Pragmatic Factorial Trial of Hydroxychloroquine, Azithromycin, or Both for Treatment of Severe SARS-CoV-2 Infection
Phase: PHASE2
Conditions: SARS-CoV-2
Sponsor: Duke University

Pragmatic trial of hydroxychloroquine/azithromycin for COVID 
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TITLE 
Pragmatic Factorial Trial of Hydroxychloroquine, Azithromycin, or both for treatment of 
SARS-CoV-2 Infection  
 
 
 
Funding agencies: 
Duke University School of Medicine 
ClinicalTrials.gov Identifier 
NCT04335552 
 
Version Number: 1.0 
7 April 2020 
 
 

Pragmatic trial of hydroxychloroquine/azithromycin for COVID 
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Statement of Compliance 
This trial will be conducted in compliance with the protocol, International Conference on Harmonisation 
Good Clinical Practice E6 (ICH-GCP), U.S. Code of Federal Regulations 45 CFR 46 and 21 CFR, and 
applicable site-specific regulatory requirements.   
 
 

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Contents 
1. 
Protocol Synopsis .................................................................................................................................. 5 
2. 
Background Information and Scientific Rationale ................................................................................ 7 
2.1 Need for therapeutic agents for SARS-CoV-2 ..................................................................................... 7 
2.2 Scientific rationale .............................................................................................................................. 7 
2.2.1  Mechanism of action of chloroquine and hydroxychloroquine in SARS-CoV-2 .......................... 7 
2.2.2 Mechanism of action of azithromycin in SARS-CoV-2 ................................................................. 8 
2.3 Evidence for chloroquine and hydroxychloroquine ............................................................................ 8 
2.3.1 In vitro studies.............................................................................................................................. 8 
2.3.2 In vivo studies .............................................................................................................................. 8 
2.3.4 Evidence for hydroxychloroquine and azithromycin combination therapy ................................ 9 
2.3.5  How this study complements other planned work ................................................................... 10 
3. 
Descriptions of Study Drugs ................................................................................................................ 11 
3.1 
Hydroxychloroquine .................................................................................................................... 11 
3.2 Azithromycin ..................................................................................................................................... 12 
4. 
Objectives............................................................................................................................................ 14 
2.1 
Definitions ................................................................................................................................... 14 
2.2 
Primary ........................................................................................................................................ 14 
2.3 
Secondary .................................................................................................................................... 15 
5. 
Study Design........................................................................................................................................ 15 
6. 
Study Population ................................................................................................................................. 16 
7. 
Enrollment and Randomization .......................................................................................................... 18 
7.1 
Enrollment Procedures ............................................................................................................... 18 
7.2 
Randomization ............................................................................................................................ 18 
8. 
Study Procedures ................................................................................................................................ 19 
8.1 
Clinical Evaluations......................................................................... Error! Bookmark not defined. 
8.2 
Concomitant Medications ........................................................................................................... 22 
8.3 
Laboratory Evaluations ............................................................................................................... 23 
8.4 
Loss to Follow Up ........................................................................................................................ 23 
8.5 
Management of Participants Who Discontinue Study Treatment ............................................. 23 
9. 
Study Schedule .................................................................................................................................... 23 
10. 
Study Intervention/Investigational Drugs ....................................................................................... 24 

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10.1 
Study Drugs ................................................................................................................................. 24 
10.2 
Study Drug Acquisition ................................................................................................................ 25 
10.3 
Administration and Dosage of Study Drugs ................................................................................ 25 
10.4 
Criteria for Discontinuation of Study Drugs ................................................................................ 25 
11. 
Assessment of Safety ...................................................................................................................... 25 
11.1.1  Definition of Adverse Events ....................................................................................................... 25 
11.1.2  Definition of Serious Adverse Events .......................................................................................... 26 
11.1.3  Assessment of Adverse Events .................................................................................................... 26 
11.1.4  Adverse Event Reporting............................................................................................................. 27 
11.1.5  Reporting of pregnancy outcome ............................................................................................... 28 
12. 
Statistical Considerations ................................................................................................................ 29 
12.1 Study Hypotheses ........................................................................................................................... 29 
12.2  Analysis Groups .............................................................................................................................. 30 
12.3  Analysis Plan ................................................................................................................................... 30 
12.3.1.  Primary efficacy analysis ........................................................................................................ 30 
12.3.2  Secondary efficacy analyses .................................................................................................... 31 
12.3.3  Safety analyses ........................................................................................................................ 31 
12.4 Sample Size Considerations ............................................................................................................ 32 
13. 
Protection of Human Subjects ........................................................................................................ 38 
13.1  Institutional Review Board ............................................................................................................. 38 
13.2  Informed consent process ............................................................................................................. 38 
13.3  Participant confidentiality .............................................................................................................. 38 
13.4  Study discontinuation .................................................................................................................... 39 
14. 
Data Handling and Record Keeping ................................................................................................ 39 
15. 
Publications and Dissemination of Study Results ........................................................................... 39 
 
 
 

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1.  Protocol Synopsis 
Protocol Title: Pragmatic Factorial Trial of Hydroxychloroquine, Azithromycin, or both for 
treatment of SARS-CoV-2 Infection 
Treatment Indication: SARS-CoV-2 Infection requiring hospitalization 
Trial Objective: Assess the impact of hydroxychloroquine, azithromycin, or both on clinically 
relevant outcomes in a real-world population of hospitalized persons with confirmed SARS-CoV-
2 infection. 
Trial Design: Open-label, factorial randomized clinical trial with two factors and nested 
randomization.  Patients hospitalized with confirmed SARS-CoV-2 infection will be initially 
randomized to one of two arms in a 1:1 ratio: 1) Standard of care treatment or 2) Standard of 
care treatment plus 5 days of hydroxychloroquine.  Patients with no contraindication to 
azithromycin who have not received azithromycin in the seven days prior to enrollment will 
undergo a second randomization in a 1:1 ratio to either receive 1) No azithromycin or 2) 5 days 
of azithromycin.  Randomization will be stratified by hospital and by whether the patient is 
receiving mechanical ventilation at the time of randomization or not.  Participants will be 
followed for 28 days after randomization or until hospital discharge, whichever is later.   
Patient population: A total of 500 hospitalized male and female patients (≥ age 12 years or 
older), newly diagnosed with SARS-CoV-2 infection by a validated nucleic acid amplification 
assay and hospitalized due to symptoms of SARS-CoV-2 disease, who do not have 
documented allergy or intolerance to hydroxychloroquine and who have not received 
hydroxychloroquine in the 180 days prior to enrollment. 
Treatment arms: Study participants will be randomized 1:1 to receive one of the following: 
Randomization 1 Treatment 
1) 
Standard of care 
2) 
Standard of care plus hydroxychloroquine 800 mg on day 1, followed by 600 mg 
daily for a total of 5 days 
Participants who have not received azithromycin in the 7 days prior to enrollment and who do 
not have documented contraindication to azithromycin will undergo a second randomization 1:1 
for concurrent treatment: 
Randomization 2 Treatment: 
1) 
Treatment dictated by Randomization 1 alone 
2) 
Treatment dictated by Randomization 1 plus azithromycin 500 mg on day 1, 
followed by 250 mg daily for 4 additional days 
 
Criteria for evaluation: 
Primary endpoint: 
(1) WHO ordinal scale measured at 14 days after enrollment: The ordinal scale is an 
assessment of the clinical status at the first assessment of a given study day. The scale is 
as follows: 1) Death; 2) Hospitalized, on invasive mechanical ventilation or extracorporeal 
membrane oxygenation (ECMO); 3) Hospitalized, on non-invasive ventilation or high flow 
oxygen devices; 4) Hospitalized, requiring supplemental oxygen; 5) Hospitalized, not 

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requiring supplemental oxygen - requiring ongoing medical care (COVID-19 [coronavirus 
disease 2019] related or otherwise); 6) Hospitalized, not requiring supplemental oxygen - no 
longer requires ongoing medical care; 7) Not hospitalized, limitation on activities and/or 
requiring home oxygen; 8) Not hospitalized, no limitations on activities. 
Secondary endpoints: 
(1) Death during the index hospitalization 
(2) Number of days on mechanical ventilation  
(3) Proportion of patients not receiving mechanical ventilation at baseline who progress to 
requiring mechanical ventilation during the index hospitalization 
(4) WHO ordinal scale measured at 28 days after enrollment 
(5) Hospital length of stay in days for the index hospitalization 
(6) Days of fever (temperature ≥38.0) after randomization 
(7) Days on supplemental oxygen after randomization 
(8) All-cause study medication discontinuation 
(9) Drug-associated adverse events of special interest 
Study sites: 
Study participants will be recruited from Duke University Hospital, Duke Regional Hospital, Duke 
Raleigh Hospital, and the Durham VA Medical Center. 
Study duration: 
It is anticipated that three months will be required for recruitment and enrollment of study 
participants.  The estimated duration of participation for each study participant is approximately 
28 days, including treatment and follow-up. 
 
 

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2. Background Information and Scientific Rationale 
2.1 Need for therapeutic agents for SARS-CoV-2 
On December 31, 2019, China reported an outbreak of an unknown respiratory illness in Hubei 
Province, now identified as SARS-CoV-2 (1). The WHO declared a Public Health Emergency of 
International Concern on January 30, 2020, and a global pandemic on March 12, 2020. As of 
April 2, 2020, there have been 941,949 cases of COVID-19 and 47,522 deaths globally. In the 
United States, cases have rapidly risen with the total number of cases currently 216,722, 
including 5,137 deaths (Johns Hopkins Coronavirus Resource Center accessed at 
https://coronavirus.jhu.edu/map.html on April 2, 2020). 
To date, no randomized, controlled trials (RCTs) exist that demonstrate the efficacy of any 
therapeutic agent for treatment of SARS-CoV-2. However, several therapeutic agents have 
been postulated to have some effect against SARS-CoV-2, including remdesivir, 
lopinivir/ritonavir, tocilizumab, chloroquine/hydroxychloroquine, interferon-1b, and convalescent 
plasma.  Data to support any of these treatments are sparse at best (1).  Therefore, in the 
context of a pandemic, it is critically important that therapeutic agents are investigated in 
scientifically and ethically sound studies. Large clinical trials are currently underway globally to 
establish an evidence base therapies used to target COVID-19. 
2.2 Scientific rationale 
We are proposing to examine the effect of hydroxychloroquine and azithromycin, alone and in 
combination, on clinically relevant outcomes in hospitalized patients with SARS-CoV-2 infection.  
Due to its in-vitro activity against SARS-CoV-2 and its wider availability in the United States 
compared with chloroquine, hydroxychloroquine has frequently been administered to 
hospitalized COVID-19 patients on an uncontrolled basis. No data from RCTs currently exist to 
inform clinical guidance on the use, dosing, or duration of hydroxychloroquine for treatment of 
SARS-CoV-2 infection. Azithromycin has been used in combination with hydroxychloroquine in 
limited case series and has been postulated to provide additional efficacy. 
2.2.1  Mechanism of action of chloroquine and hydroxychloroquine in SARS-CoV-2 
Hydroxychloroquine is an anti-malarial 4-aminoquinoline approved by the FDA for treatment in 
rheumatoid arthritis, systemic lupus erythematosus, and other inflammatory conditions. A 
derivative of chloroquine, it was first synthesized in 1946 by introducing a hydroxyl group to 
chloroquine. Hydroxychloroquine has pharmacokinetics similar to that of chloroquine with rapid 
gastrointestinal absorption and renal elimination, but it has a more favorable risk-benefit ratio 
than chloroquine due to a wider therapeutic index and better side-effect profile (2, 3).  
 
Investigators have postulated several mechanisms to explain potential activity of chloroquine 
and hydroxychloroquine against SARS-CoV-2 (4):  
1) Inhibition of viral entry. Hydroxychloroquine inhibits biosynthesis of sialic acids and 
interferes with protein glycosylation, which may disrupt interactions necessary for viral 
attachment and entry (5, 6). 
2) Inhibition of viral release into the host cell. Hydroxychloroquine blocks endosomal 
acidification, which activates endosomal proteases. These proteases are required to 
initiate coronavirus/endosome fusion that releases viral particles into the cell (7). 
3) Reduction of viral infectivity. Hydroxychloroquine has been shown to inhibit protein 
glycosylation and proteolytic maturation of viral proteins. Studies on other RNA viruses 

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have shown a resulting accumulation of non-infective viral particles, or an inability of viral 
particles to bud out of the host cell (8, 9). 
4) Immune modulation. Hydroxychloroquine reduces toll-like receptors and cGAS-STING 
signaling. It has been shown to reduce release of a number of pro-inflammatory 
cytokines from several immune cell types (10), such as interleukin-6 thereby blocking the 
pathway that subsequently leads to acute respiratory distress syndrome (ARDS) (11). By 
this mechanism, hydroxychloroquine might show benefit in treating cytokine storm that 
occurs in some critically ill patients with SARS-CoV-2 (12). 
 
2.2.2 Mechanism of action of azithromycin in SARS-CoV-2 
In addition to antibacterial effects, azithromycin has anti-inflammatory effect, particularly in the 
lungs.  A mouse model of viral bronchiolitis demonstrated that azithromycin decreased 
inflammatory mediators in bronchoalveolar lavage fluid and enhanced resolution of chronic 
airway inflammation (13).  Azithromycin has also been associated with improved lung function in 
patients with cystic fibrosis and diffuse panbronchiolitis, independent of bacterial eradication 
(14, 15).  Macrolides have also been associated with improved survival in patients with 
community acquired pneumonia when paired with a beta lactam antibiotic, regardless of 
pneumonia etiology, suggesting an additional effect beyond bacterial killing (16). In a 
nonrandomized, retrospective analysis of 349 critically ill MERS patients, macrolide therapy 
(primarily azithromycin) was associated with a nonsignificant reduction in 90-day mortality (odds 
ratio 0.84, 95% confidence interval 0.47-1.51) (17). 
2.3 Evidence for chloroquine and hydroxychloroquine 
2.3.1 In vitro studies 
Chloroquine has been shown to have in vitro activity against a diverse group of RNA viruses 
(18-20). Furthermore, chloroquine and hydroxychloroquine have been demonstrated to be 
active against a number of coronaviruses in vitro, including SARS-CoV-1 (21, 22). 
 
In one study, chloroquine appeared to block SARS-CoV-2 at low-micromolar concentration in a 
Vero E6 cells assay (23). This study also demonstrated the proof-of-concept that that 
chloroquine functioned at both entry and post-entry stages of the SARS-CoV-2 infection in Vero 
E6 cells. Hydroxychloroquine (EC50=0.72 μM) was shown to be more potent that chloroquine 
(EC50=5.47 μM) in an in vitro anti-SARS-CoV-2 study (12). This study also demonstrated 
through a PBPK model that lung concentrations of hydroxychloroquine rapidly increased and 
reached steady state after a loading dose and maintenance doses, and reached levels three 
times the potency of chloroquine. 
2.3.2 In vivo studies of hydroxychloroquine 
In February 2020, early results from a number of Chinese clinical trials on chloroquine used for 
SARS-CoV-2 pneumonia demonstrated that in 100 patients, chloroquine was superior to 
standard of care in inhibiting the exacerbation of pneumonia, improving lung imaging findings, 
promoting earlier viral conversion, and shortening the disease course (24). Severe adverse 
reactions to chloroquine were not noted. Based on these data, the National Health Commission 
of the People's Republic of China recommended inclusion of chloroquine in the Guidelines for 
the Prevention, Diagnosis, and Treatment of Pneumonia Caused by COVID-19 (25).  
 
On March 25, 2020, a small randomized study from China compared the use of 
hydroxychloroquine (400 mg per day for 5 days) with standard of care in 30 patients with non-

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severe SARS-CoV-2 infection randomized to HCQ or control. There was no difference in 
respiratory pharyngeal swab viral conversion or clinical outcome at Day 7 between the two 
groups, but patients received multiple treatments including antivirals (26). Several larger clinical 
studies are planned or underway, designed to establish the role of chloroquine and 
hydroxychloroquine for COVID-19 treatment (27) (see section 2.3.5). 
 
A randomized, unblinded study of 62 patients in Wuhan, China (not yet peer-reviewed) with mild 
SARS-CoV-2 illness examined the impact of hydroxychloroquine 200 mg twice-daily for 5 days, 
and found a 1-day reduction in fever duration among patients receiving hydroxychloroquine and 
a higher rate of radiographic resolution at day 6 in the hydroxychloroquine group (25/31, 80.6% 
improved in the hydroxychloroquine group vs. 17/31, 54.8% in the control group) (28). 
2.3.3 In vivo studies of hydroxychloroquine and azithromycin combination therapy 
A recent non-randomized study from France of hydroxychloroquine (200 mg every 8 hours x 10 
days) (n=26) for patients hospitalized with SARS-CoV-2 who had symptoms of upper and lower 
tract infections without oxygen requirement found that hydroxychloroquine decreased viral 
shedding at Day 6 compared to the untreated group (n=16) (70% vs 12.5%, p= 0.001). Six 
patients in the hydroxychloroquine group were receiving concomitant azithromycin for 
prevention of secondary bacterial pneumonia. On Day 6, all patients treated with the 
combination (hydroxychloroquine and azithromycin) demonstrated virologic clearance, 
compared to 57.1% of patients treated with hydroxychloroquine alone (n= 20) (29). 
This study had several limitations, including the following:  
1) Six patients in the hydroxychloroquine arm with poor outcomes (including death, ICU 
transfer, treatment intolerance) were excluded from the analysis.  
2) Use of an arbitrary, unvalidated virologic clearance on Day 6 was used as the primary 
outcome. For example, the authors reported that one patient tested negative for the virus 
on Day 6 but subsequently tested positive on Day 8, indicating that a single, negative 
polymerase chain reaction (PCR) test on a nasopharyngeal swab may not indicate 
persistent virologic clearance.  
3) The control group was not systematically selected, introducing allocation bias. 
 
A follow-up manuscript from the same authors (published online on March 28, not yet peer 
reviewed) reported a non-comparative study of 80 patients hospitalized with SARS-CoV-2 
infection without oxygen requirement (only 15% had fevers and 4 patients were asymptomatic) 
who were treated with the same doses of hydroxychloroquine and azithromycin for 3-10 days. 
One patient died; 3 transferred to ICU; 15% progressed to oxygen requirement; and 81.3% were 
improved at discharge. 83% of patients had negative PCR tests of nasopharyngeal swabs by 
Day 7, and 93% had negative PCR studies by Day 8. 97.5% of patients had negative viral 
cultures by Day 5. Mean hospital stay was 4.6 days. Toxicity data were not reported. ECG was 
performed before and 2 days after treatment, but QTc data were not reported (30).  A 
subsequent, smaller study of 11 patients treated with hydroxychloroquine/azithromycin at a 
different site in France did not suggest any benefit of the combination on viral clearance or 
clinical outcomes, and one patient had medications discontinued because of QTc prolongation 
(31). 
 

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Given the substantial limitations of existing in vivo studies, further trials are urgently needed to 
better understand the role of hydroxychloroquine and azithromycin in the treatment of SARS-
CoV-2.  
2.3.4  How this study complements other planned work 
Investigators across the U.S. have recently designed multiple clinical trials to evaluate impact of 
several experimental pharmacologic agents and other therapies when administered to patients 
with COVID-19. For example, three multicenter studies are actively enrolling to analyze efficacy 
and safety of remdesivir for hospitalized patients with moderate or severe disease from COVID-
19 (NCT04292730, NCT04292899, and NCT04280705). Each of these studies excludes 
patients with mild disease, moderate renal insufficiency, or moderate transaminitis. Other U.S. 
studies actively recruiting patients are assessing efficacy and safety of sarilumab in hospitalized 
patients with severe COVID-19 (NCT04315298) or nitric oxide inhalational therapy for 
mild/moderate (NCT04305457) or severe (NCT04306393) disease. Investigators have designed 
additional studies to analyze the potential impact of numerous other experimental agents, 
including losartan (NCT04312009), intravenous aviptadil (NCT04311697), PUL-042 inhalation 
solution (NCT04312997), CD24Fc (NCT04317040), and convalescent plasma (NCT04325672); 
however, these COVID-19 studies are not yet recruiting patients.  
While providers commonly prescribe hydroxychloroquine and/or azithromycin to treat 
hospitalized patients with COVID-19, no high-quality data exists to demonstrate that either of 
these treatments are effective. Investigators associated with a single U.S. study plan to evaluate 
efficacy of hydroxychloroquine versus azithromycin for hospitalized patients with COVID-19 
(NCT04329832); however, this study is not yet recruiting patients and does not randomize 
patients to a control arm where patients receive neither experimental therapy.  A randomized, 
placebo-controlled study in Alberta, Canada will randomize patients to 5 days of 
hydroxychloroquine versus placebo to examine whether hydroxychloroquine reduces the 
incidence of severe disease (NCT04329611).  The AIDS Clinical Trials Group is planning a 
multicenter study to examine the combination of hydroxychloroquine/azithromycin in outpatients 
and plans to collaborate with Novartis to do a similar treatment trial in inpatients; however, it is 
not clear when those studies will begin, and the Novartis study will reportedly exclude critically ill 
patients.  The NHLBI-funded PETAL network is also planning an inpatient trial of 
hydroxychloroquine versus placebo (NCT04332991). 
Our proposed study is unique and complements other planned studies because our study will 
allow systemic comparison of clinical outcomes for patients who receive hydroxychloroquine 
versus azithromycin, as well as for patients who receive neither or both experimental agents. 
Other studies evaluating these agents to date have not randomized patients to a control group 
that receives standard of care (i.e., neither experimental agent); however, inclusion of a 
randomized control group is critical to understanding efficacy and safety of 
hydroxychloroquine and/or azithromycin when used to treat COVID-19. Finally, many 
hospitalized patients in the proposed study population would not qualify for treatment via 
existing clinical trials due to insufficient disease severity, renal dysfunction, or transaminitis. 
Therefore, this study provides a mechanism for interested patients who would not otherwise 
qualify for a clinical trial to participate in a study and receive protocolized pharmacologic therapy 
targeting COVID-19. 
 
 

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3.  Descriptions of Study Drugs 
3.1 Hydroxychloroquine 
3.1.1 Formulation, Appearance, Packaging, and Labeling 
Hydroxychloroquine sulfate tablets are provided as a white/off-white, film-coated, peanut-
shaped tablets.  Each tablet contains 200 mg of the active ingredient (HCQ sulfate).  The non-
active ingredients in the formulation include black ink, calcium hydrogenophosphate, carnauba 
wax, cornstarch, magnesium stearate, Opadry White YS-I-7443 and polyethylene glycol 400. 
3.1.2 Dosing and administration 
Participants will receive an 800 mg loading dose the first day, followed by 600 mg daily for 4 
more days.  All study drug doses will be administered orally.  Study drug will be supplied as 200 
mg tablets. 
3.1.3 Rationale for selection of dose 
Hydroxychloroquine is an aminoquinolone antimalarial agent that is also approved and used for 
the treatment of autoimmune diseases including rheumatoid arthritis and systemic lupus 
erythematosus.  
Both chloroquine and hydroxychloroquine have been reported to have in vitro antiviral activity 
against SARS-CoV-2 (23, 32). In vitro testing using Vero cells vary; Yao et al demonstrated an 
EC50 of 23.90 and 5.47 uL for chloroquine at 24 and 48 hours respectively, compared to 6.14 
and 0.72 for HCQ (12). Liu et al reported on four multiplicities of infection (MOIs) (0.01, 0.02, 
0.2, and 0.8); the 50% maximal effective concentration (EC50) for chloroquine was (2.71, 3.81, 
7.14, and 7.36 μM) and for hydroxychloroquine was (4.51, 4.06, 17.31, and 12.96 μM) (32). 
Hydroxychloroquine may exhibit higher activity to chloroquine in protecting Vero cells from 
infection (“prophylaxis”); when Vero cells were pre-treated with drug before SARS-CoV-2 
challenge, EC50 for chloroquine was >100 and 18.01 uL at 24 and 48 hours, respectively, 
compared to 6.25 and 5.85 for HCQ (32). In animals, both drugs share similar tissue distribution 
patterns, with high concentrations in the liver, spleen, kidney, and lung reaching levels of 200–
700 times higher than those in the plasma (33).  
The optimal dose of hydroxychloroquine for treatment or prophylaxis is unknown. Absorption 
after oral dosing is incomplete and variable (~70% [range: 25 to 100%]) (2, 34). Metabolism is 
primarily hepatic with metabolites including bidesethylchloroquine, desethylhydroxychloroquine, 
and desethylchloroquine (35). The drug has long half-life elimination of approximately 40 days 
(2). Standard dosing in other medical conditions include: systemic lupus erythematosus (200-
400 mg daily given once or divided BID), malaria treatment (800 x1, followed by 400 mg at 6, 24 
& 48 h after initial dose), Q fever (600 mg/day in 1 to 3 divided doses) and rheumatoid arthritis 
(200-400 mg daily in 1-2 divided doses). The recommended maximum dose of HCQ is 5 mg/kg 
actual body weight daily. 
The dose chosen for this study is 800 mg orally once daily on day 1 as a loading dose followed 
by 600 mg QD for an additional 4 days. This dose was chosen for several reasons:  
 
current in vitro studies report a wide range of EC50 for SARS-CoV-2,  
 
variability of absorption and of tissue distribution into the lung,  

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 
due to a lack of phase 1b data for this drug in SARS-CoV-2 infection, the optimal 
pharmacokinetic/pharmacodynamic target is unknown.  
Yao et al made assumptions regarding the absorption and tissue distribution that were optimistic 
and recommended as an optimal treatment dose 400 mg BID on day 1 as a loading dose 
followed by 200 mg BID daily for the remainder of the treatment course (12). There is concern 
that this dose will not meet the necessary pharmacokinetic/pharmacodynamic targets for 
efficacy, in which case a negative study would not be due to failure of the antiviral efficacy but of 
suboptimal dosing. Yao et al also noted that the drug exposure was the same for 200 mg BID or 
400 mg QD daily dosing. We propose a higher loading dose of 800 mg once daily on day 1 to 
ensure more rapid steady state tissue distribution and 600 mg daily (versus 200 mg three times 
daily) dosing for ease of dosing and adherence. 
 
3.2 Azithromycin 
Azithromycin is an azalide antibacterial that is FDA-approved for treatment of acute bacterial 
exacerbations of chronic obstructive pulmonary disease, acute bacterial sinusitis, community-
acquired pneumonia, pharyngitis/tonsillitis, uncomplicated skin and skin structure infections, 
urethritis and cervicitis, and genital ulcer disease.  Between 2013-2015, it was one of the most 
commonly prescribed antibiotics in the United States, with an average of 443 prescriptions per 
1,000 Express Scripts beneficiaries (~15 million prescriptions per year) (36). 
3.2.1  Formulation, appearance, labeling and packaging 
Azithromycin tablets are supplied as pink film-coated tablets containing azithromycin dihydrate 
equivalent to 250 mg of azithromycin. 
3.2.2.  Dosing and administration 
Azithromycin will be dosed at 500 mg orally on day 1 of treatment, followed by 250 mg once 
daily on days 2-5 of treatment.   
3.2.3  Rationale for dose selection 
This dosing is a commonly used dose for community-acquired pneumonia; as the potential 
mechanism of action and pharmacodynamics of azithromycin when used to treat SARS-CoV-2 
are unknown, we chose a dose that has been demonstrated safe in a large number of patients.  
Mechanism of Action / Potential activity in COVID19: 
- 
Antibacterial effect: Binds bacterial 50S ribosomal subunit and inhibits RNA-dependent 
protein synthesis (37). Can potentially serve to prevent bacterial super infection in 
patients with COVID19. 
- 
Immunomodulatory effect (studied mainly in COPD and Cystic Fibrosis): Inhibits NFB 
activation and expression of activator protein-1, reduces number and function of 
neutrophils leading to lower concentrations of neutrophil elastase, IL-8, and IL-1. This 
in turn leads to a reduction in cellular damage/tissue injury (38, 39). Adjunct therapy in 
moderate to severe ARDS has been shown to improve survival and reduce days on 
mechanical ventilation, which may be related to this immunomodulatory effect (40). 

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- 
Direct antiviral effect: invitro studies; Suppresses viral replication of Rhinovirus in 
bronchial epithelial cells (exact mechanism unclear) (41-43), inactivates endocytic 
activity of A(H1N1)pdm09 blocking internalization in early infection (exact mechanism 
unclear) (44). Azithromycin has no studied direct antiviral effect with the Coronaviridae 
family. 
- 
Combination of Hydroxychloroquine (HCQ) and Azithromycin studied in a small 
prospective non-randomized trial could suggest that Azithromycin might augment the 
effects of HCQ, but this is still unclear in true effect, clinical significance as well as 
underlying mechanism (29) 
 
Oral Pharmacokinetics:  
- 
Immediate release, bioavailability 34 – 52% with a half-life of 68 – 72 hours.  
- 
Extended release bioavailability, 28 – 43% with a half-life of 59 hours.  
- 
Metabolized hepatically to inactive metabolites and excreted in both bile and urine with 
50% and 6 – 14% being unchanged, respectively.  
 
Drug-drug interactions:  
- 
CYP3A4 substrate 
- 
P-glycoprotein/ABCB1 inhibitor  
- 
Potentially drug-drug interactions relevant to our study include:  
o Chloroquine; QT prolongation and ventricular arrythmias 
o Other QT prolonging drugs such as antiemetics, antidepressants, antipsychotics, 
and other antimicrobials  
 
Safety and tolerability (Oral, Adults):  
- 
Single dose regimens tend to be associated with higher frequency of adverse events, 
however this is not relevant to our study given that the dosing of azithromycin will be a 
daily oral dose for 5 days.  
- 
The majority of adverse events are gastrointestinal with an incidence of approximately 
>10%: Loose stools (≤14%), diarrhea (2% - 9%), nausea (≤7%), and vomiting (≤2%). 
- 
Other (systems based) adverse events occur at lower rates of 1 – 10%: Cardiovascular: 
Chest pain (≤1%), palpitations (≤1%); Central nervous system: Dizziness (≤1%), 
drowsiness (≤1%), fatigue (≤1%), headache (≤1%), vertigo (≤1%); Dermatologic: Skin 
rash (≤5%), pruritus (≤2%), skin photosensitivity (≤1%); Endocrine & metabolic: 
Increased lactate dehydrogenase (1% – 3%), increased gamma-glutamyl transferase 
(1% –2%), increased serum potassium (1% – 2%), decreased serum bicarbonate (≥1%), 
decreased serum glucose (>1%); Gastrointestinal: Abdominal pain (1% – 7%), anorexia 
(≤2%), dysgeusia (≤1%), dyspepsia (≤1%), flatulence (≤1%), gastritis (≤1%), melena 
(≤1%), mucositis (≤1%), oral candidiasis (≤1%); Genitourinary: Vaginitis (≤3%), genital 
candidiasis (≤1%); Hematologic & oncologic: decreased hematocrit (>1%), decreased 
hemoglobin (>1%), increased neutrophils (>1%), thrombocythemia (adults: >1%), 
eosinophilia (≥1%), lymphocytopenia (≥1%); Hepatic: Increased serum ALT (≤6%), 

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increased serum AST (≤6%), increased serum bilirubin (≤3%), cholestatic jaundice 
(≤1%); Neuromuscular & skeletal: Increased creatine phosphokinase (1% – 2%); Renal: 
Increased serum creatinine (≤6%), increased blood urea nitrogen (≤1%), nephritis 
(≤1%); Respiratory: Bronchospasm (≤1%). 
- 
Other (rare) adverse events potentially relevant to our study include: 
o QT prolongation; Azithromycin can inhibit the rapid component of the delayed 
rectifier K+ current channels and induce QT prolongation, however several 
studies and analyses have shown that Azithromycin-induced QT prolongation 
does not lead to Torsade de Pointes (45, 46). 
o Increased risk of death from cardiovascular events; Some studies have 
suggested that Azithromycin has been associated with death from cardiovascular 
events in high risk individuals, however several subsequent studies and analyses 
have not shown an association (47, 48). 
- 
Pregnancy: Azithromycin crosses the placenta but is generally considered safe in 
pregnancy (Category B) (37, 49, 50); a recent study did associate macrolides with an 
increased incidence of genital malformations, but azithromycin was not specifically 
studied (51). 
- 
Breast Feeding: Azithromycin is present in breast milk and has been shown to have a 
half-life of ~15 hours, with some studies detecting Azithromycin in breast milk up to 30 
days after single dose administration (52, 53). There is conflicting data of the 
incidence/nature of macrolide-induced infantile hypertrophic pyloric stenosis in infants 
exposed in the first 2 weeks of life (54, 55). Some infants may experience adverse 
gastrointestinal events such as diarrhea and should be monitored while the mother is 
receiving therapy (55). It is generally considered safe in breast feeding mothers 
(Category B). 
- 
Contraindications: Hypersensitivity to macrolides, history of macrolide induced 
hepatitis/cholestatic jaundice 
4.  Objectives 
4.1  Definitions 
The World Health Organization ordinal scale is defined as an assessment of the clinical status 
of a study participant at the first assessment of a given study day.  For purposes of this study, 
this assessment will be considered to have occurred at 9 a.m. on the study day. 
The ordinal scale is an assessment of the clinical status at the first assessment of a given study 
day. The scale is as follows: 1) Death; 2) Hospitalized, on invasive mechanical ventilation or 
extracorporeal membrane oxygenation (ECMO); 3) Hospitalized, on non-invasive ventilation or 
high flow oxygen devices; 4) Hospitalized, requiring supplemental oxygen; 5) Hospitalized, not 
requiring supplemental oxygen - requiring ongoing medical care (COVID-19 related or 
otherwise); 6) Hospitalized, not requiring supplemental oxygen - no longer requires ongoing 
medical care; 7) Not hospitalized, limitation on activities and/or requiring home oxygen; 8) Not 
hospitalized, no limitations on activities. 
4.2 Primary 
(1) To compare the efficacy of each experimental treatment (azithromycin and 
hydroxychloroquine) to the efficacy of supportive care, using the World Health Organization 
ordinal scale measured on day 14 after enrollment 

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4.3 Secondary 
(1) To compare the rates of death during the index hospitalization among participants receiving 
each experimental treatment and participants receiving supportive care alone 
(2) To compare the number of days on mechanical ventilation among participants receiving 
each experimental treatment and participants receiving supportive care alone 
(3) To compare the proportion of participants not receiving mechanical ventilation at baseline 
who progress to requiring mechanical ventilation during the index hospitalization among 
participants receiving each experimental treatment and participants receiving supportive 
care alone 
(4) To compare the World Health Organization ordinal scale measured on day 28 after 
enrollment between participants receiving each experimental treatment and participants 
receiving supportive care alone 
(5) To compare the total length of hospitalization during the index hospitalization among 
participants receiving each experimental treatment and participants receiving supportive 
care alone 
(6) To compare the duration of fever (defined as temperature ≥38.0 C measured at any site) 
after randomization among participants receiving each experimental treatment and 
participants receiving supportive care alone 
(7) To compare the duration of supplemental oxygen use after randomization among 
participants receiving each experimental treatment and participants receiving supportive 
care alone 
(8) To describe the rates of all-cause discontinuation of study medication among patients 
receiving experimental treatment 
(9) To compare the rates of drug-associated adverse events of special interest among patients 
receiving each experimental treatment and participants receiving supportive care alone 
during the first fourteen days after enrollment 
5.  Study Design 
This will be a pragmatic, randomized, open-label, incomplete factorial with nested randomization 
clinical trial evaluating the efficacy and safety of two potential treatments for hospitalized 
patients with confirmed SARS-CoV-2 infection.  Participants who are hospitalized and have a 
positive nucleic acid amplification test for SARS-CoV-2 will undergo an initial randomization in a 
1:1 ratio to one of the following regimens: 
Arm 1: Supportive care alone 
Arm 2: Supportive care plus hydroxychloroquine 
Participants who meet eligibility criteria to receive azithromycin will undergo a second 
randomization in a 1:1 ratio to receive additional concurrent therapy.  This will effectively result 
in four treatment groups: 
1) Supportive care alone 
2) Supportive care plus hydroxychloroquine  
3) Supportive care plus azithromycin 
4) Supportive care plus hydroxychloroquine plus azithromycin 

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6.  Study Population 
500 hospitalized male and female patients (≥ age 12 years or older) diagnosed with SARS-CoV-
2 related infection, and who meet all study eligibility criteria will be enrolled in the study. 
Participants will be enrolled at all three DUHS hospitals: Duke University Hospital, Duke 
Regional Hospital and Duke Raleigh Hospital, as well as the Durham VA Medical Center. We 
anticipate that most participants enrolled in the study will be residents of the 12-county 
Raleigh/Durham Combined Statistical Area. Estimated time of screening to end of study for an 
individual subject will be 28 days, including treatment and follow up.  
 
Participant eligibility will be confirmed by one of the study co-investigators prior to enrollment.  
 
Inclusion Criteria 
 
To participate in the study, participants must meet ALL of the following criteria:  
1) Admitted to participating hospital with symptoms suggestive of SARS-CoV-2 infection 
OR develop symptoms of SARS-CoV-2 during hospitalization  
2) Subject (or legally authorized representative) can provide written informed consent (in 
English or Spanish) affirming intention to comply with planned study procedures prior to 
enrollment  
3) Male or female aged 12 years or older at the time of enrollment  
4) Has laboratory-confirmed SARS-CoV-2 infection determined by a validated nucleic acid 
amplification assay (public health or commercial) in any respiratory specimen collected 
within 14 days of randomization 
5) Illness of any duration that includes 
 
Radiographic evidence of pulmonary infiltrates (chest X-ray or CT scan) OR 
 
Clinical documentation of lower respiratory symptoms (cough, shortness of 
breath, or wheezing) OR  
 
Any documented SpO2 ≤ 94% on room air OR 
 
Any inpatient initiation of supplemental oxygen regardless of documented cause 
Exclusion Criteria  
Participants with ANY of the following criteria will be excluded from the study:  
1) Participating in any other clinical trial of an experimental agent for SARS-CoV-2   
2) On hydroxychloroquine at any time during hospitalization, or within 180 days of 
hospitalization for COVID-19 regardless of indication  
3) History of cirrhosis, long QT syndrome or porphyria of any classification 
4) Most recent ECG prior to time of screening with QTc of ≥500 msec  
5) Known hypersensitivity to hydroxychloroquine or 4-aminoquinoline derivatives 
6) Weight less than 40 kg 
7) Death anticipated within 48 hours of enrollment 
8) Inability to obtain informed consent from the patient or designated medical decision 
maker 
 
Inclusion of Vulnerable Subjects 
Hydroxychloroquine has been shown to permeate across the placenta, with cord blood 
concentrations equivalent to those found in maternal blood. Despite theoretical concerns of 
retinal toxicity and ototoxicity, numerous cohort studies have demonstrated its safety in 

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pregnant women exposed to the medication. Hydroxychloroquine has been used extensively as 
an immunomodulatory agent in pregnant women and the American College of Rheumatology 
recommends continuation of the drug among pregnant women with SLE who were initiated prior 
to pregnancy. After careful assessment of available data, consultation with local experts, and 
the lack of alternative therapeutic options for persons with SARS-CoV-2, the study team 
deemed that inclusion of pregnant women in the trial was warranted. 
Persons under the age of 18 are significantly less likely present with symptoms warranting 
hospitalization than the rest of the population. As a result, little clinical trial data exist on 
therapeutics for COVID-19 in this subset of the population. This trial presents an opportunity to 
systematically collect data on the impact of two widely available, low-cost agents on a 
potentially life-threatening infection, with no known therapeutic options. Both azithromycin and 
hydroxychloroquine have been shown to be safe in adolescents. Based on the risk/benefit 
analysis for prospective hospitalized patients in this age group, inclusion of adolescents (age 
12-18) in the trial is warranted.  
Prisoners will not be enrolled in this study because it does not involve one of the Health and 
Human Services permissible categories of research involving prisoners. 
 
Lifestyle Considerations 
Study participants will be asked to avoid participation in other trials of pharmacologic agents 
with possible activity against SARS CoV-2. 
  
Screen Failures 
At completion of prospective participant screening, the principal investigator (or designee) will 
review eligibility criteria to determine the patient’s suitability for study inclusion. Data on reason 
for ineligibility will be collected for all screen failures. Persons who fail study screening will be 
notified about the reason for their ineligibility. Patients who are deemed ineligible due to 
eligibility criteria based on clinical findings/data can be re-screened again at a subsequent time 
based on investigator’s discretion.  
Strategies for Recruitment and Retention  
Recruitment 
All patients admitted to study site hospitals for SARS-CoV-2 will be screened for participation in 
the study. No external recruitment efforts will occur beyond this subset of patients. Study 
coordinators will track the clinical course of every hospitalized SARS-CoV-2 patient in DUHS, 
with particular attention to whether or not they were offered enrollment into a trial involving 
another investigational agent. Through review of the electronic health record, the study 
coordinator will determine the reason patient was not enrolled. If failure to enroll was not due to 
one of this protocol’s exclusion criteria, or a broad declination of participation in SARS-CoV-2 -
associated clinical trials (available in OnCore), the patient will be screened and approached for 
inclusion in the study. Targeted groups of clinicians within the health system will be notified 
about the trial prior to the study’s commencement and on the day the study is open to 
enrollment via e-mail. Clinician groups to be targeted include: emergency department faculty, 
trainees and staff; critical care faculty, trainees and staff; hospitalist physicians; infectious 
diseases faculty trainees and staff; transplant faculty, trainees, and staff; Internal Medicine, 
General Surgery and Anesthesia house staff. Study details will also be available to all DUHS 
faculty, trainees and staff via the COVID-19 clinical trials section of CustomID.  
 
The IRB will approve the recruitment process and all materials prior to any recruitment to 
prospective subjects directly.  

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To minimize unnecessary exposure to SARS-CoV-2, study staff will be instructed to conduct as 
much screening for eligibility by electronic health record review as possible, and informed 
consent will be preferentially obtained by telephone. 
 
Retention  
Significant participant attrition is not expected given that this study primarily follows patients 
while hospitalized.  Study investigators will be available to the participant’s clinical team to 
discuss alterations in clinical course that may influence their decision to keep the patient in the 
trial.  
 
Compensation 
Participants will not be compensated for their participation in the study.  
 
Costs 
Participants will be charged for study drugs as part of their hospital care.  Participants will be 
informed of this by the study coordinator prior to obtaining informed consent.  
 
7.  Enrollment and Randomization 
7.1 Enrollment Procedures 
Individuals who are hospitalized with a positive nucleic acid amplification test for SARS-CoV-2 
and who are not enrolled in another therapeutic trial for SARS-CoV-2 will be invited to 
participate in this study.  Interested individuals will be provided with information about the study 
including risks and potential benefits of all study procedures.  If site staff are satisfied that the 
potential participant understands the information and the potential participant is willing, the 
potential participant will be asked to participate in the study.  Study-specific procedures will be 
initiated only after the participant has provided written, informed consent.  The consent process 
will be performed by providing the patient or designated healthcare provider a copy of the 
consent form (this will be given to an individual who is going into the room; the study coordinator 
will not enter the room).  The patient will be provided an opportunity to review the consent, and 
when they are ready, the consent process will occur by e-consent on study iPads.    If the 
patient is unable to provide written, informed consent and they have a designated medical 
decision maker, that person will be approached either in person or by telephone with information 
about the study.  The consent process with the medical decision maker will occur using e-
consent also. 
 
7.2 Randomization 
This will be a randomized trial.  Randomization and treatment arm assignment will be generated 
with randomly permuted blocks of size 2, 4, 6, and 8.  Participants will undergo two 
randomizations.  The first randomization (to standard of care or hydroxychloroquine) will be 
stratified by whether or not the patient is receiving mechanical ventilation at the time of 
enrollment and by hospital site.  Eligible patients who have provided informed consent will be 
randomized with equal probability to each arm.  Participants who meet inclusion criteria for the 
second randomization (to add or not add azithromycin) will be randomized with equal probability 
to each of these treatments.  Random assignment sequences will be generated in a way that 

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limits the imbalance between arms within strata while ensuring that the sequence is not 
predictable based on previous assignments. 
8.  Study Procedures 
8.1 Ascertainment of potential participants 
Potential participants will be ascertained in one of two ways: 1) Daily review of all positive 
SARS-CoV-2 tests from the relevant hospital laboratories and 2) Referral from treating 
providers. 
8.2 Baseline screening and eligibility determination 
By chart review prior to patient encounter:  
 
Confirm positive SARS-Cov2 test  
 
Confirm patient meets demographic criteria  
 
Confirm patient meets clinical criteria  
 
Ascertain in OnCore research portal that patient is not enrolled in any other COVID-19 
related clinical trial  
 
Review chart for azithromycin within 7 days and hydroxychloroquine use within 180 days 
of planned baseline encounter (including outpatient records)  
 
Review medical history for exclusionary conditions 
 
Review allergy record for evidence of hypersensitivity to azithromycin or 
hydroxychloroquine or related drugs  
 
Locate and have investigator review EKG in electronic health record obtained within 7 
days of planned encounter for evidence of prolonged QTc  
At enrollment encounter (Day 1):  
 
Locate hospital and unit where patient is located 
 
Verify with treating clinical team that patient is appropriate for the trial  
 
Determine whether the patient can provide informed consent. If not, identify patient’s 
legally authorized representative.  
 
If the patient cannot provide informed consent, patient’s legally authorized representative 
should be contacted and the informed consent process should be executed through 
them. 
 
The patient or legally authorized representative should be provided a copy of the written 
informed consent document to review. 
 
Informed consent should be preferentially obtained by e-consent as outlined above 
rather than in person. 
8.3  Initial assessment and treatment assignment 
Study Intervention Procedures 
Immediately post enrollment (Day 1):  
 
Conduct randomization procedure  
 
Inform treating team of patient’s enrollment. Study coordinator (in conjunction with on-
call investigator) will advise the team on their patient’s treatment assignment, and will 
advise the team to write treatment orders accordingly. 

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 
Inform team to perform ECG (if none available in the preceding 7 days). For women of 
childbearing potential (and no clear evidence of pregnancy), inform team to obtain urine 
qualitative HCG.  
Other activities on Day 1:  
 
Complete case report form (CRF) in a timely manner. Obtain the following data from 
EHR to complete CRF including the following:  
o Demographic data  
o Most recent height and weight  
o Chronic medical conditions related to study eligibility  
o Date of onset of COVID-19 symptoms  
o Data on hospitalization including date of admission, date of ICU transfer (if 
applicable), presenting complaint, vitals on presentation, selected laboratory data 
on presentation  
o Medications administered (with dates) for index illness prior to presentation and 
during hospitalization to date  
o Medication allergies on record  
o Findings of last documented lung examination  
o Results of chest X-ray or CT scan on presentation  
o Most recent vital signs prior to time of enrollment  
o Most recent complete blood count, basic metabolic panel and hepatic function 
panel obtained prior to the time or enrollment  
o QTc of most recent ECG 
o Urine pregnancy test (if applicable)  
o Respiratory support status (ECMO, invasive mechanical ventilation, non-invasive 
ventilation, high-flow nasal cannula (with oxygen flow rate), standard nasal 
cannula (with oxygen flow rate) 
o Level of care (ICU, stepdown, standard)  
o WHO ordinal scale value (in conjunction with study investigator)  
For patients enrolled at DUHS, all of this data will be automatically captured in Redcap as part 
of the overall COVID research plan, so the coordinator will not need to extract any data.  For 
patients at other sites, data will need to be entered into the Redcap database. 
8.4  Assessment during study 
Daily Assessment (Days 2-13):  
 
Study coordinator will complete CRF based on clinical data abstracted from chart with 
the following information:  
o Vital signs conducted most proximal to CRF entry  
o Maximum temperature in prior 24 hours (9am-8:59 am) 
o New medications administered (with dates) for index illness during interval since 
last assessment  
o Interim complete blood count, basic metabolic panel and hepatic function panel  
o QTc of most recent EKG conducted in interim  
o Respiratory support status (ECMO, invasive mechanical ventilation, non-invasive 
ventilation, high-flow nasal cannula (with oxygen flow rate), standard nasal 
cannula (with oxygen flow rate) 

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o Level of care (ICU, stepdown, standard)  
o Adverse event documentation  
 
For patients enrolled at DUHS, all of this data will be automatically captured in Redcap 
as part of the overall COVID research plan, so the coordinator will not need to extract 
any data.  For patients at other sites, data will need to be entered into the Redcap 
database. 
 
Day 14 Assessment:  
 
Study coordinator will complete CRF based on clinical data abstracted from chart with 
the following information: 
o Vital signs conducted most proximal to CRF entry  
o Medications related to index illness still being administered, or written for 
discharge  
o Most proximal chest X-ray or CT scans  
o Most proximal complete blood count, basic metabolic panel and hepatic function 
panel  
o QTc of most recent ECG conducted in interim  
o Review for possible drug-related adverse events of special interest 
o Need for supplemental oxygen on discharge (mode of delivery, and supplemental 
oxygen requirement) 
o Discharge disposition  
o If hospitalized, level of care (ICU, stepdown, standard)  
o WHO ordinal scale value (in conjunction with investigator)  
 
For patients enrolled at DUHS, all of this data will be automatically captured in Redcap 
as part of the overall COVID research plan, so the coordinator will not need to extract 
any data.  For patients at other sites, data will need to be entered into the Redcap 
database. 
 
Day 28 Assessment  
 
Study coordinator will complete CRF based on clinical data abstracted from chart as 
follows:  
 
If hospitalized:  
o Number of days since enrollment on which the participant had a measured 
temperature ≥38.0 C 
o Number of days since enrollment on which the participant was on mechanical 
ventilation 
o Number of days since enrollment on which the participant was on noninvasive 
ventilation 
o Number of days since enrollment on which the participant was on supplemental 
oxygen 
o Level of care (ICU, stepdown, standard)  
o WHO ordinal scale value (in conjunction with investigator)  

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If discharged:  
o Interim visit to emergency department or urgent care  
o Interim admission to hospital  
o Interim admission into ICU 
o Supplemental oxygen requirement  
o Level of long-term care required (skilled nursing facility, assisted living, home)  
o Functional status reported by outpatient provider (when available)  
o Recurrence of fever or respiratory symptoms  
o Lung examination reported by provider (when available)  
o Follow up lung imaging (when available)  
o WHO ordinal scale value (in conjunction with investigator)  
Day 45 Assessment: 
o Review of medical record to determine vital status and hospitalization status 
o Count of total hospital length of stay 
WHO Ordinal Scale  
 
The WHO Ordinal Scale for Clinical Improvement is an assessment developed by the 
World Health Organization as part of their COVID-19 therapeutic trial (World Health 
Organization. “WHO R&D Blueprint: Novel Coronavirus COVID-19 Therapeutic Trial 
Synopsis” 2020, Geneva, Switzerland URL: https://www.who.int/blueprint/priority-
diseases/key-action/COVID-
19_Treatment_Trial_Design_Master_Protocol_synopsis_Final_18022020.pdf. Accessed 
3/31/2020).  The assessment tool is intended to report on the clinical status of COVID-19 
patients in therapeutic trials on any given day. Details of the scale are below:  
Patient State
Descriptor 
Score 
Uninfected  
No clinical or virologic evidence of infection  
0 
Ambulatory 
No limitation of activities 
1
Limitation of activities 
2
Hospitalized, Mild Disease  
Hospitalized, no oxygen therapy  
3 
Oxygen by mask or nasal cannula 
4
Hospitalized, Severe Disease 
Non-invasive ventilation or high-flow oxygen  
5 
Intubation and mechanical ventilation
6
Mechanical ventilation + additional organ support 
(pressors, renal replacement therapy, ECMO) 
7 
Death  
Death  
8 
 
 
 
All designations of WHO ordinal scale will be made by the study PI or co-investigators.  
 
A study physician licensed to make medical diagnoses and listed on the delegation log 
will be responsible for all trial-related decisions.  
 
8.5 Concomitant Medications 
The use of all non-study drugs from enrollment until 14 days after enrollment will be monitored 
and recorded.  Per standard of care, all potential drug interactions with hydroxychloroquine and 
azithromycin will be reviewed prior to administration and discussed with the treating team.  If a 

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potential interaction between hydroxychloroquine and a concurrent medication is deemed 
potentially harmful by the investigator or treating team, a formal discussion of risks and benefits 
of hydroxychloroquine will take place between the investigator and the treating team.  If either 
the investigator or the treating team deem that the risk of the concurrent medication outweighs 
potential benefit, hydroxychloroquine will not be administered and the patient will be withdrawn 
from the study. 
8.6 Laboratory Evaluations 
Laboratory evaluations will be performed per standard of care under the direction of the treating 
team.   
ECG testing will be recommended as a standard of care intervention prior to and during study 
drug administration for participants with baseline cardiac conduction abnormalities or electrolyte 
abnormalities that might predispose to cardiac arrhythmias (see section 11.6). 
8.7 Loss to Follow Up 
Patients who are discharged from the hospital prior to 14 days after study enrollment will be 
contacted to ascertain their status on day 14 after enrollment.  Patients who cannot be 
contacted after at least three separate attempts will be assigned a World Health Organization 
ordinal scale based on health status at the time of hospital discharge. 
8.8 Management of Participants Who Discontinue Study Treatment 
Participants who prematurely discontinue study treatment will be followed as for other 
participants and will not receive any additional study treatment during the hospitalization.  
Participants assigned to receive two study drugs who only discontinue one drug should 
complete the full course of the other study drug unless there is a contraindication.    
If the study is terminated early, study treatment may be continued at the discretion of the 
investigator and treating team if it is deemed in the participant’s best interest. 
8.9 Collection of specimens for pharmacokinetic analysis 
While no specific specimens will be collected from study participants, we will request permission 
to use blood left over from routine laboratory blood draws for potential future 
pharmacokinetic/pharmacodynamic analyses.  Blood will either be stored at -80C or will be 
stored at room temperature as dried blood spots.  The future pharmacokinetic/phamacodynamic 
analyses would consist of measuring the concentrations of study drug (hydroxychloroquine and 
azithromycin) and correlating these concentrations with clinical outcomes. 
9  Study Schedule 
The schedule of study activities is summarized in Table 1 below. 

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Table 1.  Schedule of study activities. 
 
 
In addition to the activities outlined above, there will be a single evaluation at the time of hospital 
discharge to assess length of hospital stay and WHO ordinal scale, if discharge occurs later 
than 28 days after enrollment. 
10 Study Intervention/Investigational Drugs 
10.1 Study Drugs 
The study drugs are hydroxychloroquine and azithromycin.  Each of the study drugs is 
described in section 3. 
The study drugs will be combined into the following study treatment regimens: 
Arm 1 (control) Standard of care 
Arm 2: Hydroxychloroquine orally for 5 days 
Arm 3: Azithromycin orally for 5 days 
 
Screen 
Baseline
Treatment 
Period  
End of 
Treatment 
Inpatient  
Follow-up 
Day 28 follow-
up  
Day +/- Window -1 or 1  
1 
1 to 5  
5 
6-14 
28 ± 2 
ELIGIBILITY  
Demographics & Medical History 
Review
X 
 
 
 
 
 
Review SARS CoV-2 results 
X
Hospital Course Review 
X
Clinical Data Review (including 
EKG for QTc screen) 
X 
 
 
 
 
 
Informed consent 
X
STUDY INTERVENTION  
Randomization 
X
Study medication administration
X
STUDY PROCEDURES  
Vital signs including SpO2
X
Daily
Daily 
Clinical data collection 
X
Daily
X
Daily
Review of laboratory/ECG data
X
Daily
Daily
Targeted medication review 
X
Daily
Daily
Adverse event evaluation 
X
Daily
Daily
X
WHO ordinal scale adjudication 
X
Daily 
X
Daily 
X
EHR review for re-admission or 
death 
 
 
 
 
 
X 
SAFETY INVESTIGATIONS 
Pregnancy test for females of 
childbearing potential (if no clear 
clinical evidence/history of 
pregnancy) 
 
X 
 
 
 
 

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Arm 4: Hydroxychloroquine orally for 5 days plus azithromycin orally for 5 days 
10.2 Study Drug Acquisition 
As this is a pragmatic trial using readily available FDA-approved medications, study drug will be 
supplied by the pharmacy of the hospital where the patient is receiving treatment. 
10.3 Administration and Dosage of Study Drugs 
Hydroxychloroquine will be administered orally or via feeding tube at a dosage of 800 mg on 
day 1, followed by 600 mg daily on days 2-5. 
 
Patients who do not tolerate the 600 mg daily dose may be dose-reduced to a 400 mg 
daily dose at the discretion of the study investigator 
Azithromycin will be administered orally or via feeding tube at a dosage of 500 mg on day 1, 
followed by 250 mg daily on days 2-5.  If oral azithromycin is not available, intravenous 
azithromycin at the same dosages may be substituted.  
10.4 Criteria for Discontinuation of Study Drugs 
Participants who meet any one or more of the following criteria will be discontinued from study 
treatment: 
 
Participant request for premature discontinuation of study treatment 
 
Any clinical adverse event, laboratory abnormality, or other medical condition occurs 
such that continued administration of study treatment is deemed not in the best interest 
of the patient 
11 Assessment of Safety 
11.1  Definition of Adverse Events 
An adverse event (AE) is any unfavorable and unintended sign (including an abnormal 
laboratory finding), symptom, or disease that occurs to a study participant during the course of 
the study.  
Adverse Event Grading 
 
Mild (Grade 1): Transient or mild symptoms; no limitation in activity; no intervention 
required. The AE does not interfere with the participant’s normal functioning level.   
 
 
Moderate (Grade 2): Symptom results in mild to moderate limitation in activity; no or 
minimal intervention required. The AE produces some impairment of functioning, but it is 
not hazardous to health.   
 
 
Severe (Grade 3): Symptom results in significant limitation in activity; medical 
intervention may be required. The AE produces significant impairment of functioning or 
incapacitation. 
 
 
Potentially life-threatening (Grade 4): Extreme limitation in activity, significant assistance 
required, significant medical intervention or therapy required, or hospitalization required. 
 
[Note: ‘Life-threatening’ as a Grade 4 severity refers to a ‘potential’ threat to life whereas ‘life 

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threatening’ as a ‘serious adverse event’ refer to an ‘immediate’ threat to life.] 
11.2  Definition of Serious Adverse Events  
 
An AE is considered to be ‘serious’ if it results in one of the following outcomes. An AE 
needs to meet only one of the above criteria to be considered serious.  
 
Death, 
 
Life-threatening event (the participant is at immediate risk of death), 
 
Requires inpatient hospitalization or prolongation of existing hospitalization 
 
Results in persistent or significant disability/incapacity (a substantial disruption of a 
person's ability to conduct normal life functions),  
 
Results in congenital anomaly/birth defect 
 
Important medical event that may not be immediately life-threatening or results in 
death or hospitalization but may jeopardize the participant or may require 
intervention to prevent one of the other outcomes listed in the definition above.  
11.3  Assessment of Adverse Events 
Both hydroxychloroquine and azithromycin have a track record of safety since their FDA-
approval in 1955 and 2002, respectively. There is extensive experience with these drugs in 
clinical practice. However, because the medicines have not been used in COVID-19 patients, 
and the use of hydroxychloroquine and azithromycin in combination can increase the risk for 
major cardiac toxicity including QT prolongation, only drug-specific AEs that are listed in the 
FDA Warning List for the study drugs and only AEs Grade 3 or higher will be collected.  
Hydroxychloroquine has been used for decades for the treatment and prevention of malaria 
and for autoimmune conditions including rheumatoid arthritis and lupus. The following system 
abnormalities have been listed under the FDA Warning List for Hydroxychloroquine (mostly with 
long-term use for autoimmune conditions) 
Hypoglycemia: may be severe necessitates close serum glucose monitoring. 
Dermatologic: Erythema multiforme, may precipitate a severe attack of psoriasis and porphyria 
Nervous system: Dizziness, vertigo, tinnitus, proximal myopathy and muscle weakness, 
extrapyramidal symptoms, seizure, and suicidal behavior. Close observation in patients with 
neurological or seizure disorders.  
Ocular: irreversible retinal damage has occurred with only with prolonged use (> 5 years) and in 
individuals with subnormal glomerular filtration, and those with concurrent macular disease (up 
to 7.5%) 
Cardiac: risk of QT prolongation and torsades de pointes, AV block, and ventricular arrythemia. 
Risk is increased in in patients with underlying QT-prolongation and/or on concurrent QT-
prolonging agents.  
G6PD deficiency: Although warning exists regarding the potential hemolysis in G6PD-deficient 
patients, studies indicate the risk is very low for hydroxychloroquine and treatment can be 
initiated while awaiting G6PD test results. 
Risk of fetal toxicity: no increase in rate of birth defects has been reported; however as the data 
are considered limited, birth outcomes will be collected at the end of pregnancy. 
Drug-drug interactions: See University of Liverpool drug interaction checkers: 
http://www.covid19-druginteractions.org 

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The following drugs should not be co-administered: amiodarone, bepridil, mexiletine, rifampicin, 
and rifapentine 
The following drugs may require dose monitoring, alteration of dosage or timing of 
coadministration: digoxin, propofol, sevoflurane, tizanidine, hydrocodone, methadone, 
disopyramide, dofelitide, propafenone, quinidine, azithromycin, clarithromycin, erythromycin, 
levofloxacin, moxifloxacin, ofloxacin, clofazimine, delamanid, bedaquiline, pyrazinamide, 
telithromycin, mefloquine 
Use in children: safety has not been established in children. Fatalities have been reported 
following accidental ingestion of chloroquine but not of hydroxychloroquine. 
Azithromycin has been used extensively for treatment of acute bacterial sinusitis, traveler 
diarrhea, and community-acquired pneumonia. The following system abnormalities have been 
listed under the FDA Warning List for arithromycin: 
Hypersensitivity: angioedema, anaphylaxis, and dermatologic reactions including Stevens 
Johnson Syndrome and toxic epidermal necrolysis have been reported rarely 
Cardiac: potential risk of QT prolongation and torsades de pointes have been seen, usually in 
settings of proarrhythmic conditions such as hypokalemia, hypomagnesemia, bradycardia, or 
those receiving hydroxychloroquine, or antiarrhythmic drugs (quinidine, procainamide, defetilide, 
amiodarone, sotalol), Risk is also higher in elderly patients. Monitoring for QT prolongation, 
particularly in setting of concomitant drugs with risk of QT prolongation.  
Hepatotoxicity: abnormal liver function, hepatitis, cholestatic jaundice, hepatic necrosis, and 
hepatic failure have been reported 
Clostridium difficile-associated diarrhea: this risk is associated with all antibiotic use 
Risk of adverse pregnancy outcome: A recent study suggested that macrolides may confer a 
higher risk of birth defects compared to penicillins according to a recent study (51). This study 
compared 100,000 women who had either a single course of a macrolide antibiotic or a 
penicillin during gestation. The prevalence of major congenital malformations (primarily 
cardiovascular) associated with first trimester exposure was 28 per 1000 live births compared to 
18 in the penicillin group. Second and third trimester prescriptions were not associated with 
malformations. Macrolide prescriptions in any trimester were associated with genital 
malformations (mainly hypospadias) in 5 vs 3 per 1000 live births. The absolute numbers of 
azithromycin compared to other macrolides (clarithromycin and erythromycin) were too small to 
analyze separately and so it is not clear that the conclusions apply to azithromycin specifically.  
 
 
11.4  Adverse Event Reporting 
Non-serious adverse events will not be collected with the exception of adverse events of special 
interest possibly related to hydroxychloroquine, which are as follows: 
 
Arrhythmias (ventricular) 
 
Hepatic Failure 
 
Bone marrow failure 
 
Aplastic anemia 
 
Prolonged QT interval 

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 
Angioedema 
 
Exfoliative dermatitis  
 
Acute generalized exanthematous pustulosis (AGEP) 
 
Psychosis 
 
Suicidal Ideation 
 
Seizure 
Serious adverse events which occur between initial dose of study medication and day 14 will be 
recorded in the electronic case report form. These events, along with the adverse events of 
special interest listed above, will be entered into the study database and provided to the Data 
and Safety Monitoring Board upon safety review as required.  
Hydroxychloroquine and azithromycin are FDA-approved medicines being used at standard 
dosing. Reporting to the IRB will occur in a summary format after each Data and Safety 
Monitoring Board review,  
11.5  Reporting of pregnancy outcome 
For participants who are pregnant women, we will obtain permission to review the medical 
record of both the participant and the infant to assess for any complications of the pregnancy 
and the outcome of the pregnancy.  Specific pregnancy-associated outcomes that will be 
collected are as follows: 
 
miscarriage in the first trimester 
 
any fetal deaths and gestational age 
 
spontaneous preterm birth 
 
premature rupture of membranes 
 
fetal growth restriction 
 
gestational diabetes 
 
indicated preterm birth 
 
preeclampsia and severity 
 
gestational age at delivery 
 
mode of delivery 
 
neonatal birth weight 
 
fetal malformation 
 
APGAR scores  
 
neonatal length of stay 
 
Neonatal intensive care unit admission 
 
Neonatal death 
 
11.6  Management of adverse events 
For adverse events that in the investigator’s judgment may be due to study drugs, one of the 
protocol investigators will discuss the adverse event with the treating physician, and a joint 
decision will be made whether to discontinue study therapy.  If study therapy has already been 
completed at the time of the adverse event, the patient will be followed per protocol by the study 
team to assess for resolution of the adverse event.  If resolution of the adverse event has not 
occurred by the day 14 assessment, the adverse event will be reassessed at the day 28 
assessment to determine whether resolution has occurred. 

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Participants who experience adverse events requiring temporary or permanent discontinuation 
of study drug will still considered to be part of the study and will be followed as outlined above. 
11.6.1  Monitoring and management of QTc interval prolongation 
The management will be according to the recommendations of the Duke COVID-19 Therapeutic 
Group as follows: 
All patients who are randomized to hydroxychloroquine and/or azithromycin will be placed on 
telemetry 
After 2nd dose of study drug (approx. >24h), repeat ECG 
a. If QTc change is <50ms, continue telemetry, no further ECGs 
b. If QTc change >50ms or QTc>500, the patient is considered a ‘QT reactor’: continue 
telemetry, mitigate QT prolonging factors: including correcting any K+ or Mg+ abnormality 
and stop azithromycin (if patient is in the azithromycin group), repeat ECG in 24h 
c. If further prolonged, or remains >500ms, stop hydroxychloroquine 
11.7  Data and Safety Monitoring Board 
A data and safety monitoring board (DSMB) will be assembled consisting of three individuals 
with complementary expertise.  At least one member will have expertise in infectious diseases 
or critical care, and at least one member will have expertise in clinical trials.  DSMB members 
will not otherwise be involved in the study.  The DSMB will be apprised of ongoing study results 
and will meet approximately monthly, or more often as needed. 
11.8  Interim Monitoring and Analyses by the Data and Safety Monitoring Board 
Evaluation of potential adverse events and tolerability will be performed as close to “real time” 
as possible by the study team and will be regularly reviewed by the DSMB.  As the planned 
Bayesian analysis permits “real-time” looks at the efficacy results without adjustment, an interim 
efficacy analysis will be available at each DSMB meeting.  The DSMB may recommend early 
closure of either experimental arm or the trial if, in their judgment, interim evidence is sufficiently 
strong that one of the trial interventions is clearly indicated/contraindicated because of 
differences in safety and/or efficacy.  The interim results will be presented as described in 
section 12.3. 
12 Statistical Considerations 
12.1 Study Hypotheses 
 
1) The World Health Organization ordinal scale measured at day 14 after enrollment will be 
higher (better) in participants receiving hydroxychloroquine than in participants who do not 
receive hydroxychloroquine. 
2) The World Health Organization ordinal scale measured at day 14 after enrollment will be 
higher (better) in participants receiving azithromycin than in participants who do not receive 
azithromycin. 
3) The proportion of participants not on mechanical ventilation at baseline who progress to 
requiring mechanical ventilation during the index hospitalization will be lower in participants 
receiving hydroxychloroquine than in participants who do not receive hydroxychloroquine. 

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4) The proportion of participants not on mechanical ventilation at baseline who progress to 
requiring mechanical ventilation during the index hospitalization will be lower in participants 
receiving azithromycin than in participants who do not receive azithromycin 
5) The World Health Organization ordinal scale measured at day 28 after enrollment will be 
higher (better) in participants receiving hydroxychloroquine than in participants who do not 
receive hydroxychloroquine. 
6)  The World Health Organization ordinal scale measured at day 28 after enrollment will be 
higher (better) in participants receiving azithromycin than in participants who do not receive 
azithromycin. 
7) The median length of hospitalization during the index hospitalization will be lower among 
participants receiving hydroxychloroquine than in participants who do not receive 
hydroxychloroquine. 
8) The median length of hospitalization during the index hospitalization will be lower among 
participants receiving azithromycin than in participants who do not receive azithromycin. 
12.2  Analysis Groups 
Intention to treat (ITT): Includes all participants who receive a treatment assignment 
Modified intention to treat (mITT): Includes all participants who received at least one dose of 
study medication 
Per protocol (PP): Includes all participants who received all assigned study medication 
12.3  Analysis Plan 
All major treatment group comparisons will be performed according to the principle of "intent-to-
treat;" that is, participants will be analyzed (and outcomes attributed) according to the 
randomized treatment, regardless of actual treatments received. The primary analysis of all 
primary and secondary endpoints will be performed in the modified intent-to-treat population. 
Treatment group comparisons will involve the following randomized treatment groups: 
 
A. Standard of care plus hydroxychloroquine 
   A1. Standard of care + hydroxychloroquine + sub-randomization to azithromycin 
   A2. Standard of care + hydroxychloroquine + sub-randomization to no azithromycin 
   A3. Standard of care + hydroxychloroquine, not eligible for azithromycin sub-randomization 
B. Standard of care without hydroxychloroquine 
   B1. Standard of care + azithromycin 
   B2. Standard of care + no azithromycin 
   B3. Standard of care, not eligible for azithromycin sub-randomization 
 
The two major treatment group comparisons to be performed are:   
 
Comparison #1: Hydroxychloroquine versus no hydroxychloroquine.  
o Aggregate of (A1 + A2 + A3) versus (B1 + B2 + B3) 
 
Comparison #2: Azithromycin versus no azithromycin.  
o Aggregate of (A1 + B1) versus (A2 + B2) 

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12.3.1.  Primary efficacy analysis 
The primary analysis of the primary efficacy endpoint, the World Health Organization scale 
measured on day 14 after enrollment, will be performed using the proportional odds model for 
ordered categorical data.  The proportional odds methodology is an extension of binary logistic 
regression to accommodate 𝑘> 2 ordered outcome categories. The model considers all 
possible ways of dichotomizing 𝑘 categories into a binary outcome and assumes that the set of 
all possible binary outcomes is related to covariates through a set of ordinary binary logistic 
regression models. Treatment group differences in the proportional odds model are expressed 
as odds ratios with an odds ratio less than 1 suggesting that treatment is better than the control 
and an odds ratio greater than 1 suggesting that treatment is worse than the control. The 
magnitude of the odds ratio describes the extent to which treated and untreated patients tend to 
have outcomes shifted toward better or worse ranking categories. To increase power and 
enhance interpretability, the proportional odds model may be adjusted for a set of relevant 
covariates, such as enrolling site and ventilator status at baseline. Covariates will be carefully 
selected and defined in the trial's Statistical Analysis Plan document. The comparison of 
hydroxychloroquine versus no hydroxychloroquine will be further adjusted for azithromycin 
assignment and the comparison of azithromycin versus no azithromycin will be adjusted for 
hydroxychloroquine assignment.  Interaction (effect modification) between hydroxychloroquine 
and azithromycin will be specifically examined, and estimates of the potential magnitude and 
credible intervals around that interaction term will be generated.  
Parameters of the proportional odds model will be estimated in a Bayesian statistical framework 
by specifying a prior probability distribution for unknown parameters. The output of a Bayesian 
analysis is a posterior probability distribution describing the relative likelihood of different 
numerical estimates for the unknown parameter values. This posterior distribution can be used 
to determine the likelihood of a clinically important treatment benefit or harm in light of the study 
data. The Bayesian approach provides an especially useful perspective for interpreting results 
of small studies that are likely to be accompanied by substantial statistical uncertainty. The 
Bayesian approach allows decisions to be made based on the estimated probability of a 
clinically important treatment benefit, as opposed to an arbitrary dichotomization of results as 
"significant" or "non-significant." 
12.3.2  Secondary efficacy analyses 
Mechanical ventilation.  
The proportion of participants not on mechanical ventilation at baseline who progress to 
requiring mechanical ventilation will be compared for patients belonging to the following 
treatment groups as defined above at the beginning of section 12.3: 
1) Hydroxychloroquine versus no hydroxychloroquine. 
2) Azithromycin versus no azithromycin.  
The comparison will be performed using logistic regression and will be adjusted for the same 
covariates as the primary endpoint proportional odds model.  
World Health Organization ordinal scale measured at day 28 
The World Health Organization ordinal scale at day 28 will be compared for the following 
treatment groups as defined above at the beginning of section 12.3:  

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1) Hydroxychloroquine versus no hydroxychloroquine. 
2) Azithromycin versus no azithromycin. 
 
Analysis will be based on the proportional odds model and will be similar to the analysis of the 
primary endpoint as described above.  
Length of hospitalization 
The distribution of days from randomization to hospital discharge will be compared for the 
following treatment groups as defined above at the beginning of section 12.3: 
1) Hydroxychloroquine versus no hydroxychloroquine. 
2) Azithromycin versus no azithromycin. 
 
The comparison will be based on a semi-parametric or rank based analysis. For participants 
who die during the enrollment hospitalization, the time from randomization to discharge will be 
regarded as infinite.  
12.3.3  Safety analyses 
There will be two comparisons for the primary safety analysis; both will consider the modified 
intention to treat population during the first fourteen days after enrollment.  Comparison 1 will 
compare the rate of severe adverse events in participants who received hydroxychloroquine 
versus the rate in participants who did not receive hydroxychloroquine.  Comparison 2 will 
compare the rate of severe adverse events in participants who received azithromycin versus the 
rate in participants who did not receive azithromycin.  An experimental arm will be deemed less 
safe than the standard of care arm if the observed rate of severe adverse events is greater than 
the control arm and the 95% credible interval (using a neutral prior distributions) around the 
difference in rates excludes zero. 
The secondary safety analyses will examine the rates of all-cause study drug 
(hydroxychloroquine and azithromycin) discontinuation in the modified intention to treat 
population, and will describe them as point estimates and 95% credible intervals (using a 
neutral prior distribution). 
12.4 Sample Size Considerations 
Table 2 summarizes frequentist considerations of sample size, based on comparison of the 
primary outcome between the group of participants who received hydroxychloroquine and those 
who did not receive hydroxychloroquine.  This assumes that the aggregate probabilities of all 
trial participants to have a day 14 World Health Organization ordinal scale score of 1-8 are as 
follows: Death 5%, mechanical ventilation/ECMO 10%, noninvasive ventilation/high flow oxygen 
15%, hospitalized on supplemental oxygen 35%, hospitalized not requiring supplemental 
oxygen 5%, hospitalized not requiring ongoing medical care 5%, not hospitalized, limitation on 
activities and/or requiring home oxygen 5%, not hospitalized, no limitations on activities 20%. 
Table 2.  Frequentist estimates of power with given odds/win ratios and sample sizes. 
Two-Sided
Odds
Win
Power  (Two-Group Trial With Total Sample Size N)
Alpha
Ratio
Ratio
N = 200
N = 300
N = 400
N = 500

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0.05
0.75
1.26
20%
29%
36%
44%
0.70
1.33
29%
41%
51%
61%
0.65
1.41
40%
55%
67%
77%
0.60
1.50
52%
70%
82%
89%
0.55
1.61
66%
83%
92%
96%
0.50
1.73
78%
92%
97%
99%
0.45
1.89
88%
97%
99%
>99%
0.40
2.07
95%
99%
>99%
>99%
0.10
0.75
1.26
31%
40%
49%
56%
0.70
1.33
41%
53%
64%
72%
0.65
1.41
52%
67%
78%
85%
0.60
1.50
64%
80%
89%
94%
0.55
1.61
76%
89%
96%
98%
0.50
1.73
86%
96%
99%
>99%
0.45
1.89
93%
99%
>99%
>99%
0.40
2.07
98%
>99%
>99%
>99%
 
The table above is based on the Wilcoxon rank sum test. Group-specific probabilities are 
assumed to be described by a proportional odds model i.e. randomization to 
hydroxychloroquine decreases the odds of being in one of the k worst outcome categories and 
the same odds ratio applies to all possible choices of k. When OR=0.5 the group-specific 
probabilities are HCQ: 3.41%, 7.28%, 12.14%, 34.39%, 5.61%, 5.79%, 5.95%, 25.43% and 
standard care: 6.59%, 12.72%, 17.86%, 35.61%, 4.39%, 4.21%, 4.05%, 14.57%. The Win Ratio 
implied by these two sets of probabilities is 1.73. 
 
To examine sample size from a Bayesian perspective, we used Monte Carlo simulations to 
study trial operating characteristics under a range of hypothetical true treatment effects. The 
operating characteristics of interest were frequentist power, type-I error, and expected study 
duration. Simulations are based on a two-arm trial that enrolls patients until declaring the 
treatment of interest to be efficacious, stopping for evidence of harm or inefficacy, or reaching a 
pre-specified maximum sample size, which we assumed to be N = 200, 300, 400, or 500. The 
primary endpoint is the WHO 8-level ordinal scale and the primary measure of treatment effect 
is the win ratio. Treatment group differences were assessed by fitting a proportional odds model 
with the WHO scale as an ordinal outcome and treatment group as a covariate. Properties of a 
fully Bayesian analysis were approximated by assuming that the maximum likelihood estimate 
of the treatment effect coefficient was normally distributed. The prior distribution assigned 2.5% 
probability to the hypothesis of a true odds ratio below 0.5 (highly efficacious) and 2.5% 
probability to the hypothesis of a true odds ratio exceeding 2.0 (highly harmful). In each 
simulated trial we assumed that interim analyses were performed sequentially after each patient 
beginning after collecting data from the first 20 patients. Signals for harm and efficacy were 
defined as follows:  
 
Moderate efficacy: The win ratio exceeds 1.25 with at least 80% Bayesian probability 

Pragmatic trial of hydroxychloroquine/azithromycin for COVID 
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 
Any efficacy: The win ratio exceeds 1.00 with at least 95% Bayesian probability 
 
Moderate harm: The win ratio is less than 0.90 with at least 75% Bayesian probability 
 
Inefficacy/any Harm: The win ratio is less than 1.00 with at least 80% Bayesian 
probability 
For simplicity, the simulation assumes that each patient's outcome is known before the next 
patient is randomized. In each simulated trial, the true probabilities for the 8 WHO categories in 
aggregate across two treatment groups were assumed to be 5%, 10%, 15%, 35%, 5%, 5%, 5%, 
20%. Data were generated under scenarios with true odds ratios ranging from 0.4 (benefit) to 
2.0 (harm). We simulated 500 trials per odds ratio scenario and estimated operating 
characteristics by averaging across simulated trial results.  
 
 
 
 
 
Table 3. Probability of reaching threshold for moderate or any efficacy as function of true 
odds ratio if enrollment is capped at 200, 300, 400, or 500 patients.  
True
Max N = 200
Max N = 300
Max N = 400
Max N = 500
Odds 
Ratio 
Moderate
Any
Moderate
Any
Moderate
Any
Moderate
Any
Power
OR = 0.75
19%
27%
27%
43%
35%
55%
40%
66%
OR = 0.70
24%
36%
39%
55%
46%
71%
55%
80%
OR = 0.65
41%
51%
56%
70%
65%
81%
71%
88%
OR = 0.60
52%
64%
70%
84%
80%
93%
87%
96%
OR = 0.55
62%
71%
81%
90%
89%
96%
93%
99%
OR = 0.50
76%
84%
90%
96%
96%
99%
99%
>99%
OR = 0.45
86%
93%
95%
98%
98%
99%
>99%
>99%
OR = 0.40
93%
96%
99%
99%
99%
>99%
>99%
>99%
Type-I 
Error 
OR = 1.00
4%
8%
5%
10%
5%
13%
6%
14%
OR = 1.05
4%
5%
5%
7%
5%
8%
5%
9%
OR = 1.10
1%
2%
2%
4%
2%
4%
2%
4%
OR = 1.15
1%
1%
1%
1%
1%
2%
1%
2%
OR = 1.25
1%
1%
1%
1%
1%
1%
1%
1%
OR = 1.50
<1%
<1%
<1%
<1%
<1%
<1%
<1%
<1%
OR = 2.00
<1%
<1%
<1%
<1%
<1%
<1%
<1%
<1%
 
 
 

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Table 4. Probability of reaching threshold for inefficacy or harm as function of true odds 
ratio if enrollment is capped at 200, 300, 400, or 500 patients.  
True
Max N = 200
Max N = 300
Max N = 400
Max N = 500
Odds 
Ratio 
Harm
Inefficacy
Harm
Inefficacy
Harm
Inefficacy
Harm
Inefficacy
Power
OR = 1.00
22%
37%
28%
43%
31%
47%
33%
49%
OR = 1.05
32%
45%
37%
52%
40%
55%
42%
59%
OR = 1.10
32%
45%
39%
55%
43%
61%
47%
65%
OR = 1.15
43%
56%
49%
64%
54%
71%
58%
77%
OR = 1.25
55%
67%
65%
76%
70%
81%
74%
87%
OR = 1.50
73%
84%
85%
93%
91%
96%
94%
98%
OR = 2.00
97%
98%
>99%
>99%
>99%
>99%
>99%
>99%
Type-II 
Error 
OR = 0.75
6%
10%
6%
11%
6%
11%
6%
11%
OR = 0.70
4%
8%
4%
8%
4%
8%
4%
8%
OR = 0.65
2%
5%
2%
5%
2%
5%
2%
5%
OR = 0.60
1%
2%
1%
2%
1%
2%
1%
2%
OR = 0.55
1%
3%
1%
3%
1%
3%
1%
3%
OR = 0.50
<1%
1%
<1%
1%
<1%
1%
<1%
1%
OR = 0.45
<1%
1%
<1%
1%
<1%
1%
<1%
1%
OR = 0.40
<1%
<1%
<1%
<1%
<1%
<1%
<1%
<1%
 
Table 5. Expected looks until reaching threshold for moderate efficacy or reaching 
enrollment cap as function of true odds ratio 
Odds Ratio
Max N = 200
Max N = 300
Max N = 400
Max N = 500
OR = 0.75
182
259
327
390
OR = 0.70
181
249
306
355
OR = 0.65
167
218
257
289
OR = 0.60
155
193
218
234
OR = 0.55
145
173
187
197
OR = 0.50
133
150
156
158
OR = 0.45
115
124
127
128
OR = 0.40
101
104
105
105
 
 

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Table 6. Expected looks until stopping for futility or harm or reaching enrollment cap as 
function of true odds ratio 
Odds Ratio
Max N = 200
Max N = 300
Max N = 400
Max N = 500
OR = 1.00
159
219
274
326
OR = 1.05
149
200
246
289
OR = 1.10
146
196
238
275
OR = 1.15
131
170
203
228
OR = 1.25
117
146
168
184
OR = 1.50
96
107
113
116
OR = 2.00
59
60
60
60
 
12.5  Interim Analysis 
For ethical reasons, interim examination of clinical endpoints and key safety events will be 
performed at regular intervals (approximately monthly) during the course of the trial. An 
independent Data and Safety Monitoring Board (DSMB) will be appointed to monitor participant 
safety and to review overall trial performance. The primary objective of these interim analyses 
will be to ensure the safety of trial participants and evaluate the accumulating evidence for 
safety and efficacy. In addition, interim monitoring will involve a review of participant 
recruitment, compliance with the study protocol, status of data collection, and other factors 
which reflect the overall progress and integrity of the study.   
Interim efficacy analyses will focus on the primary endpoint (World Health Organization ordinal 
scale) and will be performed both for the comparison of hydroxychloroquine versus no 
hydroxychloroquine and for the comparison of azithromycin versus no azithromycin. At each 
interim analysis, we will use the Bayesian proportional odds model to evaluate the probabilities 
for each of the following hypotheses in light of accruing study data: 
Hydroxychloroquine 
 
Moderate efficacy. 
o Defined as an odds ratio of 0.80 or less favoring treatment hydroxychloroquine 
over no hydroxychloroquine 
 
Any efficacy 
o Defined as an odds ratio of less than 1.0, favoring treatment hydroxychloroquine 
over no hydroxychloroquine 
 
Inefficacy 
o Defined as an odds ratio greater than 1.0, favoring treatment no 
hydroxychloroquine over hydroxychloroquine 
 
Moderate harm 
o Defined as an odds ratio greater than 1.1, favoring treatment no 
hydroxychloroquine over hydroxychloroquine 
 
Azithromycin 
 
Moderate efficacy. 

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o Defined as an odds ratio of 0.80 or less favoring treatment azithromycin over no 
azithromycin 
 
Any efficacy 
o Defined as an odds ratio of less than 1.0, favoring treatment azithromycin over 
no azithromycin 
 
Inefficacy 
o Defined as an odds ratio greater than 1.0, favoring treatment no azithromycin 
over azithromycin 
 
Moderate harm 
o Defined as an odds ratio greater than 1.1, favoring treatment no azithromycin 
over azithromycin 
 
The following events will be regarded as signals for review and consideration of early stopping.  
 
Probability of at least moderate efficacy is >80%.  
 
Probability of any efficacy is >95% 
 
Probability of inefficacy is >80% 
 
Probability of moderate harm is >75%  
 
If any of the above signals are triggered, the DSMB will be notified and enrollment will be 
temporarily suspended while the DSMB considers whether to recommend continuing or 
terminating the study. 
It is important to emphasize the triggers listed above are intended as a guide for interpreting the 
interim analyses and not as a strict rule for early termination.  The DSMB will consider a variety 
of results, including efficacy analyses of secondary endpoints, results from other ongoing and 
recently completed studies, and all aspects of safety.  
12.6 Multiplicity Considerations 
With the various primary and secondary endpoints, and two major treatment group 
comparisons, we recognize that there is a multiplicity of analyses to be performed, which leads 
to an increased probability that at least one of the comparisons could be statistically "significant" 
by chance. The Bayesian statistical framework addresses multiplicity by focusing on the 
probability that a treatment is beneficial or harmful as opposed to the probability of observing a 
significant difference if the treatment effect is literally zero. If the prior distribution is chosen to 
be a faithful reflection of the prior evidence in favor or against a clinically important treatment 
benefit, then posterior probabilities are an appropriate reflection of the posterior evidence and 
can be used for decision making without the need for an ad hoc multiplicity adjustment. 
Importantly, the results of the trial will not be explicitly tied to a labeling decision or 
recommendation for practice but will be reported in an objective manner in order to provide 
evidence to be used by multiple stakeholders. Although the analysis will not be explicitly 
adjusted to account for multiple comparisons, we will be appropriately conservative in the 
interpretation of these multiple comparisons, taking into account the magnitude of observed 
differences and looking for consistency across endpoints and subgroups. Publications 
presenting trial results will present a balanced interpretation and will call attention to the 
potential for results to be impacted by random sampling variation.  

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13 Protection of Human Subjects 
This study will be conducted in conformity with the ethical standards set out in the latest version 
of the Declaration of Helsinki.  
 
13.1  Institutional Review Board 
Each participating institution will provide for the review and approval of this protocol and the 
associated informed consent documents by an appropriate institutional ethics committee (IEC) 
or Institutional Review Board (IRB). Any amendments to the protocol or consent materials must 
also be approved before they are placed into use. 
 
13.2  Informed consent process 
Only individuals who provide written informed consent, or in the absence of ability to provide 
consent have consent provided by their appointed medical decision-maker, will be enrolled in 
this study.  Written informed consent is required before any study-specific procedures or receipt 
of study medications. Potential participants and/or their medical decision makers will have the 
conditions of the study explained to them, including potential harms and benefits, the nature and 
timing of study procedures, alternatives to study participation, that study participation is 
voluntary, that a decision to not participate in the study will not affect the quality of their future 
medical care, and that they may withdraw from participation at any time.  Literate individuals will 
be provided with a language-appropriate document to read; illiterate individuals (i.e. individuals 
who speak and understand, but do not read and write, the language in which the consent 
discussion is conducted) will have the contents of the document explained to them by a trained 
study staff member; such individuals can be enrolled by ‘making their mark’ on the consent 
document.  Potential participants will have the opportunity to ask questions of the site 
investigator or delegate, and to discuss participation with their family and/or friends or think 
about the study prior to deciding whether or not to participate.  A copy of the signed informed 
consent document will be given to the participant for his/her records.  
 
13.3  Participant confidentiality 
All records identifying the participant will be kept confidential and, to the extent permitted by the 
applicable laws and regulations, will not be made publicly available without sufficient de-
identification procedures.  
 
All paper study records will be stored in a locked office and electronic study records will be 
stored on password-protected computers; only designated trained study staff will have access to 
study records.   
 
Clinical data will be maintained in a secure, HIPAA compliant, password-protected RedCap 
database managed by the Duke Office of Clinical Research (DOCR) Research Management 
Team. The database resides on Duke Health Technology Services (DHTS) managed servers 
behind a Duke firewall and is accessible only by designated study staff with a password 
protected account.  
 
Study monitors and/or regulatory authorities may inspect all documents and records required to 
be maintained by the Investigator, including but not limited to, medical records, primary 
laboratory data, and pharmacy records for study participants; this information will be provided to 

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participants during the Informed Consent process.  The clinical study site will permit access to 
such records. 
 
13.4  Study discontinuation 
The Data and Safety Monitoring Board may recommend study termination, termination of a 
study arm, or termination of a study site. If the study is closed or enrollment is terminated at a 
site, the DUHS Institutional Review Board will be notified.  Participants experiencing an adverse 
event at the time of early termination should be followed until resolution or stabilization of the 
event.  Participants who were receiving study treatment at the time of early termination will 
continue care per the preferences of their treating medical providers.  
 
14 Data Handling and Record Keeping 
Information from the medical record will be directly downloaded into a secure RedCap 
database.  The directly downloaded data will be supplemented with electronic medical record 
review by one of the investigators, the results of which will also be entered into the RedCap 
database.  After study completion, the secure data will be retained for six years per DUHS 
policy. 
15 Publications and Dissemination of Study Results 
Study results will be analyzed and published promptly after approval by all investigators.  No 
identifying information regarding individual participants will be included in any publication. 
 
 

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REFERENCES 
- 
1. 
Huang C, Wang Y, Li X, Ren L, Zhao J, Hu Y, et al. Clinical features of patients 
infected with 2019 novel coronavirus in Wuhan, China. Lancet. 2020;395(10223):497-
506. 
- 
2. 
Rainsford KD, Parke AL, Clifford-Rashotte M, Kean WF. Therapy and 
pharmacological properties of hydroxychloroquine and chloroquine in treatment of 
systemic lupus erythematosus, rheumatoid arthritis and related diseases. 
Inflammopharmacology. 2015;23(5):231-69. 
- 
3. 
Sahraei Z, Shabani M, Shokouhi S, Saffaei A. Aminoquinolines Against 
Coronavirus Disease 2019 (COVID-19): Chloroquine or Hydroxychloroquine. Int J 
Antimicrob Agents. 2020:105945. 
- 
4. 
Devaux CA, Rolain JM, Colson P, Raoult D. New insights on the antiviral effects 
of chloroquine against coronavirus: what to expect for COVID-19? Int J Antimicrob 
Agents. 2020:105938. 
- 
5. 
Vincent MJ, Bergeron E, Benjannet S, Erickson BR, Rollin PE, Ksiazek TG, et al. 
Chloroquine is a potent inhibitor of SARS coronavirus infection and spread. Virol J. 
2005;2:69. 
- 
6. 
Olofsson S, Kumlin U, Dimock K, Arnberg N. Avian influenza and sialic acid 
receptors: more than meets the eye? Lancet Infect Dis. 2005;5(3):184-8. 
- 
7. 
Yang ZY, Huang Y, Ganesh L, Leung K, Kong WP, Schwartz O, et al. pH-
dependent entry of severe acute respiratory syndrome coronavirus is mediated by the 
spike glycoprotein and enhanced by dendritic cell transfer through DC-SIGN. J Virol. 
2004;78(11):5642-50. 
- 
8. 
Savarino A, Lucia MB, Rastrelli E, Rutella S, Golotta C, Morra E, et al. Anti-HIV 
effects of chloroquine: inhibition of viral particle glycosylation and synergism with 
protease inhibitors. J Acquir Immune Defic Syndr. 2004;35(3):223-32. 
- 
9. 
Klumperman J, Locker JK, Meijer A, Horzinek MC, Geuze HJ, Rottier PJ. 
Coronavirus M proteins accumulate in the Golgi complex beyond the site of virion 
budding. J Virol. 1994;68(10):6523-34. 
- 
10. 
Schrezenmeier E, Dorner T. Mechanisms of action of hydroxychloroquine and 
chloroquine: implications for rheumatology. Nat Rev Rheumatol. 2020;16(3):155-66. 
- 
11. 
Savarino A, Boelaert JR, Cassone A, Majori G, Cauda R. Effects of chloroquine 
on viral infections: an old drug against today's diseases? Lancet Infect Dis. 
2003;3(11):722-7. 

Pragmatic trial of hydroxychloroquine/azithromycin for COVID 
Version 1.0 7 April 2020 
41 
 
- 
12. 
Yao X, Ye F, Zhang M, Cui C, Huang B, Niu P, et al. In Vitro Antiviral Activity and 
Projection of Optimized Dosing Design of Hydroxychloroquine for the Treatment of 
Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). Clin Infect Dis. 
2020. 
- 
13. 
Beigelman A, Mikols CL, Gunsten SP, Cannon CL, Brody SL, Walter MJ. 
Azithromycin attenuates airway inflammation in a mouse model of viral bronchiolitis. 
Respir Res. 2010;11:90. 
- 
14. 
Schultz MJ. Macrolide activities beyond their antimicrobial effects: macrolides in 
diffuse panbronchiolitis and cystic fibrosis. J Antimicrob Chemother. 2004;54(1):21-8. 
- 
15. 
Equi A, Balfour-Lynn IM, Bush A, Rosenthal M. Long term azithromycin in 
children with cystic fibrosis: a randomised, placebo-controlled crossover trial. Lancet. 
2002;360(9338):978-84. 
- 
16. 
Garcia Vazquez E, Mensa J, Martinez JA, Marcos MA, Puig J, Ortega M, et al. 
Lower mortality among patients with community-acquired pneumonia treated with a 
macrolide plus a beta-lactam agent versus a beta-lactam agent alone. Eur J Clin 
Microbiol Infect Dis. 2005;24(3):190-5. 
- 
17. 
Arabi YM, Deeb AM, Al-Hameed F, Mandourah Y, Almekhlafi GA, Sindi AA, et al. 
Macrolides in critically ill patients with Middle East Respiratory Syndrome. Int J Infect 
Dis. 2019;81:184-90. 
- 
18. 
Keyaerts E, Vijgen L, Maes P, Neyts J, Van Ranst M. In vitro inhibition of severe 
acute respiratory syndrome coronavirus by chloroquine. Biochem Biophys Res 
Commun. 2004;323(1):264-8. 
- 
19. 
Keyaerts E, Li S, Vijgen L, Rysman E, Verbeeck J, Van Ranst M, et al. Antiviral 
activity of chloroquine against human coronavirus OC43 infection in newborn mice. 
Antimicrob Agents Chemother. 2009;53(8):3416-21. 
- 
20. 
Tan YW, Yam WK, Sun J, Chu JJH. An evaluation of Chloroquine as a broad-
acting antiviral against Hand, Foot and Mouth Disease. Antiviral Res. 2018;149:143-9. 
- 
21. 
Colson P, Rolain JM, Lagier JC, Brouqui P, Raoult D. Chloroquine and 
hydroxychloroquine as available weapons to fight COVID-19. Int J Antimicrob Agents. 
2020:105932. 
- 
22. 
Biot C, Daher W, Chavain N, Fandeur T, Khalife J, Dive D, et al. Design and 
synthesis of hydroxyferroquine derivatives with antimalarial and antiviral activities. J Med 
Chem. 2006;49(9):2845-9. 

Pragmatic trial of hydroxychloroquine/azithromycin for COVID 
Version 1.0 7 April 2020 
42 
 
- 
23. 
Wang M, Cao R, Zhang L, Yang X, Liu J, Xu M, et al. Remdesivir and 
chloroquine effectively inhibit the recently emerged novel coronavirus (2019-nCoV) in 
vitro. Cell Res. 2020;30(3):269-71. 
- 
24. 
Gao J, Tian Z, Yang X. Breakthrough: Chloroquine phosphate has shown 
apparent efficacy in treatment of COVID-19 associated pneumonia in clinical studies. 
Biosci Trends. 2020;14(1):72-3. 
- 
25. 
Gao ZC. [Efficient management of novel coronavirus pneumonia by efficient 
prevention and control in scientific manner]. Zhonghua Jie He He Hu Xi Za Zhi. 
2020;43(0):E001. 
- 
26. 
Chen X. Hydroxychloroquine and chloroquine. Clin Infect Dis. 2020. 
- 
27. 
Cortegiani A, Ingoglia G, Ippolito M, Giarratano A, Einav S. A systematic review 
on the efficacy and safety of chloroquine for the treatment of COVID-19. J Crit Care. 
2020. 
- 
28. 
Chen Z, Hu J, Zhang Z, Jiang S, Han S, Yan D, et al. Efficacy of 
hydroxychloroquine in patients with COVID-19: results of a randomized clinical trial. 
medRxiv. 2020. 
- 
29. 
Gautret P, Lagier JC, Parola P, Hoang VT, Meddeb L, Mailhe M, et al. 
Hydroxychloroquine and azithromycin as a treatment of COVID-19: results of an open-
label non-randomized clinical trial. Int J Antimicrob Agents. 2020:105949. 
- 
30. 
Gautret P, Lagier JC, Parola P, Hoang VT, Meddeb L, Sevestre J, et al. Clinical 
and microbiological effect of a combination of hydroxychloroquine and 
- 
azithromycin in 80 COVID-19 patients with at least a six-day follow up: an 
- 
observational study Int J Antimicrob Agents. 2020;Online only(Pre-print). 
- 
31. 
Molina JM, Delaugerre C, Le Goff J, Mela-Lima B, Ponscarme D, Goldwirt L, et 
al. No evidence of rapid antiviral clearance or clinical benefit with the combination of 
hydroxychloroquine and azithromycin in patients with severe COVID-19 infection. 
Medecine et Maladies Infectieuses. 2020. 
- 
32. 
Liu J, Cao R, Xu M, Wang X, Zhang H, Hu H, et al. Hydroxychloroquine, a less 
toxic derivative of chloroquine, is effective in inhibiting SARS-CoV-2 infection in vitro. 
Cell Discov. 2020;6:16. 
- 
33. 
Popert AJ. Chloroquine: a review. Rheumatol Rehabil. 1976;15(3):235-8. 
- 
34. 
Lim HS, Im JS, Cho JY, Bae KS, Klein TA, Yeom JS, et al. Pharmacokinetics of 
hydroxychloroquine and its clinical implications in chemoprophylaxis against malaria 
caused by Plasmodium vivax. Antimicrob Agents Chemother. 2009;53(4):1468-75. 

Pragmatic trial of hydroxychloroquine/azithromycin for COVID 
Version 1.0 7 April 2020 
43 
 
- 
35. 
McChesney EW, Conway WD, Banks WF, Jr., Rogers JE, Shekosky JM. Studies 
of the metabolism of some compounds of the 4-amino-7-chloroquinoline series. J 
Pharmacol Exp Ther. 1966;151(3):482-93. 
- 
36. 
Durkin MJ, Jafarzadeh SR, Hsueh K, Sallah YH, Munshi KD, Henderson RR, et 
al. Outpatient Antibiotic Prescription Trends in the United States: A National Cohort 
Study. Infect Control Hosp Epidemiol. 2018;39(5):584-9. 
- 
37. 
Lexi-Drugs: Azithromycin (Systemic) Drug Information [Internet]. Wolters Kluwer 
Clinical Drug Information, Inc. 2020 [cited March 27, 2020]. 
- 
38. 
Zimmermann P, Ziesenitz VC, Curtis N, Ritz N. The Immunomodulatory Effects 
of Macrolides-A Systematic Review of the Underlying Mechanisms. Front Immunol. 
2018;9:302. 
- 
39. 
Kanoh S, Rubin BK. Mechanisms of Action and Clinical Application of Macrolides 
as Immunomodulatory Medications. Clinical Microbiology Reviews. 2010;23(3):590-615. 
- 
40. 
Kawamura K, Ichikado K, Takaki M, Eguchi Y, Anan K, Suga M. Adjunctive 
therapy with azithromycin for moderate and severe acute respiratory distress syndrome: 
a retrospective, propensity score-matching analysis of prospectively collected data at a 
single center. Int J Antimicrob Agents. 2018;51(6):918-24. 
- 
41. 
Gielen V, Johnston SL, Edwards MR. Azithromycin induces anti-viral responses 
in bronchial epithelial cells. Eur Respir J. 2010;36(3):646-54. 
- 
42. 
Menzel M, Akbarshahi H, Bjermer L, Uller L. Azithromycin induces anti-viral 
effects in cultured bronchial epithelial cells from COPD patients. Sci Rep. 2016;6:28698. 
- 
43. 
Schogler A, Kopf BS, Edwards MR, Johnston SL, Casaulta C, Kieninger E, et al. 
Novel antiviral properties of azithromycin in cystic fibrosis airway epithelial cells. Eur 
Respir J. 2015;45(2):428-39. 
- 
44. 
Tran DH, Sugamata R, Hirose T, Suzuki S, Noguchi Y, Sugawara A, et al. 
Azithromycin, a 15-membered macrolide antibiotic, inhibits influenza A(H1N1)pdm09 
virus infection by interfering with virus internalization process. J Antibiot (Tokyo). 
2019;72(10):759-68. 
- 
45. 
Ohara H, Nakamura Y, Watanabe Y, Cao X, Yamazaki Y, Izumi-Nakaseko H, et 
al. Azithromycin Can Prolong QT Interval and Suppress Ventricular Contraction, but Will 
Not Induce Torsade de Pointes. Cardiovasc Toxicol. 2015;15(3):232-40. 
- 
46. 
Milberg P, Eckardt L, Bruns HJ, Biertz J, Ramtin S, Reinsch N, et al. Divergent 
proarrhythmic potential of macrolide antibiotics despite similar QT prolongation: fast 

Pragmatic trial of hydroxychloroquine/azithromycin for COVID 
Version 1.0 7 April 2020 
44 
 
phase 3 repolarization prevents early afterdepolarizations and torsade de pointes. J 
Pharmacol Exp Ther. 2002;303(1):218-25. 
- 
47. 
Svanstrom H, Pasternak B, Hviid A. Use of azithromycin and death from 
cardiovascular causes. N Engl J Med. 2013;368(18):1704-12. 
- 
48. 
Almalki ZS, Guo JJ. Cardiovascular events and safety outcomes associated with 
azithromycin therapy: a meta-analysis of randomized controlled trials. Am Health Drug 
Benefits. 2014;7(6):318-28. 
- 
49. 
Sarkar M, Woodland C, Koren G, Einarson AR. Pregnancy outcome following 
gestational exposure to azithromycin. BMC Pregnancy Childbirth. 2006;6:18. 
- 
50. 
Ramsey PS, Vaules MB, Vasdev GM, Andrews WW, Ramin KD. Maternal and 
transplacental pharmacokinetics of azithromycin. Am J Obstet Gynecol. 
2003;188(3):714-8. 
- 
51. 
Fan H, Gilbert R, O'Callaghan F, Li L. Associations between macrolide antibiotics 
prescribing during pregnancy and adverse child outcomes in the UK: population based 
cohort study. BMJ. 2020;368:m331. 
- 
52. 
Salman S, Davis TM, Page-Sharp M, Camara B, Oluwalana C, Bojang A, et al. 
Pharmacokinetics of Transfer of Azithromycin into the Breast Milk of African Mothers. 
Antimicrob Agents Chemother. 2015;60(3):1592-9. 
- 
53. 
Sutton AL, Acosta EP, Larson KB, Kerstner-Wood CD, Tita AT, Biggio JR. 
Perinatal pharmacokinetics of azithromycin for cesarean prophylaxis. Am J Obstet 
Gynecol. 2015;212(6):812 e1-6. 
- 
54. 
Lund M, Pasternak B, Davidsen RB, Feenstra B, Krogh C, Diaz LJ, et al. Use of 
macrolides in mother and child and risk of infantile hypertrophic pyloric stenosis: 
nationwide cohort study. BMJ. 2014;348:g1908. 
- 
55. 
Goldstein LH, Berlin M, Tsur L, Bortnik O, Binyamini L, Berkovitch M. The safety 
of macrolides during lactation. Breastfeed Med. 2009;4(4):197-200.