associated pneumonia 
or nosocomial pneumonia (defined as sputum, BAL or ETA culture with new 
pathogen associated with new antibiotic +/-48 hours from culture). Any 
antibacterial used to treat bacteremia or antifungal used to treat fungemia should be 
reported. 
o Apache III at baseline only 
o COMS survey (see Section 9.10) at Day 180 (± 6 weeks) 
o Bilateral versus unilateral infiltrates on chest radiograph: To be assessed as close as 
possible to, but not prior to, randomization. Chest radiograph at admission is 
acceptable if it is the only one available. If the word "bilateral" is not on the report 
of the chest radiograph, then the coordinator will ask the site PI to assign unilateral, 
bilateral, or "no infiltrates" to the patient's chest radiograph.  If the word bilateral is 
in the report, then the coordinator will assign bilateral.   
• 
For women of childbearing potential, a serum pregnancy test will be performed at day 7 
or  at the time of hospital discharge if discharged before study day 7. 
• 
In patients discharged from the hospital before day 28, a follow-up call will be completed 
on day 28 (±4 days) to assess reportable adverse events, vital status, and pregnancy. Vital 

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status and pregnancy within 30 days of last dose of study drug will also be assessed 
during the day 180 follow-up phone call. 
• 
Because ganciclovir carries a black box warning for tumors in lab animals, at the Day 
180 follow-up call subjects will be asked if there is any known new development of a 
malignant tumor.  If a new tumor is reported, records will be requested from the primary 
care physician or hospital.  
Follow up for this study population has been historically difficult. Despite effort by sites to obtain 
all study specimens, it is expected that there may be missed blood draws after discharge from the 
hospital.  Because these missed labs are expected, they will not be considered to be unanticipated 
problems or protocol violations. In the event a patient cannot be reached for the 180 Day follow 
up, survival data may be determined through death registry records. 
 
 Monitoring of renal function 
Renal function will be monitored at least weekly throughout the active study drug dosing period. 
Monitoring will be performed by the research team at each site for all participants, with any 
reportable AE reporting per the protocol. These research personnel will be blinded. Study drug 
dose will be adjusted based on the calculated creatinine clearance according to the package insert.  
 
 Monitoring for and managing neutropenia 
Suggested Management of Neutropenia. Short-term neutropenia is a potential reportable 
adverse event of ganciclovir, although the incidence is projected to be low in the ICU setting 
and was not reported in the phase 2 ICU study.   
1. Neutropenia will be monitored at least weekly in all participants while they are in the 
hospital. Monitoring will be performed by the research team at each site, who will be 
blinded.   
2. If ANC drops below 1000/mm3, study drug will be temporarily held. 
3. Concomitant drugs should be reviewed and adjusted as feasible. 
4. ANC monitoring should continue (i.e. approximately twice a week without G-CSF; once 
a week with GCSF) until the ANC is > 1000/mm3.   
5. A dose of G-CSF may be administered (5 microgram/kg) at the discretion of the treating 
physician.  
6. If the ANC increases > 1000/mm3 study drug may be resumed.  
7. If the neutropenia recurs at levels of < 1000/mm3, study drug should be discontinued 
permanently, but the patient should continue to undergo all other study procedures & be 
followed for safety & other endpoints. 
8. If the duration of neutropenia (ANC <500/mm3) is ≥ 5 days (with or without GCSF), the 
event should be reported as a reportable SAE (see SAE reporting section).  
 

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 Pregnancy 
Participants who are women of childbearing potential will be advised to use effective 
contraception during treatment and for at least 30 days following treatment with study drug. 
Similarly, participants who are men will be advised to practice barrier contraception during and 
for at least 90 days following treatment with study drug. 
If a patient becomes pregnant during the course of the study, no further administration of study 
drug should be given but other procedures should be completed unless medically contraindicated.  
The investigator will submit a pregnancy report form to the coordinating center.  The Reporting 
Plan and timeline is described in the table in Section 11.3.  If the subject terminates from the 
study prior to the pregnancy outcome, the site must keep in touch with the patient in order to 
ascertain the pregnancy outcome. Pregnancy status for all participants who are women of 
childbearing potential will also be assessed at the Day 28 (in participants who are no longer 
hospitalized) follow up phone call, and pregnancy within 30 days after the last dose of study drug 
will be assessed during the Day 180 follow up phone call. 
 
 Unblinding  
9.15.1 Unblinding criteria  
Unblinding may be precipitated either by conclusion of the study or an emergency situation, in 
discussion between the site PI and protocol chair(s) (Drs. Stapleton, Boeckh, Limaye or 
Rubenfeld).  All patients or family members can be informed of their treatment assignment at the 
conclusion of the study, after all key analyses are complete, and upon written request.  
In the event of an emergency situation, patients may be unblinded prematurely. Emergency 
unblinding decisions will be made by the site PI only after discussion with one of the protocol 
chairs (Drs. Stapleton, Boeckh, Limaye or Rubenfeld). Additionally, if a reportable serious 
adverse event (SAE) occurs which qualifies for expedited reporting to one or more regulatory 
agencies, the patient’s treatment assignment will be unblinded, if specifically requested by the 
regulatory agencies, the institutional review board (IRB), or the DSMB. All cases of unblinding 
should be discussed with one of the protocol chairs (Drs. Stapleton, Boeckh, Limaye or 
Rubenfeld).  
9.15.2  Unblinding procedures 
After one of the protocol chairs (Drs. Stapleton, Boeckh, Limaye or Rubenfeld) agrees with the 
site PI to unblind the patient’s treatment assignment, the protocol chair will request the 
coordinating site’s statistical center (SCHARP) to send a password-protected email to the site PI 
containing the treatment assignment for the particular patient. The code should not be broken 
except in an emergency where knowledge of the patient’s treatment assignment is absolutely 
necessary for the further management of the patient, or in the context of review of an expedited 
reportable adverse event as described in the adverse event section of the protocol. If the treatment 
assignment is unblinded under any other circumstances, it will be considered a protocol violation. 
This information should also be recorded in the patient’s CRF. 

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10 
LABORATORY PROCEDURES 
Routine clinical laboratory tests will be performed through the hospital-based clinical laboratory. 
In this critically ill population, laboratory tests shall be those deemed necessary based upon 
clinical indications of the patient; others will be ordered as per protocol. 
 
 Laboratory procedures 
Laboratory procedures include but are not limited to:  
• 
Baseline whole blood sample for immunity and biomarker studies. 
• 
ETA at baseline (+ 1 day of randomization) and then twice weekly (± 2 days) while the 
patient is intubated for CMV viral load, inflammatory biomarkers, characterization of 
cellular content. 
• 
Aliquot of BALF and/or lung biopsy done for clinical purposes. For patients who 
undergo lung biopsy or autopsy, a sample of lung tissue (frozen or fresh) is requested.  
• 
Blood samples at baseline, then: 
o  Weekly (twice weekly in intensified monitoring subset) until day 28 or hospital 
discharge, whichever happens first. 
o For CMV viral load, cytokine analysis, and safety labs (CBC with neutrophil 
count and platelets, and creatinine). 
o Samples for studies to assess CMV-specific immunity and transcriptomics will 
be collected at selected sites that have the capability for sample processing 
• 
Bacteremia/fungemia and nosocomial pneumonia (including ventilator-associated 
pneumonia) will be assessed by clinical testing. 
 
 Future use of stored specimens 
The investigators intend to store specimens from patients. These samples will be used for future 
testing and research related to furthering the understanding of CMV and other viral infections and 
immunologic control, which may require additional IRB approval as specified in the informed 
consent form. Other testing on specimens will only occur after review and approval by the IRB of 
the researcher requesting the specimens and at the central IRB at the coordinating site.  
 
 Biohazard containment 
As the transmission of CMV and other blood-borne pathogens can occur through contact with 
contaminated needles, blood, and blood products, appropriate precautions will be employed by all 
personnel in the drawing of blood and shipping and handling of all specimens for this study, as 
currently recommended by the CDC and the NIH or other locally appropriate agencies. 
All dangerous goods materials, including Biological Substances, Category A or Category B, must 
be transported according to instructions detailed in the International Air Transport Association 
(IATA) Dangerous Goods Regulations.  

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11 
ADVERSE EVENT REPORTING 
 Reportable Adverse Events  
In this trial, we will not collect data on or report every adverse event (AE) by the commonly used 
standard definition because critically ill patients have innumerable abnormal signs and symptoms 
associated with their disease that are expected.  Similar to other critical care clinical trials, we will 
utilize a modified definition of reportable AEs.   
For this trial, a reportable adverse event is defined as:  
1. Any clinically important untoward medical occurrence in a patient receiving study drug 
or undergoing study procedures which is different from what is expected in the clinical 
course of a patient with acute respiratory failure/ARDS, 
  
OR,  
2. Any clinically important, untoward medical occurrence that is thought to be associated 
with the study drug or procedures, regardless of the “expectedness” of the event for the 
course of a patient with acute respiratory failure.  
Expected events for patients with acute respiratory failure are clinical occurrences that are 
perceived by the investigator to occur with reasonable frequency in the day to day care of patients 
with acute respiratory failure treated in an intensive care unit with respiratory support. Examples 
of adverse events that are expected in the course of acute respiratory failure include transient 
hypoxemia, agitation, delirium, nosocomial infections, skin breakdown, and gastrointestinal 
bleeding.  Such events, which are often the focus of prevention efforts as part of usual ICU care, 
will not be considered reportable adverse events unless the event is considered by the investigator 
to be associated with the study drug or procedures, or unexpectedly severe or frequent for an 
individual patient with acute respiratory failure.  Examples of unexpectedly frequent adverse 
events would be repeated episodes of unexplained hypoxemia.  This would be in contrast to an 
isolated episode of transient hypoxemia (e.g. SpO2 ~85%), related to positioning or suctioning.  
This latter event would not be considered unexpected by nature, severity or frequency.  Expected 
events for patients with acute respiratory failure will not be or reported in this trial. 
The research team at each site will determine if any clinical adverse experiences (while 
hospitalized) occur during the period from randomization through the last dose of study drug. The 
site research team will evaluate any changes in laboratory values and physical signs and will 
determine if the change is clinically important and different from what is expected in the course 
of  patients with acute respiratory failure. If reportable adverse events occur, they will be 
recorded on the reportable adverse event case report form.  All reportable AEs, per the modified 
definition above, will be graded according to CTC guidelines. The severity of each event should 
be classified into one of five defined categories as follows: 
• Grade 1 Mild  
• Grade 2 Moderate  
• Grade 3 Severe  
• Grade 4 Life Threatening or Disabling  
• Grade 5 Death 

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The Safety and Protocol Adherence Review Team (SPART, see section 14.3) will also review 
laboratory safety and reportable adverse event data reports monthly.  The DSMB will review all 
reportable AEs during their regularly scheduled meetings. 
Note: Study drug specific laboratory events (e.g. hematologic values, renal function) will be 
collected as secondary safety endpoints. 
 
Note: See additional information outlined in Section 5.4 on definitions of AEs for the purpose of 
post hoc analyses.  
 
 Reportable Serious Adverse Events 
In this trial, we will also utilize a modified definition of reportable serious adverse events (SAEs). 
Investigators will report all events that are serious AND unexpected AND study-related, as 
defined in the reporting guidelines found in the next section, to the Fred Hutch by fax or email 
within 7 business days of becoming aware of event.  Sites must notify their local Institutional 
Review Board (IRB) in a timely manner, according to local IRB guidelines. 
The following will also be reported within 7 business days, even if not meeting expedited 
reportable SAE reporting criteria:  
• 
ANC < 500/mm3 for a period > 5 days 
• 
Death in the presence of neutropenia (ANC< 500/mm3 for any duration) 
Of note, a large fraction of ICU patients with acute respiratory failure experience other organ 
failures and/or die, and these events are expected as a result of the usual course of their critical 
illness.  These organ failures and deaths related to acute respiratory failure or the patient’s 
underlying critical condition should not be reported as reportable SAEs unless they are 
considered to be study related AND unexpected.   
While not all deaths are reported as SAEs in this trial, all deaths occurring during this study 
will be documented in the Study Termination CRF that is completed when any participant 
leaves the study for any reason, including death. As such, deaths are reported to the central 
database in a timely fashion and are reported to the DSMB during the regular biannual meetings. 
Fred Hutch will report all serious, unexpected, and study-related reportable SAEs to the DSMB 
and NHLBI by fax or email within 7 business days of being notified of the event.  Reportable 
SAE forms received by Fred Hutch will be sent to participating sites for submission to their 
respective IRBs, according to their local IRB guidelines.  The DSMB will also review all 
reportable SAEs and all deaths during scheduled interim analyses and at each regularly scheduled 
6-month meeting.  Fred Hutch will distribute the written summary of the DSMB’s periodic 
review of reportable AEs and SAEs to investigators for submission to their respective 
Institutional Review Boards in accordance with NIH guidelines. 
When a reportable SAE arises, one of the protocol chairs not involved in subject enrollment at the 
relevant study site will also determine if the reportable SAE is unexpected for ganciclovir. 
Unexpected for ganciclovir is defined as any event not listed in the package insert. 
Investigators must also report Unanticipated Problems, regardless of severity, associated with the 
study drug or study procedures to Fred Hutch, the DSMB, and NHLBI within 7 business days 
after becoming aware of the event, and to site IRBs according to local guidelines.  An 
unanticipated problem is defined as follows: 

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Unanticipated Problem (UP): any incident, experience, or outcome that meets all of the 
following criteria:  
• 
Unexpected, in terms of nature, severity, or frequency, given the research procedures that 
are described in the protocol-related documents, such as the IRB-approved research 
protocol and informed consent document; and the characteristics of the subject 
population being studied;  
• 
Related or possibly related to participation in the research, in this guidance document, 
possibly related means there is a reasonable possibility that the incident, experience, or 
outcome may have been caused by the procedures involved in the research;  
• 
Suggests that the research places subjects or others at a greater risk of harm (including 
physical, psychological, economic, or social harm) than was previously known or 
recognized.  
Note: See additional information outlined in Section 5.4 on definitions of SAEs for the 
purpose of post hoc analyses.  
 
 Reporting Adverse Events 
1. Assuring patient safety is an essential component of this protocol.  Each participating 
investigator has primary responsibility for the safety of the individual participants under 
his or her care.  The site Principal Investigator will evaluate all local reportable adverse 
events and reportable severe adverse events.  The Study Coordinator must view patient 
records for possible reportable AEs and SAEs throughout the study period.  All 
reportable adverse events occurring within the study period must be reported in the 
participants’ case report forms.  
2. Investigators will report all serious, unexpected, AND study-related adverse events to the 
Fred Hutch within 7 business days by fax or email.  Sites must notify their local 
Institutional Review Board in a timely manner, according to local IRB guidelines. 
3. Definitions of Adverse Events  
a. A modified definition of a reportable AE (see Section 11.1 above) is being used for 
this trial. 
b. A serious adverse event is any event that is fatal or immediately life threatening, is 
permanently disabling, or severely incapacitating, or requires or prolongs inpatient 
hospitalization.  Important medical events that may not result in death, be life 
threatening, or require hospitalization may be considered serious adverse events 
when, based upon appropriate medical judgment, they may jeopardize the patient or 
subject and may require medical or surgical intervention to prevent one of the 
outcomes listed above.  Note that the modified definition of reportable SAE (see 
Section 11.2 above) is being used for this trial. 
i. 
Life-threatening means that the patient was, in the view of the investigator, at 
immediate risk of death from the reaction as it occurred.  This definition does not 
include a reaction that, had it occurred in a more serious form, might have caused 
death.  Assessment of the cause of the event has no bearing on the assessment of 
the event’s severity. 

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c.  An unexpected event is any experience not identified by the type, severity, or 
frequency in the current study protocol or an event that is unexpected in the course of 
treatment for acute respiratory failure or ARDS. 
d. Reportable adverse events will be considered to be study-related if the event follows 
a reasonable temporal sequence from a study drug or procedure and could readily 
have been produced by the study drug or procedure. 
e. Organ failures or death related to acute respiratory failure or ARDS or the patient’s 
underlying condition that are systematically captured by the protocol should not be 
reported as reportable SAEs unless they are considered to be study related and 
unexpected, but deaths will be reported to the central database in a timely fashion via 
the Study Termination CRF. 
All reportable SAEs must be reported to the Fred Hutchinson Cancer Center in a timely fashion to 
allow expedited reporting to the DSMB and other entities (see Figure 11-1: safety reporting 
chart). The following table summarizes the reporting timelines:  
 
Type of Event 
Definition of  
Reportable Event 
Reporting Plan 
Reporting Timeline 
(after becoming aware 
of event) 
Reportable Serious 
Adverse Events 
(SAE) 
Any untoward medical event 
that is: 
 
Serious 
  AND 
Unexpected 
  AND  
Related to study drug or 
procedure 
Site to local IRB 
According to local IRB 
guidelines 
Site to coordinating 
center 
Initial report within 7 
business days 
Coordinating center to 
NHLBI executive 
secretary and DSMB 
chair 
 
DSMB Chair to 
determine if full meeting 
is necessary 
Within 7 business days 
of receipt of initial 
report from site 
Within 72 hours after 
Chair receives report 
from coordinating center 
Coordinating center to 
NHLBI & participating 
sites 
Within 7 business days 
of receiving initial report 
Coordinating center to 
report to FH IRB 
Within 7 business days 
of receipt of initial 
report from site 
Neutropenia 
ANC < 500/mm for > 5 days 
SAME AS ABOVE 
SAME AS ABOVE 
Death 
Death in the presence of 
neutropenia (ANC< 500/mm3 
for any duration) 
OR 
Death that meets the 
requirements for a reportable 
SAE as defined above 
 
SAME AS ABOVE 
 
SAME AS ABOVE 

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Death not meeting reporting 
definition above 
Site to local IRB 
According to local IRB 
guidelines 
 Site to coordinating 
center 
As part of Study 
Termination CRF 
Coordinating center to 
NHLBI executive 
secretary and DSMB 
Included in report 
prepared for each 
DSMB meeting 
Coordinating center to 
NHLBI & participating 
sites 
Annually 
Coordinating center to 
report to FH IRB 
Annually 
 
Unanticipated 
problem 
Any untoward event that is: 
• 
Unexpected, in terms of 
nature, severity, or 
frequency 
AND 
• 
Related or possibly related 
to participation in the 
research 
AND 
Suggests that the research places 
subjects or others at a greater 
risk of harm (including physical, 
psychological, economic, or 
social harm) than was 
previously known or 
recognized. 
Site to local IRB 
According to local IRB  
 Site to coordinating 
center 
Within 7 business days 
via memo  
Coordinating center to 
NHLBI executive 
secretary and DSMB 
Chair 
Within 7 business days 
of receipt of information 
from site 
Coordinating center to 
NHLBI & participating 
sites 
Within 7 business days 
of receipt of information 
from site 
Coordinating center to 
FH IRB 
Within 7 business days 
of receipt of information 
from site 
Pregnancy 
ALL 
Site to local IRB 
 
According to local IRB 
guidelines 
 
Site to coordinating 
center 
Within 7 business days 
(via pregnancy report 
form) 
Coordinating center to 
NHLBI executive 
secretary and DSMB 
Within 7 business days 
of receiving pregnancy 
report form 
Coordinating center to 
NHLBI & participating 
sites 
Within 7 business days 
of receiving pregnancy 
report form 
Coordinating center to 
FH IRB 
Within 7 business days 
of receiving pregnancy 
report form 
Reportable Adverse 
Event 
Any untoward medical event that 
is considered by the investigator 
to be: 
Site to local IRB 
According to local IRB 
guidelines  
Site to coordinating 
center 
Reportable AEs 
reported as required on 
CRFs.  CRFs to be 
completed on a timely 
basis.  

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• 
Unexpectedly severe 
or more frequent than 
typical course of ALI 
OR 
• 
Have any relationship 
to study drug or 
procedures 
Coordinating center to 
NHLBI executive 
secretary and DSMB 
Included in report 
prepared for each 
DSMB meeting 
Coordinating center to 
NHLBI & participating 
sites 
Annually - summarized 
from CRFs in database 
Coordinating center to 
FH IRB 
Annually – summarized 
from CRFs in database 
 
All sites will be responsible for compliance with local safety reporting guidelines.  
 
 
Figure 11-1: Safety reporting chart. 
 
The reportable SAE Report will include the following information (as available):  
o Patient ID  
o Description of the reportable SAE (onset date, severity, causal relationship)  
o Basic demographic information  
o Outcomes attributed to the event  
o Summary of relevant test results, laboratory data, and other relevant history  
o The first and last dates of study drug administration  
o Statement whether study drug was discontinued or schedule modified  
o Statement whether the event abated after study drug was discontinued/modified  

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o Statement whether the event recurred after reintroduction of the study drug if it had 
been discontinued or held  
Participating sites will be provided with reportable SAE report forms and contact numbers for 
transmitting the reports. 
 
 Relationship to study drug 
All reportable AEs will have a causality assessment performed at the time of reporting the event 
to document the Investigator’s perception of causality. There is currently no standard 
international nomenclature to define causality. For the purposes of this study, causality will be 
assigned using the following criteria: 
 
Definitely 
related 
The event cannot be attributed to the patient’s underlying 
medical condition or other concomitant therapy and there is a 
compelling temporal relationship between the onset of the 
events and study drug administration that leads the 
Investigator to believe that there is a causal relationship. 
Probably 
related 
There is a clinically plausible time sequence between the 
onset of the AE and the study drug administration. The AE is 
unlikely to be caused by a concurrent/underlying illness, 
other drugs or procedures. 
Possibly 
related 
There is a clinically plausible time sequence between the 
onset of the AE and study drug administration, but the AE 
could also be attributed to a concurrent/underlying disease, 
other drugs, or procedures. “Possibly related” should be used 
when the study drug administration is one of several 
biologically plausible causes of the AE. 
Not related 
The patient’s underlying medical condition or concomitant 
therapy can easily be identified as the cause of the event and 
there is no temporal relationship between the event and the 
study drug. 
 
 Pregnancy 
A pregnancy is not an adverse event. If a patient becomes pregnant while enrolled in the study 
following administration of study drug, administration of study drug will be discontinued 
immediately and the patient will be followed through the outcome of the pregnancy.  The 
investigator will submit a pregnancy report form to the coordinating center.  The Reporting Plan 
and timeline is described in the table in Section 11.3. 
 
 Breaking the blind 
The blind will not routinely be broken for reportable SAE’s. Decisions on whether or not to break 
the blind will be made as described in Section 9.15,1, and unblinding will follow procedures 
outlined in Section 9.15.2. 
 

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 Stopping rules 
The study may be stopped prematurely if an excess rate of toxicity is observed. The DSMB will 
monitor throughout the study and there will be scheduled interim analyses for safety (see 
Statistical section). 

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12 
DATA MANAGEMENT CONSIDERATIONS 
 Overview 
The Data Coordinating Center will utilize the existing Validation Study Information Management 
System (VSIMS) to facilitate Trial collaborative activities via the SCHARP Data Management 
System (CDMS). VSIMS/CDMS provides online, end-to-end data management solutions, 
including investigator and study coordinator communications, regulatory compliance, remote 
subject registration, clinical data capture, biospecimen sample management, and document 
management. VSIMS/CDMS can provide online visibility of analytical datasets for all 
participating researchers, and statistical and informatics tools relevant to Trial research, and will 
be specifically configured to support this trial. 
 Data Collection 
Each patient will be assigned an identification number to be used for all patient data. Links to 
patient name and identifiers will be maintained and stored in files on computers protected by 
password and in locked office cabinets. Research staff and physicians will remain blinded until 
the study is completed. De-identified data collected during this study may be used for future 
research. 
Chart abstraction for demographic, laboratory, and physiologic data will occur at study entry, 
daily on study days 1-7, then twice weekly through day 28 while the patient is hospitalized, and 
again at hospital discharge or death. While patient remains hospitalized, review of the hospital 
record will occur daily throughout the hospitalization (to Day 28) to identify any reportable 
adverse events.  
All information will be in Medidata RAVE.   
 Data Management 
Participating sites can enter clinical data into VSIMS, in real time. The Data Coordinating Center 
will assist the Clinical Coordinating Center with monitoring data completion by reporting data 
entry status. Sites will receive queries to reconcile inconsistencies. 
 Quality Control and Quality Assurance 
By signing this protocol, the Investigator/Sponsor agrees to be responsible for implementing and 
maintaining quality control and quality assurance systems with written Standard Operating 
Procedures (SOPs) to ensure that the study is conducted and data are generated, documented, and 
reported in compliance with the protocol, accepted standards of Good Clinical Practice, and all 
applicable federal, state, and local laws, rules, and regulations relating to the conduct of the 
clinical study. 
By signing this protocol, the investigators agree to conduct the study in an efficient and diligent 
manner and in conformance with this protocol; to follow generally accepted standards of Good 
Clinical Practice; and to follow all applicable federal, state, and local laws, rules, and regulations 
relating to the conduct of the clinical study. 
The investigator also agrees to allow monitoring, audits, Institutional Review Board review and 
regulatory agency inspection of study-related documents and procedures and provide for direct 
access to all study-related source data and documents.  
The investigator shall prepare and maintain complete and accurate study documentation in 
compliance with Good Clinical Practice standards and applicable federal, state, and local laws, 
rules, and regulations. 

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The investigator has the responsibility of explaining the correct use of the study drug to the site 
personnel, ensuring that instructions are followed properly, and maintaining accurate records of 
study drug dispensing and collection. 
 Study monitoring  
Because of the risk profile of the study drug, which carries a black box warning on its package 
insert and has the potential of hematologic toxicity, we will perform study monitoring of 
intermediate intensity. Study data and regulatory aspects at study sites will be monitored by a 
study monitor, managed by the clinical coordinating center. The study monitor will perform 
monitoring of the first patient at each site as well as a random selection of 10% of participants (to 
be determined by DCC)  across the whole study. The study monitor will conduct remote 
monitoring via EPIC if feasible, and by uploads into Florence eReg platform if not (with site PI 
review of charts for unreported AEs), and by Vestigo, according to industry standard (GCP) and 
as outlined in the monitoring plan. For non-domestic sites, alternative monitoring plans may be 
designed.  The study monitor will create monitoring reports, submit them to site investigators and 
the Executive Committee, and conduct the necessary follow-up. A detailed monitoring plan, 
including the scope of monitoring and deadlines for entering the data, is shown in Appendix F. 

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13 
ETHICAL CONSIDERATIONS & HUMAN SUBJECTS PROTECTIONS 
 Ethical Review 
This study will be conducted in accordance with the ethical principles stated in the Declaration of 
Helsinki (1996) and applicable guidelines on Good Clinical Practice.  
The investigator will obtain approval of the protocol and the informed consent from the single 
Institutional Review Board before the study may begin. IRB approval will also be obtained 
locally from each additional clinical site before the study commences at that site. The investigator 
will supply the following to the Institutional Review Board and Data Safety and Monitoring 
Board: 
• 
Study protocol and appendices. 
• 
Informed consent document and updates. 
• 
Safety alerts. 
This study will be registered with the U.S. NIH’s clinical trials registry ClinicalTrials.gov. 
 
 Potential risks of study drugs and procedures 
The following table presents common, less common, and uncommon risks based on experience 
with this drug in humans and animal data. This information will be communicated to patients in 
the sample informed consent form. 
Less common 
Blood: leucopenia, neutropenia, anemia  
Uncommon or rare 
Central nervous system: fever, headache, insomnia, paresthesia, and 
peripheral neuropathy.  
Ocular: retinal detachment.  
Effects on the fetus and on pregnancy (which is why pregnant women will be 
excluded from participating). 
Unknown frequency 
or theoretical risks 
Cancer 
 
 Risks of Endotracheal Aspirates 
Endotracheal aspiration is routinely performed on intubated patients by respiratory therapy as part 
of  their clinical care routine to help clear respiratory secretions.  There are no known risks to this 
procedure and would be considered inappropriate care if this procedure were not performed. 
 
 
 Risks of blood collection 
Blood collection may cause some discomfort during the insertion of the needle. Bleeding, 
infection, or hematoma could occur. These risks will be low magnitude, low probability, short 
duration, and reversible. These risks will be mitigated by using a trained phlebotomist to perform 
the blood draw. 
 
Table 13-1 Summary of potential risks of study medication and administration  

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 Risk of loss of confidentiality 
Risk of breach of confidentiality will be minimized by keeping all research forms and electronic 
databases in locked and protected environments, and through training staff to be especially 
sensitive to confidentiality whenever interacting with participants. 
 
 Potential benefit of enrollment 
Respiratory failure and sepsis carry a high mortality and consume millions of health care dollars 
each year. Any treatment that is found to impact outcomes in sepsis-associated respiratory failure 
could have a substantial societal benefit. Ganciclovir is not routinely administered to respiratory 
failure patients, so individual patients participating in this trial have an opportunity to receive this 
treatment through the study. If ganciclovir is ultimately found to positively affect outcomes, such 
as increased respiratory-support-free days, individuals in this study may benefit. It is possible, 
though, that an individual may not derive any direct benefit from participating in this trial, or 
even experience toxicities or adverse outcomes. 
 
 

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14 
PROTOCOL OVERSIGHT AND GOVERNANCE 
 Principal investigator 
The PI will adhere to requirements of the Code of Federal Regulations. Additionally, the primary 
Principal Investigator/Sponsor will sign the final clinical study report for this study, confirming 
that to the best of her/his knowledge the report accurately describes the conduct and results of the 
study. 
 Protocol Leadership Team 
The Protocol Leadership Team will be responsible for administrative oversight of the study, 
provides the overall operational direction for the trial, and is responsible for the conduct of the 
trial according to the highest scientific and ethical standards, as well as approving revisions and 
amendments to the protocol. The Protocol Leadership Team will remain blinded to the treatment 
group assignment of individual patients during the course of the study. 
 Safety and protocol adherence review team 
The safety and protocol adherence review team (SPART) will, on a monthly basis, review all 
clinical and laboratory safety data during the study. The SPART will include CCC PIs (Drs. 
Boeckh, Stapleton, Rubenfeld, and Limaye), Clinical Site Manager Sara Ardren, and Clinical 
Trial Project Manager/Site Monitoring Manager Dr. Louise Kimball. Meetings of the SPART (by 
videoconference or teleconference) will occur regularly and will not require attendance of all 
team members at each meeting. The SPART is responsible for the review of the clinical safety 
reports, discordant CMV serostatus test results (LFA versus CLIA test), and protocol violations, 
as well as communication with the IRB, NHLBI executive secretary, and DSMB, as outlined 
above.  
 Data Safety and Monitoring Plan (outlined briefly below and in more detail in Appendix F) 
Investigators are responsible for monitoring the safety of patients who have entered this study. 
While hospitalized, patients will be assessed daily for evaluation of reportable adverse events by 
the research nurse/coordinator and principal investigator at study sites. Drs. Rubenfeld, Boeckh, 
Stapleton and Limaye will act as the investigator on-call for urgent issues and questions; Dr. 
Rubenfeld or Dr. Boeckh will serve in this role for any questions about eligible patients at Dr. 
Limaye or Dr. Stapleton’s study sites. 
The investigator remains responsible to follow, through an appropriate health care option, 
reportable adverse events (AEs) that are serious, cause the patient to discontinue before 
completing the study, or are ongoing at the time of study completion. The site investigators will 
maintain responsibility for forwarding of reportable SAEs to the coordinating center and local 
Institutional Review Board. The patient will be followed until the event resolves or stabilizes. 
Frequency of follow-up is left to the discretion of the investigator. 
 Data and Safety Monitoring Board  
A Data and Safety Monitoring Board (DSMB) will be established. This DSMB will assess the 
effects of the study drug during the trial and is advisory to NHLBI. The members of the 
committee are independent of the University of Washington, Fred Hutchinson Cancer Center, and 
clinical investigators participating in this trial, and will not have any other involvement in the 
study, nor will they have any relation to study subjects. 
Prior to beginning patient accrual, the DSMB will review the research protocol and identify any 
potential problems with randomization and implementation of the protocol. At this early phase, 
the DSMB will also review plans for data and safety monitoring to ensure that the frequency of 
monitoring is appropriate for the ganciclovir intervention.  

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During patient accrual, all reported serious adverse events will be reported according to Figure 
11.1. The DSMB may recommend any steps to ensure the safety of study subjects and the 
integrity of the trial.  
The DSMB will be involved with planned interim analyses. The interim monitoring guidelines 
that the DSMB will follow will be described in the Statistical Analysis Plan. The DSMB minutes 
will summarize the actions and deliberations of the DSMB and will be made available at the 
conclusion of the trial. At the time of interim analyses, the DSMB will aid in identifying 
problems surrounding patient accrual and randomization, data collection, and follow-up. At this 
time the DSMB will evaluate safety through a comparison of reportable adverse events across 
study arms.  
The DSMB may recommend that specific groups be withdrawn from the study, if any subgroup 
manifests serious or widespread side effects, or that the trial be terminated altogether. To 
guarantee the unrestricted performance of its task, the DSMB may receive the individual study 
morbidity and mortality data from an unblinded statistician.  
 Study termination 
This study may be terminated by the determination of the US NIH or US Office for Human 
Research Protections (OHRP). In addition, the conduct of this study at an individual site may be 
terminated by the determination of the local IRB. 
The study may be terminated in the following situations: 
• 
All patients have been accrued and have completed follow-up. 
• 
If the interim analysis conducted by the DSMB at midpoint demonstrates a highly 
significant difference in treatment groups, as defined above.  

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15 
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Boeckh, M., D. Myerson, and R.A. Bowden, Early detection and treatment of cytomegalovirus 
infections in marrow transplant patients: methodological aspects and implications for therapeutic 
interventions. Bone Marrow Transplantation, 1994. 14(Suppl 4): p. S66-70. 
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Boeckh, M., et al., Valganciclovir for the prevention of complications of late cytomegalovirus 
infection after allogeneic hematopoietic cell transplantation: a randomized trial. Ann Intern Med, 
2015. 162(1): p. 1-10. 
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Boeckh, M., et al. Prevention of Late CMV Disease after HCT: A Randomized Double-Blind 
Multicenter Trial of Valganciclovir (VGCV) Prophylaxis versus PCR-Guided GCV-VGCV 
Preemtive Therapy. in Annual Meeting of the American Society for Blood and Marrow 
Transplantation. 2008. San Diego, CA. 
65. 
Merigan, T.C., et al., A controlled trial of ganciclovir to prevent cytomegalovirus disease after 
heart transplantation. N Engl J Med, 1992. 326(18): p. 1182-6. 
66. 
Spector, S.A., et al., Oral ganciclovir for the prevention of cytomegalovirus disease in persons 
with AIDS. Roche Cooperative Oral Ganciclovir Study Group. N Engl J Med, 1996. 334(23): p. 
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Papazian, L., et al., Preemptive ganciclovir for mechanically ventilated patients with 
cytomegalovirus reactivation. Ann Intensive Care, 2021. 11(1): p. 33. 
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Winston, D.J., et al., Randomised comparison of ganciclovir and high-dose acyclovir for long-term 
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in a subset of patients with chronic fatigue syndrome. J Med Virol, 2013. 85(12): p. 2101-9. 
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Stragliotto, G., et al., Effects of valganciclovir as an add-on therapy in patients with 
cytomegalovirus-positive glioblastoma: a randomized, double-blind, hypothesis-generating study. 
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With Worse Clinical Outcome in Critically Ill Adults. Crit Care Med, 2015. 43(7): p. 1415-22. 
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Singer, M., et al., The Third International Consensus Definitions for Sepsis and Septic Shock 
(Sepsis-3). JAMA, 2016. 315(8): p. 801-10. 
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Goddard, S.L., et al., The randomized educational acute respiratory distress syndrome diagnosis 
study: a trial to improve the radiographic diagnosis of acute respiratory distress syndrome. Crit 
Care Med, 2018. 46(5): p. 743-748. 
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Hamprecht, K., et al., The lung as a central compartment of active CMV infection. Inflammation 
Research, 2007. 56(Abstract A383): p. S242-S242. 
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von Muller, L., et al., Active cytomegalovirus infection in patients with septic shock. Emerg Infect 
Dis, 2006. 12(10): p. 1517-22. 
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von Muller, L., et al., Cellular immunity and active human cytomegalovirus infection in patients 
with septic shock. J Infect Dis, 2007. 196(9): p. 1288-95. 
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Einsele, H., et al., Polymerase chain reaction monitoring reduces the incidence of 
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transplantation. Blood, 1995. 86(7): p. 2815-20. 
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Ferreyro, B.L., et al., Association of Noninvasive Oxygenation Strategies With All-Cause Mortality 
in Adults With Acute Hypoxemic Respiratory Failure: A Systematic Review and Meta-analysis. 
JAMA, 2020. 324(1): p. 57-67. 
79. 
Shankar-Hari, M. and G.D. Rubenfeld, Just Because Things Are Not Different, Does Not Mean 
They Are the Same: Biomarker Patterns in Acute Respiratory Distress Syndrome*. Critical Care 
Medicine, 2017. 45(11). 
80. 
Matthay, M.A., B.T. Thompson, and L.B. Ware, The Berlin definition of acute respiratory distress 
syndrome: should patients receiving high-flow nasal oxygen be included? Lancet Respir Med, 
2021. 9(8): p. 933-936. 
81. 
Contentin, L., S. Ehrmann, and B. Giraudeau, Heterogeneity in the definition of mechanical 
ventilation duration and ventilator-free days. Am J Respir Crit Care Med, 2014. 189(8): p. 998-
1002. 
82. 
National Heart, L., et al., Comparison of two fluid-management strategies in acute lung injury. N 
Engl J Med, 2006. 354(24): p. 2564-75. 
83. 
Laterre, P.F., et al., Effect of Selepressin vs Placebo on Ventilator- and Vasopressor-Free Days 
in Patients With Septic Shock: The SEPSIS-ACT Randomized Clinical Trial. Jama, 2019. 
322(15): p. 1476-1485. 
84. 
Angus, D.C., et al., Effect of Hydrocortisone on Mortality and Organ Support in Patients With 
Severe COVID-19: The REMAP-CAP COVID-19 Corticosteroid Domain Randomized Clinical 
Trial. JAMA, 2020. 324(13): p. 1317-1329. 
85. 
Goligher, E.C., et al., Therapeutic Anticoagulation with Heparin in Critically Ill Patients with Covid-
19. N Engl J Med, 2021. 385(9): p. 777-789. 
86. 
Finkelstein, D.M. and D.A. Schoenfeld, Combining mortality and longitudinal measures in clinical 
trials. Stat Med, 1999. 18(11): p. 1341-54. 
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cannot undergo surgery. N Engl J Med, 2010. 363(17): p. 1597-607. 
88. 
Schoenfeld, D.A., G.R. Bernard, and A. Network, Statistical evaluation of ventilator-free days as 
an efficacy measure in clinical trials of treatments for acute respiratory distress syndrome. Crit 
Care Med, 2002. 30(8): p. 1772-7. 
89. 
Davidian, M., A.A. Tsiatis, and S. Leon, Semiparametric Estimation of Treatment Effect in a 
Pretest-Posttest Study with Missing Data. Stat Sci, 2005. 20(3): p. 261-301. 
90. 
Zhang, B., et al., Estimating Optimal Treatment Regimes from a Classification Perspective. Stat, 
2012. 1(1): p. 103-114. 
91. 
Shepherd, B.E., Does Finasteride Affect the Severity of Prostate Cancer? A Causal Sensitivity 
Analysis. Journal of the American Statistical Association, 2009. 484: p. 1392-1404. 

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Cox, D.R., Some Remarks on Overdispersion. Biometrika, 1983. 70(1): p. 269-274. 
93. 
Hinde, J. and C.G.B. Demetrio, Overdispersion: Models and estimation. Computational Statistics 
& Data Analysis, 1998. 27(2): p. 151-170. 
94. 
Hall, D.B., Zero-inflated Poisson and binomial regression with random effects: A case study. 
Biometrics, 2000. 56(4): p. 1030-1039. 
95. 
Vieira, A.M.C., J.P. Hinde, and C.G.B. Demetrio, Zero-inflated proportion data models applied to 
a biological control assay. Journal of Applied Statistics, 2000. 27(3): p. 373-389. 
96. 
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Psychological Measurement, 1979. 39(3): p. 527-535. 
97. 
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Annals of the Institute of Statistical Mathematics, 1994. 46(2): p. 317-331. 
98. 
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Oncol, 2012. 30(26): p. 3297-303. 
100. 
DeMets, D.L. and K.K. Lan, Interim analysis: the alpha spending function approach. Stat Med, 
1994. 13(13-14): p. 1341-52; discussion 1353-6. 
101. 
Kieser, M. and T. Friede, Simple procedures for blinded sample size adjustment that do not affect 
the type I error rate. Stat Med, 2003. 22(23): p. 3571-81. 
102. 
Zucker, D.M., et al., Internal pilot studies II: comparison of various procedures. Stat Med, 1999. 
18(24): p. 3493-509. 
103. 
Hanson, K.E., et al., 2031. False-positive Serologic Results attributable to IVIG therapy. Open 
Forum Infectious Diseases, 2018. 5(Suppl 1): p. S591-S592. 
104. 
The Acute Respiratory Distress Syndrome Network, Ventilation with lower tidal volumes as 
compared with traditional tidal volumes for acute lung injury and the acute respiratory distress 
syndrome. The Acute Respiratory Distress Syndrome Network. 2000. 342(18): p. 1301-1308. 
105. 
Brower, R.G., et al., Higher versus lower positive end-expiratory pressures in patients with the 
acute respiratory distress syndrome. N Engl J Med, 2004. 351(4): p. 327-36. 
106. 
Grieco, D., et al., Comfort during high-flow oxygen therapy through nasal cannula in critically ill 
patients: effects of gas temperature and flow. Intensive Care Med, 2013. 39(512). 
107. 
Antonelli, M., et al., New treatment of acute hypoxemic respiratory failure: noninvasive pressure 
support ventilation delivered by helmet--a pilot controlled trial. Crit Care Med, 2002. 30(3): p. 602-
8. 
108. 
Chiumello, D., et al., Effect of a heated humidifier during continuous positive airway pressure 
delivered by a helmet. Crit Care, 2008. 12(2): p. R55. 
109. 
Ueta, K., et al., Influence of humidification on comfort during noninvasive ventilation with a 
helmet. Respir Care, 2013. 58(5): p. 798-804. 
110. 
Mojoli, F., et al., An optimized set-up for helmet noninvasive ventilation improves pressure 
support delivery and patient-ventilator interaction. Intensive Care Med, 2013. 39(1): p. 38-44. 
111. 
Ferrone, G., et al., A bench study of 2 ventilator circuits during helmet noninvasive ventilation. 
Respir Care, 2013. 58(9): p. 1474-81. 
112. 
Taccone, P., et al., Continuous positive airway pressure delivered with a "helmet": effects on 
carbon dioxide rebreathing. Crit Care Med, 2004. 32(10): p. 2090-6. 
 

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16 
INVESTIGATORS STATEMENT/PROTOCOL SIGNATURE PAGE 
I have read and understood the contents of this protocol and all study documents, and agree to carry 
out all of its terms in accordance with Good Clinical Practice. 
 
I agree to permit trial related monitoring, audits, Institutional Review Board review and regulatory 
agency inspection of study-related documents and procedures, and to provide for direct access to all 
study-related source data and documents. 
 
I agree that all the test article(s) supplied will be used solely for the purpose of conducting this study. 
 
 
 
Principal Investigator (printed name) 
 
 
 
 
 
Principal Investigator (Signature) 
 
Date 
 
This protocol version 5.0 has been approved by the Protocol Leadership Team.  The following 
signatures document this approval. 
 
 
 
 
 
Signature 
 
 
 
 
 
 
Date 
Michael Boeckh, MD 
Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Center, Seattle, WA 
 
 
 
 
 
Signature 
 
 
 
 
 
 
Date 
Renee Stapleton, MD, PhD, Division of Pulmonary and Critical Care Medicine, University of 
Vermont, Burlington, VT 
 
 
 
 
 
Signature 
 
 
 
 
 
 
Date 
Ajit Limaye, MD 
Division of Infectious Diseases, Univ. of California San Francisco, San Francisco, CA 
 
 
 
 
 
Signature 
 
 
 
 
 
 
Date 
Gordon Rubenfeld, MD, MSc 
Sunnybrook Medical Centre, Univ. of Toronto, Toronto, Canada                                    

 
 
 
 
Page 81 
APPENDIX A: TIME AND EVENTS SCHEDULE 
Visit 
Screening 
01 
02 
03 
04 
05 
06 
07 
08 
09 
10 
Assay Location 
Day 
-4 to 1 
1 
4 
7 
11 
14 
18 
21 
25 
28 
180 
 
Window (+/- days) 
0 
1 
1 
1 
1 
1 
1 
1 
1 
4 
42 
 
 
Informed consent 
X 
- 
- 
- 
- 
- 
- 
- 
- 
- 
- 
 
Administer Study Drug 
 
Patient receives 5 days of ganciclovir (or placebo) intravenously TWICE daily, 
then up to 23 additional days of ganciclovir (or placebo) intravenously ONCE 
daily in hospitalized patients. 
- 
 
 
L = local test; S=Seattle, WA; A= ARUP in Utah 
a  Pregnancy tests (serum or urine) performed within 120 hours before enrollment are acceptable.  
b  At day 7 or discharge, whichever is earlier (if day 7 is missed, at discharge is acceptable). Only serum test is acceptable. 
Blood collection (ml)  vol. estimated: 
Pregnancy testing 
3a  
 
 
3b 
 
 
 
L 
CMV Serology  
3  
- 
- 
- 
- 
- 
- 
- 
- 
- 
- 
L/A 
Genomic analysis samplec 
- 
5 
- 
- 
- 
- 
- 
- 
- 
- 
- 
S 
T cell immunity samplesc,e,m 
- 
(30) 
- 
- 
- 
- 
- 
- 
- 
- 
- 
S 
Serum Creatinined,n  
1 
1 
1 
1 
1 
1 
1 
1 
1 
1 
- 
L 
CMV PCR (plasma)d,e 
- 
5 
(5) 
5 
(5) 
5 
(5) 
5 
(5) 
5 
- 
S 
CBC, w/diff, plateletsd 
1 
3 
3 
3 
3 
3 
3 
3 
3 
3 
- 
L 
Blood volume (estimate) 
8  
14 (44) 
4 (9) 
9 
4 (9) 
9 
4 (9) 
9 
4 (9) 
9 
- 
 
Endotracheal aspirate (ETA) 
CMV PCRe, f 
- 
X 
(X) 
(X) 
(X) 
(X) 
(X) 
(X) 
(X) 
(X) 
- 
S 
Clinical Assessments: 
 
 
 
 
 
 
 
 
 
 
 
 
SOFAg  and, fluid balance  
 
Daily on Days 1-7 
X 
X 
X 
X 
X 
X 
 
 
Ventilator (including static compliance),f 
NIV, HFNC parameters, and FiO2 
 
Daily 
 
 
Antiviral, antibacterial or antifungal 
medicationsh 
 
Daily on  Days 1-7 
X 
X 
X 
X 
X 
X 
 
 
Apache III 
 
X 
 
 
 
 
 
 
 
 
 
 
AE/SAE monitoring 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
 
ECMO, prone positioning,  steroids, 
immunosuppressive medications, and  
neuromuscular blockers 
 
X 
At any time while on study; collected at discharge 
 
 
Vital status 
 
 
ICU and hospital mortality collected at discharge 
X 
 
Bacteremia, fungemia, nosocomial 
pneumonia (including ventilator-
associated pneumonia) 
 
 
At any time while on study; collected at discharge 
 
 
Contact (e.g. phone) 
 
 
 
 
 
 
 
 
 
Xj 
Xk 
 
NHLBI COMS survey 
 
Xl 
 
 
 
 
 
 
 
 
Xl 
 

 
 
 
 
Page 82 
c  All samples will be stored for analysis at the end of the study 
d  All blood draws will occur only during hospitalization, blood chemistry, CBC and platelet values can be obtained from clinical testing result if available, thereby reducing required blood volume 
e  Time points in ( ) only in patients with intensified monitoring (n=150).  
f   Can only be collected if subject is ventilated 
g  While in the ICU  
h Use of antibacterials or antifungals will be collected in the event of ventilator-associated pneumonia or nosocomial pneumonia (defined as sputum, BAL or ETA culture with new pathogen 
associated with new antibiotic +/-48 hours from culture). Any antibacterial used to treat bacteremia or antifungal used to treat fungemia should be reported. 
 
 
i  In patients who were discharged before day 28, visit window ± 4 days,  to assess reportable adverse events, vital status, and pregnancy within 30 days of study drug (see text). 
j   A reminder contact (e.g. by mail, email, text) will be done ~3 months before the day 180 assessment. 
k   A reminder contact (e.g. by mail, email, text) will be done if the participant does not appear for their day 180 assessment. 
l    Katz ADL and Lawton IADL at baseline and Day 180; HADS, EQ-5D-5L and IESR only at Day 180. 
m  A different window for sample processing may apply; see the MOP for additional information. 
n  Serum Creatinine is not required if the patient is receiving renal replacement therapy 

 
 
Page 83 
APPENDIX B: NCI COMMON TOXICITY CRITERIA (CTC) 
 
A. The NCI CTC criteria will be used for reportable Adverse Event reporting. The NCI CTC criteria can 
be downloaded at: 
https://ctep.cancer.gov/protocolDevelopment/electronic_applications/ctc.htm#ctc_50.  A hard copy of 
the NCI CTC can be found in the study reference manual.  
 
B. For this study the CTC guideline categories have been assigned numbers as follows: 
CATEGORY 
CODE 
BLOOD AND LYMPHATIC SYSTEM DISORDERS 
 
01 
CARDIAC DISORDERS 
 
02 
CONGENITAL, FAMILIAL AND GENETIC DISORDERS  
 
03 
EAR AND LABYRINTH DISORDERS 
 
04 
ENDOCRINE DISORDERS 
 
05 
EYE DISORDERS 
 
06 
GASTROINTESTINAL DISORDERS  
 
07 
GENERAL DISORDERS AND ADMINISTRATION SITE CONDITIONS  08 
HEPATOBILIARY DISORDERS 
 
09  
IMMUNE SYSTEMS DISORDERS 
 
10 
INFECTIONS AND INFESTATIONS 
 
11 
INJURY, POISONING AND PROCEDURAL COMPLICATIONS  
12 
INVESTIGATIONS 
 
13  
LYMPHATICS  
 
14 
METABOLIC AND NUTRITION DISORDERS 
 
15 
MUSCULOSKELETAL AND CONNECTIVE TISSUE DISORDERS 
16 
NEOPLASMS BENIGN, MALIGNANT AND UNSPECIFIED 
 
17 
NERVOUS SYSTEM DISORDERS 
 
18 
PSYCHIATRIC DISORDERS 
 
19 
RENAL AND URINARY DISORDERS 
 
20 
RESPIRATORY, THORACIC AND MEDIASTINAL DISORDERS 
21 
SKIN AND SUBCUTANEOUS TISSUE DISORDERS 
 
22 
SOCIAL CIRCUMSTANCES 
 
23 
SURGICAL AND MEDICAL PROCEDURES 
 
24 
VASCULAR DISORDERS 
 
25 

 
 
Page 84 
APPENDIX C:  COMMONLY PRESCRIBED IMMUNOSUPPRESSIVE AGENTS WITH 
KNOWN EFFECT ON CMV REACTIVATION 
       
   Generic Name 
 
 
 
 
Trade Name 
Antithymocyte Globulin (Equine)  
ATG, ATGAM 
Antithymocyte Globulin (Rabbit)  
Thymoglobulin 
Alemtuzumab 
Campath 
Prednisonea 
 
 
a  An average of >20mg/day of prednisone for the past 30 days prior to hospital admission. The total 
prednisone intake should be added and averaged over 30 days.  For example, a patient taking 40mg/day 
for 14 days would not be excluded because (40x14)/30=18.7.  Other steroids should be converted into 
prednisone equivalents. Additionally, any dose of corticosteroids given for any reason on or after hospital 
admission is acceptable. 
    
 

 
 
Page 85 
APPENDIX D: LUNG PROTECTIVE  VENTILATION PROTOCOL RECOMMENDATIONS 
Note:  The following are guidelines that are recommended for applicable patients.  However, each study 
site should use their own best judgment, and consult with the coordinating center if there are any 
questions.  
Ventilator Management 
A modified, simplified version of the ARDS Network lung protective lower tidal volume strategy will be 
used in this trial.  This strategy, which was associated with low mortality rates in three previous ARDS 
Network trials (ARMA, ALVEOLI, and FACTT), will ensure that study subjects receive the beneficial 
effects of lung protection while participating in this trial [104, 105].  ARDS Network personnel have 
substantial experience in the application of this protocol from the three completed trials noted above. 
1. Any mode of ventilation capable of delivering the prescribed tidal volume (VT, 6ml/kg predicted body 
weight, +/- 2ml/kg) may be used, provided the VT target is monitored and adjusted appropriately.  If 
airway pressure release ventilation (APRV) is used, tidal volume is defined as the sum of the volume 
that results from the ventilator pressure-release and an estimation of the average spontaneous VT.  In 
the spirit of providing lung protective ventilation, high frequency oscillatory ventilation will also be 
allowed in this trial. 
2. VT Goal:  6 ml / kg predicted body weight.  
3. Predicted body weight (PBW) is calculated from age, gender, and height (heel to crown) according to 
the following equations: 
a. Males: PBW (kg) = 50 + 2.3 [height (inches) – 60] 
b. Females:  PBW (kg) = 45.5 + 2.3 [height (inches) – 60] 
4. Measure and record inspiratory plateau pressure (Pplat) according to ICU routine (at least every four 
hours and after changes in VT and PEEP recommended) 
5. If Pplat > 30 cm H2O, reduce VT to 5 ml / kg and then to 4 ml / kg PBW if necessary to decrease Pplat 
to ≤ 30 cm H2O. 
6. If VT < 6 ml/kg PBW and Pplat < 25 cm H2O, raise VT by 1 ml / kg PBW to a maximum of 6 ml/kg. 
7. If “severe dyspnea" (more than 3 double breaths per minute on volume-cycled ventilator or airway 
pressure remains at or below PEEP level during inspiration), then raise VT to 7 or 8 ml/kg PBW if Pplat 
remains below 30 cm H2O.  If Pplat exceeds 30 cm H2O with VT of 7 or 8 ml/kg PBW, then revert to 
lower VT and consider more sedation. 
8. If pH < 7.15, VT may be raised and Pplat limit suspended (not required). 
9. Oxygenation target: 55 mm Hg < PaO2 < 80 mm Hg or 88% < SpO2 < 95%.  When both PaO2 and 
SpO2 are available simultaneously, the PaO2 criterion will take precedence.   
10. Minimum PEEP = 5 cm H2O 
11. Adjust FIO2 or PEEP upward within 5 minutes if there are consistent measurements below the 
oxygenation target range 
12. Adjust FIO2 or PEEP downward within 30 minutes if there are consistent measurements above the 
oxygenation target range.   
13. There are no requirements for maintaining a specific PEEP to FIO2 ratio.  The lower PEEP/higher FIO2 
table represents a consensus approach developed by ARDS Network investigators in 1995.  The higher 
PEEP/lower FIO2 table (ALVEOLI) yielded equivalent results in a randomized trial [105] and would be 
acceptable and perhaps preferable in patients who appear to respond with a substantial increase in 
arterial oxygenation in the transition from lower to higher PEEP. 
Lower PEEP/Higher FIO2 Treatment Group 
FIO2 
.30 
.40 
.40 
.50 
.50 
.60 
.70 
.70 
.70 
.80  
.90 
.90 
.90 
1.0 
PEEP 
5 
 5 
  8 
  8 
 10 
10 
10 
12 
14 
14 
14 
16 
18 
18-24 

 
 
Page 86 
 
Higher PEEP/Lower FIO2 Study Group 
FIO2 
.30 
.30 
.30 
.30 
.30 
.40 
.40 
.50 
.50 
.50 – .80 
.80 
.90 
1.0 
1.0 
PEEP 
5 
8 
10 
12 
14 
14 
16 
16 
18 
20 
22 
22 
22 
24 
 
Note: Levels of PEEP in these FIO2/ PEEP tables represent levels set on the ventilator, not levels of total-
PEEP, auto-PEEP, or intrinsic-PEEP. 
14.  No specific rules for respiratory rate. It is recommended that the respiratory rate be increased in 
increments to a maximum set rate of 35 if pH < 7.30. 
15.  No specific rules about I:E.  It is recommended that duration of Inspiration be ≤ duration of Expiration. 
16.  Bicarbonate is allowed (neither encouraged nor discouraged) if pH < 7.30. 
17.  Changes in more than one ventilator setting driven by measurements of PaO2, pH, and Pplat may be 
performed simultaneously, if necessary. 
 
D.2. Weaning 
Note:  Commencement of Weaning is occurring at the clinician’s discretion. 
Commencement of Weaning (applicable to patients ventilated invasively) 
Patients will be assessed for the following weaning readiness criteria each day between 0600 and 1000.  If a 
patient procedure, test, or other extenuating circumstance prevents assessment for these criteria between 
0600 and 1000, then the assessment and initiation of subsequent weaning procedures may be delayed for up 
to six hours. 
1. At least 12 hours since enrollment in the trial 
2. FIO2 £ 0.40 and PEEP £ 8 cm H2O or FIO2 £ 0.50 and PEEP = 5 cm H2O 
3. Values of both PEEP and FIO2 £ values from previous day  
4. Not receiving neuromuscular blocking agents and without neuromuscular blockade 
5. Patient exhibiting inspiratory efforts. If no efforts are evident at baseline, ventilator set rate will be 
decreased to 50% of baseline level for up to 5 minutes to detect inspiratory efforts. 
6. Systolic arterial pressure ³ 90 mm Hg without vasopressor support (£ 5 mcg/kg/min dopamine or 
dobutamine will not be considered a vasopressor) 
 
Spontaneous Breathing Trial Procedure and Assessment for Unassisted Breathing 
If criteria 1-6 above are met, then initiate a trial of up to 120 minutes of spontaneous breathing with FIO2 < 
0.5 using any of the following approaches: 
1. Pressure support (PS) ≤ 5 cm H2O, PEEP ≤ 5 cm H2O  
2. CPAP ≤ 5 cm H2O  
3. T-piece  
4. Tracheostomy mask  
 
The clinical team may decide to change mode during spontaneous breathing (PS = 5, CPAP, tracheostomy 
mask, or T-piece) at any time during the spontaneous breathing trial. 
 

 
 
Page 87 
Monitor for tolerance using the following: 
1. SpO2 ³ 90% and / or PaO2 ³ 60 mm Hg 
2. Mean spontaneous tidal volume ³ 4 ml/kg PBW (if measured) 
3. Respiratory Rate £ 35 / min 
4. pH ³ 7.30 (if measured) 
5. No respiratory distress (defined as 2 or more of the following): 
a. Heart rate ≥ 120% of the 0600 rate ( ≤ 5 min at > 120% may be tolerated) 
b. Marked use of accessory muscles 
c. Abdominal paradox 
d. Diaphoresis 
6. Marked subjective dyspnea 
 
If any of the goals 1-6 are not met, revert to previous ventilator settings or to PS greater than or equal to 10 
cm H2O with Positive End-expiratory Pressure and FIO2 = previous settings and reassess for weaning the 
next morning.  The patient will be reassessed for weaning (Section D2) the following day. 
 
Decision to remove ventilator support: 
If tolerance criteria for spontaneous breathing trial (1-6 above) are met for at least 30 minutes, the clinical 
team may decide to discontinue mechanical ventilation. However, the spontaneous breathing trial can 
continue for up to120 minutes if tolerance remains in question.  
 
D.3. Definition of Unassisted Breathing  
1. Spontaneously breathing with face mask, nasal prong oxygen, or room air, OR 
2. T-tube breathing, OR 
3. Tracheostomy mask breathing, OR 
4. CPAP £ 5 without PS or IMV assistance 
5. Use of CPAP or BIPAP solely for sleep apnea management  
 
D.4. Definition of Extubation 
1. Removal of an oral or nasotracheal tube 
2. If a patient receives a tracheostomy, the time of extubation is defined as the time when the patient 
achieves unassisted breathing as defined in Section D.3 
 
D.5. Completion of Ventilator Procedures 
Patients will be considered to have completed the study ventilator procedures if any of the following 
conditions occur: 
1. Death 
2. Hospital discharge 
3. Alive 28 days after enrollment 

 
 
Page 88 
 
If a patient requires positive pressure ventilation after a period of unassisted breathing, the study ventilator 
procedures will resume unless the patient was discharged from the hospital or > 28 days elapsed since 
enrollment. 
 
D.6. Removal from the Ventilator Management Protocol 
Patients may be removed from the 6 ml/kg PBW tidal volume ventilation requirement if they develop 
neurologic conditions where hypercapnia would be contraindicated (e.g., intracranial bleeding, GCS < 8, 
cerebral edema, mass effect [midline shift on CT scan], papilledema, intracranial pressure monitoring, fixed 
pupils). 
 
 
 
 
 

 
 
Page 89 
APPENDIX E: CONSERVATIVE FLUID MANAGEMENT 
Note: The following are guidelines that are recommended for applicable patients.  However, each study 
site should use their own best judgment, and consult with the coordinating center if there are any 
questions.   
This fluid protocol captures the primary positive outcome of the FACTT trial on increasing ventilator free 
days.  This protocol should be initiated within four hours of randomization in enrolled patients when 
applicable, and continued until UAB or study day 7, whichever occurs first. 
1.  Discontinue maintenance fluids. 
2. Continue medications and nutrition. 
3. Manage electrolytes and blood products per usual practice. 
4. For shock, use any combination of fluid boluses# and vasopressor(s) to achieve MAP ≥ 60 mmHg as fast as 
possible. Wean vasopressors as quickly as tolerated beginning four hours after blood pressure has 
stabilized. 
5. Withhold diuretic therapy in renal failure § and until 12 hours after last fluid bolus or vasopressor given. 
 
 
CVP 
(recommended) 
 
 
PAOP 
(optional) 
 
MAP ≥ 60 mm Hg AND off vasopressors for ≥  12 hours 
 
Average urine output < 0.5 ml/kg/hr 
 
Average urine output ≥ 0.5 ml/kh hr 
 
>8 
 
> 12 
 
Furosemide* 
Reassess in 1 hour 
 
 
Furosemide* 
Reassess in 4 hours 
 
4-8 
 
8-12 
 
Give fluid bolus as fast as possible* 
Reassess in 1 hour 
 
No intervention 
Reassess in 4 hours 
 
< 4 
 
< 8 
§ Renal failure is defined as dialysis dependence, oliguria with serum creatinine > 3mg/dl, or oliguria with serum 
creatinine 0-3 with urinary indices indicative of acute renal failure. 
# Recommended fluid bolus = 15 mL / kg crystalloid (round to nearest 250 mL) or 1 Unit packed red cells or 25 grams 
albumin 
*Recommended Furosemide dosing = begin with 20 mg bolus or 3 mg/hr infusion or last known effective dose.  
Double each subsequent dose until goal achieved (oliguria reversal or intravascular pressure target) or maximum 
infusion rate of 24 mg or 160 mg bolus reached.  Do not exceed 620 mg/day. Also, if patient has heart failure, 
consider treatment with dobutamine. 
 
NIH ARDS Network 
Revision date: March 9, 2009 
 
 

 
 
Page 90 
APPENDIX F: DATA AND SAFETY MONITORING PLAN  
 
Overview Prospective safety monitoring will be performed. Specific emphasis will be on hematotoxic effects as 
ganciclovir is known to be associated with these effects. The duration of treatment selected in this study is relatively 
short, thus we do not expect high rates of neutropenia. Relevant studies in this regard in the literature are two 
comparative studies of valganciclovir or ganciclovir in solid organ transplant recipients. Based on these studies, 
the rate of neutropenia should be approximately 2% with a 28-day course as proposed in this proposal. Most 
relevantly, our phase 2 RCT showed no neutropenia events. Numerous randomized trials do not show any evidence 
of ganciclovir causing thrombocytopenia. Mild anemia has been seen in some studies but the majority of trials did 
not show an association. A theoretical concern is the carcinogenicity in animal models. This effect has not been 
reported in humans; however, we have included a late follow-up time point to assess if there are any new diagnoses 
of tumors. Additional discussion of the safety profile is included in Section 3. 
Safety monitoring will be by standard CTC criteria. Also, specific expected adverse effects will be tracked. 
• 
Number and severity of reportable AEs and SAEs 
• 
Time to neutropenia (absolute neutrophil count [ANC] < 1000, <500 per mm3); use of G-CSF 
• 
Time to renal insufficiency (creatinine clearance < 60, < 30 ml/min) 
• 
Time to thrombocytopenia (platelet count < 50,000, < 20,000 per mm3) 
Long-term follow-up. The final protocol will include follow-up contact after hospital discharge. This is to assess 
secondary efficacy endpoints. The subject will be contacted over the phone at days 28 (if discharged from the 
hospital before this date) and 180 to assess vital status, pregnancy, and reportable adverse events. 
 
1.1. Monitoring for Safety by Study Sites 
 
Investigators are responsible for monitoring the safety of patients who have entered this study. Monitoring 
of weekly safety labs will be performed by the research team at each site for all participants, while 
hospitalized, with any reportable AE reporting per the protocol.  These research personnel will be blinded.  
 
The investigator remains responsible to follow, through appropriate health care options, reportable adverse 
events (AEs) that are serious, cause the patient to discontinue before completing the study, or are ongoing 
at the time of study completion.  The investigator will maintain responsibility for forwarding reportable 
SAEs to the coordinating site and their institutional review board.   
 
1.2. Monitoring by the Safety and Protocol Adherence Review Team 
The safety and protocol adherence review team (SPART) will, on a monthly basis, review all clinical and 
laboratory safety data during the study. The SPART will include CCC PIs (Drs. Boeckh, Stapleton, 
Rubenfeld, and Limaye), Clinical Site Manager Sara Ardren, and Clinical Trial Project Manager Dr. Louise 
Kimball. Meetings of the SPART (by videoconference or teleconference) will occur regularly, and will not 
require attendance of all team members at each meeting. The SPART is responsible for the review of the 
clinical safety reports, discordant CMV serostatus test results (LFA versus CLIA test), and protocol 
violations, as well as communication with the IRB, NHLBI executive secretary, and DSMB, as outlined 
above. 
 
1.3. Monitoring of Safety by an Independent Study Monitor 
Study data and regulatory aspects at study sites will be monitored by a study monitor. The study monitor 
will be managed by Dr. Louise Kimball, and will perform study monitoring remotely and according to 
industry standard, using EPIC if feasible, and via Florence eReg platform if not, and via Vestigo. The study 
monitor will perform monitoring of the first patient at each site as well as a random selection of 10% of 
participants (to be determined by DCC) across the whole study. The scope of monitoring is detailed below. 
Annually, they will monitor delegation of authority logs and training logs for each site, via Florence. The 
study monitor will create monitoring reports, submit them to the site investigators and the Executive 
Committee, and conduct the necessary follow-up. The study monitor will not have a direct reporting 

 
 
Page 91 
relationship with the DSMB. If an unreported reportable AE or SAE is found during the course of routine 
study monitoring, the usual processes for reporting the event will occur (see Section 11.3). 
 
Deadlines for data to be put in: 
• 
4-6 weeks after randomization (except for day 180 data) 
• 
Payments not made until data are in 
 
Scope of Monitoring: 
• 
Informed Consent Form 
• 
Inclusion/Exclusion criteria 
• 
CMV Lateral Flow Assay results and confirmatory IgG ELISA results 
• 
Study drug delivery to confirm intervention accuracy (dose and time of first dose) 
• 
Pharmacy logs to confirm appropriate temperature of drug 
• 
Serious adverse events, with special attention to known complications of ganciclovir 
o Protocol-defined reportable SAEs  
o Hospital discharge or death summary for possible reports of a serious unexplained events 
that the team associated with study drug administration 
• 
Outcome data, including data that might be used for adjustment in statistical models 
o Respiratory-support-free days (confirm start and stop times, including any separate 
episodes that might have occurred) 
o Death 
o The following parameters will be monitored randomly in 5% of subjects (selected at 
random from among the 10% of participants selected for monitoring above):  
§ 
Oxygenation (PaO2/FiO2 ratio) on study days 1-7 
§ 
ICU-free days by day (confirm ventilation start and stop times, including any 
separate episodes of ICU admission that might have occurred) 
§ 
Static respiratory system compliance at randomization, day 4 and day 7 
§ 
Invasive bacterial and fungal infections 
§ 
SOFA variables at baseline, day 4, and day 7 
§ 
The NHLBI-endorsed Acute Respiratory Failure Core Outcome Measurement Set 
(COMS) in survivors at day 180 
§ 
Risk factors for CMV reactivation 
• 
Sex 
• 
Age 
• 
Race/ethnicity 
• 
Co-morbidities 
• 
APACHE III score at baseline 
• 
SOFA score and individual components 
• 
Lymphocyte count 

 
 
Page 92 
• 
Time of hospital admission 
§ 
Use of ECMO; if yes, start and stop time 
§ 
Occurrence of neuromuscular blockade; if yes, start and stop time 
§ 
Prone positioning; if yes, start and stop time 
 
Protection against Risks 
Study procedures (blood draw, ETA) will be conducted in a clinical setting by medical staff trained to perform the 
various procedures. Medical attention will be promptly provided to patients who experience reportable adverse 
events resulting from study procedures. 
 
Safety labs will be monitored regularly by the research team at each site for any adverse reactions to study drug. In 
order to address the black box warning for ganciclovir, we have included an extended follow-up period of six 
months. 
 
Maintaining confidentiality 
Risk of breach of confidentiality will be minimized by keeping all research forms and electronic databases in 
locked and protected environments, and through training staff to be especially sensitive to confidentiality whenever 
interacting with participants. 
 
 .  

 
 
Page 93 
 
APPENDIX G:  SEPSIS CRITERIA  
 
Sepsis is defined according to the recent Sepsis-3 consensus definition [72], as life-threatening organ dysfunction 
caused by a dysregulated host response to infection.  
Organ dysfunction can be identified as an acute change in total SOFA score ≥2 points consequent to the infection. 
• 
The baseline SOFA score can be assumed to be zero in patients not known to have preexisting organ 
dysfunction. 
• 
A SOFA score ≥2 reflects an overall mortality risk of approximately 10% in a general hospital population 
with suspected infection. Even patients presenting with modest dysfunction can deteriorate further, 
emphasizing the seriousness of this condition and the need for prompt and appropriate intervention, if not 
already being instituted. 
 
Septic shock is a subset of sepsis in which underlying circulatory and cellular/metabolic abnormalities are profound 
enough to substantially increase mortality. 
Patients with septic shock can be identified with a clinical construct of sepsis with persisting hypotension requiring 
vasopressors to maintain MAP ≥65 mm Hg and having a serum lactate level >2 mmol/L (18 mg/dL) despite 
adequate volume resuscitation. With these criteria, hospital mortality is in excess of 40%. 
 
Abbreviations: MAP, mean arterial pressure; SOFA: Sequential [Sepsis-related] Organ Failure Assessment. 

 
 
Page 94 
APPENDIX H:  LIST OF INITIALLY SELECTED SITES 
Final listing will be on clinicaltrials.gov  NCT04706507 
 
Site 
PI 
Henry Ford Hospital (HFH) 
Mayur Ramesh 
Ohio State University (OSUMC) 
Matthew Exline 
Cleveland Clinic (CCF) 
Abhijit Duggal 
Brigham & Women’s Hospital (BWH) 
Rebecca Marlene Baron 
Harborview & University of Washington Medical Center (UW) 
Ajit Limaye 
University of Vermont (UVM) 
Renee Stapleton 
University of Pittsburgh Medical Center (UPMC) 
Scott Gunn 
University of Maryland (MARYLAND) 
Giora Netzer 
Washington University, St. Louis (UWSTL) 
Christina Vazquez Guillamet 
Wake Forest (WAKEHEALTH) 
Clark Files 
Johns Hopkins University (JHU) 
Dale Needham 
University of Michigan (UMICH) 
Robert Hyzy 
University of Colorado Denver (UCDENVER) 
Ellen Burnham 
University of Cincinnati 
Duncan Hite 
Duke University (DUKE) 
Christopher Cox 
Medical University of South Carolina (MUSC) 
Nandita Nadig 
Intermountain Medical Center 
Samuel Brown 
Vanderbilt University (VANDERBILT) 
Todd Rice 
Montefiore Medical Center (MONTEFIORE) 
Michelle Gong 
 
 

 
 
Page 95 
APPENDIX I: RECOMMENDATIONS FOR HFNC AND NIV 
Excerpt from the HENIVOT protocol (NCT04502576) 
 
Note: The following are guidelines that are recommended for applicable patients.  However, each study 
site should use their own best judgment, and consult with the coordinating center if there are any 
questions.   
 
High-Flow Nasal Cannula (HFNC) Oxygen  
HFNC therapy will be delivered with the Optiflow system.  
Initial set flow will be ≥ 50 /min, and flows will be decreased in case of intolerance and/or according to patients’ 
requirements: flows ≥30 L/min will be mandatory in all enrolled patients. Humidification chamber (MR860, Fisher 
and Paykel healthcare, New Zealand) will be set at 37 °C or 34 °C according to patient’s comfort [106]. FiO2 will be 
titrated to obtain an SpO2≥92% and ≤98%. 
Weaning the patient from HFNC will be considered only after 48 hours from enrolment.  
Weaning from HFNC within the first 2 days of the study will be allowed only whether the patient is considered for 
ICU discharge, according to the decision of the attending physician.  
All enrolled patients will be discharged from the ICU while undergoing low-flow oxygen, according to the 
prescription of the attending physician and the clinical practice of each participating institution (VenturiMask, nasal 
prongs, non-rebreathing oxygen mask). As suggested by Maggiore (NCT02107183), weaning from HFNC will be 
allowed when FiO2<40% and respiratory rate<25/min. Oxygen flow will be lowered to 10 L/min, keeping FiO2 
unchanged. Weaning from HFNC will be considered successful if the SpO2 remains between 92% and 98% and the 
respiratory rate ≤ 25/min with an oxygen flow of 10 L/min. In this case, the HFNC device will be replaced by the 
low-flow oxygen and oxygen flow or FiO2 will be set to obtain the same SpO2 target.  
HFNC treatment can be resumed any time if the patient is experiencing respiratory distress and hypoxemia, according 
to the prescription of the attending physician. 
 
Non-Invasive Ventilation (NIV) 
Patients in PSV group will receive continuous helmet pressure support ventilation for at least 16 hours/day the first 
2 calendar days. Continuous NIV without interruptions will be strongly encouraged in the first 24 hours of treatment. 
When NIV is interrupted, patients will receive low flow oxygen therapy or nasal high flow oxygen therapy, according 
to physician’s decision. Dedicated helmets for NIV (Dimar, Italia, or Intersurgical, UK) and size will be chosen 
according to neck circumference, as suggested by Antonelli et al. [107], or according to manufacturer 
recommendations, if present. 
 
Table 1. Helmet size according to neck circumference. 
17-27 
Extra small 
27-34 
Small 
34-40 
Medium 
40-47 
Large 
> 45 
Extra large 
  
Each patient will be connected to an ICU compressed gas based ventilator through a bitube circuit with no 
humidification.  
The ventilator will be set in PSV (the choice to use NIV modes will be left to the decision of the physician in charge 
of the patient), with the following suggested settings [108-112]:  
1. initial pressure support≥8-10 cmH2O and adequate to permit of a peak in the inspiratory flow of 100 l/min;  
2. positive end-expiratory pressure ≥10 cmH2O and increased to achieve the oxygenation target according 
to the choice of the attending physician.  
3. FiO2 will be titrated to obtain an SpO2≥92% and ≤98%.  

 
 
Page 96 
4. Inspiratory flow trigger = 1 l/min or according to the practice of each institution;  
5. fastest pressurization time;  
6. expiratory trigger: 10-50% of the maximum inspiratory flow;  
7. maximum inspiratory time 1.2 second.  
 
The use of earplugs to mitigate noise-related discomfort will be allowed according to the decision of the attending 
physician and will be encouraged especially overnight.  
Any modification in the ventilator settings and in the interface set-up to optimize comfort and patient-ventilator 
interaction will be allowed at the discretion of the attending physicians. However, maintenance of PEEP ≥10 during 
the treatment is mandatory. 
Weaning from NIV will be discouraged within the first 48 hours from enrolment. Weaning from NIV at any time 
will be attempted only whether FiO2 ≤40%, respiratory rate ≤25%: to assess the readiness for interrupting NIV, PEEP 
will be lowered to 8 cmH2O with pressure support=8 cmH2O, keeping FiO2 unchanged. If the patient maintains SpO2 
≥92% and respiratory rate ≤25 during the following 30 minutes with these settings, NIV weaning will be considered 
successful. After weaning from NIV and between two NIV sessions, patients will undergo low-flow, VenturiMask 
or HFNC, according to the choice of the attending physician: oxygen flow or FiO2 will be set to obtain the same SpO2 
target.  
NIV will be resumed at any time if the respiratory rate is more than 25 breaths per minute and SpO2 is less than 92% 
with and/or anytime deemed necessary by the attending physician.