Clinical trial NCT02544984
Conditions: Chronic Lung Disease
Keywords: prophylactic macrolides RSV chronic lung disease respiratory infections
Sponsor: The University of Texas Health Science Center, Houston
Start date: 2015-10-01; Completion date: 2017-03-31
Enrollment: 60 participants; Allocation: RANDOMIZED; Intervention model: PARALLEL

Brief summary:
The purpose of this study is to determine if the prophylactic use of azithromycin will reduce the total number of days when unscheduled treatment is given outside of the home in a clinic, urgent care, emergency room or hospital setting between the respiratory illness season months (October 1-March 31) and subsequent 2 month follow-up (April and May)

Detailed description:
STUDY DESIGN AND METHODS

Overview:

This pilot randomized control trial (RCT) will enroll 92 children from 6 months to 6 years of age that have chronic lung disease (CLD) such as bronchopulmonary dysplasia during two pulmonary illness seasons. If the minimum number of patients (n=92) in Season 1 is achieved, we will then perform an interim analysis of the data. However, if recruitment does not reach 92patients during the first season we will continue to recruit in Season 2 (2016-2017). These children will all be patients who receive their primary medical care from either the High Risk Children Clinic or the similar High Risk Infant Clinic that cares for premature infants from discharge until age 2. Clinic records will be screened to determine eligibility. Patients that have parental consent will be given a baseline EKG, a nasal aspirate, an oscillometer reading (over 2 years of age only), and a six month supply of either the medication or the placebo at an initial study or office visit. The medication/placebo will be taken once a day for three days a week: Monday, Wednesday and Friday. The azithromycin medication will be dosed at 5 mg/kg/day. Adjustments in dosage amount will not be made during the 6 month trial. Any child that is eligible to receive Synagis will be given this every 28-30 days in clinic. Patients will be followed on a monthly basis and closely monitored for adverse reactions; this will occur by phone, in clinic during their regularly scheduled appointments, and/or during any necessary illness visits. Any children with adverse reactions will discontinue the medication, but will continue to be followed clinically. At any clinic visit in which a child presents with respiratory infections, including pneumonia, upper respiratory illness, bronchiolitis, etc., he/she will have an additional nasal aspirate and an oscillometer reading performed. At the completion of the six month treatment phase, each child will have a final EKG, final nasal aspirate and an oscillometer reading performed. Data will continue to be collected for the following twelve months, to monitor for respiratory illnesses.

Study Procedures:

Patients will be randomized by the REDCap statistical program in a double blind manner. Parents will then be provided with an entire 6 months of medication or placebo. Half of the patients will receive azithromycin at a dose of 5 mg/kg to be given once a day on Monday, Wednesday, and Friday. The dosage will not be adjusted during the trial period. The other half, the control group, will be provided with a placebo medication of similar taste, color, texture, and consistency, also to be taken once a day on Monday, Wednesday, and Friday. Both the study medication and the placebo will be dispensed from Corner Compounding Pharmacy, mixed with a fish-oil base to ensure a shelf life of more than six months, and flavored with citrus to improve palatability. Parents will be contacted monthly, either in clinic or by phone, to monitor for any adverse reactions, including rash, nausea, vomiting, diarrhea, or abdominal cramping. If a significant adverse reaction occurs, the medication will be discontinued. If an allergic reaction (such as rash or shortness of breath) is noted, the blind will be broken by Claudia Pedroza, the statistician, who is not involved with the project. This un-blinding will be done to note if it is an allergy to the medication.

After the initial appointment, at any face to face encounter (unscheduled sick visit or hospital admission/ER admission Monday through Friday) in which the patient presents with respiratory symptoms, the patient will be evaluated by the research coordinator or one of the co-investigators. Specifically, if a patient presents with the following symptoms: cough, wheeze, tachypnea, rhinorrhea, increased respiratory secretions, hypoxemia, and/or an increased oxygen requirement, an additional nasal aspirate sample and an oscillometry reading (for patients over 2 years of age) will be performed. These samples will also be stored in the office of Dr. Piedra. At the conclusion of the six month treatment phase, a final nasal aspirate sample, an oscillometer reading and EKG will be performed while the patients are in clinic for an office visit. There will be no expected study visits and no compensation will be provided for parents or patients.

Research Aims

Primary Aim: To determine if the prophylactic use of azithromycin will reduce the total number of days when one or more unscheduled medical encounters occur in a clinic, urgent care, emergency room (ER) or hospital setting during the 3-6 month study period and subsequent 2 months.

Secondary Aims:

1. To determine if the prophylactic use of azithromycin will reduce the total number of days when one or more unscheduled medical encounters occur for acute respiratory illness in a clinic, urgent care, ER or hospital setting during the 3-6 month study period and subsequent 2 months.
2. To determine if the administration of azithromycin will have a similar number of adverse side effects, or adverse events leading to unscheduled face-to-face clinic, urgent care, ER or hospital visit as compared to placebo during the 3-6 month study period and subsequent 2 months.
3. To determine if the prophylactic use of azithromycin will reduce healthcare costs during the 3-6 month study period and subsequent 2 months.

Exploratory Outcomes:

1. To determine if the prophylactic use of azithromycin will reduce the level of pro-inflammatory cytokines and additional biomarkers of disease including Lactate Dehydrogenase and Myeloid Peroxidase, by 10%.
2. To determine if the prophylactic use of azithromycin during the 3-6 months intervention will reduce the level of airway resistance of patients with CLD during respiratory illness from their baseline when compared to the placebo group as measured by an Airwave Oscillometry System
3. To determine if prophylactic use of azithromycin will reduce the total number of unscheduled face-to-face provider visits for respiratory related illness in a clinic, urgent care, emergency room or hospital setting in the 12 months after the intervention phase.

Data Analysis Plan:

Patients will be randomized to one of two different branches by the REDCap data base randomization program. This will be a double-blind placebo controlled study. Standard frequentist and Bayesian analyses will be performed using an intent-to-treat approach. Total hospital days, total ER visits (counting one day for each ER visit), and unscheduled clinic visits (counting one day for each visit) will be analyzed and related to treatment (Azithromycin vs Placebo), with logistic regression models and the treatment group as a covariate and random intercept to account for within patient correlation (due to multiple ED visits). To assess the probability of benefit, we will use Bayesian hierarchical models with interaction terms between treatment groups (Azithromycin vs Placebo) and predefined potential moderators. The groups will be stratified by synagis use and if they have a tracheostomy.

Sample size and power:

Based on data from our HRCC, we expect the control group to have 1.6 encounters per child-year (SD=1.66). Assuming a two-sided alpha = 0.05, a sample size of 92 (46/group) will have 80% power to detect a difference of 1 in the encounter rate between placebo and azithromycin groups (i.e., 1.6 vs 0.6 in encounter rate or 38% reduction). Power will be more limited for secondary outcomes but Bayesian analyses will provide an estimate of the probability of benefit in these outcomes.

Arms: arm 1: The control group will be provided with a placebo medication of similar taste, color, texture, and consistency as the study medication, and will be dispensed once a day on Monday, Wednesday, and Friday. arm 2: Patients will receive azithromycin at a dose of 5 mg/kg to be given once a day on Monday, Wednesday, and Friday. The dosage will be not be adjusted if a new weight is obtained during the trial period.
Interventions: intervention 1: Azithromycin intervention 2: placebo
Intervention descriptions: intervention 1: Patients will receive azithromycin at a dose of 5 mg/kg to be given once a day on Monday, Wednesday, and Friday. The dosage will not be adjusted if a new weight is obtained during the trial period. intervention 2: Patients will receive placebo at a dose of 5 mg/kg to be given once a day on Monday, Wednesday, and Friday. The dosage will not be adjusted if a new weight is obtained during the trial period.

Protocol documents:
Prot_SAP_000.pdf: 
IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
 
 
 
 
 
 
 
 
The anti-inflammatory effect of prophylactic macrolides on children with chronic lung 
disease: a double blinded RCT 
 
NCT02544984 
 
Version date: 10/09/2015 
 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
PROTOCOL 
 
 
 
The anti-inflammatory effect of prophylactic macrolides on children with 
chronic lung disease: a double blinded RCT 
 
 
 
 
Principal Investigator: 
 
 
Ricardo A. Mosquera, MD 
UT Health Science Center 
 
 
 
Co-Investigators: 
 
 
Cheryl Samuels, RN PNP                                                UT Health Science Center 
Susan Wootton, MD                                                         UT Health Science Center 
Cindy Jon, MD                                                                    UT Health Science Center 
Claudia Pedroza, PhD                                                     UT Health Science Center 
Elenir Avritscher, MD, PhD, MBA                                UT Health Science Center 
Aravind Yadav, MD                                                          UT Health Science Center 
Tomika Harris, PNP                                                         UT Health Science Center 
 
Research Nurse/Coordinators: 
 
Ana Maria Gomez-Rubio 
UT Health Science Center 
 
 
Contact Person(s): 
Ricardo A. Mosquera, MD 
Division of Pulmonology 
Department of Pediatrics, University of Texas 
 
6410 Fannin St 
 
Houston, TX 77030 
 
 (office) 
 
 (pager) 
 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
Research Aims 
Primary Aims: 
 
1. To determine if prophylactic use of azithromycin will reduce the total number of 
unscheduled face-to-face physician visit for respiratory related illness in a clinic, urgent 
care, emergency room or hospital setting during the 3-6 month study period, and 
subsequent 2 months follow-up. 
Secondary Aim: 
 
1. To determine if the administration of azithromycin will have similar number of adverse 
side effects, or adverse events leading to unscheduled clinic, urgent care, emergency room 
or hospital visit as compared to placebo during the 3-6 month study period and the 
following 2 months. 
2. To determine if the use of azithromycin will reduce healthcare cost associated with 
 
respiratory illness during the 3-6 month study period and 2 month follow-up period. 
 
 
Exploratory Outcomes: 
 
1. Using a standardize nasal wash procedure, respiratory samples will be collected at 
enrollment, at end of study, and during acute respiratory illness requiring face-to-face 
provider interaction. Children with a tracheostomy will have both a nasal wash sample and 
a tracheal aspirate sample collected. The respiratory samples will be stabilized with a 
universal transport media, processed and stored at -80 C for future testing. Testing will be 
performed for cytokines/chemokines; other biomarkers of disease such as LDH, MPO, and 
caspase; viral and bacterial respiratory pathogens; and microbiome. 
 
2. To determine if prophylactic use of azithromycin will reduce level of airway resistance as 
measured by an Airwave Oscillometry System in subjects above 2 years of age, at the time 
of respiratory illnesses, during the 3-6 months intervention. 
 
 
3. To determine if prophylactic use of azithromycin will reduce the total number of 
unscheduled face-to-face physician visit for respiratory related illness in a clinic, urgent 
care, emergency room or hospital setting during the following 12 months after the 
intervention. 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
 
 
 
 
Background 
 
 
 
For the past 3 years, the High Risk Children’s Clinic at UTP (HRCC) has been 
providing a medical home for medically complex children. We have demonstrated 
major benefits (e.g., 50% fewer serious illnesses & 55% fewer hospital days) and 
savings (~$10K/child/year) from the comprehensive care provided in our 
enhanced medical home to high-risk chronically ill children--including patients with 
chronic lung disease (CLD). These benefits have not been previously shown for 
medical homes for patients of any kind or age.1,2 These benefits result primarily 
from 24/7 access by phone to dedicated and experienced caregivers directed by a 
pediatric pulmonologist. The clinic offers same day appointments and provides 
comprehensive coordination of care for this population. We now aim to further cut 
morbidity rates by developing specific outpatient interventions to augment 
comprehensive care for each major disorder that we treat. 
 
 
A significant portion (44%) of our patients in the High Risk Children clinic are 
chronically ill children who have some form of CLD including patients with 
bronchopulmonary dysplasia (BPD). Chronic lung disease, as defined by the ATS 
statement from 2002, is “a heterogeneous group of respiratory diseases of infancy 
that usually evolves from an acute respiratory disorder experienced by a newborn 
infant.” Specifically, infants with bronchopulmonary dysplasia, defined as the need 
for supplemental oxygen therapy in children past 28 days born under 32 weeks 
gestation.3    These infants often incur long-term pulmonary function abnormalities, 
including oxygen dependency after discharge, recurrent respiratory infections, and 
other reactive airway diseases. From our data, we have learned that many of the 
hospital admissions in our group of patients were related to respiratory infections 
(37%) during the winter season. Despite vaccination rates of nearly 100%, 
administration of Palivizumab to all of those eligible patients and access to our 
comprehensive care clinic, viral respiratory illnesses continue to cause considerable 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
morbidity and high healthcare costs. Innovative new prophylactic treatments are 
needed. 
 
 
Macrolides have received considerable attention for their anti-inflammatory and 
immunomodulatory actions beyond the antibacterial effect. Such properties may ensure 
some efficacy against a wide spectrum of respiratory viral infections.4 Recent studies, 
including a study performed in our lab with elderly BALB/c mice infected with RSV, 
have shown that the high mortality rate of respiratory virus infections is a result of a 
neutrophilic overactive inflammatory response.5 A recently published study examined 
the inflammatory response in hospitalized infants with RSV and evaluated the 
predictive value of cytokines in nasopharyngeal aspirate in comparison to disease 
severity and found an increase in Th1 and Th2 cytokines.6 Respiratory viral infections 
are characterized by the appearance of cytokine storms which are an extreme 
production and secretion of numerous pro-inflammatory cytokines. Severity of infection 
is closely related to virus-induced cytokine dysregulation, which is responsible for the 
development of fatal clinical symptoms, such as massive pulmonary edema, acute 
bronchopneumonia, alveolar hemorrhage, and acute respiratory distress syndrome.4 
Macrolides down-regulate the inflammatory cascade, attenuate excessive cytokine 
production in viral infections, and may reduce virus-related exacerbation. Clinical trials 
have demonstrated controversial results in the effects of macrolides in respiratory viral 
infections.7  To date, studies have only evaluated macrolide use as a treatment, not as a 
prophylactic therapy. Long-term therapy with the macrolide antibiotic erythromycin 
was shown to alter the clinical course of diffuse pan bronchiolitis in the late 1980s.8 
Since that time, macrolides have been found to have a 
large number of anti-inflammatory properties in addition to being antimicrobials. These 
observations provided the rationale for many studies performed over the last decade to 
assess the usefulness of macrolides in other inflammatory airways diseases, such as 
cystic fibrosis, asthma, COPD, and bronchiolitis obliterans syndrome.9  One RCT looked at 
the daily use of macrolides for up to six weeks to prevent bronchopulmonary dysplasia 
in premature infants in a NICU setting and found the neonates to have better outcomes 
without an increase in adverse effects.10 However, chronic use of macrolides 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
has not been studied to prevent respiratory infection complications in patients with 
 
CLD of infancy. We will test the hypothesis that prophylactic macrolides are effective in 
reducing the severity of respiratory viral illness and that this is associated with the 
prevention of the full activation of an inflammatory cascade. 
 
 
 
 
Study Design and Methods 
 
 
 
 
 
Overview: 
 
This pilot randomized control trial (RCT) will enroll 92 children from 6 months to 
 
<6 years of age that have chronic lung disease (CLD) such as bronchopulmonary 
dysplasia during two seasons of respiratory illnesses defined as 1st October to 31st 
March of each year (2015-2016 Season and 2016-2017 Season). If the minimum 
number of patients (n=92) in Season 1 is achieved, we will then perform an interim 
analysis of the data. These children will all be patients who receive their primary 
medical care from either the High Risk Children Clinic or the similar High Risk Infant 
Clinic that cares for premature infants from discharge until age 2. Clinic records will 
be screened to determine eligibility. Patients that have parental consent will be 
given a baseline EKG, a nasal aspirate, an oscillometer reading (over 2 years of age 
 
only), and a three to six month supply of either the medication or the placebo at an 
initial study visit. Both medications will be taken once a day for three days a week: 
Monday, Wednesday and Friday. The azithromycin medication will be dosed at 5 
mg/kg/day. Adjustments in dosage amount will be made with any new weights 
obtained during the trial period. Any child that is eligible to receive Palivizumab 
will be given this every 28-30 days in clinic as per usual care. Patients will be 
followed on a monthly basis and closely monitored for adverse reactions; this will 
occur by phone, in clinic during their regularly scheduled appointments, and/or 
during any necessary illness visits. Any children with adverse reactions will 
discontinue the medication, but will continue to be followed clinically. At any clinic 
visit in which a child presents with respiratory infections, including pneumonia, 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
upper respiratory illness, bronchiolitis, etc., he/she will have an additional nasal 
aspirate and/or tracheal aspirate if applicable and an oscillometer reading (only for 
children >2 years) performed. At the completion of the 3-6 months treatment phase, 
each child will have a final nasal aspirate and/or tracheal aspirate if applicable, and 
an oscillometer reading performed. Data will continue to be collected for the 
following 2 months (April 1st to May 31st), to monitor for respiratory illnesses and 
possible side effects. 
 
 
Study Population: 
 
High risk children between the chronological age of 6 months and 6 years who attend 
either the High Risk Children’s Clinic or the High Risk Infant Clinic at UTP will be 
screened by the research nurse, and additional help will be provided by the clinic 
providers. The high risk infant clinic follows premature infants born before 32 weeks 
gestation for their first 2 years of life; the high risk pediatric clinic follows medically 
complex children who have had at least 3 ED visits, 2 hospitalizations, and/or 1 PICU 
visit within the last year for a chronic health condition. 
 
 
Inclusion/Exclusion Criteria: 
 
Children who belong to either the High Risk Infant Clinic or the High Risk Children’s 
Clinic and that have been diagnosed with a chronic lung disease as defined by ATS by a 
medical provider will be included.3 Chronic lung disease (CLD) is the final common 
pathway of a heterogeneous group of pulmonary disorders that start in the neonatal 
period. Often the inciting factor is bronchopulmonary dysplasia (BPD), a chronic 
condition that can develop after premature birth and respiratory distress syndrome 
due to surfactant deficiency. Myriad other conditions can also cause airway and 
parenchymal inflammation that leads to chronic airflow obstruction, increased work of 
breathing, and airway hyper reactivity. Usually the inciting factors are not only the 
underlying disorders, but also the effects of the supportive treatment, including 
mechanical ventilation, barotrauma, and oxygen toxicity. These aggressive 
interventions for serious neonatal and infant lung diseases are often responsible for 
much of the chronic pulmonary abnormalities that follow. 3Exclusion criteria include 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
children with cystic fibrosis or bronchiectasis,11 because the prophylactic use of 
macrolides has already demonstrated value and become usual care for these patients. 
Children with cardiac arrhythmias will be excluded, due to the potential increase in 
cardiovascular death that has been shown in the adult population12. Patients with 
cyanotic heart disease would be excluded. Children with colitis will also be excluded 
due to the potential effects to the gastrointestinal flora.  In addition, any child with a 
known macrolide allergy or child who is taking any medication that has a known 
interaction with macrolides, and any child with kidney or liver failure will also be 
excluded. Patients with short-bowel syndrome would also be excluded due to 
malabsorption of medication. 
 
 
Once patients are screened for eligibility, families will be approached, either over the 
phone or at a regularly scheduled office visit.  Informed written consent will be 
obtained in the clinic from the parent or legal guardian of each eligible child by any of 
the co-investigators or the research nurse at the time of enrollment. Either the research 
nurse or another clinical member of the HRCC team will then collect a nasal aspirate 
sample at the first study visit (description in Laboratory section). The nasal aspirate 
will be stored and studied after the conclusion of the treatment phase for its levels of 
 
myleoperoxydase, cytokines, respiratory virology, and micro biome. In addition, all 
patients 2 and older will have a spirometry reading performed using a TremoFlo airway 
oscillometry system (iOS) manufactured by Thorasys. With the iOS, the child breathes 
normally into the machine and lung function is measured by a gentle multi-frequency 
airway against the patient’s respiratory airflow.  A baseline EKG will be obtained on all 
patients. 
 
 
Study Procedures: 
 
Patients will be randomized by the REDCap statistical program in a double blind 
manner. Parents will then be provided with a medication or placebo for the duration of 
the study. Patients would be recruited for this study on a rolling basis from October 1st 
to December 31st, and all patients completing the intervention part of the protocol on 
31st March at the end of the season. Half of the patients will receive azithromycin at a 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
dose of 5 mg/kg to be given once a day on Monday, Wednesday, and Friday. The other 
half, the control group, will be provided with a placebo medication of similar taste, 
color, texture, and consistency, also to be taken once a day on Monday, Wednesday, and 
Friday. Both the study medication and the placebo will be dispensed from Corner 
Compounding Pharmacy, mixed with a fish-oil base to ensure a shelf life of more than 
six months, and flavored with citrus to improve palatability. Parents will be contacted 
biweekly, either in clinic or by phone, to monitor for any adverse reactions, including 
rash, nausea, vomiting, diarrhea, or abdominal cramping. If a significant adverse 
reaction occurs, the medication will be discontinued. If an allergic reaction (such as 
rash or shortness of breath) is noted, the blind will be broken by Research Coordinator 
Ana Gomez-Rubio, who is not involved with the project.  This un-blinding will be done 
to note if it is an allergy to the medication. 
 
 
After the initial appointment, at any face to face encounter (unscheduled sick visit or 
hospital admission Monday through Friday) in which the patient presents with 
respiratory symptoms, the patient will be evaluated by the research nurse or one of the 
co-investigators. Specifically, if a patient presents with the following symptoms: cough, 
wheeze, tachypnea, rhinorrhea, increased respiratory secretions, hypoxemia, and/or an 
increased oxygen requirement, an additional nasal aspirate sample/and tracheal 
aspirate if applicable, and an oscillometry reading (for patients over 2 years of age) will 
be performed. These samples will also be stored in the office of Dr. Piedra. At the 
conclusion of the 3-6 months treatment phase, a final nasal aspirate/and tracheal 
aspirate if applicable sample. Oscillometer reading will also be performed for those 
above 2 years of age while the patients are in clinic for an office visit at the end of the 
study visit and during each sick clinic visit for respiratory illness. There will be no 
expected/additional study visits and no compensation will be provided for parents or 
patients. 
 
 
Risks: 
 
Potential risks include an allergic reaction or adverse reaction to the medication or 
placebo. Examples of potential side effects include nausea, vomiting, diarrhea, and skin 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
reactions. In addition, there may be an increased risk of diaper rash and/or oral thrush 
with the increased use of antibiotics. Another adverse outcome of azithromycin has 
been an increase in pyloric stenosis with both prenatal and infant exposure for the first 
4 months of life, and benefit in this age group must outweigh this potential risk. 13 
 
However, our population for this project will begin at 6 months of age. Each of these 
risks and any other unexpected outcomes will be monitored at any visits to the clinic, or 
with biweekly phone calls if no visits have been scheduled in the previous two weeks. 
The primary benefit we hope to see is a reduction in the number of respiratory illness 
related office visits. With the anti-inflammatory properties of the macrolide, we predict 
an overall reduction in the severity of respiratory illnesses during the study period. 
Additionally, we will monitor data for a 2 month period following last azithromycin 
administration. This reduction may lead to a decrease in the number of unscheduled 
office visits, emergency room visits, and hospital admissions. With less face to face 
provider encounters, there will be less opportunity for potential exposure to other 
viruses, as well as less time away from home or work for the patients’ parents and, 
potentially, decreased health care related costs. 
 
 
Study Outcomes 
 
1.   In the treatment group, we expect a 20% total reduction in face to face 
encounters (defined as unscheduled sick visits, emergency room visits, and 
hospital admissions) for respiratory symptoms with the prophylactic use of 
azithromycin, as compared to the control group. 
a.   In the treatment group, we expect to have a 20% decrease in the number 
of unscheduled sick visits due to respiratory illness during the 3-6 month 
study period with the prophylactic use of azithromycin, as compared to 
the control group . 
b.   In the treatment group, we expect to have a 20% decrease in the number 
of Emergency Room visits due to respiratory illness during the 3-6 month 
study period with the prophylactic use of azithromycin, as compared to 
the control group. 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
c. In the treatment group, we expect to have a 20% reduction in the number 
of hospital admissions due to respiratory illness during the 3-6 month 
study period with the prophylactic use of azithromycin, as compared to 
the control group. 
 
 
2.   In the treatment group, we expect a 10% total reduction (as compared to 
baseline) in the level of myeloperoxidase in the nasopharyngeal secretions 
collected during respiratory illnesses that require a face to face provider 
encounter, as compared to the control group. 
 
 
3.   In the treatment group, we expect a 10% total reduction in the level of pro- 
inflammatory cytokines in the nasopharyngeal secretions collected during 
respiratory illnesses that require a face to face provider encounter, as 
compared to the control group. 
 
 
4.   In the treatment group, we expect a 10% reduction from baseline in the level 
of airway obstruction as measured by a tremoFlo Airwave Oscillometry 
System during the 3-6 month study period, respiratory illness and at the 
conclusion of the intervention phase. 
 
 
5.   We expect no significant difference in adverse side effects between the 
treatment group receiving the macrolide and the control group receiving the 
placebo during the 4 month study. 
 
 
6.   We expect the intervention to be cost-effective from a healthcare system 
perspective. 
 
 
Laboratory Evaluation 
 
 
 
Microbiologic studies for the secondary hypotheses will be conducted under the 
direction of Dr. Pedro A. Piedra, the Clinical Director of the Respiratory Virus Diagnostic 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
Laboratory at Baylor College of Medicine in Houston, Texas. The study involves 
collecting 2-8 nasal aspirates from each patient. Nasal aspirate samples will be tested 
to measure the levels of myeloperoxidase; we will also use ELISA tests to measure the 
levels of specific pro-inflammatory cytokines, including IL-8 and IL-6, and PCR, in order 
 
to screen for several major respiratory viral pathogens. 
 
 
 
Nasal aspirate specimens will be collected at the first visit, the final visit, and during any 
episodic respiratory sick visits at clinic or at Children Memorial Hermann Hospital. 
These samples will all be frozen and stored until the completion of the trial.  For the 
collection, the child will be placed supine, 2 ml of normal saline will be instilled into one 
nostril, and then an 8 or 10 French suction catheter (depending on the size of the 
patient) will be used to remove the mucus. This procedure will be performed once on 
each nostril. After the specimen collection from both nostrils, 2 ml of normal saline will 
be suctioned through the catheter to clear the tubing and to insure that a standard 
volume of aspirate is obtained. All specimens will be placed on ice in a bio-hazard bag 
and then collected by study personnel. After collection by study personnel, the 
specimen will be immediately diluted (1:1 solution) with viral transport medium (15% 
glycerol in Iscove’s media).   The final solution (aspirated specimen mixed with viral 
transport medium) will contain a maximum of 9 mls (with a range of 3-9 mls). After 
dilution, the specimens will be immediately refrigerated in the HRCC refrigerator at 37 
degrees, until transportation to the freezers at the Department of Pathology at Baylor, 
where they will be stored at -80oC. 
 
 
Specimen Handling / Storage 
 
Nasal aspirate specimens will be transported by study personnel from the HRCC 
refrigerators to Dr. Piedra’s laboratory.  Once received, the dilution will be separated 
into 0.5 – 1 ml aliquots and snap frozen to -80º C. Samples will be thawed and 
centrifuged at 1,000g for 10 minutes at room temperature. This clarified 
nasopharyngeal lavage fluid supernatant will be used to measure the levels of 
myeloperoxidase, will undergo cytokine analysis, and will be screened for up to 5-6 
major respiratory viruses. Remaining volumes will be stored for future studies, 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
including studies for additional biomarkers, such as Flu A, Flu B or other viruses  All 
specimens will be transported to Dr. Piedra’s laboratory Monday – Thursday between 
8am and 3pm and Friday between 8am and 1pm by involved research personnel 
 
walking to the laboratory.  Specimens received Friday afternoon will be stored in clinic 
freezer until Monday morning. For during the weekend episodes, samples will be 
collected on Monday and properly marked with date and time of ER/Hospital visit and 
date and time of sample collection. Samples will be stored for one year after the 
completion of the study at Dr. Piedra’s laboratory, and will be destroyed after the one- 
year period. Neither samples nor data will be shared outside of the University of Texas. 
 
 
Labeling 
 
 
 
Specimens collected for this study must be easily distinguishable from other laboratory 
specimens stored in the Department of Pathology refrigerators and laboratory freezers. 
ID numbers will be pre-assigned and pre-printed on a label for each specimen. The date 
(mm/dd/yr) and time (hh:mm) of specimen collection must be written on the label. The 
ID number used for the case report form must match the ID number used on the 
specimen (eg, first patient 001 should be typed on the specimen labels and case report 
form). Specimens will also be labeled with date (mm/dd/yr) and time (hh:mm) of 
symptoms onset. 
 
 
Laboratory Procedures 
 
 
 
The level of myeloperoxidase will be measured using reagent tests, and several cytokines 
that are markers of inflammation will be tested using ELISA tests.  These tests are 
routinely performed in Dr. Piedra’s laboratory and details of the primers and probes to 
be used are found in a 2005 article by Beckham et al. 14 In addition, RT-PCR assays will 
be performed using TaqMan-based primers and probes to detect the presence of 5- 
6 major respiratory viruses, and the remaining volume of aspirate will be stored for 
future studies of additional biomarkers. Nasal aspirate samples will be collected for 
assay of up to 4 cytokines and chemokines. Samples will be tested according to the 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
manufacturer’s instructions.  Samples and serial dilutions of the cytokine standards will 
 
be incubated with anti-human cytokine-coated beads in a 96-well filtration plate. 
 
 
 
Data Analysis Plan 
 
 
 
Patients will be randomized to one of two different branches by the REDCap data 
base randomization program. This will be a double-blind placebo controlled study. 
Standard frequentist and Bayesian analyses will be performed using an intent-to- 
treat approach. Total hospital days, total ER visits (counting one day for each ER 
visit), and unscheduled clinic visits (counting one day for each visit) for respiratory 
symptoms will be analyzed and related to treatment (Azithromycin vs Placebo), 
with logistic regression models and the treatment group as a covariate and random 
intercept to account for within patient correlation (due to multiple ED visits). To 
assess the probability of benefit, we will use Bayesian hierarchical models with 
interaction terms between treatment groups (Azithromycin vs Placebo) and 
predefined potential moderators. The groups will be stratified by palivizumab use 
and by tracheostomy on chronic mechanical ventilation. The conservative Bayesian 
approach of Dixon and Simon allows us to shrink the subgroup estimates to the 
overall mean treatment effect.  For all Bayesian analyses, prior distributions for 
main effects will be neutral ~N(0,1000), and Uniform (0,1000) for SD parameters. 
Neutral and skeptical priors will be used for interaction terms. Point estimates of 
treatment effect and 95% credible intervals will be reported along with probability 
of treatment benefit. 
 
 
Sample size and power 
 
Based on data from our HRCC, we expect the control group to have 1.6 encounters 
per child-year (SD=1.66). Assuming a two-sided alpha = 0.05, a sample size of 90 
(45/group) will have 80% power to detect a difference of 1 in the encounter rate 
between placebo and azithromycin groups (i.e., 1.6 vs 0.6 in encounter rate or 38% 
reduction). Power will be more limited for secondary outcomes but Bayesian 
analyses will provide an estimate of the probability of benefit in these outcomes. 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
 
 
Limitations 
 
 
 
Parents may opt out of allowing their child to participate in the study due to the extent 
of the sampling. The standard of care for respiratory illnesses may include a nasal 
aspirate, but the additional sampling at the baseline and conclusion of the study is a 
deviation from standard procedure. We will, however, emphasize the ease and low- 
invasive nature of both the nasal aspirate specimen collections and the oscillometer 
readings. In addition, parents may opt out of participating if their child’s medical 
diagnosis or current health status is too emotionally overwhelming, or if they have 
concerns regarding the prophylactic administration of an antibiotic for 3-6 months. We 
will obtain respiratory samples from patients that are admitted Monday through Friday 
during business hours, but it will be difficult to obtain nasal samples from the 
Emergency Department, on weekend admissions, and afterhours. For sick episodes 
occurring weekends and afterhours we will collect samples on the next business day 
morning. Parents may also not adhere to the administering the medication/placebo 
three days a week. 
 
 
Confidentiality 
 
 
 
Every effort will be made to protect the privacy of the subjects. Unique identifiers will 
not be stored in the final database that will be used for analytic purposes. Information 
such as age, sex, and race will be collected to examine the patterns of interaction in 
different demographic subgroups. 
 
 
Specimens will be sent to Baylor College of Medicine for processing and testing. 
Specimens will be coded with study ID numbers and will not have any personal 
information. 
 
 
Upon completion of the study, electronic data will be retained by the principal 
investigator (R. Mosquera) at his University of Texas Medical School office. All data 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
forms will be kept in a locked file cabinet and electronic files will be password 
protected. Subjects will not be directly identifiable. The PI, co-investigators, and 
designated study staff will use the information for data analysis and manuscript 
preparation. 
 
 
 
 
Interpretation and Potential Importance of Findings 
 
 
 
A substantial portion of our high risk chronically ill children have some form of 
chronic lung disease, including patients with tracheostomies, bronchopulmonary 
dysplasia, and chronic respiratory failure requiring mechanical ventilation. Most of 
the hospital admissions in this group of patients were related to respiratory 
infections in children less than 6 years of age (37% during the winter season for the 
previous 3 years). Despite vaccination rates of nearly 100% and 24/7 access to our 
clinic, viral respiratory illnesses continue to cause considerable morbidity and high 
healthcare costs. Innovative new prophylactic treatments are needed. 
 
 
With this proposal, we will determine if the prophylactic use of azithromycin will 1) 
reduce the number of days when unscheduled medical treatment health care related 
encounters was given outside the home, 2) reduce the number of emergency room 
visits, hospitalizations and clinic visits due to respiratory illness, 3) reduce the level of 
myeloperoxydase and pro-inflammatory cytokines during viral illnesses requiring face 
to face physician interaction, 4) demonstrate a reduction in airway obstruction as 
measured by an oscillometer 5) have a similar safe profile compared to the placebo, and 
6) demonstrate cost-effectiveness of macrolides use. Understanding the anti- 
inflammatory effects of azithromycin when used as a prophylactic drug will provide 
important insight into the prevention of more serious sequelae of respiratory 
infections. In particular, this proposal will contribute to understanding disease in 
children ages 6 months to 6 years with chronic lung disease, a population that has a 
higher rate of hospitalizations for respiratory symptoms. 


IRB NUMBER: HSC-MS-14-0476 
IRB APPROVAL DATE: 10/09/2015 
 
 
 
 
 
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