Clinical trial NCT02620761
Conditions: Patent Ductus Arteriosus (PDA) Acute Kidney Injury (AKI)
Sponsor: Medical College of Wisconsin
Start date: 2019-02-06; Completion date: 2020-01-31
Enrollment: 1 participants; Allocation: RANDOMIZED; Intervention model: PARALLEL

Brief summary:
The investigators will conduct a prospective, blinded, randomized, placebo-controlled trial with a sample size of 20 patients in each of the two arms (fenoldopam vs placebo) based upon a difference in serum creatinine by one standard deviation. Fluid and salt intake will be held constant within clinical parameters and carefully measured. Fenoldopam will be started at 0.1 ug/kg/min. If, after 6 hrs there is no decrease in blood pressure, the dose will be increased to 0.2 ug/kg/min. This dose will be continued throughout the remainder of the study. A study of pediatric patients previously provided to the FDA showed no hypotension at a dose of 0.2 ug/kg/min. Fenoldopam will be started 12 hrs before the first dose of indomethacin and discontinued 12 hrs after the 3rd dose of indomethacin. Study samples will include both blood and urine. The primary outcome will be a reduction in renal dysfunction, as determined by creatinine and urine output over the course of treatment. Additional outcomes will include determination of known and novel metabolomic urine markers of renal dysfunction.

Detailed description:
Hypotheses

* The investigators primary hypothesis is that fenoldopam reduces renal dysfunction associated with indomethacin administration for closure of patent ductus arteriosus in preterm infants. A secondary endpoint or measured outcome will be the determination of fenoldopam pharmacokinetics in the premature population. Lastly, the investigators hypothesize that urine and serum acute kidney injury (AKI) biomarkers will be superior to contemporary neonatal AKI definitions in their ability to identify renal injury.

Specific Aims

* Evaluate the effect of fenoldopam on renal function in preterm infants administered indomethacin
* Determination of fenoldopam pharmacokinetic and pharmacodynamic profiles in preterm infants
* Define whether newly identified biomarkers of renal dysfunction are more sensitive markers of renal dysfunction following indomethacin than traditional markers including urine output and serum creatinine.

Study design

* The study will be a prospective, blinded, randomized, placebo controlled trial. Fenoldopam will be started at 0.1 ug/kg/min. If, after 6 hrs there is no decrease in blood pressure, the dose will be increased to 0.2 ug/kg/min and continued throughout the remainder of the study. The previous study in pediatric patients showed no hypotension at a dose of 0.2 ug/kg/min. Fenoldopam will be started 12 hrs before the first dose of indomethacin and discontinued 12 hrs after the 3rd dose of indomethacin.

Describe study population or sample material

* preterm infants born at less than or equal to 28 weeks gestation with patent ductus arteriosus in whom attempted medical closure with indomethacin is indicated as decided upon by the attending physician Sample size/power of primary endpoint
* Sample size is 20 patients in each of the two arms (fenoldopam vs placebo) based upon an improvement in serum creatinine by one standard deviation.

Arms: arm 1: Infants in the Placebo arm will receive 0.9% sodium chloride (0.1 ml/hr). If, after 6 hrs there is not a clinically concerning decrease in blood pressure, as determined by attending physician, the rate of infusion (in this arm the placebo) will be increased to 0.2 ml/kg/hr. This rate will be continued throughout the remainder of the study. arm 2: Infants in the experimental arm will receive fenoldopam (60 ug/ml; 0.1 ml/hr to provide 0.1ug/kg/min). If, after 6 hrs there is not a clinically concerning decrease in blood pressure, as determined by attending physician, the rate of infusion will be increased to 0.2 ml/kg/hr (0.2 ug/kg/min for infants receiving fenoldopam). This rate will be continued throughout the remainder of the study.
Interventions: intervention 1: Fenoldopam intervention 2: 0.9%NS
Intervention descriptions: intervention 1: Randomized to receive Fenoldopam or 0.9%NS intervention 2: Randomized to receive Fenoldopam or 0.9%NS

Protocol documents:
Prot_SAP_000.pdf: 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Fenoldopam to Prevent Renal Dysfunction in 
Indomethacin Treated Preterm Infants 
 
 
Principal Investigator Dr. Jeffrey Segar 
IND Number: 127782 
ClinicalTrials.gov number: NCT02620761 
Version Date 11-26-18 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


Specific Aims 
Failure of the ductus arteriosus to close after birth results in significant morbidity in premature infants.  As 
many as 60% of newborns delivered prior to 28 weeks gestation require therapy to close their patent ductus 
arteriosus (PDA).  Administration of non-steroidal antiinflammatory agents, such as indomethacin, is the 
preferred initial therapy.  However, impairment of renal function commonly occurs as a result of indomethacin 
and may result in 1) fluid retention that may inhibit ductal closure; 2) limitation of nutritional fluid intake; 3) 
interruption of the three-dose course of treatment with resultant treatment failure; and 4) increased risk for long 
term renal injury. Alleviating the severity of renal impairment associated with indomethacin would significantly 
reduce neonatal morbidity.  To date, no effective approach to attenuate the risk for renal impairment has been 
identified.  Fenoldopam is a selective dopamine type 1 receptor agonist with the potential to dilate the renal 
vasculature, as it does not have the effect on vasoconstricting adrenergic receptors seen with dopamine. In 
critically ill adults and postoperative pediatric cardiac surgical patients, fenoldopam preserves renal function 
compared to placebo or dopamine. Studies in neonates are limited and suffer from retrospective design, small 
sample size and heterogeneous study populations. In late gestation fetal sheep, an animal model with a pattern 
of renal development stage similar to humans, we demonstrated fenoldopam significantly increased glomerular 
filtration, urine flow and sodium excretion.  Demonstration of similar effects in the preterm infant would suggest 
fenoldopam as an important therapeutic intervention in this population. 
Our primary hypothesis is that administration of fenoldopam, in doses previously demonstrated in preterm 
infants to not have significant systemic hemodynamic effects, will preserve renal function in preterm infants 
treated with indomethacin for pharmacologic closure of PDA.  A secondary objective will be to determine 
fenoldopam pharmacokinetics in the premature population, which we hypothesize will differ from those 
published in older pediatric populations.  Lastly, we hypothesize that urine and serum acute kidney injury (AKI) 
biomarkers will be superior to contemporary neonatal AKI definitions in their ability to identify renal injury.  To 
address these hypotheses, we propose to undertake a randomized, blinded, placebo-controlled trial to pursue 
the following aims: 
Specific Aim 1: Evaluate the effect of fenoldopam on renal function in preterm infants administered 
indomethacin.  Specifically, we will study the effects of fenoldopam on indomethacin induced renal 
dysfunction using classic markers of acute kidney injury according to the neonatal specific AKI definition 
modified from the 2012 Kidney Disease / Improving Global Outcomes (KDIGO) criteria.  
Specific Aim 2: Determination of fenoldopam pharmacokinetic (PK) and pharmacodynamic (PD) 
profiles in preterm infants. We propose to use a novel population based approach, in which the PK profile of 
fenoldopam can be determined with a limited number of serum samples from each infant. The fenoldopam PK 
profile will be used as the driving force to further build exposure-response models (i.e. PK/PD modeling), which 
will be used to examine the relationships between fenoldopam concentrations and drug effects, including 
systemic hemodynamics, urine output and markers of AKI. Efficacy of fenoldopam in promoting closure of the 
ductus arteriosus will be assessed. 
Specific Aim 3: Define whether newly identified biomarkers of acute kidney injury are more sensitive 
markers of renal dysfunction following indomethacin than traditional markers such as urine output and 
serum creatinine. We will take advantage of commercially available sensitive and precise biomarker platforms 
(Meso Scale Discovery) for quantitative determination of urine and serum levels of 7 biomarker analytes. 
Additionally, we will define the molecular signature differences in the urine metabolome in infants before and 
after indomethacin therapy, and we will identify the effects of fenoldopam on the metabolic profile. 
Collectively, these studies will 1) identify a medication that preserves renal function in the preterm infant, likely 
having an immediate impact on reducing short term morbidities associated with prematurity; 2) develop a 
PK/PD model of fenoldopam in this population to determine the most effective dosage regimen; and 3) define 
novel and early metabolomic markers of AKI and pathways of injury and repair.  Avoidance of acute kidney 
injury in neonates will positively impact clinical care in this population and lessen the burden of chronic kidney 
disease later in life. 
 
 
 


B. Significance 
Patent ductus arteriosus (PDA) affects up to 60% of infants less than 28 weeks gestation.5 While the 
indications for pharmacological closure are debated, avoiding the need for surgical closure is optimal.6 The 
effectiveness of indomethacin, a non-steroidal anti-inflammatory agent (NSAID) and the most commonly used 
medication for PDA closure, is variably with 30% or more of PDAs failing to close with initial therapy. 
Furthermore, greater than 25% reopen after initial therapy, necessitating multiple courses of therapy.7-9 There 
is concern regarding the use of indomethacin given its toxicity to the neonatal kidney.10 In preterm infants 24 to 
32 weeks gestation, a single course of indomethacin results in oliguria in up to 30% of patients.9, 11 Even in 
infants without frank oliguria, there are often significant decreases in urine output and increases in serum 
creatinine. To avoid fluid overload, the volume of parenteral nutritional provided is often reduced before or 
during indomethacin therapy, resulting in a reduction in energy, protein and carbohydrate intake. 
 
Treatment of PDA with indomethacin significantly reduces renal blood flow and glomerular filtration through 
inhibition of cyclo-oxygenase and decreased production of prostaglandin E, an important renal arterial 
vasodilator. To date, efforts to maintain renal blood flow following administration of indomethacin have failed. A 
meta-analysis reviewing the effects of dopamine in indomethacin-treated infants found no evidence to support 
to use of dopamine to prevent renal dysfunction.12 The diuretic furosemide is known to increase renal 
prostaglandin production and has also been investigated in indomethacin-treated infants. Surprisingly, use of 
furosemide with indomethacin increased the incidence of acute kidney injury (AKI).13 
 
Causes of neonatal AKI are multifactorial and include nephrotoxic agents such as indomethacin.14 The 
incidence of AKI among neonatal intensive care unit patients varies widely, but is estimated to range between 
8 and 24%; with rates even higher among premature infants.15, 16 Neonatal AKI has been clearly identified as 
an independent risk factor for poor outcome, with associations existing between AKI and mortality and 
morbidity, including chronic kidney injury.15, 17 Data from the Pediatric Health Information System database, 
representing 44 tertiary care children’s hospitals, suggest that NICU patients exposed to NSAIDs had a 2.3-
fold risk of AKI compared to non-exposed patients (J Misurac, personal communication). In view of the recent 
report that very low birth weight infants with AKI are at two-fold higher risk of death compared to those without 
AKI, efforts to prevent or ameliorate development of AKI in this population are urgently needed.18 
 
There is increasing concern regarding the risk of chronic kidney disease later in life for infants born 
premature.4, 19, 20 Disruption of normal nephrogenesis and a reduction in nephron number likely contribute to 
increased susceptibility to renal injury and increased risk of renal disease later in life.19, 21, 22 Recent evidence 
suggests that although nephrogenesis continues after birth in preterm infants, kidney growth is altered in terms 
of nephron number and size. In autopsy samples from human preterm infants, a large number of glomeruli 
appear abnormal with scant capillarization and enlarged Bowman spaces.23 In baboons and mice delivered 
prematurely, nephron number is significantly decreased compared to term-delivered, postconceptional age-
matched controls.24, 25 Additionally, preterm infants are at significant risk for acute kidney injury related to birth 
associated events such as maternal chorioamnionitis, episodes of hypoxemia, intravascular volume depletion, 
and exposure to nephrotoxins such as aminoglycosides and indomethacin.19, 21 While classically taught that 
acute kidney injury is reversible, recent epidemiological studies link acute kidney injury in adults and children to 
the later development of chronic kidney disease.26, 27 Although data are lacking in neonates, it is reasonable to 
assume that in the face of abnormally developed kidneys resulting from prematurity, there are deleterious 
effects of AKI on long-term kidney health. In 3-10 year old children born prematurely and diagnosed with and 
without neonatal AKI, those with AKI had significantly decreased renal volume.4 Thus, identification of novel 
markers of AKI and measures to avoid kidney injury in the preterm infant population is critical. 
 
Studies of pharmacologic agents in pediatric patients, let alone neonates, are lacking.  We propose to study 
the renal sparing effects of fenoldopam and its pharmacokinetic/pharmacodynamics profile of fenoldopam. Our 
proposal, which is carefully designed and for which FDA Investigation New Drug (IND) approval has been 
obtained, will address a troubling information gap in the care of this unique and vulnerable population. 
 
C. Innovation 
The innovation of this proposal exists at multiple levels. First, we are using a currently marketed 
pharmaceutical with a well-defined profile in adults for a new and unique population. Fenoldopam mesylate is a 
selective dopamine receptor (D1 receptor) agonist which induces vasodilation of renal, mesenteric and other 
arteries. Unlike dopamine, it has no affinity for vasoconstricting α-adrenoreceptors and thus improves blood 


flow to the renal cortex and medulla. Approved as a short-term antihypertensive agent, fenoldopam has also 
been widely promoted and studied in the adult population for prevention and treatment of AKI.28-30 A meta-
analysis of randomized placebo-controlled studies involving over 400 patients undergoing cardiac surgery 
found that fenoldopam reduced the risk of acute kidney injury by more than half (OR = 0.41; CI,0.23-0.74; 
p = 0.003).31 A multicenter randomized controlled trial to evaluate the effects of fenoldopam on the need for 
renal replacement therapy in cardiac surgery patients who had already developed AKI was stopped early for 
futility, as no effect was seen.28 Such findings emphasize the importance of starting fenoldopam before the 
development of AKI or its inciting event occurs. Second, the design of our studies allows for exact timing of the 
inciting risk for AKI—indomethacin administration—and for obtaining important study samples both before and 
after indomethacin.  We will initiate fenoldopam therapy prior to the development of AKI or indomethacin 
administration, taking a preventive rather than therapeutic approach. Third, we will use a commercially 
available, sensitive and precise biomarker platform (Meso Scale Discovery) for quantitative determination of 
urine and serum levels of 7 biomarker analytes. We will determine whether these markers, singly or in 
combination, are superior in identifying AKI compared to conventional measures. Finally, we will define the 
molecular signature differences in the urine metabolome in infants before and after indomethacin therapy, and 
we will identify the effects of fenoldopam on the metabolic profile. These studies will identify novel and early 
metabolomic markers of AKI, which we anticipate will lead to identification of pathways of injury and repair. 
 
D. Approach 
D.1 Previous Work 
Studies of fenoldopam for reducing the risk or severity of AKI in the pediatric population are limited. High-dose 
fenoldopam infusion (1 µg/kg/min) was found to significantly reduce the levels of AKI biomarkers in children 
undergoing cardiopulmonary bypass with a strong trend towards reduction in AKI (p = 0.08).32 In contrast, a 
study of low-dose fenoldopam (0.1 µg/kg/min) in newborn patients undergoing bypass found no significant 
effect on AKI incidence.33 A retrospective review of fenoldopam use in a single NICU in patients with 
established oliguria or anasarca found patients not on extracorporeal membrane oxygenation at the time of 
treatment (0.1µg/kg/min over 24 hours) had a significant increase in urine output.2 Use specifically in the 
extremely preterm population has not been described. However, in fetal sheep, commonly used to model 
human kidney development, fenoldopam significantly increased urine flow rate, glomerular filtration rate and 
sodium excretion.34 Taken together, the above findings substantiate the need for determining whether 
fenoldopam can prevent renal dysfunction in indomethacin-treated preterm infants. 
 
D.2 Study Design 
We propose to conduct a randomized, blinded, placebo-controlled trial to determine whether administration of 
fenoldopam reduces renal dysfunction associated with indomethacin administration for closure of patent ductus 
arteriosus in preterm infants. 
Patient Selection: Infants born between 23 0/7 and 27 6/7 weeks gestation (by obstetrical dating) and admitted 
to the University of Iowa Children’s Hospital NICU are routinely screened for PDA by echocardiography during 
first few weeks of the postnatal period. If a PDA is present, the decision to attempt pharmacologic closure is at 
the discretion of the attending physician. If the decision is made to use indomethacin, the infant will be deemed 
eligible to be screened for enrollment in the study, and the clinical team will notify the designated research 
nurse. Echocardiographic screening for PDA will not be performed specifically for the purpose of this study. 
Inclusion criteria include: 
1) 
Gestational age at birth 23 0/7 to 27 6/7 weeks by best obstetrical dating 
2) 
No previous exposure to indomethacin 
3) 
Clinical determination made to use indomethacin to attempt closure of PDA 
4) 
No known congenital abnormalities involving the kidneys, heart or lungs 
5) 
No preexisting renal dysfunction, defined as serum creatinine >1.0 mg/dl, or oliguria, defined as urine     
 
output <1.0 ml/kg/hour over the previous 24 hours. 
Exclusion criteria: 
1) 
Enrollment in concurrent study in which interventions may contribute confounding variables 
2) 
Infants with diagnosed or suspected renal or urinary tract abnormalities 
3) 
Infants with umbilical cord or infant blood pH below 7.0 at any time before enrollment 
4) 
Attending physician unwilling to have infant participate in study 
 


Figure. Timeline of study  
Infants will be screened for eligibility for enrollment by a designated research nurse. Before being approached, 
the parents will have been informed by the care team about the presence of a PDA and the decision to attempt 
PDA closure using indomethacin. A log will be maintained of all screened infants indicating which are eligible 
and which were enrolled. Upon receipt of consent, treatment assignments will be made electronically using the 
Research Electronic Data Capture (REDCap) treatment randomization module using a randomized block 
design to assign treatment group and stratified for gestational age (23 - 25 6/7 and 26 - 27 6/7 weeks 
gestation). A block randomization strategy will be employed to achieve balance between study arms and 
reduce the opportunity for bias and confounding. Infants from multiple gestations will be randomized 
independently. Subjects will be analyzed by intention to treat, with the group to which they were randomized, 
regardless of subsequent events. Fenoldopam will not be used outside of the study. 
 
Determination of status of ductus arteriosus: 
After an echocardiogram is obtained at the discretion of the attending neonatologist as part of routine care, a 
decision regarding whether to treat or not is made.  If the decision to treat with indomethacin is made, an 
echocardiogram will be obtained immediately prior to the first dose of indomethacin and again 24 hours after 
completion of the course of indomethacin therapy (routine in our NICU). The echocardiographer and 
cardiologist interpreting the echocardiogram will be blinded to the treatment group. 
 
Time course for study (Figure): 
Urine will be collected in 6-hour increments by extracting from special diapers (Cuddle-Buns Preemie Diapers, 
Trace Medical Equipment, Lisle, IL), measured, and frozen for later determination of urinary metabolomics as 
described below. Serum creatinine, which in 
our NICU is always measured within 12 
hours prior to initiating indomethacin, will be 
recorded. Infants will receive either placebo 
(0.9% sodium chloride, 0.2 ml/kg/hr) or 
fenoldopam (30 µg/ml; 0.2 ml/kg/hr to 
provide 0.1 µg/kg/min). If, after 6 hrs there is 
not a clinically concerning decrease in blood 
pressure as determined by attending 
physician, the rate of fenoldopam infusion 
(or placebo) will be increased to 0.2 
µg/kg/min (30 µg/ml, 0.4 ml/kg/hr). 
Increased urine output has been reported in 
neonates at median doses of 0.1-0.3 
ug/kg/min while decreased blood pressure 
has been at doses above 0.2 ug/kg/min.1, 2, 
35 Thus to avoid systemic hemodynamic changes, we will limit dose to 0.2 ug/kg/min.  This rate will be 
continued throughout the remainder of the study. Fenoldopam will be started 12 hours before the first dose of 
indomethacin and discontinued 12 hours after the third dose of indomethacin. Blood pressure will be monitored 
15, 30, 45 and 60 minutes after starting fenoldopam or increasing the dose and hourly throughout the infusion. 
Infants will receive indomethacin therapy (3 doses, 0.2, 0.25, 0.25 mg/kg iv, every 12 h, respectively), in 
accordance with our current medical practice. Provision of parenteral and/or enteral nutrition, including fluid, 
electrolyte, lipid and energy intake will be at the discretion of the attending physician, though attempts will be 
made to hold these constant during the study period. Except for administration of fenoldopam or placebo, all 
clinical care decisions, including respiratory care, will be at the discretion of the attending physician. Infants will 
be weighed daily per our NICU routine. 
 
Determination of patient demographics: Routine clinical data will be abstracted from the infants’ medical 
records by study personnel. We will use the standardized NICHD Neonatal Research Network (of which we are 
a part) Generic Database forms for data collection; this is routinely used for all <1500 g infants admitted to 
Network centers. These forms will be used as the source for maternal and infant demographic data, 
maternal/pregnancy history, including exposure to antenatal indomethacin, MgSO4, antenatal steroids, and 
clinical outcome data including potential confounding variables. We will ensure sex, gestational age, 
appropriateness of weight for gestational age, daily weight, daily fluid intake, daily energy intake, ventilator 
settings, supplemental oxygen requirements, urine output, presence of documented or suspected sepsis, age 
at time of treatment and treatment outcome are obtained during the course of the study as will the occurrence 


Table.  Neonatal AKI definition 
Stage 
SCr criteria 
Urine output (UOP criteria) 
0 
No change or rise < 0.3 mg/dl 
UOP > 1ml/kg/h 
(over previous 24 h) 
1 
↑ SCr of ≥0.3 mg/dl or     
↑ SCr to 150-199% of baseline 
UOP > 0.5 ml/kg/h and ≤ 1 ml/kg/h 
 (over previous 24 h) 
2 
↑ SCr to 200%-299% of baseline 
UOP > 0.1 ml/kg/h and ≤ 0.5 ml/kg/h 
(over previous 24 h) 
3 
↑ SCr to ≥ 300% of baseline or   
SCr ≥ 2.5 mg/dl or  
Receipt of dialysis 
UOP ≤ 0.1 ml/kg/h 
( over previous 24 h) 
Baseline SCr will be defined as the SCr value prior to indomethacin 
of any attendant medical conditions, including intracranial bleeding, change in respiratory status, necrotizing 
enterocolitis, renal failure and development of retinopathy of prematurity. 
 
Specific Aim 1: Evaluate the effect of fenoldopam on renal function in preterm infants administered 
indomethacin. 
 
Urine output will be recorded 
and expressed as ml/kg/h for the 
12 hours prior to administration 
of fenoldopam, the 12-h during 
fenoldopam administration prior 
to starting indomethacin, and for 
each subsequent 12-h period 
until fenoldopam has been 
discontinued for 12 hours 
(Figure). Serum creatinine, 
measured by our hospital central 
laboratory, will be determined immediately prior to the administration of the initial and third dose of 
indomethacin as well as 24 hours after the final dose. The stage of AKI (0 – 3) will be determined according to 
the neonatal specific AKI definition as modified from the 2012 KDIGO definition (Table).36 
 
Specific Aim II: Determination of fenoldopam pharmacokinetic and pharmacodynamic profiles in 
preterm infants. 
 
To our knowledge, only a single study has examined pharmacokinetic (PK) and pharmacodynamic (PD) 
characteristics and the side-effect profile of fenoldopam in children.35 In this study of 77 children from 3 weeks 
to 12 years of age, only doses above 0.8 µg/kg/min were associated with significantly reduced blood pressure. 
Understanding fenoldopam pharmacokinetics is critical for optimizing fenoldopam dosing regimens as well as 
establishing the relationship between the exposure of fenoldopam (i.e., pharmacokinetics) and the clinical 
outcome of renal function protection (i.e. pharmacodynamics). This study will provide important, novel 
information about this pharmacologic agent in the preterm population. 
 
Pharmacokinetic/Pharmacodynamic (PK/PD) Approach 
Because of their limited circulating blood volume and ethical restrictions placed on removal of blood from 
premature infants, it is not feasible to perform a traditional pharmacokinetic study. To evaluate the PK of 
fenoldopam, we propose to use a novel population-based approach, in which the PK profile of fenoldopam is 
determined using relatively few samples (1-3 samples) from each infant. In this approach, samples are 
collected randomly (i.e., not at predetermined time points) and subjects contribute different numbers of 
samples. This approach permits an estimation of PK parameters of fenoldopam and an explanation of 
variability using the nonlinear mixed-effects modeling approach.37 Blood samples (100-200 µL) will be obtained 
from discarded blood from other clinical lab tests or collected during this routine clinical sampling.38 We will 
target obtaining 1-2 samples per patient for time periods 0-48 hours and 2-3 samples per patient out to 60 
hours, 12 hours after stopping fenoldopam. Fenoldopam concentrations will be measured using LC/MS/MS 
based on a sensitive method reported by Wang et al.39. A nonlinear mixed mathematic model (i.e. population-
based approach) using specialized PK software NONMEM will be employed to fit the data to characterize the 
PK profile as previously performed by one of us (GA).40 In this model, the first-order conditional estimation 
method with the interaction and a user-defined subroutine (ADVAN6) will be used to estimate the typical 
population PK parameters, random inter-individual variability, and residual variability between observed and 
individually predicted plasma fenoldopam concentrations. Final PK model selection will be based on the visual 
inspection of goodness-of-fit plots, the objective function value, and the precision of parameter estimation. The 
likelihood ratio test will be used for comparing nested models. 
 
Pharmacodynamic effects of constant rate, fixed-dose fenoldopam infusion compared to placebo infusion on 
heart rate, mean blood pressure and urine output will be determined after 12, 24 36 hours of infusion and 12 
hours after completion of infusion. Comparisons of hemodynamic and urine output data before, during, and 
after fenoldopam infusion will be made for each patient by linear regression analysis. Ductal patency will be 
assessed 24 hours after completion of the course of indomethacin to allow for determination of benefit of 


fenoldopam on this outcome.  Fenoldopam PK will be used as the driving force to further build exposure-
response models (i.e. PK/PD modeling), which will be used to examine the relationships between fenoldopam 
concentrations and drug effects, including hemodynamics and urine output. The exposure-response analyses 
will be performed modeled using NONMEM software (NONMEM Project Group, UCSF, San Francisco, CA). 
 
Specific Aim III: Define whether newly identified biomarkers of renal dysfunction are more sensitive 
markers of renal dysfunction following indomethacin than traditional markers such as urine output and 
serum creatinine. 
 
We will take a two-faceted approach to the exploration of potential new biomarkers of renal dysfunction 
following indomethacin treatment in preterm infants. First, we will examine urinary levels of previously reported 
biomarkers in the preterm infant population. Askenazi et al recently reported increased urinary levels of NGAL 
and osteopontin in a population of VLBW infants (500-1500 grams) with AKI.41 A prospective study of 140 
pediatric patients (ages 1 month – 21 years) found urinary NGAL levels rose 48 hours before identification of 
AKI.42 This proposed study provides a novel opportunity to evaluate the utility of potential biomarkers in this 
population since the timing of the inciting cause of AKI (indomethacin administration) is known and we will 
have pre- and post-administration urine samples, allowing longitudinal comparisons. Second, in parallel with 
determination of pre-described urine biomarkers, we will undertake an exploratory urinary metabolomic 
approach to identify previously suggested and novel markers of AKI in this population. 
 
Urine and Serum Biomarker Identification 
Seven biomarkers, including albumin, beta 2 macroglobulin, Cystatin C, EGF, NGAL, osteopontin and 
uromodulin, will be measured in urine and serum using the Meso Scale Discovery (MSD) Sector imager 2400 
and MSD multiplex human kidney injury biomarker panel 5 assay. Urinary determinations will be made in 
samples obtained 6-12 hours after starting fenoldopam (prior to indomethacin), 6-12 hours after the second 
dose of indomethacin and 18-24 hours after the third dose of indomethacin while serum samples will be 
obtained immediately prior to the first and third doses of indomethacin, as well as 24 hours after the third dose 
of indomethacin (see figure). Analysis of these samples is as described below. Dr. Askenazi will serve as a 
consultant related to the identification of biomarkers using the MSD assay (see letter of support). 
 
Determination of urinary metabolomic markers of kidney injury 
Using high throughput technology, metabolomics examines thousands of low molecular weight metabolites that 
may change according to the pathologic state of the organism. Metabolites represent the downstream 
expression of genome, transcriptome and proteome, reflecting the phenotype at a defined time. The clinical 
utility involves biomarker discovery, defining as yet unrecognized biological therapeutic targets, linking 
metabolites to relevant standard indices and clinical outcomes.  Urinary metabolomics will be performed on 
samples obtained 6-12 hours after starting fenoldopam (prior to indomethacin), 6-12 hours after the second 
dose of indomethacin and 18-24 hours after the third dose of indomethacin while serum samples will be 
obtained immediately prior to the first and third doses of indomethacin, as well as 24 hours after the third dose 
of indomethacin. Mass spectrometry (MS) is used to identify metabolites after separation by either gas 
chromatography (GC), or high-performance liquid chromatography (HPLC) as described by our group.10 
Evaluation of the urine metabolome will identify pathways of renal cell injury and novel diagnostic and 
therapeutic targets, filling a large unmet need in this area of medicine. 
 
Biomarker data analysis 
For the two primary measures of AKI, namely SCr and urine output, univariate analysis will be performed and 
the sensitivity, specificity, and positive and negative predictive value for each individual AKI biomarker will be 
calculated, as will receiver operator curves. Logistic regression models will also be created to determine the 
ability of multiple biomarkers to predict AKI. Urine biomarker studies in premature infants must be adjusted for 
gestational age, as levels are inversely related to gestational age independent of AKI status. The longitudinal 
design of our study, which includes determination of biomarker levels prior to and following the AKI-inducing 
event (indomethacin administration), allows us to examine changes from baseline in each patient, rather than 
relying on population normative values adjusted for gestational age. 
 
Metabolomic data analysis 
The MS spectra processing will be carried out using the XCMS software.43 Peak detection, alignment and 
retention time correction are performed sequentially. Absolute value changes between the pre and post-


indomethacin administration means will be compared by fold-change analysis. Unsupervised principal 
component analysis (PCA) will performed using the prcomp software. Supervised partial least squares 
discriminates analysis (PLS-DA) will be performed to identify and characterize metabolic perturbations 
signatures induced by indomethacin. To assess the significance of class discrimination, a permutation test will 
be performed. Variable Importance in Projection (VIP) will be measured in PLS-DA, representing a weighted 
sum of squares of the PLS loadings.44 Metabolic pathway analysis will be performed using Ingenuity Pathway 
Analysis (IPA) MetPA as previously described by us.45 
 
E. Power Calculation, Data Capture and Data Analysis Plan 
In 2014 and 2015, our NICU had on average 78 infants per year admitted between 23 0/7 and 27 6/7 weeks 
gestation. 70% of these infants receive at least one course of indomethacin. Based upon historical consent 
rates in our NICU and using conservative estimates, we will be able to enroll at least 20 patients per treatment 
group over an 18 month period. Review of infants in our NICU over the past 3 years as well as data from the 
published literature revealed an increase in serum creatinine of about 2 standard deviations following 
indomethacin treatment. 42  Based on a sample of 20 patients per group with α= 0.05, the power to detect an 
increase in serum creatinine in the fenoldopam group compared to placebo by one standard deviation (0.3 
mg/dl) is 86.9%.  A REDcap database will be developed capturing demographic and prospective data (outlined 
above). The REDcap application interface allows direct importing to SAS® 9.4 software, which will be used for 
performing statistical analyses. Descriptive statistics for the study populations by treatment group will be 
generated, using means and standard deviations for the continuous measures (non-normally distributed 
variables will be normalized using a transformation) and percentages for categorical measures. For the three 
outcomes in Aim 1 (largest change in serum creatinine, largest decrease in 24h urine output, AKI stage), we 
will fit generalized linear models using the canonical link function for the distribution of each outcome. AKI 
stage will also be dichotomized by collapsing stages 1-3 together. The two predictor variables that will be used 
in every model are treatment and gestational age groups. Additional demographic predictors will be considered 
as control variables in the models. In order to determine the most appropriate collection of predictor variables 
for each outcome, we will fit all possible subsets of the control variables with each model fit having a 
corresponding corrected Akaike model information criteria (AICc).46, 47 This measure is an extension of the 
Akaike information criterion (AIC) and is more appropriate when the sample size is small.48, 49 The model with 
the smallest AICc will be deemed the most favorable in the collection and used for interpretation of its effects. 
The use of AICc is beneficial in that we can compare model fits of predictor sets as a whole, rather than 
assessing the significance of each independent variable in an attempt to specify the best model. This method 
ensures we select a statistically viable model by including important predictors that may impact the relationship 
between treatment or gestational age groups and the outcomes, while omitting spurious variables. 
 
F. Expected Outcomes, Potential Pitfalls and Alternative Approach 
We do not expect any major difficulties with our experimental approach. The investigators are well versed with 
the described methodologies. We recognize that with the projected sample size, and the published rates of AKI 
after indomethacin therapy that the absolute number of infants developing Stage 1 or higher AKI will likely be 
low. The lack of sensitivity and specificity of the current definitions of AKI have been well acknowledged.  
However, we anticipate that the incidence of AKI, as determined by significant increases in levels of urinary 
biomarkers will be much higher than that identified by SCr values and/or changes in urine output. The 
investigation of novel biomarkers will provide us with the opportunity to contribute to the discussion regarding 
new and refined definitions of neonatal AKI with important clinical implications. We further anticipate that 
fenoldopam with preserve urine output, obviating the need to reduce fluid intake, and thus nutritional intake. 
Additionally, we expect fenoldopam to increase the success rate of closure of PDA with indomethacin, as 
preservation of renal function will allow for completion of the 3-dose course of indomethacin, rather than 
interruption due to renal concerns. Development of hypotension may lead investigators/clinicians to suspect 
treatment with fenoldopam, though hemodynamic changes have not been reported at the doses used. The 
clinical team has the option to stop infusion of the investigative drug at any point. 
 
G. Recruitment Strategy 
Our team of research nurses is in constant communication with the NICU care team and will be informed when 
the decision is made to treat an infant diagnosed with a PDA with indomethacin. They will screen patients and 
present to the parents the details of the study and obtain informed consent. The extensive experience of the 
nurses results in effective communication about protocols and success in recruitment. Our team is highly 
successful in recruiting subjects for clinical trials in our NICU. Based on this extensive experience and absence 


of invasive procedures in this study, we anticipate a consent rate of approximately 70% of eligible infants and 
an enrollment period of approximately 18 months. The dropout rate is likely to be minimal given the short 
duration of the study and lack of anticipated side effects from the intervention. 
 
 
 


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