Extending CPAP Therapy in Stable Preterm Infants to Increase Lung Growth and Function: A Randomized Controlled Trial
Phase: NA
Conditions: Neonatal, Premature Birth, Respiratory Distress Syndrome
Sponsor: Cynthia McEvoy

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Extending CPAP Therapy in 
Stable Preterm Infants to 
Increase Lung Growth and 
Function: A Randomized 
Controlled Trial 
 
 
 
 
 
Principal Investigator:  Cindy McEvoy, MD, MCR 
 
Funded by: National Heart, Lung, and Blood Institute 
Clinical Trials #NCT04295564 
 
 
 
 
 
 
 
 
 
 
 
Version 2.0 
 
 
 
 
 
 
 

 
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TABLE OF CONTENTS 
Contents 
TABLE OF CONTENTS ..........................................................................................................................................2 
PROTOCOL AMENDMENT HISTORY………………………………………………………………………………………………………………………………….4 
1. Objectives ........................................................................................................................................................................ 5 
Specific Aim 1 / Primary Outcome ................................................................................................................................. 5 
Specific Aim 2 / Secondary Outcomes .......................................................................................................................... 5 
Specific Aim 3 / Exploratory Outcomes ........................................................................................................................ 5 
2. Background ..................................................................................................................................................................... 6 
a. Introduction ................................................................................................................................................................. 6 
b. Background Studies ................................................................................................................................................. 7 
b1. Mechanical Stretch and Lung Growth ........................................................................................................... 7 
b2. Preclinical Preliminary Data ............................................................................................................................. 8 
b3. Clinical Preliminary Data ................................................................................................................................... 8 
3. Rationale for a Randomized Clinical Trial ............................................................................................................. 11 
4. Study Design ................................................................................................................................................................. 12 
a. Overview .................................................................................................................................................................... 12 
5. Study Population ......................................................................................................................................................... 12 
a. Number of Subjects ................................................................................................................................................ 12 
b. Inclusion Criteria ..................................................................................................................................................... 12 
c. Exclusion Criteria .................................................................................................................................................... 12 
d. Vulnerable Populations.......................................................................................................................................... 13 
e. Setting......................................................................................................................................................................... 13 
f. Recruitment Methods .............................................................................................................................................. 13 
g. Consent Process ..................................................................................................................................................... 14 
h. Randomization Method and Blinding ................................................................................................................. 14 
i. Assessment of Treatment Adherence ................................................................................................................. 15 
j. Removal of Patients from Allocated Treatment Arm or from the Study .................................................... 15 
6. Study Procedures Involved ....................................................................................................................................... 15 
a. Clinical Data Collection in NICU .......................................................................................................................... 15 
b. Blood for Epigenetics and Proteomics .............................................................................................................. 16 
c. Inpatient Infant Pulmonary Function Tests (PFTs) ......................................................................................... 16 
d. Outpatient Infant Pulmonary Function Tests ................................................................................................... 16 
e. Respiratory Questionnaires .................................................................................................................................. 17 
f. Neurodevelopmental Testing ................................................................................................................................ 18 
g. Cohort Retention Methods .................................................................................................................................... 18 

 
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h. Parent Questionnaire and Future Contact ........................................................................................................ 18 
i. Adverse Event and Protocol Violation Reporting ............................................................................................ 18 
i1. Definitions ............................................................................................................................................................ 18 
i2. Reporting of Adverse Events .......................................................................................................................... 19 
i3.Periodic Summary Reporting ........................................................................................................................... 19 
i4.Expected Risks ..................................................................................................................................................... 19 
7. Data Analysis ................................................................................................................................................................ 20 
a. Sample Size Estimation and Power Considerations ...................................................................................... 20 
b. Statistical Analysis ................................................................................................................................................. 21 
b1. Specific Aim #1 (Primary Outcome) ............................................................................................................. 21 
b2. Specific Aim 2 (Secondary Outcomes) ........................................................................................................ 22 
b3.  Specific Aim 3 (Exploratory Outcomes) ..................................................................................................... 22 
c. Missing Data ............................................................................................................................................................. 22 
d. Interim analysis and stopping rules ................................................................................................................... 22 
8. Data and Specimens ................................................................................................................................................... 22 
a. Handling of Data and Specimens ........................................................................................................................ 22 
b. Sharing of Results with Subjects ........................................................................................................................ 22 
c. Data and Specimen Banking ................................................................................................................................ 23 
d. Performance Monitoring ........................................................................................................................................ 23 
9. Privacy, Confidentiality, and Data Security .......................................................................................................... 23 
a. Provisions to Monitor the Data to Ensure the Safety of Subjects .............................................................. 23 
b. Risks and Benefits .................................................................................................................................................. 23 
b1. Risks to Subjects .............................................................................................................................................. 23 
b2. Potential Benefits to Subjects ....................................................................................................................... 23 
10. Study Administration................................................................................................................................................ 24 
a. Funding and Participating Center ....................................................................................................................... 24 
b1. eCPAP Steering Committee ............................................................................................................................ 24 
b2. eCPAP Executive Committee ......................................................................................................................... 25 
b3. Data and Safety Monitoring Board ............................................................................................................... 25 
c. Subject Stipend or Payments ............................................................................................................................... 25 
d. Study Timeline ......................................................................................................................................................... 25 
e. Training and Certification ...................................................................................................................................... 25 
f. Recruitment and Data Collection Period ............................................................................................................ 26 
g. Final Analysis ........................................................................................................................................................... 26 
Appendix A ........................................................................................................................................................................ 33 
 
 

 
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Extending CPAP Therapy in Stable Preterm Infants to Increase Lung Growth and 
Function: A Randomized Controlled Trial 
 
 
PROTOCOL AMENDMENT HISTORY 
 
 
Version number, date 
Alterations 
Version 2.0, 3/9/2020 
Clarification of measurement of FRC before 
randomization, addition of PHSW site 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
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1. Objectives 
 
    The objective of this randomized clinical trial is to determine the efficacy of extended continuous positive 
airway pressure (eCPAP) to improve lung growth and lung function in preterm infants, measured at six months 
of age. Prematurity occurs in 10% of births and is the most common cause of impaired lung development and 
increased respiratory morbidity1. In the early 1970’s, CPAP was used to acutely increase lung volumes and 
improved survival of preterm infants with respiratory distress. The subsequent addition of antenatal steroids, 
surfactant, and intubation with mechanical ventilation (MV) became the standard of care for several decades2. 
Despite these advances, prematurity still caused impaired lung development3 with decreased airway4 and 
parenchymal function5, and increased respiratory morbidity lasting into adulthood. No current  therapies exist 
to stimulate lung growth, improve lung function, and thus decrease respiratory morbidity after preterm birth. 
      Nasal CPAP at birth is now standard care to stabilize and maintain functional residual capacity (FRC), to 
avoid atelectrauma, volutrauma, intubation, and to reduce bronchopulmonary dysplasia (BPD)2;6-9. Although 
early CPAP use has been well studied, there are few studies defining the optimal time to discontinue CPAP in  
preterm infants10. Early CPAP use may benefit the preterm lung by minimizing injury; but, clinical and animal 
data indicate that mechanical stretch of the lung with CPAP may provide an important benefit of stimulating 
lung growth. Congenital diaphragmatic hernias (CDH) compress the lung in-utero causing lung hypoplasia11;12, 
and increasing intra-tracheal pressure in-utero stimulates lung growth in CDH models13.  We found 1-week of 
CPAP increased lung volume in ferrets14. We also reported in human 
preterm infants without BPD, those treated with CPAP had larger 
alveolar volumes and lung diffusion versus non-CPAP treated infants 
when studied after discharge15. These data suggest a physiologic 
rationale that CPAP may stimulate lung growth and development.  
     Our recent data16 strongly suggest that CPAP in stable, convalescing 
preterm infants stimulates lung growth. Preterm infants receiving CPAP 
and meeting criteria for stopping CPAP were allocated to an extra 2-
weeks of CPAP versus CPAP discontinuation (usual care). Both groups 
met CPAP stability criteria at ~ 32 wks of corrected gestational age 
(CGA). Infants assigned to the eCPAP demonstrated a greater increase 
in FRC compared to those who had CPAP stopped (Fig. 1). Importantly,  
the larger FRC in the eCPAP group persisted at discharge (~36.1 wks  
in both groups),although CPAP had been off > 2 weeks. There were no  
adverse events during treatment. This exciting data suggests that eCPAP in stable preterm infants is a novel 
non-pharmacologic therapeutic strategy to stimulate lung growth. In this study, lung growth and function were 
not measured after discharge. 
    We hypothesize that eCPAP in stable preterm infants in the neonatal intensive care unit (NICU) will 
increase alveolar volume (VA), lung diffusion (DL), and forced expiratory flows (FEFs) measured at 6 
months CGA compared to stable preterm infants who had CPAP discontinued by clinical criteria. 
Infants stratified by gestational age will be allocated to CPAP discontinuation (dCPAP, usual care) or eCPAP 
for 2-weeks to address the following Specific Aims (Fig 2):  
 
Specific Aim 1 / Primary Outcome: 
     Establish that 2 additional weeks of CPAP in the NICU for stable preterm infants increases alveolar volume 
at 6 months of age compared to infants who have CPAP discontinued, usual care.   
 
Specific Aim 2 / Secondary Outcomes: 
     Establish that 2 extra weeks of CPAP in stable preterm infants increases lung diffusion and forced 
expiratory flows at 6 months of age versus infants who have CPAP discontinued, usual care.   
 
Specific Aim 3 / Exploratory Outcomes: 
     Evaluate whether 2 extra weeks of CPAP in stable preterm infants results in lower respiratory morbidity 
and/or improved neurodevelopmental outcomes through 12 months of age compared to infants who had CPAP 
discontinued, usual care. This will provide pilot data for a future multi-center randomized trial. 
Figure 1. FRC in Randomized Patients; 
Values Mean ± SEM; *p<0.05 ∆FRC b/w  
groups during treatment; **p<0.05 ∆FRC 
b/w groups randomization to discharge 

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   Abbreviations: FRC= functional residual capacity; VA= alveolar volume; DL= lung diffusion; FEFs= forced expiratory flows 
     The clinical implications of our studies are highly significant as results could provide a non-pharmacologic 
approach to stimulate lung growth, improve lung function, and therefore decrease respiratory morbidity in 
preterm infants. Our proposal capitalizes on our novel hypothesis, strong pre-clinical and clinical pilot data, 
unique respiratory physiologic measurements to be obtained in infants, and clinical investigators with expertise 
in neonatology and pediatric pulmonary physiology and a record of productive collaboration. 
2. Background
a. Introduction 
      
     The primary function of the lung is gas exchange, which requires the parenchyma to develop a large alveolar 
surface area coordinated with pulmonary capillary blood volume. The lung must develop airways that conduct 
air between the environment and the lung parenchyma for gas exchange. These two different lung components 
have differing patterns of growth and development in utero. During fetal development, the airways are present 
at 16 weeks gestation and formation is nearly complete by 24 weeks gestation. However, development of a 
rudimentary gas exchanging region of the lung (saccules) does not begin until 23 weeks gestation, which defines 
the current limit for extra-uterine viability. In-utero lung parenchyma development occurs with an increase in 
capillary density and alveolar surface area, thinning of the interstitial space, and the presence of alveoli in late 
gestation17-20. Alveolar number and lung volume increase rapidly during the first 2-years of life and then continues 
at a far slower rate17;21;22. 
     Preterm delivery is the most common cause of altered lung development with potential lifelong sequelae, 
and prematurity is a global health crisis23;24. The origins of adult respiratory disease can be traced back to the 
fetal/early childhood period when the lung undergoes rapid development. Our laboratory has shown that 
infants born at 30-35 weeks GA with no clinical respiratory disease after birth had decreased airway function 
with lower forced expiratory flows (FEFs) in the first year of life, which lasted into the second year of life4. This 
confirms findings of other birth cohorts that individuals track along the lung function percentile that is 
established early in life25. This emphasizes the need to optimize lung function/growth in fetal and early 
postnatal life. Multiple risk factors for prematurity have been identified, but little progress has been made to find 
interventions to stimulate lung growth1. We, and others14-16;26-28 have strong pilot animal and human data 
that mechanical strain of the lung from continuous positive airway pressure (CPAP) may be a simple, 
safe, and non-pharmacologic stimulus to promote lung growth, improve lung function, and decrease 
airway reactivity. Although CPAP is the standard of care for preterm infants with respiratory distress, there is 
little evidence defining the optimal time to discontinue CPAP in stable preterm infants10. We have promising 
data randomizing stable preterm infants (≤ 32 weeks GA treated with CPAP for respiratory distress) to 
extended CPAP [(eCPAP) extra 2 weeks] versus CPAP discontinuation. The eCPAP resulted in increased lung 
volumes at the end of treatment and at discharge16. This study did not perform pulmonary function tests (PFTs) 
after discharge. Our goals in this proposal are to test the hypothesis that eCPAP in stable premature infants 
will stimulate lung growth and improve lung function as measured at 6 months of age.   
      The incidence of prematurity is a health care crisis with significant economic and emotional costs in the 
United States (US) and worldwide. One of every 9-10 babies in the US29, and an estimated 15 million babies 
worldwide29;30 are born preterm each year. Complications from preterm births are the leading cause of neonatal 
mortality and an important contributor to child and adult morbidities.  In the US, the yearly estimated costs 
associated with preterm births are $26.2 billion31-33.  All preterm infants have altered lung development with a 
high incidence of wheezy respiratory illnesses, hospitalizations, and asthma compared to full term infants34. In 
preterm infants <37 weeks with and without BPD in the first year of life, the incidence of cough was reported to 

 
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be 79%, wheezing 44%, and respiratory re-hospitalizations 25%35. A  recent study in 537 infants born at < 28 
wks gestation reported that 65% had post discharge respiratory disease and 71%  had frequent wheeze or 
cough for ≥ 6 months in the first year of life36.   If eCPAP is demonstrated to stimulate lung growth and 
improve lung function, it could improve the respiratory health of up to 500,000 preterm infants yearly in 
the US and millions of preterm infants worldwide. 
      Although CPAP is standard therapy for preterm infants with respiratory distress, it is not viewed as a 
potential therapeutic stimulus to promote lung growth and function. There is data that delivery room CPAP is 
feasible and effective in stabilizing preterm infants, even those very preterm2;7-9. Previously it was standard for 
very preterm infants to be intubated quickly after delivery and be given prophylactic surfactant. However, a 
2016 Cochrane meta-analysis concluded that compared to mechanical ventilation, prophylactic CPAP in very 
preterm infants reduces the incidence of BPD and death or BPD37. The consensus is that BPD is decreased in 
part due to the avoidance of volutrauma from intubation and mechanical breaths. However we and others have 
shown that CPAP used in small animal models can stimulate lung growth and airway size14;38, so the reduced 
BPD may be partially due to CPAP’s ability to promote lung growth and function in preterm infants. This has 
been shown in large preterm animal models. Preterm baboons quickly weaned from mechanical ventilation to 
CPAP did not show an alveolar arrest and had a normal internal surface area and surface to volume ratio39.  In 
a preterm BPD lamb model, Albertine et al showed that high frequency nasal ventilation (to mimic CPAP) is as 
effective as mechanical ventilation but led to better alveolarization27;28. Of note, Columbia University has been 
known for its low rate of BPD for decades and in addition to minimizing intubation, they continue CPAP until at 
least 34 weeks of gestation40. Our proposed study will provide physiologic evidence as to the optimal 
duration of CPAP therapy in preterm infants to stimulate lung growth and function.  
 
Our group has created two robust translational models for the fetal origins of childhood respiratory disease. 
The first is an ongoing trial in preterm primates with pilot data showing evidence of increased alveolar growth in 
the group randomized to CPAP versus sham CPAP41. This supports the findings of our pilot clinical eCPAP 
trial16 and the importance of our proposed study. The second is a model of in-utero nicotine in which we 
demonstrated that nicotine crosses the placenta, up regulates nicotinic receptors in the fetal lung42;43,and 
causes significant decreases in the offspring’s FEFs42-44. In this model we demonstrated that vitamin C blocked 
some of the negative effects of prenatal nicotine45. We translated these findings in a randomized controlled trial 
(RCT) showing offspring of pregnant smokers randomized to vitamin C had significantly improved newborn 
PFTs and less wheeze compared to the infants of pregnant smokers randomized to placebo46.  
 
     b. Background Studies 
 
     b1. Mechanical Stretch and Lung Growth 
 
   
     Congenital diaphragmatic hernias (CDH) encroach upon the lung during fetal development and prevent in-
utero lung expansion, which results in a hypoplastic lung at birth with high neonatal morbidity and mortality11;12. 
In contrast, complete congenital airway obstruction by laryngeal or tracheal atresia prevents lung fluid from 
leaving the trachea and produces a large, hyperplastic lung13. As an approach to treating CDH in utero, animal 
models have shown that experimental occlusion of the fetal trachea increases the lungs distending pressure 
and results in increased fetal lung growth47;48. Increased fetal lung distension stimulates capillary growth and 
epithelial branching in the fetal lung49-52.  
      While one approach to stimulate lung growth has focused on the fluid filled fetal lung, mechanical distention 
of air-filled lungs can also increase lung growth. Resection of lung tissue early in life can stimulate 
compensatory lung growth, which may occur due to the increased distending force in the remaining lung 
tissue53-55. Delivery of CPAP to young tracheostomized ferrets for 1– 2 weeks increased lung volume and DNA 
without altering the lung’s elastic properties38. Using young tracheostomized ferrets, we showed that 2 weeks 
of CPAP at 6 cmH2O increased both lung volume and airway size compared to non-distended lungs at CPAP 
= 1 cmH2O14. Therefore, chronic lung distention (in-utero or extra-utero) stimulates lung growth in preterm and 
term lungs, which may occur due of mechanical strain stimulating mechano-sensitive angiogenic pathways.  
  
 
 

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b2. Preclinical Preliminary Data   
b2a. Chronic CPAP increases Lung Volume and Airway Size in Young Ferrets 
     Eight-week old ferrets 
were tracheostomized and 
maintained at High (H) CPAP 
of 5-6 cmH2O or Low (L) 
CPAP of 0-1 cmH20 for 2 
weeks while mobile in their 
cages. Volumetric CT scans 
were obtained at distending 
airway pressures of 0, 10 and 
20 cmH2O to measure lung 
volume and airway size, pre 
and post H vs L CPAP 
treatment (Figures 3 A, B). 
There were no differences 
between groups prior to treatment; however, H-CPAP treatment significantly increased lung volume and airway 
size at all distending pressures14. 
      
b2b. Chronic CPAP Decreases Airway Reactivity in Young Ferrets 
     Mechanical strain imposed on the lungs during breathing is an important modulator of airway 
responsiveness in vivo56-60. Our laboratory has demonstrated that chronic mechanical strain produced in vivo 
by imposing CPAP dramatically reduces airway reactivity in 
vivo14;61 both in animal models and humans. Following CPAP 
treatment for 2 weeks, animals underwent in vivo bronchial 
challenge with increasing concentrations of acetylcholine. 
Animals treated with H-CPAP had markedly lower airway 
reactivity compared to animals treated with L-CPAP (Figure
4). We have also reported that adults with asthma treated with 
nocturnal CPAP (10 cmH2O) for 1-week resulted in a 
significant reduction in airway reactivity compared to adult 
asthmatics sham treated (CPAP = 0 cmH2O)26. As infants and 
children who were born preterm often have increased airway 
reactivity and are treated with asthma medications, extended 
use of CPAP very early in life to stimulate lung growth may 
suppress airway reactivity in premature infants62-65.  
b3. Clinical Preliminary Data 
Figure 4. Airway reactivity after low versus high 
CPAP. 
Figure 3: A) lung volumes and B) airway size, pre and post treatment with high vs. low 
CPAP 

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  b3a. Lung Growth and Development in Human Infants
     Lung Parenchyma: The primary function of the lung is gas 
exchange, which requires the parenchyma to develop a large alveolar 
surface area coordinated with pulmonary capillaries. This process 
begins during the saccular stage of gestation (>23 weeks) with an 
increase in capillary density, an increase in lung volume, and a 
thinning of the interstitial space. Alveolarization, which dramatically 
increases the pulmonary surface area and capillary density, begins in 
late gestation (36-weeks) and continues until at least 2-years of 
age5;66;67. All preterm birth may impair ongoing parenchymal 
development. Our laboratory developed the methodology to assess 
alveolar volume (VA) and DL in human infants5;68. We demonstrated that infants with BPD have significantly 
lower DL/VA, (Figure 5) which is consistent with morphometric findings from autopsy studies of infants born 
very preterm that indicates impaired alveolar development with fewer and larger alveoli69-72.   
While in human infants we are not able to demonstrate a direct relationship between alveolar structure and 
the physiologic measurement of DL, we have done so using a murine hyperoxia model of BPD. Figure 6
illustrates the impaired alveolar development following neonatal hyperoxia for 7 days, while Figure 7 illustrates 
the very strong correlations between structural measures of impaired alveolar development (increased mean 
linear intercept and decreased vessel density) and in-vivo measurements of pulmonary diffusion (DF) prior to 
sacrifice73. Therefore, physiologic measurements of pulmonary diffusion provides an excellent in vivo 
assessment of parenchymal growth and development. 
Airways: Our laboratory also developed the methods to measure FEFs 
in infants using the raised volume rapid thoracic technique, which 
provides comparable information about airway function as maneuvers 
obtained in cooperative older children and adults74;75. We demonstrated 
that infants with BPD have decreased FEFs compared to infants born 
full term76. In addition, we have also reported that infants born preterm 
(30-35 weeks) and who didn’t require significant respiratory support also 
have lower FEF75 in the first year of life compared to full term infants, 
which persisted into the second year of life4 (Figure 8). These findings 
demonstrate our ability to assess airway function in infants, and the 
significant and persistent effect of preterm birth upon airway function.  
 
b3b. Preterm Human Infants treated with any CPAP vs no CPAP in NICU had Higher VA and DL  
     
     Using measures of VA and DL, we assessed lung growth and development of 48 stable outpatient infants 
who had been born preterm between 27 and 36 weeks gestational age (GA), but did not develop BPD versus  
88 healthy full term infants15. The preterm infants had a mean GA at birth of 31.7 weeks, and were studied at a 
mean of 13.7 months of age. Term infants had a GA of 39.3 weeks and were studied at an average of 14.4 
 
Fig 8. FEF75 were significantly lower 
in preterm versus term infants when 
measured in both year 1 and 2 of life. 
Figure 5: Pulmonary diffusion (DL) was 
significantly lower for Infants with BPD 
than those born Full term (FT) 
adjusting for body length (p < 0.0001). 
 
0
1
2
3
4
FT
CLDI
DLCO (mL/min/mmHg)
Fig.7. Alveolar development after room air 
exposure and hyperoxia for 7 days. 
Fig. 6. Alveolar development after room air 
exposure and hyperoxia for 7 days. 

 
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months. VA and DL were not associated with GA, mechanical ventilation or oxygen therapy suggesting that in 
the absence of extreme prematurity and BPD, prematurity itself does not impair lung parenchymal 
development. However, further analysis of the preterm infants demonstrated that both VA and DL were 
significantly higher (by about 16%) for infants who received any CPAP in the NICU compared to those who had 
not received any CPAP (Table 1). This was not a controlled trial of CPAP, but our findings suggest that 
mechanical stretch from CPAP in preterm infants may produce larger, more functional lungs. 
Table 1. VA and DL in Healthy Preterm Infants According to CPAP Treatment 
Lung Function Measures 
No CPAP  (n=29) 
CPAP (n=19) 
P value 
% difference 
VA (mL) 
606.8 ± 171.5 
694.6± 205.2 
0.046 
16% 
DL (mL/min/mmHg) 
3.86  ± 0.97 
4.49± 1.42 
0.007 
16% 
Values are Mean ± SD; p values adjusted for length and gender 
 
 
b3c. Pilot Study: The Effect of eCPAP on FRC through NICU discharge, a RCT 
 
     We recently published a pilot study randomizing stable preterm infants ≤ 32 weeks of gestation at birth and 
requiring ≥ 24 hours of CPAP for respiratory distress  to 2 extra weeks of CPAP (eCPAP) versus CPAP 
discontinuation when CPAP stability criteria were met16. CPAP stability criteria included: on CPAP of 4-5 cm 
H2O; on room air; respiratory rate ≤ 70 breaths/minute, retraction score ≤ 1; < 3 self resolving apneas (< 20 
seconds) and/or bradycardia (< 100 bpm) and/or desaturations (pulse oximeter oxygen saturation, SpO2 <86%) 
in 1 hour for past 6 hours; average SpO2 >86% for 90% of the time over the past 24 hours, not being treated  for 
a PDA or sepsis, and tolerating time off CPAP during routine care (up to 15 minutes). 
       
 
Critical to the pilot’s success was the extensive in- servicing with all 
caregivers on the rationale for the study prior to the study start, 
establishing bedside and parent advocates, and emphasis that 
participation in the study would not prolong the infant’s hospitalization or 
increase patient costs. In the pilot, 66% of the families whose infant met 
all of the inclusion and exclusion criteria consented to the study. The 
primary reason parents refused consent was the desire to see their 
infant’s face unobstructed by CPAP.  Additional recruitment approaches 
developed during the pilot included: professional grade framed photo for 
the parent done during short periods infant was taken off CPAP for  
“CPAP cares”;  emphasizing the ability for parents to hold their baby on 
their chest skin to skin while infant on CPAP.  Only 1 parent out of 50 
withdrew consent after randomization. 
 
As shown in Table 2, both groups of infants fulfilled CPAP stability 
criteria at ~32 wks CGA. Infants randomized to the eCPAP had a 
significantly increased change in FRC at the end of the 2-week treatment 
period and excitingly also at discharge (Figure 1, Table 2). Participation in the study did not extend the infant’s 
duration of stay which is usually dictated by the infant’s oral maturation to adequately nipple all of their feeds 
(usually occurs at 35-36 weeks CGA). The CGA at full nipple feeds and at discharge (35.8 ± 0.6 wks in the 
eCPAP and 36.0 ±1.0 wks in the CPAP discontinuation) was comparable between groups. This data 
supports the hypothesis that eCPAP in stable preterm infants will stimulate lung growth. 
 
Table 2. Characteristics of Randomized Infants 
 
Birth 
GA(wks) 
CGA at 
Rnd (wks) 
Weight at 
Rnd (g) 
FRC at Rnd 
(mL) 
Avg FRC∆, Rnd 
to 2 wks (mL) 
Avg FRC∆, Rnd 
to Discharge (mL) 
eCPAP (n=22) 
29.5±1.9 
32.2±0.7 
1567±293 
40.5± 11.8 
12.6 ±11.4* 
27.2±12.5* 
Off CPAP (n=22) 
28.7±2.2 
32.0±0.8 
1482±323 
41.5±10.6 
6.4 ± 10.1 
17.1± 11.7 
Values are Mean ± SD; eCPAP= extended CPAP; CGA= corrected gestational age; RND= randomization; Avg=Average; 
*p<0.05 for the difference in the change in FRC between the CPAP and no CPAP infants by multiple linear regression with 
repeated measures and time treatment interaction adjusting for gender, twins, and weight at randomization. 
Figure 1. FRC in Randomized 
Patients; Values Mean ± SEM; 
*p<0.05 ∆FRC b/w groups during 
treatment; **p<0.05 ∆FRC b/w groups 
randomization to discharge;  solid 
line is treatment period and dotted 
is through discharge.

 
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     Extended CPAP may constrain developmental therapies77 and may affect intermittent hypoxic episodes 
which are common in preterm infants, underestimated by clinically used pulse oximeter settings, but have been 
associated with long-term neurodevelopmental problems78-80. The pilot eCPAP study had limited funding for 
the collection of detailed clinical respiratory and neurodevelopmental outcomes after discharge from the NICU. 
These outcomes will be explored in greater detail in the proposed study to provide data for a planned multi-
center trial. In the pilot eCPAP study, 89% of the infants had a respiratory questionnaire done at 12 months of 
age with wheeze or cough without cold as per parent report35 occurring in 58% of the infants in the eCPAP arm 
versus 73% in the CPAP discontinuation. This will be further studied with monthly respiratory questionnaires in 
the proposed study. Available Bayley neurodevelopmental scores at 6 months CGA from 29 of the 44 infants in 
the pilot study demonstrated the infants in the eCPAP arm had an average cognitive percentile score of 61.5% 
versus 46.7% in the CPAP discontinuation arm.  
 
b3d. Sedation with Chloral Hydrate 
 
     Sedation with chloral hydrate is the standard for infant PFTs after 44 weeks of CGA as per the American 
Thoracic Society81;82. Dr. McEvoy has IRB approval for sedation for PFTs at Oregon Health & Science 
University (OHSU) for term/preterm infants (current eIRB #386). In our recent RCT (Drs. McEvoy, Morris Co-
PIs; Dr. Tepper Co-I) “Vitamin C to Decrease the Effects of Smoking in Pregnancy on Infant Lung Function 
(VCSIP)83;84, chloral hydrate sedation was done in 220 infants at 3 and 12 months of age for a total of about 
440 total pulmonary function tests. There were no serious adverse events due to sedation.  
 
3. Rationale for a Randomized Clinical Trial 
 
 
Our pilot study which showed a significantly increased change in FRC at the end of the two week treatment 
period and at discharge in the infants randomized to the additional two weeks of CPAP versus CPAP 
discontinuation is very encouraging16. However the measurements in the pilot study were limited to those done 
in the NICU and did not measure lung diffusion which currently cannot be measured in the NICU. This new 
randomized trial will allow us to measure the persistence of the effect of eCPAP on lung volumes and to 
measure the effect on lung diffusion at six months of age. It will also allow us to measure the effect of eCPAP 
on forced expiratory flows and other exploratory outcomes including clinical respiratory and 
neurodevelopmental outcomes. 
  

 
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4. Study Design 
     a. Overview 
     This is a randomized single-blind Phase 2 trial of eCPAP in 
100 stable preterm infants born at 24 to < 32 weeks of 
gestational age (Figure 9, Study Overview). Once infants are 
consented and, meet CPAP stability criteria  they will be 
randomized to one of two groups: 
 
1. 2 additional weeks of CPAP (eCPAP) 
2. CPAP discontinuation as per usual NICU care 
(dCPAP) 
 
     Prior to randomization and at the end of the 2 weeks of 
treatment, a pulmonary function test (PFT) will be performed. 
Infants will return to OHSU at approximately 6 months of 
corrected gestational age (CGA) for pulmonary function 
(primary outcome of alveolar volume). Neurodevelopmental 
testing will be done at 6-12 months of age. Infants will be 
followed up to 1 year of age with monthly respiratory 
questionnaires that will be completed via phone. The study will 
be performed at OHSU and at PeaceHealth SouthWest 
Washington Medical Center (PHSW) and build on Dr. McEvoy’s 
strong pilot data16. It is not feasible or ethical to blind the 
caregivers and researchers to the randomized intervention in 
the NICU; however, the 6 month PFTs will be directed by 
personnel blinded to treatment allocation in the NICU and 
blinded personnel will administer the monthly respiratory 
questionnaires and neurodevelopmental assessments. The 
individual PFTs will be reviewed for acceptability, reproducibility, 
and completeness by Dr. Tepper, (Co-PI of study) and Christina 
Tiller, RRT, an experienced infant pulmonary function 
technologist. They will be blinded to the treatment allocation as 
will the statisticians analyzing the final data. 
5. Study Population 
      
     a. Number of Subjects 
 
• 
We anticipate screening 300 infants to recruit the cohort  
• 
Up to 130 infants will be consented once inclusion/exclusion criteria have been verified  
• 
100 infants will be randomized  
 
     b. Inclusion Criteria  
1) Infants born at >24 to ≤ 32 weeks gestation  
2) Treated with CPAP for ≥ 24 hours for respiratory distress (either as initial therapy or following 
extubation)  
 
     c. Exclusion Criteria  
1) Significant congenital heart disease 
2) Major malformations 
3) Chromosomal anomalies  
4) Culture proven sepsis at consent  
Figure 9. Study Overview 

 
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5) Complex maternal medical conditions  
6) Clinical instability  
7) Multiple gestations > twins  
8)  <3rd or >97th percentile for weight85  
9) Participating in another neonatal randomized clinical trial with a competing outcome 
10) Mother/legal guardian without stable method of communication 
 
Additional criteria that must be met prior to randomization: 
1) CPAP stability criteria met for at least 12 hours 
 
• 
The following CPAP stability criteria16;86 has to be met for more than 12 hours by 34 weeks of CGA:  
o CPAP of 5 cmH2O 
o On room air  
o Respiratory rate ≤70 bpm without significant retractions 
o <3 self-resolving apneas or bradycardias or desaturations in 1 hour in last 6 hours 
o Average SpO2 >86% for 90% of the last 24 hours 
o Not being treated for a patent ductous or sepsis 
o Tolerating CPAP “cares” 
 
 
 
     d. Vulnerable Populations 
      
     Neonates will be included in this research. The inclusion of neonates is critical to the research because we 
have pilot data in preterm infants demonstrating that eCPAP may stimulate lung growth. However, our initial 
trial did not include follow-up testing after discharge to determine the potential duration of the effect of eCPAP 
on lung volumes and did not include measurements of lung diffusing capacity.  Parent or guardian permission 
will be obtained before the child will be enrolled in the study, as per IRB guidelines.     
 
     e. Setting 
 
     Infants will be recruited in the 44 bed NICU at OHSU where Dr. McEvoy is the Director of Neonatal 
Research and the 20 bed NICU at PHSW where the OHSU neonatology division also provides NICU services. 
The OHSU Neonatal Division has extensive experience with bubble CPAP which has been the standard 
therapy of preterm infants for > 15 years at OHSU. The Neonatal Division follows extensive clinical consensus 
guidelines (CCG) with regards to important aspects of patient care including: enteral feeds, transfusions, septic 
workups among others. This will provide critical consistency to the proposed eCPAP study for the multiple 
aspects of patient care. Most importantly, the CPAP CCG since 2014 establishes bubble CPAP via Hudson 
prongs as the only CPAP delivery system used in the OHSU NICU and outlines the following practice as usual 
care at OHSU: when an infant meets CPAP stability criteria for at least 12 hours, the CPAP is discontinued and 
the infant is placed on room air. This CCG was established after review of limited published evidence to 
provide consistent weaning of preterm infants on CPAP. In NICUs worldwide, there are many different 
approaches to weaning patients off CPAP87;88 even within the same NICU. The consistent approach at OHSU 
and PHSW is an exception and will minimize potential confounders to our results. 
 
     f. Recruitment Methods 
 
     Recruitment will be maximized by having a specific NICU research coordinator screen daily for subjects, 
maintain a screening log, and consent subjects. Daily review of eligibility and CPAP stability criteria for 
individual patients will be performed with the neonatologist and bedside nurse. The study will be introduced to 
parents whose infant meets the screening criteria (24 to ≤ 32 weeks gestation and on CPAP). The infant will be 

 
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consented after inclusion and exclusion criteria are verified and randomized when CPAP stability criteria are 
met for at least 12 hours.  
 
 
     We will do extensive in-servicing with all caregivers working in the both NICUs prior to starting the study 
outlining the study’s rationale. We will present data from our initial study16 which demonstrated that the infant’s 
participation in the study did not affect length of stay or the time taken to achieve full nipple feeds. We will also 
discuss the fact that participation in the study will not increase hospitalization costs for the infant (level of 
hospital charges are based on need for gavage feeds regardless of whether infant is on or off CPAP). These 
same discussions will be done with the parents, starting far in advance of the infant being consented and 
meeting randomization criteria.  We will also assemble a patient advisory group of a projected four parents who 
will meet at least twice per year to give advice and feedback on the study elements. These parents will be 
recruited from the current OHSU NICU family advisory council that meets monthly. 
 
     g. Consent Process 
 
     Written informed consent will be obtained from each subject’s parent/legal guardian prior to any study 
procedures taking place. Once an eligible infant is identified, individuals authorized to obtain written informed 
consent (PI, co-investigators, and study staff specifically designated by the PI) will begin the informed consent 
discussion with the subject’s parent(s)/legal guardian. The informed consent process will take place in the 
NICU or in a quiet, private area.   
 
     The information provided in the consent will cover the elements in the CFR Part 50.25 and be approved by 
the site Institutional Review Board (IRB). This includes the investigational nature and objective of the trial; the 
procedures and treatments involved and their attendant risks, discomforts, and benefits; and the potential 
alternative therapies, alternative to not participate and right to withdraw without penalty, all of which will be 
explained to the parent/legal guardian in detail. All of the parent/legal guardian’s questions will be answered 
before signing the consent form. If the parent/legal guardian wishes to take the consent form home to consult  
with other family members or health care providers, or to allow more time for consideration, they will be 
allowed to do so.  A copy of the signed consent form will be given to the parent/legal guardian. 
 
     The informed consent process will be an ongoing active process of sharing information between the 
investigator and the parent/legal guardian(s).  If a protocol change requires a change to the consent form, 
parents/legal guardian(s) will be notified in a timely manner and the new informed consent form will be signed. 
Parent(s)/guardian(s) can withdraw from the study at any time. The study PI can discontinue a subject from the 
study at any time for safety concerns. Assent will not be obtained as the subjects in this study are neonates. 
 
     Permission may be obtained from legally authorized representatives, described as an individual who is 
authorized under applicable state or local law to consent on behalf of a child to general medical care when 
general medical care includes participation in research. Non-English speaking subjects may be enrolled. We 
will utilize a translated consent in Spanish and the IRB approved short forms for other languages. We will 
obtain an “Investigator’s Certification for Activities Preparatory to Research” from the OHSU IRB to examine 
records of infants preparatory to research that contain protected health information. The medical records will 
be reviewed to assess eligibility for this trial prior to approaching the parents to discuss consent. 
 
     h. Randomization Method and Blinding 
 
     Subjects will be randomized after meeting inclusion/exclusion criteria, being consented, and meeting the 
additional randomization criteria: 
1) CPAP stability criteria met for at least 12 hours 
The following CPAP stability criteria16;86 has to be met for more than 12 hours:  
o CPAP of 5 cmH2O 
o On room air  
o Respiratory rate ≤70 bpm without significant retractions 

 
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o <3 self-resolving apneas or bradycardias or desaturations in 1 hour in last 6 hours 
o Average SpO2 >86% for 90% of the last 24 hours 
o Not being treated for a patent ductous or sepsis 
o Tolerating CPAP “cares” 
 
 
 
     Randomization will be done via REDCap by the data management team within strata defined by gestational 
age at delivery (28 and 6/7 and 29 and 0/7 weeks).  If twins are both enrolled they will be allocated to the same 
treatment arm89. A computer generated randomization scheme with stratum using random block sizes of 2 or 4 
will be uploaded into REDCap for retrieval. The NICU team will know the treatment allocation but the follow-up 
research team assessing outcomes will be blinded to the treatment allocation. The electronic medical record 
will note the infant was in the study, but will not state allocation arm. 
 
     i. Assessment of Treatment Adherence 
 
     After randomization, the infant will be assessed daily by research staff for the 14 days on the assigned trial 
arm. As done in the pilot eCPAP study16, failure criteria for the respective randomization arms will be as 
follows: Infants randomized to dCPAP (usual care) will be defined as failing if they demonstrate:  
 
o significantly increased work of breathing with a persistent respiratory rate > 75 breaths/ minute, 
o increased apnea and/or bradycardia and/or desaturations > 2 significant  episodes in 1 hour in 
the past 6 hours,  
o oxygen need to maintain oxygen saturations > 86%,  
o a blood gas pH of < 7.2, PaCO2 > 65 mm Hg, or  
o major apnea or bradycardia needing resuscitation16;86 
  
     Subjects who meet the criteria for failing dCPAP will be resumed on their previous level of CPAP. Another 
attempt to discontinue the subject’s CPAP will occur within the next 5 days when they again meet the 
predefined stability criteria. If the subject does not meet the stability criteria or fails a second attempt at 
discontinuation, further weaning/management will be by the clinical team. The infant will continue to be studied 
as intention to treat. In our pilot study16, only 1 out of 26 subjects failed CPAP discontinuation.  
     Infants randomized to the eCPAP arm will have the CPAP stopped during the treatment period if there is 
significant abdominal distension with feeding intolerance or significant nasal breakdown (checked and 
documented every 3-4 hours as outlined in the OHSU NICU’s CCG). The CPAP will be discontinued for up to 5 
days and if symptoms resolve, the CPAP will be reapplied. If symptoms reoccur, the CPAP will be removed 
and further management will be by the clinical team. The infant will continued to be studied as intention to 
treat. In the pilot CPAP study16, all 22 patients tolerated eCPAP without adverse events.  
 
     j. Removal of Patients from Allocated Treatment Arm or from the Study 
 
 
     The investigator or the parents of the preterm infant may discontinue treatment arm allocation or withdraw 
the infant early from the study at any time for safety or administrative issues. The infant may be withdrawn from 
the treatment arm per parent request, protocol violation, worsening clinical condition, or investigator decision.  
Infants who discontinue study arm/treatment allocation will remain in the study for outcome measurements, 
unless the consent is withdrawn.  
 
6. Study Procedures Involved 
 
     a. Clinical Data Collection in NICU        
 

 
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     Pertinent maternal and infant data through discharge from the NICU will be collected, including: maternal: 
age; parity; smoking history; antenatal steroid treatment; etiology of preterm delivery; birth weight and 
gestational age; surfactant dosing; duration of: oxygen, mechanical ventilation and CPAP (prior to study 
randomization); respiratory medications; gestational age at full nipple feeds and at discharge. 
 
     During the 2-week treatment period detailed data will include daily weights; weekly lengths; calories per 
day; apneic episodes; oxygenation needs; parental interactions; developmental interactions such as frequency 
and duration of time spent skin to skin with a parent. 
 
     In addition, downloads of continuous oxygen saturation will be done during the two week treatment period 
with a separate pulse oximeter (will not be used for clinical care so monitor will be covered from caretaker and 
alarms disabled) with a 2 second averaging time and a 2 second sample rate (Radical; Masimo Corp., Irvine, 
California) monitors. Intermittent hypoxemia (IH) events will be defined as an oxygen saturation ≤ 80% for ≥ 20 
seconds and ≤ 180 seconds as previously described78-80. Episodes of IH and other patterns of oxygen 
saturation will be compared between the groups. 
 
     b. Blood for Epigenetics and Proteomics 
 
      Alterations in epigenetics including DNA methylation have been associated with perinatal origins of lung 
disease after different scenarios including in-utero smoke exposure90 and in association with decreased lung 
disease in preterm infants managed on CPAP versus mechanical ventilation91.  At the end of the two week 
treatment period of randomization to eCPAP versus dCPAP, about 2 mL of blood will be collected at the time 
of a blood draw for clinical care. Blood or DNA will be stored for future analysis of targeted and global DNA 
methylation and for proteins shown to be important in lung development in our primate model of CPAP such as 
YAP1, SCGB1A1, and STAT192.  Proteomic analysis will give a non-invasive assessment of the types and 
quantity of circulating proteins since samples of lung tissue and washings are not feasible. The targeted DNA 
methylation will be done using bisulfite amplicon sequencing or polymerase chain reaction based methods 
based on pre-existing knowledge from the literature such as IGF1 or TGFB pathways. Global methylation 
patterns will be investigated using ELISA based method (Imprint® Methylated DNA Quantification Kit, Sigma). 
This blood sampling and analyses are an optional part of the consent and will be stored in Dr. McEvoy’s locked 
research freezer in the CDRC-P at OHSU.  
 
 
     c. Inpatient Infant Pulmonary Function Tests (PFTs) 
 
     Infant pulmonary function testing will be performed when CPAP stability criteria is met prior to 
randomization and at the end of the two week treatment period. Studies will be performed with the infant 
supine, quietly asleep and at least 30 minutes after a feed.  All measurements will be obtained using a 
computerized infant pulmonary function cart (SensorMedics 2600; SensorMedics Inc, Yorba Linda, CA; 
Jaeger/Viasys Master Screen Baby Body) in the OHSU NICU at the infant’s bedside. Standard testing criteria 
outlined by the American Thoracic Society / European Respiratory Society will be met 81;93,94.  Tidal breathing 
will be recorded with up to 50 flow-volume loops46. Passive respiratory mechanics (respiratory compliance 
[Crs] and respiratory resistance [Rrs]) will be obtained using the single breath occlusion technique95. Functional 
residual capacity (FRC) will be obtained with the nitrogen washout technique with acceptance criteria as per 
ATS/ERS guidelines94. Vital signs (respiratory rate, heart rate, oxygen saturation) will be continuously 
monitored during the testing. If a PFT does not meet the pre-specified technically acceptable criteria it can be 
reattempted in 24-48 hours.  
 
     d. Outpatient Infant Pulmonary Function Tests 
 
     Infant pulmonary function testing will be performed at 6 months of CGA (range of 5-8 months corrected age) 
at OHSU for all randomized patients.  These will be primarily directed and performed by Dr. MacDonald who 
will be blinded to the NICU treatment allocation. Dr. MacDonald has experience with measurements of VA, DL, 

 
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and FEFs as these are the PFTs that were performed in the study at the Oregon National Primate Research 
Center examining the effect of CPAP on lung development. Keith Jackson RRT will also be a part of the 
research team performing the 6 month PFTs and he will be blinded as well. He was an integral part of the 
successful VCSIP study83,84 which measured FEFs at 3 and 12 months of age in offspring of pregnant smokers 
randomized to vitamin C versus placebo during pregnancy. Standard acceptance criteria will be applied. The 
PFT results will be reviewed off site for acceptability, reproducibility, and completeness by Dr. Tepper and/or 
Christina Tiller, RRT who has over ten years of experience in Dr. Tepper’s PFT laboratory performing and 
interpreting these tests. They will be blinded to the treatment allocation in the NICU as will the statisticians at 
Indiana University performing the data analysis.  
 
     No testing will be done within 3 weeks of a respiratory illness. A test may be repeated if the quality was not 
acceptable. This testing will include the measurement of alveolar volume (VA), pulmonary diffusing capacity 
(Dl),5;15 forced expiratory flows (FEFs),84 and functional residual capacity93;94 following the guidelines of the 
American Thoracic Society and European Respiratory Society which were co-written by Dr. Tepper 74;81. The 
pulmonary function testing equipment and operational procedures will be rigorously calibrated according to the 
manufacturer’s directions and outlined explicitly in the operations manual. The OHSU Pediatric Sedation 
Protocol will be followed using oral chloral hydrate.74;81  The infant will be given 50 to 100 milligrams per 
kilogram of chloral hydrate by mouth, with a maximum dose of 1 gram.  The parents will be counseled about 
possible side effects including temporary disorientation, nervousness, excitement, dizziness, coughing, 
nausea, mild desaturations, obstruction of airway, apnea and will be asked to sign a separate consent for 
sedation. They also will also be given patient education-discharge instructions. Adverse events will be reported 
as outlined below. For pulmonary function measurements with published normative data, the parents will be 
told whether the results of the pulmonary function test were within normal limits or outside of normal limits, and 
this report will be faxed to the patient’s physician.  The hemoglobin level will also be shared with the parent and 
if outside of normal limits for the CGA will be relayed to the infant’s physician.   
      
     Alveolar Volume (VA ) (Primary Outcome of Study) and: Pulmonary Diffusing Capacity (DL)(Secondary 
Outcome): Measurements will be obtained using an induced respiratory pause technique at an elevated lung 
volume (30 cm of H2O) that we developed and have previously described5;68 as approved in OHSU eIRB# 386. 
The inspiratory test gas contains 0.5% Ne (neon), 0.5% CO (carbon monoxide), 21% O2 (oxygen) and balance 
N2 (nitrogen). Concentrations of CO and Ne are measured with a gas chromatograph. During passive 
expiration following the 4-second induced respiratory pause for gas exchange, CO and Ne concentrations are 
used to calculate VA and DL. Results are expressed as averages of 2-3 measurements within 10%, adjusting 
for Hemoglobin from a finger stick. 
 
      
     Forced Expiratory Flows (FEF50) (Secondary Outcome) Measurements in triplicate will be obtained using 
the raised volume thoracic compression technique that we developed, and previously described74;75;83.  
 
     Chloral hydrate sedation: Infant PFTs will be performed at Doernbecher Children’s Hospital in sleeping 
infants following chloral hydrate sedation, as previously described68;74;84. For testing, infants are examined by a 
physician and continuously monitored by a sedation nurse or health care provider certified in sedation and a 
respiratory therapist. The parents will be counseled about possible side effects, sign a separate sedation 
consent, and be given the patient education-discharge instructions. Safety will be monitored as outlined in the 
DSMB charter. 
 
     e. Respiratory Questionnaires 
 
After discharge, a validated respiratory questionnaire will be administered monthly to the infant’s primary 
caregiver 83;96 by research personnel blinded to the treatment allocation of eCPAP versus dCPAP in the NICU.. 
Caregivers are asked about new episodes of wheezing in the previous month, as well as administration of 
prescribed medications, other illnesses, trips to the emergency room, and hospitalizations. If an infant has 
wheezing associated with an illness lasting up to seven days, this will be counted as one episode. These 
questionnaires will be administered in person at the PFT done at approximately 6 months of CGA while the 

 
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remainder of the monthly questionnaires will be administered by phone. A small time reimbursement will be 
available for those patients after each questionnaire is completed. 
  
     f. Neurodevelopmental Testing 
 
All preterm infants born at <34 GA are examined in the OHSU Developmental Evaluation Clinic (DEC)  if born 
at OHSU or the PHSW DEC clinic if born at PHSW.at 6 -12 months CGA. This assessment is the standard 
practice of their care. We will document the Bayley IV neurodevelopmental scores for cognitive, receptive 
language, expressive language, fine motor, gross motor, and audiology results from these visits97;98.  The 
personnel performing this testing will be blinded to the treatment allocation of eCPAP versus dCPAP in the 
NICU. 
 
 
     g. Cohort Retention Methods 
 
     Cohort retention will be maximized with the application of novel and evidence based retention techniques83 
as used in our previous VCSIP study which had a cohort retention of 93% at attempted 3 month PFTs84 and 
94% of infants with completion of respiratory questionnaires through 12 months of age as per protocol 84. 
Techniques include regular multimodal contact such as phone calls and text messages, social media private 
messaging, and pre-arranged check-ins at mother or child’s appointments. In instances where contact is 
difficult, alternative retention strategies are: searching databases for updated contact information; calling at 
different times of the day and on weekends; and contacting close friends and/or relatives who were listed by 
the participant as back-up contacts, or the participant’s work if permission was obtained during consent. 
 
     h. Parent Questionnaire and Future Contact 
 
 
     At the completion of the study the parent will be asked to complete a series of questions adapted from the 
standardized Research Participant Perception Survey 99 consisting of Likert scale items to assess their 
experience and satisfaction with the research study, team, and process. 
     We may contact subjects in the future if there are things we need to discuss or ask regarding this study or 
for future study opportunities, if they agreed on the consent form to this future contact. We will also review the 
subject’s medical records in the future to look at breathing and behavior outcomes up to 24 months of age. We 
may also review and obtain other health information which may be important for the study. We may obtain this 
information so that we can be sure to have all of the important information we need for the study. 
 
     i. Adverse Event and Protocol Violation Reporting 
 
     Safety events requiring expedited reporting will be sent to the DSMB chair within 7 calendar days of 
learning of the event. The DSMB chair will respond to Dr. Cindy McEvoy (study PI) with recommendations 
within 14 calendar days.  
     
All adverse events after randomization through discharge from the NICU and any adverse events after 
discharge deemed related or possibly related to the study (performance of the pulmonary function test done at 
6 months of age with required sedation) will be recorded on the Adverse Events Log. Events will be reported to 
the IRB as per IRB policy and procedures.  Adverse events (AEs) will be identified by review of the subject’s 
electronic medical record (EMR), and by physical exam / observations (including vital signs monitoring during 
sedation). AEs will be graded as to their expectedness and attribution (unrelated, possibly, probably or 
definitely related to the protocol).  The following standard definitions will be used: 
 
 
 
i1. Definitions 
 

 
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Adverse Event (AE):  Any untoward or undesirable, although not necessarily unexpected, event experienced 
by a human subject that may be a result of: 
• 
The interventions and interactions use in the research 
• 
The collection of identifiable private information in the research 
• 
An underlying disease, disorder, or condition of the subject 
• 
Other circumstances unrelated to the research or any underlying disease, disorder, or condition of the 
subject 
 
Serious Adverse Event (SAE):  Any AE that: 
 
• 
Is fatal 
• 
Is life-threatening 
• 
Is persistent or significantly disabling or incapacitating 
• Results in inpatient hospitalization or prolongation of hospitalization 
• 
Results in psychological or emotional harm requiring treatment 
• 
Creates a persistent or significant disability 
• 
Results in a significant medical incident (considered to be a serious study related event because, based 
upon appropriate medical judgment, it may jeopardize the subject and may require medical or surgical 
intervention to prevent one of the outcomes listed in this definition) 
 
Unanticipated problems (UP):  Events that are not expected given the nature of the research procedures and 
the subject population being studied and suggest that the research places subjects or others at a greater risk 
of harm or discomfort related to the research than was previously known or recognized.  Harm to a subject 
need not occur for an event to be an unanticipated problem. 
 
     i2. Reporting of Adverse Events 
All AEs, SAEs, and UPs should be documented on the appropriate adverse event logs/forms,(see Appendix B 
of DSMB charter) entered into the REDCap database, and reported to the study PI, Dr. McEvoy, as outlined 
below. 
• 
All SAEs and UPs should be reported by to Dr. McEvoy within 1 business day. 
• 
All other AEs should be entered into REDCap within 7 days. 
Dr. McEvoy and her research team will evaluate each event and will determine reporting requirements.  Dr. 
McEvoy and/or her research team will report events to the DSMB according to the following timeframes: 
• 
All SAEs and UPs that require expedited reporting (SAEs that are deemed related, suspected to be 
related, and unexpected, all UPs) will be reported to the DSMB chair within 7 calendar days of the time 
the PI learns of the event. 
• 
All other SAEs and AEs will be tabulated and reported to the DSMB at the next biannual meeting. 
Additionally, the following events will also require expedited reporting: 
• 
Neonatal or infant death through 1 year of age 
• 
Any sedation related event requiring significant resuscitation and/or hospitalization 
     i3.Periodic Summary Reporting 
All other AE’s (that did not require expedited reporting as above) will be summarized biannually for the DSMB. 
 
     i4.Expected Risks 
As detailed in the consent form, expected risks during the course of the study may include: 
• 
Restlessness during PFTs in the NICU 
• 
Pain during heel stick 
• 
Embarrassing questions 

 
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• 
Skin breakdown around infant’s nose or significant abdominal distension with feeding intolerance during 
the 2 weeks treatment period in infants randomized to eCPAP 
• 
Increased respiratory distress with significant apnea during the 2 week treatment period in the infants 
randomized to dCPAP  
• 
Sedation with chloral hydrate may include temporary disorientation, nervousness, excitement, 
dizziness, coughing, nausea, mild desaturations, obstruction of airway, apnea 
• 
Breach of confidentiality 
 
These risks are considered moderate and are addressed in the consent form.  
 
The DSMB will meet twice per year by teleconference and review all other AEs/SAEs at that time. The DSMB 
will be composed of two neonatologists, a research ethicist (who may also serve as one of the neonatologists), 
a pediatric pulmonologist, and a statistician. Members of the NHLBI will also attend the DSMB meetings. See 
DSMB charter for further details. Minor protocol deviations will be documented in the subject’s research chart.  
Major protocol violation will be reported to the IRB, per IRB guidelines.  
 
7. Data Analysis 
 
     a. Sample Size Estimation and Power Considerations 
 
 
     No studies have compared VA at 6 month of age in stable preterm infants randomized to eCPAP versus 
dCPAP in the NICU. Data from our pilot eCPAP study showed a 12% higher FRC in stable preterm infants 
randomized to eCPAP versus dCPAP at the end of the two week treatment period16. A study by Dr. Tepper et 
al reported a 16% difference in both VA and DL in preterm infants according to CPAP treatment in the NICU 
(Table 1)15 and a 12% difference in DL between infants with BPD versus healthy term infants69. Therefore, we 
powered our randomized trial to detect a 12% difference in VA between the eCPAP and dCPAP groups.   
 
     Using an ANCOVA model and assuming a mean VA of 642 mL (SD=189) based on data of the overall mean 
and SD from the control and CPAP groups reported in Assaf et al15, a sample size of 34 infants per group is 
needed to yield 80% power for detecting a 12% difference (effect size=0.42) in VA in the eCPAP group at 
α=0.05 (Table 3). This sample size also allows us to detect a 12% difference in DL with an effect size of 0.42. 
We estimate that 18% of the study samples will be twin pairs based on our pilot eCPAP study. Conservatively 
counting just one infant out of each twin pair for the sample size estimation, we will need 38 infants per group 
(76 total infants at 6 months) with completed VA outcomes. We further conservatively project a 10% loss in the 
NICU and a 15% loss from NICU discharge to the 6 month PFT (due to preterm population and need to be at 
least 3 weeks after a respiratory infection for the performance of the PFT) and therefore we project we will  
 
Table 3. Estimated power for 34 infants per group with completed PFT data  
 
Outcome Measures 
 
Mean (SD) in 
Control Group 
Estimated Power for Group Difference 
12% 
15% 
17% 
Alveolar volume (VA) 
Primary Outcome 
642 (189) 
80 
92 
97 
Lung diffusing capacity (DL) 
Secondary Outcome 
4.1 (1.19) 
80 
92 
98 
 

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need to randomize 50 infants per group (total n=100) into the trial. This sample size will provide 80% power for 
detecting an effect size difference of 0.42 in outcome measures between the two groups in Aim 1 (primary 
outcome of VA) and Aim 2 for DL. This sample size allows us to detect a 23% difference in FEF50. Figure 12 
shows the projected study recruitment to achieve the desired sample size based on recruitment and cohort 
loss data from the pilot eCPAP study and from the VCSIP83,84 
study. In the pilot eCPAP study 66% of parents whose infants 
met all inclusion and exclusion criteria consented to enrollment 
in the study. In this follow-up eCPAP study we are projecting that 
50% of the parents whose infant meets all inclusion and 
exclusion criteria will consent due to the needed sedation at the 
6 month PFT. 
     With regards to the exploratory clinical respiratory outcome, 
published data in 48 preterm infants with an average gestational 
age of 28 weeks who did not develop BPD (similar to the group 
we project to recruit) reported a 48% incidence of wheeze and a 
79% incidence of wheeze and cough in the first year of life 35. 
These researchers also documented a 21% incidence of wheeze 
in 195 term infants during the first year of life. In our initial 
vitamin C study46 using the same questionnaire we will use in 
this proposed eCPAP study, 76 term infants born to non-smokers 
had a 27% incidence of wheeze in the first year of life. Based on 
these data, we project that 48% of the infants who have CPAP 
discontinued by the CPAP stability criteria will have wheeze in 
the first year of life.  With our projected sample size of 50 per 
group accounting for a 15% missing questionnaire rate, we will 
have 70% power to show a decrease in the incidence of wheeze from 48% to 21% and 90% power to show a 
decrease in wheeze and cough from 79% to 50%.  
b. Statistical Analysis
     To verify the comparability of the randomized groups, infants’ baseline characteristics will be compared 
using analysis of T-test or non-parametric Wilcoxon test for continuous variables and chi-squared test for 
categorical variables. We will examine the distributions of continuous variable and use alternative approaches 
such as transformation or nonparametric methods in cases of violation to the normal distribution assumption. 
We will examine the frequency distribution of all categorical variables and adopt exact inference procedures in 
cases of zero or small cell size. All analyses will be conducted using the SAS 9.4 (SAS Institute, Carey, NC).  
b1. Specific Aim #1 (Primary Outcome)
     General linear mixed models (GLMMs) will be used to compare the primary outcomes of alveolar volume 
(VA) between the eCPAP and dCPAP while adjusting for the design factor gestational age at delivery (28 and 
6/7 and 29 and 0/7 weeks) and other baseline variables that are found to be statistically different between the 
two groups.  Since the lung function measures are highly dependent on infant length and sex, we will further 
adjust for length and sex in the GLMMs.  In order to adjust for intra-twin correlations, generalized Estimating 
Equations (GEEs) with the linear link function will be used to model VA.. The GLMMs can be performed using 
the GEE estimation approach. Intention to treat analysis will be used for all aims as the primary analysis. The 
intention to treat population is defined as all randomized infants and the analysis uses the assigned 
randomization group even if infants did not complete the intervention. In addition, a per protocol analysis will be 
completed for all aims. The per protocol population is defined as all infants who successfully completed the 
treatment arm to which they were assigned. 
Figure 12: Projected Study Recruitment and 
Follow-up

 
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     b2. Specific Aim 2 (Secondary Outcomes) 
 
 
     Lung diffusion (DL) and FEF50 will be will be analyzed with the same general approaches described for the 
primary outcome in SA #1. 
      
     b3.  Specific Aim 3 (Exploratory Outcomes)  
  
     Logistic regression models will be used to compare the incidence of wheeze and the incidence of wheeze 
and cough between the two groups adjusting for GA at delivery (<29 vs ≥ 29 weeks). Neurodevelopmental 
scores will be analyzed with the same general approaches described for the primary outcome in Specific 
Aim 1. 
 
     c. Missing Data       
 
     We anticipate loss to follow-up for the lung function outcomes at 6 months and respiratory outcomes 
through 12 month of age. Our sample size estimation conservatively accounts for 10% cohort loss from 
randomization to NICU discharge and a 15% loss from discharge until the 6 month PFTs. However based on 
VCSIP, which had a 93% cohort retention for attempted of 3 month PFTs84, coupled with our retention 
techniques, we suspect this will be overestimated but want to ensure an adequate sample size for analysis of 
the primary outcome. We will compare patients’ characteristics at baseline between those with outcome 
measures and those lost to follow-up and determine whether the missing data differ between the groups. If the 
missing at random assumption is violated, we will conduct extensive sensitivity analyses to model various non-
ignorable missing data patterns to examine the robustness of our findings to the various missing data 
assumption100.          
 
     d. Interim analysis and stopping rules  
      
     No interim analysis for efficacy or futility is planned. There will be no pre-specified stopping rules, but the 
DSMB will review SAEs as they occur. Although this is a fragile population, based on our pilot CPAP study, the 
fact that infants will be convalescing by the time they are enrolled in the study, and our experience with 
sedation in the VCSIP84, we do not anticipate a large number of SAEs but will be vigilantly monitoring. The 
DSMB chair may call a DSMB meeting to specifically review safety concerns.  
 
8. Data and Specimens 
 
     a. Handling of Data and Specimens 
 
     All data will be stored in locking file cabinets and contained within a password protected database. Neonatal 
and follow-up data from all patients consented for participation into the study will be recorded on forms 
designed specifically for this study. A screening log of all preterm infants who qualify for randomization to 
eCPAP versus dCPAP will be kept to determine the generalizability of the population. All data will be entered 
electronically by the research staff into the secure REDCap system.  Data will be store indefinitely.   
 
     b. Sharing of Results with Subjects 
 
     For pulmonary function measurements with published normative data, the parents will be told whether the 
results of the pulmonary function test were within normal limits or outside of normal limits, and this report will 
be sent to the patient’s physician. The hemoglobin level will also be shared with the parent and if outside of 
normal limits for CGA will be relayed to the infant’s physician.   
 

 
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c. Data and Specimen Banking 
 
Data and blood specimen will be banked in a repository for future research as outlined in the separate 
repository protocol. 
 
d. Performance Monitoring 
 
The data center will present regular reports to the eCPAP Steering Committee and the Data and Safety 
Monitoring Board. These will include:  
• 
Monthly recruitment reports: reports of the number of preterm infants screened and enrolled by month 
will be provided weekly and monthly to the eCPAP Steering Committee.   
• 
Quarterly Steering Committee reports: a report detailing recruitment, baseline patient characteristics, 
data quality, incidence of missing data and adherence to study protocol will be provided quarterly to the 
eCPAP Steering Committee.    
• 
Data and Safety Monitoring Board reports: for every meeting of the DSMB, a report is prepared which 
includes adverse events, patient recruitment, treatment data, baseline patient characteristics, 
performance information with respect to data quality, timeliness of data submission and protocol 
adherence (in addition to safety and efficacy data). 
9. Privacy, Confidentiality, and Data Security 
 
     Upon enrollment, subjects will be assigned a code that will be used instead of their name, medical record 
number or other personally identifying information. Electronic files for data analysis will contain only the subject 
code. Codes will not contain any part of the 18 HIPAA identifiers (initials, DOB, MRN). The key associating the 
codes and the subjects personally identifying information will be restricted to the PI and study staff. Data will be 
entered into the REDCap system by site study personnel. 
 
     Standard practices will be followed to maintain the confidentiality and security of data collected in this study.  
Study staff will be trained with regard to these procedures. Paper files will be stored in locked filing cabinets in 
restricted access locations. Electronic data will be stored in a web-accessible encrypted REDCap database 
housed on an OHSU secure server. Access to data/specimens is restricted to study personnel.      
 
     a. Provisions to Monitor the Data to Ensure the Safety of Subjects 
 
A locally appointed NIH approved DSMB will monitor the study. See attached DSMB charter  
 
     b. Risks and Benefits 
 
     b1. Risks to Subjects 
See section i4 above. 
 
     b2. Potential Benefits to Subjects 
 
     The subject may or may not benefit by taking part in this study. There is no guarantee that the infant will 
receive direct benefit from his/her participation in this study. The benefits of being in this study may be a 
chance that the additional 2 weeks of CPAP may improve lung function and possibly lower respiratory 
morbidity and/or improve neurodevelopmental outcomes.  Participation may provide information that may 
benefit other premature babies by helping to determine what length of CPAP provides maximum benefits. Still, 
the infant may get no direct benefit from this study.  
 

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eCPAP Protocol, Version 2 
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10. Study Administration
     a. Funding and Participating Center
     The study is funded by the National Institute of Health’s Heart, Lung and Blood Institute. The study is being 
conducted at one clinical center at Oregon Health & Science University in Portland, Oregon. 
     b. Committees 
     The Principal Investigators (Drs. McEvoy and Tepper) of the study have agreed to abide by the study 
protocol. Execution and oversight of this trial will occur through a clinical coordinating team and a data 
coordinating team as detailed in Figure 13 along with a clear understanding of roles and responsibilities. The 
clinical team will be led by Dr. Cindy McEvoy (Co-PI of the proposed study) and the data team will be led by 
Dr. Cindy Morris. Kristin Milner is an experienced project manager who has worked with Dr. McEvoy for over 8 
years and will be a crucial part of this administrative structure as the clinical project manager. Annette Vu will 
serve as the data team project manager. 
 
b1. eCPAP Steering Committee
     This committee will provide oversight for protocol development and implementation as well as 
troubleshooting any issues that arise that may impact the overall success of the trial. The committee will meet 
quarterly in person and/or via teleconference. The Steering Committee will also act as the Publication 
Committee to review and approve all abstracts, presentations, and manuscripts prior to submission or 
presentation. 
Dr. Cindy McEvoy and Dr. Cynthia Morris will lead the eCPAP Steering Committee which will include: 
• 
Clinical management /center leader/PI, Dr. Cindy McEvoy 
• 
Data management /center leader/PI, Dr. Cynthia Morris 
• 
Dr. Robert Tepper, Co-PI, expert on measurements of infant alveolar volume and lung diffusing 
capacity 
• 
Data statistician, James Slaven, MS (Indiana) 
• 
Dr. Kelvin MacDonald, Co-Investigator 
• 
NIH representative 
eCPAP Steering 
Committee
eCPAP Executive 
Committee
DSMB
eCPAP Clinical Management Team 
Dr. Cindy McEvoy 
Dr. Robert Tepper
Kristin Milner, BS, CCRP  
eCPAP Data Management Team 
Dr. Cynthia Morris
Dr. Yi Zhao
Annette Vu, MPH
eCPAP Clinical Operations Committee
Figure 13. eCPAP Study Team 

 
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eCPAP Protocol, Version 2 
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     b2. eCPAP Executive Committee 
 
     Drs. McEvoy and Morris as well as the corresponding project managers Kristin Milner and Annette Vu will 
meet every two weeks as the executive committee to review progress in study start up during the R61 phase, 
and recruiting and retention during the R33 phase. Their role is to provide day-to-day management of the trial 
and to resolve operations issues as they arise. This group will review the study tracking as well as reports of 
recruiting, retention, data safety and data quality at every visit. In the event there is a disagreement or dispute 
between the clinical management team and the data management team, it will be resolve in conjunction with 
the NIH representative 
 
     b3. Data and Safety Monitoring Board 
     The DSMB will be approved by the NHLBI and will be a group of individuals not affiliated with any of the 
institutions.  Before the trial can begin, the protocol must be approved by the committee. During the conduct of 
the study, the committee is charged with monitoring the emerging results for efficacy and safety, in addition to 
center performance and protocol adherence. Recommendations by the committee can include protocol 
modification, early termination for efficacy, or for unexpected safety problems. Recommendations are made to 
the NHLBI and disseminated to the Steering Committee.  
     c. Subject Stipend or Payments 
 
     The infant’s legal guardian will receive the value of the following as a gift card, placed on a ClinCard, or as 
cash:  $10 per monthly questionnaire completed, $200 for time and travel reimbursements for the pulmonary 
function test at about 6 months of age and $100 for time and travel to the neurodevelopmental testing.  
 
     d. Study Timeline 
  
     We plan on a 6-month start-up period followed by patient randomization over 36 months (month 7 through 
month 42). This requires recruitment of 3 infants per month. Using an average randomization GA of 32 weeks, 
we project infants will have their 6 month PFT done during months 14 to 49 and 12 month respiratory 
questionnaires done during months 20 and 54, allowing 6 months to complete final data analysis, 
dissemination, publication, and resource sharing. 
 
 
     e. Training and Certification 
 
     Prior to the beginning of the study, research coordinators, respiratory therapists, and study staff will 
undergo training on all aspects of data collection and procedures at either OHSU in Portland, Oregon or 
Indiana University in Indianapolis.  All personnel must be trained in all applicable study procedures before 
patient recruitment can begin. 
 
 
Figure 14. Study Timeline 

 
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     f. Recruitment and Data Collection Period 
 
     As in the pilot CPAP study16, infants will be recruited in the 44 bed NICU at OHSU where Dr. McEvoy is the 
Director of Neonatal Research. She has been performing infant PFTs here for 15 years as part of research 
studies93;101;102, so the NICU is familiar with this testing. The number of patients admitted to the OHSU NICU 
meeting the gestational age inclusion criteria (>24 to ≤ 32 weeks gestation) is stable (Table 4) and enables the 
desired sample size to be randomized over 36 months. This will allow trial completion, including patient follow-
up to finish within the 5 year grant period.  
 
Recruitment will also occur at PHSW where Dr. McEvoy recruited patients for the VCSIP study. It is a 20 bed 
NICU that admits approximately 50 infants >24 to ≤ 32 weeks per year. 
     g. Final Analysis 
 
     After a two month period for completion of data entry for the primary outcome of the trial, the data set will be 
locked and available for analysis. Approximately six months will be required to complete the final report to the 
eCPAP Steering Committee and to prepare the study’s primary paper for publication. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Table 4.                  Year 
2014 
2015 
2016 2017 
2018  Number needed per year  
Infants 24 to ≤32 wks admitted at OHSU/ 
year  
113 
118 
128 
94 
120 
36 

 
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Appendix A 
 
Data Collection Schedule 
 
Study 
entry 
Beginning 
of 
treatment 
period 
After 2 week 
treatment 
period 
6 months  of 
age 
6-12 
months of 
age 
Discharge 
through 
12mo of age 
Consent and medical 
history 
X 
 
 
 
 
 
Breathing test* 
 
X 
X 
X 
 
 
Blood sample** 
 
 
X 
 
 
 
Monthly breathing 
questionnaire by 
phone 
 
 
 
 
 
X 
Neurodevelopmental 
testing 
 
 
 
 
X 
 
*A syrup to make child sleepy will be given only at the 6 month breathing test 
** Collected when a sample for clinical care is being done