Conditions: Severe Obstructive Sleep Apnea (Apnea Hypopnea Index > 30 Events/Hour) Hypertension
Keywords: Hypertension High Blood Pressure Sleep Apnea Obstructive Sleep Apnea OSA
Allocation: RANDOMIZED | Intervention model: PARALLEL
Enrollment: 87
Healthy volunteers: no

Brief summary:
The purpose of this study is to determine if two medicines (allopurinol and losartan) can influence heart and blood vessel health compared to placebo in patients with sleep apnea who are using continuous positive airway pressure (CPAP).

Detailed description:
The specific aims of this research project are: 1) Determine if treatment with losartan, an angiotensin type I receptor (AT1R) antagonist, or allopurinol, a XO inhibitor, normalize chemoreflex control of sympathetic outflow and ventilation and improve local vascular regulation and stiffness; and 2) Determine if these interventions reduce the severity of sleep disordered breathing and lower diurnal blood pressure.

Arms:
arm 1: Losartan 50 mg daily for two weeks, then increased to 100mg daily for 4 weeks if asymptomatic and blood pressure within range. arm 2: Allopurinol 300 mg daily for 6 weeks arm 3: Placebo capsule daily for 6 weeks

Interventions: intervention 1: Losartan intervention 2: Allopurinol intervention 3: Placebo
Intervention descriptions: intervention 1: Losartan 50 mg daily for two weeks, then increased to 100mg daily for 4 weeks if asymptomatic. Remain at 50 mg daily for 4 more weeks or removed from study if symptomatic. intervention 2: Allopurinol 300 mg daily for 6 weeks intervention 3: Placebo capsule daily for 6 weeks

Protocol document:
Prot_SAP_000.pdf: 
 
CONFIDENTIAL 
 
CLINICAL RESEARCH PROTOCOL 
 
 
 
 
 
Pharmacologic Interventions for Cardiovascular Disease in 
Obstructive Sleep Apnea 
 
A Phase II, randomized, double-blind, placebo-controlled, multi-center study of the effects 
of losartan and allopurinol on chemoreflex responses, sympathetic output, vascular 
function, severity of sleep apnea and blood pressure in hypertensive patients with 
obstructive sleep apnea. 
John M. Dopp, Pharm.D. 
Barbara J. Morgan, Ph.D., PT 
James Stein, M.D. 
Jerome Dempsey, Ph.D. 
Ailiang Xie, M.D., Ph.D. 
David DeMets, Ph.D. 
Christine Sorkness, Pharm.D. 
Mihaela Teodorescu, M.D. 
Scott Hetzel, M.S. 
Paul Smith, M.D. 
Debra Kruser, RN, BSN, MS 
MJ Washburn RRT, CCRC 
 
University of Wisconsin Schools of Pharmacy and Medicine and Public Health 
Institute for Clinical and Translational Research 
University of Wisconsin-Madison 
Madison, WI 
 
Jaime Boero, M.D., Ph.D. 
Marshfield Clinic Research Foundation 
Marshfield Clinic 
Marshfield, WI 
 
Daren Tobert, M.D. 
Gundersen Medical Foundation 
Gundersen Health System 
LaCrosse, WI 
 
James Gapinski, M.D. 
Aurora Bay Care Sleep Center 
Green Bay, WI 
 
Scott Sands, PhD 
Brigham and Women’s Hospital 
Boston, MA 
 
 
 
 
 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page ii 
Version: 23, 7-25-17 
CONFIDENTIAL 
Initial version: 
[01-26-10] 
 
Amended: 
[06-17-10] 
Amended: 
[09-14-10] 
Amended: 
[03-05-11] 
Amended: 
[07-20-11] 
Amended: 
[08-03-11] 
Amended: 
[08-19-11] 
Amended: 
[10-22-11] 
Amended: 
[11-29-11] 
Amended: 
[01-09-12] 
Amended:         [03-18-12] 
Amended: 
[04-20-12] 
Amended:          [07-13-12] 
Amended: 
[08-22-12] 
Amended: 
[04-26-13] 
Amended: 
[07-15-13] 
Amended:  
[08-16-13] 
Amended:  
[11-13-13] 
Amended:  
[06-25-14] 
Amended:  
[09-18-14] 
Amended:  
[12-22-14] 
 
Amended:  
[03-03-15] 
Amended: 
[12-05-16] 
 
 
 
 
 
 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page iii 
Version: 23, 7-25-17 
CONFIDENTIAL 
Table of Contents 
 
STUDY SUMMARY ...................................................................................................................................... 1 
1 
INTRODUCTION .................................................................................................................................. 2 
1.1 
BACKGROUND .......................................................................................................................... 2 
1.2 
PRECLINICAL DATA ................................................................................................................... 4 
1.3 
DOSE RATIONALE AND RISK/BENEFITS ........................................................................................ 4 
2 
STUDY OBJECTIVES ......................................................................................................................... 5 
3 
STUDY DESIGN ................................................................................................................................... 5 
3.1 
GENERAL DESIGN ..................................................................................................................... 5 
3.2 
PRIMARY STUDY ENDPOINTS ...................................................................................................... 7 
3.3 
SECONDARY STUDY ENDPOINTS ................................................................................................. 7 
4 
SUBJECT SELECTION AND WITHDRAWAL .................................................................................... 8 
4.1 
INCLUSION CRITERIA ................................................................................................................. 8 
4.2 
EXCLUSION CRITERIA ................................................................................................................ 8 
4.3 
SUBJECT RECRUITMENT AND SCREENING .................................................................................... 9 
4.4 
EARLY WITHDRAWAL OF SUBJECTS........................................................................................... 10 
4.4.1 
When and How to Withdraw Subjects ................................................................................... 10 
4.4.2 
Data Collection and Follow-up for Withdrawn Subjects ........................................................ 11 
5 
STUDY DRUG .................................................................................................................................... 11 
5.1 
DESCRIPTION ......................................................................................................................... 11 
5.2 
TREATMENT REGIMEN ............................................................................................................. 11 
5.3 
METHOD FOR ASSIGNING SUBJECTS TO TREATMENT GROUPS ..................................................... 12 
5.4 
PREPARATION AND ADMINISTRATION OF STUDY DRUG ................................................................ 12 
5.5 
SUBJECT ADHERENCE MONITORING .......................................................................................... 12 
        5.5.1     CPAP before and during study…………………………………………………………………… 13 
5.6 
PRIOR AND CONCOMITANT THERAPY ......................................................................................... 13 
        5.6.1    Pregnancy testing and birth control ……………………………………………………………… 13 
5.7 
BLINDING OF STUDY DRUG....................................................................................................... 14 
5.8 
RECEIVING, STORAGE, DISPENSING AND RETURN ...................................................................... 14 
5.8.1 
Receipt of Drug Supplies ....................................................................................................... 14 
5.8.2 
Dispensing of Study Drug ...................................................................................................... 14 
6 
STUDY PROCEDURES ..................................................................................................................... 14 
     6.1    STUDY PERSONNEL ……………………………………………………………………………….   15 
     6.2     SUMMARY of VISITS …………………………………………………………………………………. 15 
     6.3     SAMPLE COLLECTION ……………………………………………………………………………… 17 
     6.4     STUDY COMMUNICATION AND OVERSIGHT …………………………………….…………….  17 
     6.5   UNEXPECTED FINDINGS …………………………………………………………………………….18 
7 
STATISTICAL PLAN ......................................................................................................................... 18 
7.1 
SAMPLE SIZE DETERMINATION ................................................................................................. 18 
7.2 
STATISTICAL METHODS ............................................................................................................ 19 
7.3 
SUBJECT POPULATION(S) FOR ANALYSIS ................................................................................... 19 
8 
SAFETY AND ADVERSE EVENTS ................................................................................................... 19 
8.1 
DEFINITIONS ........................................................................................................................... 19 
8.2 
RECORDING OF ADVERSE EVENTS ............................................................................................ 21 
8.3 
REPORTING OF SERIOUS ADVERSE EVENTS AND UNANTICIPATED PROBLEMS ................................ 21 
 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page iv 
Version: 23, 7-25-17 
CONFIDENTIAL 
8.3.1 
Investigator reporting: notifying the IRB ................................................................................ 22 
8.3.2 
Sponsor reporting: Notifying NHLBI ...................................................................................... 23 
8.3.3 
Sponsor reporting: Notifying the FDA ................................................................................ 23 
8.4 
UNBLINDING PROCEDURES ...................................................................................................... 24 
8.5 
STOPPING RULES ................................................................................................................... 24 
8.6 
INDEPENDENT DATA AND SAFETY MONITORING BOARD ............................................................... 24 
    8.7 
    STUDY COORDINATION and DATA MONITORING ……………………………………………..25 
 
9 
DATA HANDLING AND RECORD KEEPING ................................................................................... 26 
9.1 
CONFIDENTIALITY .................................................................................................................... 26 
9.2 
SOURCE DOCUMENTS ............................................................................................................. 27 
9.3 
CASE REPORT FORMS ............................................................................................................. 27 
    9.4 
    DATA COLLECTION AT SITES ……………………………………………………………………..27 
    9.5 
    DATA ANALYSIS AT BRIGHAM AND WOMEN'S HOSPITAL…..……………………………….27 
10 
ETHICAL CONSIDERATIONS .......................................................................................................... 27 
11 
STUDY FINANCES ............................................................................................................................ 28 
11.1 
FUNDING SOURCE ................................................................................................................... 28 
11.2 
SUBJECT STIPENDS OR PAYMENTS ........................................................................................... 28 
12 
PUBLICATION PLAN ........................................................................................................................ 29 
13 
REFERENCES ................................................................................................................................... 29 
14 
APPENDICES .................................................................................................................................... 37 
 
 
 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page v 
Version: 23, 7-25-17 
CONFIDENTIAL 
 
List of Abbreviations 
 
 
A-II =   
angiotensin II 
AE = 
 
adverse event 
AT1R =  
angiotensin type 1 receptor 
ATP =   
adenosine triphosphate 
CCR =   
carotid chemoreceptor 
CIH =  
chronic intermittent hypoxia 
CO2 =  
carbon dioxide 
CPAP =  
continuous positive airway pressure 
CRU =  clinical research unit (at UW-Madison) 
CV =   
cardiovascular 
DMC =  
data monitoring committee (at UW-Madison) 
ICTR =   
institute for clinical and translational research (at UW-Madison) 
MSNA =  
muscle sympathetic nerve activity 
NO = 
 
nitric oxide 
O2 = 
 
oxygen 
OSA =   
obstructive sleep apnea 
PI =  
 
principal investigator(s) 
PRC =  
Pharmaceutical Research Center (at UW-Madison) 
ROS =  
reactive oxygen species 
SAE =  
serious adverse event 
SaO2 =  
oxygen saturation 
UIP =  
University of Iowa Pharmaceuticals 
WiNHR =  
Wisconsin Network for Health Research 
XO = 
 
xanthine oxidase 
 
 
 
 
 
 
 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 1 
Version 20 
CONFIDENTIAL 
 
Study Summary 
Title 
Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep 
Apnea 
 
Protocol Number 
 
Phase 
Phase II 
Methodology 
Randomized, double-blind, placebo-controlled, parallel group study 
Study Duration 
4 years 
Study Site(s) 
Multi-Center 
-University of Wisconsin, Madison, WI 
-Marshfield Clinic, Marshfield, WI 
-Gundersen Health System La Crosse, WI 
-Aurora Bay Care, Green Bay, WI 
-Brigham and Women’s Hospital, Boston, MA (Data analysis only) 
Objectives 
The objective of this application is to investigate the potential utility of 
angiotensin receptor blockade and xanthine oxidase inhibition in targeting and 
reversing putative mechanisms of hypertension in patients with OSA.   
Number of Subjects 
N=100 
Diagnosis and Main 
Inclusion Criteria 
Hypertensive adult patients with obstructive sleep apnea using continuous 
positive airway pressure 
Study Product, Dose, 
Route, Regimen 
Losartan (Cozaar) 50-100 mg orally daily, Allopurinol 300 mg orally daily 
Duration 
of 
administration 
6-weeks 
Reference therapy 
Placebo 
Statistical 
Methodology 
ANOVA and chi-square tests where appropriate 
 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 2 
Version: 23, 7-25-17 
CONFIDENTIAL 
1 Introduction 
 
This document is a protocol for a human research study. This study is to be conducted according to US 
and international standards of Good Clinical Practice (FDA Title 21 part 312 and International Conference 
on Harmonization guidelines), applicable government regulations and Institutional research policies and 
procedures.  
1.1 Background 
Obstructive sleep apnea (OSA) is a condition that is increasingly associated with adverse effects 
on cardiovascular (CV) health.  It is estimated that up to 18 million Americans have OSA, many 
of whom remain undiagnosed (22).  The U.S. population is becoming increasingly overweight and 
is aging, both of which are alarming trends because obesity and aging are two of the strongest 
risk factors for the development of OSA (23-25).  As more people develop OSA, the prevalence 
of CV disease is expected to increase.  Untreated OSA is associated with the development of 
hypertension and other CV diseases (26-30).  Discovery of treatable causes of CV disease in 
OSA may lead to reductions in morbidity and mortality. Increased sympathetic traffic and 
endothelial dysfunction are well-documented in OSA and are believed to play a significant role in 
the development of hypertension.  Continuous positive airway pressure (CPAP), the therapy of 
choice for OSA, has beneficial effects on sympathetic activity, endothelial function, and blood 
pressure (31-33); however, up to 30 percent of patients do not adhere to CPAP therapy (17).  
Therefore, development of other treatments that specifically target mechanisms of CV disease 
and hypertension is warranted. 
Sympathetic Nervous System in OSA:  Evidence from animal models suggests that 
carotidchemoreceptors (CCR)s (via intact sympathetic nerves) mediate changes in sympathetic 
output that result in hypertension (34-36).  Patients with OSA have elevated sympathetic nerve 
activity while exposed to intermittent hypoxia during sleep and also during daytime waking hours 
when blood oxygen levels are normal (37).  Plasma and urinary norepinephrine concentrations 
are also elevated in patients with OSA (38).  Hypoxia, rather than hypercapnia, is primarily 
responsible for increases in sympathetic nerve activity (39) that occur during apneas in patients 
with OSA.  Narkiewicz et al demonstrated that inhibiting the CCR with hyperoxia reduced mSNA 
in patients with OSA, suggesting that tonic activation of CCR afferents contributes to elevations 
in daytime mSNA (2).  Furthermore, OSA patients have CCR hypersensitivity to subsequent 
hypoxia exposure (3).  Treatment of OSA with CPAP or oxygen normalizes sympathetic activity, 
blood pressure, and CCR hypersensitivity (3,31,32).  These data suggest that intermittent 
episodes of hypoxia mediate increased sympathetic traffic in patients with OSA via sensitization 
of the carotid chemoreflex and that this alteration is reversible. 
Role of A-II and XO in Carotid Body Function:  Heightened sensitivity of carotid chemoreflex 
regulation of sympathetic discharge is one putative mechanism of sympathetic overactivity in 
patients with OSA. Changes in CCR sensitivity appear to be driven by exposure to intermittent 
hypoxia, rather than hypercapnia or arousal from sleep (40).  Experimental animal studies have 
demonstrated that chronic intermittent hypoxia (CIH) increases CCR sensitivity to subsequent 
hypoxic episodes (41-43). The signal for increased CCR sensitivity during hypoxia appears to 
involve endothelin-1 and angiotensin II (A-II) via superoxide and other reactive oxygen species 
(ROS)(44).  The importance of Angiotensin II’s (A-II) role in enhancing chemoreceptor sensitivity 
to hypoxia is becoming increasingly recognized.  Injection of A-II directly into isolated carotid 
bodies elevates nerve activity in the carotid sinus nerve (44).  Additionally, carotid afferent nerve 
activity is increased in ex vivo studies by application of physiologic concentrations of A-II (45).  
Application of an A-II antagonist, losartan reverses the sympathoexcitation of A-II, indicating 
involvement of the A-II type 1 receptor (45).  Hypoxia increases expression of carotid body A-II 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 3 
Version: 23, 7-25-17 
CONFIDENTIAL 
type 1 receptors (44) and A-II receptor mRNA (46).  Most importantly, our recent laboratory 
findings demonstrated that CIH-induced chemoreflex hypersensitivity was prevented by 
administration of losartan before and during CIH exposure (See Preliminary Studies Section).   
 
Previous studies have shown that CIH increases generation of ROS in the carotid body 
(40). Furthermore, administration of superoxide scavengers prevents CIH-induced CCR 
sensitization (40), indicating this anion plays a pivotal role in sensitization of the carotid 
chemoreflex. Likely sources of superoxide in the carotid body include NADPH-oxidase and XO, 
which can be upregulated by hypoxia (47-49). Preliminary data from our laboratory illustrates that 
XO and A-II inhibition prevent CIH-induced increases in carotid body superoxide production (see 
Preliminary Studies). Taken together, these data suggest that superoxide is a key player in the 
signaling pathway that mediates enhanced CCR sensitivity caused by CIH and that A-II and XO 
increase carotid superoxide production under these conditions.  Thus, targeting A-II with an ARB 
and XO with allopurinol would appear to be a logical strategy to reduce the chemosensitizing 
effects of CIH in the carotid body.  Evaluation of XO and A-II inhibition in human subjects with 
OSA is warranted to determine if these preliminary findings have clinical relevance.  If XO and A-
II inhibition can reduce CCR sensitivity, they may also reduce sympathetic overactivity and the 
risk of developing hypertension. 
 
Role of Nitric Oxide (NO) in Carotid Body and Vascular Endothelium:  NO decreases CCR 
sensitivity and appears to work in a counter-regulatory fashion to A-II (50). Two isoforms of nitric 
oxide synthase (eNOS and nNOS) are present in the carotid body (51). In our preliminary studies, 
exposure to CIH downregulated carotid body nNOS, potentially removing an important effect of 
NO in maintaining normal CCR sensitivity (52). Our preliminary data demonstrate that inhibition 
of XO and A-II partially prevents CIH-induced reductions in nNOS expression (see Preliminary 
Studies section).  These findings are in accordance with previous studies demonstrating that 
inhibition of XO and A-II increases NO availability in patients with OSA (53). Thus, XO and A-II 
inhibition may boost NO availability in the carotid body and reduce pathological CCR 
hypersensitivity produced in response to OSA. 
 
Reduced availability of NO is thought to play a role in the development of vascular abnormalities 
in patients with OSA (54).  These individuals demonstrate impairments in endothelium-dependent 
vasodilation in numerous vascular beds (55-57), and treatment with CPAP improves these 
impairments (32).  Animal data demonstrate that CIH reduces acetylcholine-induced endothelium-
dependent vasodilation in gracilis arteries and middle cerebral arteries (58). These impairments 
likely represent reductions in NO availability since endothelium-independent vasodilation to 
exogenous nitrates is not impaired in subjects with OSA or animals exposed to CIH (55,58).  ROS 
are a likely trigger for OSA-induced endothelial dysfunction. ROS, e.g. superoxide anion, 
produced in the vessel wall can decrease the availability of NO via scavenging and by combination 
with NO to form peroxynitrite which leads to NOS uncoupling (59).  Markers of oxidative stress, 
including neutrophil superoxide production and lipid peroxidation, are elevated in patients with 
OSA (60,61).  Angiotensin II and XO are involved in the production of ROS in the vascular 
endothelium (12,62).  Plasma concentrations of A-II are elevated in OSA and the activity of XO is 
increased by hypoxia (48,49,13).  In patients with OSA, XO inhibition improves NO-mediated 
vasodilation.  We have shown that XO inhibition and A-II inhibition both can prevent the CIH 
induced endothelial impairments and improve NO-mediated vasodilation in ex vivo animal studies 
(See Figure 5, Preliminary Studies Section). 
 
Remodeling as a cause of impaired vasodilation:  Increased carotid intima-media thickness 
(63,64) and increased arterial stiffness (64-69) are observed in individuals with OSA. Blood levels 
of NO and ET-1, endothelium-derived regulators of vascular stiffness with opposing actions 
(70,71), are decreased and increased, respectively (54,72). In rat gracilis arteries, we have shown 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 4 
Version: 23, 7-25-17 
CONFIDENTIAL 
that CIH exposure causes a leftward shift in the stress-strain curve, consistent with an increase 
in arterial stiffness (73). 
Mitogenic factors (e.g., vascular endothelial growth factor (VEGF), basic fibroblast growth 
factor, platelet-derived growth factor) are up-regulated by hypoxia (74,75) and during 
inflammation (76). In patients with OSA, elevated plasma Ang II levels (13,64) have been 
implicated in increased VEGF expression (77). 
Chronic sympathetic activation causes vascular remodeling via release of catecholamines 
that induce vascular wall growth (78). In pulmonary arteries, ATP released from adrenergic nerves 
during hypoxia-induced sympathetic stimulation causes proliferation and migration of adventitial 
fibroblasts into the intima and media (79). This mechanism is a putative cause of remodeling in 
the systemic circulation (80). In addition, surges in sympathetic outflow during episodes of apnea 
produce cyclical increases in arterial pressure and blood flow that may trigger adaptations in 
endothelial cells, vascular smooth muscle, and extracellular matrix aimed at normalizing wall 
stress (81,82). 
Heightened Chemoreflex Sensitivity and Diminished Cerebrovascular Reactivity Destabilize 
Breathing during Sleep:  Unstable ventilatory control during sleep in individuals with upper airways 
that are anatomically susceptible to collapse may precipitate OSA (7,8,83,84). Further, limited 
evidence points to a significant prevalence of unstable ventilatory control (i.e. high control system 
“loop gain”) in patients with severe OSA, secondary in part to increases in CO2 sensitivity 
(increased controller gain) (8,85). Augmented carotid chemoreflex sensitivity secondary to 
intermittent hypoxia (86) might also contribute to increased ventilatory instability and, in turn, to 
cyclical obstructions in OSA patients. Accordingly, reducing this important component of 
increased controller gain by preventing intermittent hypoxia-induced increases in carotid body 
sensitivity may also reduce the severity of sleep disordered breathing concomitantly with a 
reduction in sympathetic nervous system activity. We emphasize that prevention of ventilatory 
instability, per se, is most likely to reduce cyclical apneas in patients with mild to moderate levels 
of airway collapsibility, and less likely in those with severely compromised airway anatomy (7,8). 
We have demonstrated that OSA impairs hypercapnic vasodilation in the cerebral circulation 
(87). In health, cerebral resistance arteries are exquisitely sensitive to changes in PaCO2 (88). 
This important characteristic of the cerebral circulation minimizes the change in brain PCO2 during 
fluctuations in arterial PCO2. Reductions in cerebrovascular CO2 responsiveness could 
exacerbate breathing instability during sleep by exaggerating the accumulation and also the 
washout of CO2 from central chemoreceptors during fluctuations in ventilation and arterial PCO2. 
We have shown, in healthy humans during wakefulness, that cerebral CO2 responsiveness is an 
important determinant of eupneic ventilation and also the ventilatory response to hypercapnia 
(89). Furthermore, we have shown during sleep in healthy subjects, that reducing the cerebral 
vascular response to CO2 with indomethacin decreases the CO2 reserve below eupnea (ΔPaCO2, 
eupnea-apnea) and increases the susceptibility to apnea (90). 
In patients with OSA, attenuated cerebrovascular CO2 reactivity and exaggerated carotid 
chemoreflex sensitivity are both reversible when sleep disordered breathing is eliminated with 
CPAP (3,87,91). We hypothesize that pharmacological measures targeting the mechanisms 
involved also may reverse these impairments and lead to a reduction in the severity of sleep 
disordered breathing in selected patients. 
1.2 Preclinical Data 
N/A 
1.3 Dose Rationale and Risk/Benefits 
Drug selection and dose rationale:  In our rat studies, an allopurinol dose of 65 mg/kg/day 
(~300mg/day in humans) (134) effectively reduced AT1R and superoxide concentrations in the 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 5 
Version: 23, 7-25-17 
CONFIDENTIAL 
rat CCR (see Preliminary Studies).  Subjects in our study will take allopurinol 300 mg daily for six 
weeks (dose will remain unchanged during four-week period).  Allopurinol is quickly converted in 
vivo to its active metabolite, oxypurinol.  Oxypurinol is responsible for the majority of the effects 
of allopurinol and has a half-life of 17-21 hours (135).  Steady state blood concentrations of 
oxypurinol occur in approximately 105 hours (4.4 days).  A study demonstrating vascular benefits 
of XO inhibition in patients with OSA was two weeks in duration, and our rat study used two weeks 
of CIH.  We propose to have subjects take six weeks of study medication to increase the likelihood 
of observing significant sympathetic, vascular and blood pressure changes and to determine if 
preliminary changes seen after two weeks are sustained.  A 300 mg daily dose will sufficiently 
inhibit XO because the IC90 of oxypurinol for XO is approximately 5 µg/mL, and a single 300 mg 
dose of allopurinol produces peak levels of 5-7 µg/mL (136-138).  At steady state, peak drug 
concentrations are higher than peak concentrations from single doses, and > 90% of XO will be 
inhibited.  Losartan will be used in the doses that are clinically used to treat hypertension (50-100 
mg daily by mouth).  E-3174 is the active metabolite responsible for the clinical effects of losartan 
and E-3174 has a half-life of 4-9 hours (139).  Thus, steady-state concentrations of E-3174 are 
quickly achieved after approximately 20-45 hours (139).  Use of this dose is based on data 
demonstrating that 100 mg doses produce plasma concentrations > 1000 ng/mL of E-3174 (140).  
The ability of E-3174 to block the pressor effects of an intravenous angiotensin II challenge 
plateaus between 500-750 ng/mL (141).  Thus, we feel our proposed doses will effectively block 
the actions of A-II.  
We have chosen a six week treatment period because in our previous study, vascular endothelial 
function was improved after 6 weeks of treatment and did not further improve after 12 weeks of 
treatment.  Furthermore, three weeks of losartan therapy successfully prevented chemoreflex 
sensitization in adult male, Sprague-Dawley rats.   
 
Risks to subjects are reasonable in relation to the anticipated benefits because the two 
medications are FDA approved with significant clinical history to base safety upon.  If the study 
drugs are proven to be effective for the primary endpoint, they may undergo further trials as 
potential interventions for prevention of hypertension and cardiovascular disease in patients with 
OSA. 
2 Study Objectives 
1. Determine if treatment with losartan, an angiotensin type I receptor (AT1R) antagonist, or 
allopurinol, a xanthine oxidase (XO) inhibitor, normalize chemoreflex control of 
sympathetic outflow and ventilation and improves local vascular regulation and stiffness. 
2. Determine if these interventions reduce the severity of sleep disordered breathing and 
lower diurnal blood pressure. 
 
3 Study Design 
3.1 General Design 
 
The proposed project is a randomized, double-blind, placebo-controlled, parallel three-treatment 
arm phase II clinical evaluation of allopurinol, losartan, and placebo in hypertensive patients with 
severe OSA.  The proposed trial is based on preliminary data suggesting that both of these 
available medications may reduce chemoreflex sensitivity and endothelial function in patients with 
OSA.  Subject enrollment will be conducted on a rolling basis during the three year study period 
and subjects will be randomized (stratified by baseline CPAP adherence) to one of the three 
treatments.  Randomization will be stratified based on three categories of baseline CPAP 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 6 
Version: 23, 7-25-17 
CONFIDENTIAL 
adherence: < 1 hr, 1 – 4 hrs, and > 4hrs of CPAP use on average per night.  This will ensure 
balanced CPAP use between treatment arms.  Allopurinol will be initiated at 300 mg daily and the 
dose will remain constant for the six-week study session.  Losartan will be initiated at 50 mg daily 
and the dose will be increased to 100 mg after two weeks if systolic blood pressure is above 120 
mm Hg and as long as serum creatinine and potassium do not significantly increase.  Subjects 
with SBP between 120-100 mm Hg may increase to 100 mg if approved by physician.  If the 
subject’s serum creatinine (> 0.5 mg/dL) or serum potassium (above 5.0 mEq/L) significantly 
increase, or if the subject experiences a significant side effect from the medication (e.g. 
maculopapular rash) the subject will be discontinued from the study.  If the subject does not 
tolerate the losartan dose increase to 100 mg, his/her dose will be reduced back to 50 mg by 
study staff at request of subject. If the subject tolerates the dose increase, the losartan dose will 
then remain unchanged from the second to sixth week of the study.  Patients in the placebo group 
will receive placebo capsules daily for six weeks that are identical in appearance to the 
overencapsulated losartan and allopurinol.   
 
Hypertensive patients who have severe sleep apnea (apnea hypopnea index or respiratory 
disturbance index > 25 events/hour) will be studied to allow the greatest likelihood of detecting 
significant differences in outcome variables.  We will take patients who are and who are not using 
CPAP and will stratify their randomization based on baseline CPAP use.  This will be done to 
simulate real-world treatments as best as possible and determine if our therapies’ effectiveness 
is modified by CPAP use.  All subjects will undergo a screening visit (Visit 1) at their local site to 
determine eligibility and collect pertinent data.  Subsequently, subjects will be randomized to one 
of the three treatment groups (allopurinol, losartan, or placebo) and will have a physiology and 
vascular baseline study completed (Visit 3). 
 
Subjects will have 24-hour ambulatory blood pressure and 24-hour Holter/Heart rate variability 
measurements taken* (Visit 2) on a day/night prior to Visit 3.  Prior to starting study drug therapy, 
subjects will come to the Clinical Research Unit (CRU) at the University of Wisconsin for 
physiology/vascular study (Visit 3).  Subjects recruited from the other recruitment sites will come 
to Madison for Visit 3.  Subjects will arrive on day one of their study, and will have physiologic 
assessments taken before full overnight polysomnography and overnight Holter/Heart rate 
variability* is completed during night one (to determine severity of OSA).  Prior to 
polysomnography, subjects will have muscle sympathetic nerve activity, forearm blood flow, 
cerebral blood flow velocity, plasma catecholamines, ventilation, blood pressure, and heart rate 
measured at rest under normoxic (room air) conditions and during graded levels of hypoxia (90%, 
85% and 80% SaO2).  Following this, subjects will be allowed to return to normoxia (room air 
levels of oxygen ~ 21% oxygen) and after dinner will be hooked up for polysomnography and 
Holter/Heart rate variability*.  The morning after polysomnography, subjects will have assessment 
of brachial artery flow-mediated vasodilation and nitroglycerin induced vasodilation measured by 
ultrasound.  Then subjects will have aortic pulse wave velocity and augmentation index measured 
by arterial tonometry.  The 2nd night subjects will stay overnight again in the VA Research Unit 
and have control of breathing studies completed (Pcrit and apnea threshold).  Subjects will initiate 
study drug treatment and will return two weeks later for a blood pressure and heart rate (office 
blood pressure) measurement, CPAP assessment, and safety assessment to measure serum 
creatinine and serum potassium concentrations.  This is standard safety monitoring in patients 
taking ARBs, and to maintain blinding, all subjects will have this monitoring completed.  Subjects 
who do have clinically significant increases in serum creatinine (an increase > 0.5 mg/dL) or 
whose serum potassium increases to a concentration > 5.0 mEq/L will have their participation in 
the study terminated.  Subjects continuing in the study will then complete four additional weeks 
of drug therapy.  At the end of six weeks of study medication, subjects will have 24-hour 
ambulatory blood pressure and 24-hour Holter/Heart rate variability assessed one day and night.  


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 7 
Version: 23, 7-25-17 
CONFIDENTIAL 
On a following night, subjects will return to Madison for physiology studies, overnight 
polysomnography and Holter/heart rate variability measurements*, vascular function measures 
and control of breathing studies the 2nd night to assess the study drug effects on variables 
measured at baseline. Subjects will be instructed not to consume caffeine (both solid and liquid 
forms) after midnight the night before they are admitted to the CRU and after 8pm on night 1. 
Subjects may consume caffeine after the physiology study on Day 1 and before 8pm on Day 1. 
Subjects will also be instructed not to smoke after 10 pm once they are admitted to the CRU until 
discharged. Subjects will not be allowed to participate in the study if they are unable to refrain 
from smoking after 10 pm for up to 24 to 48 hours once they are admitted to the CRU until 
discharged.  
 
*Holter and Heart rate variability measurements are optional and will only be offered to UW 
subjects.  They can choose to opt-in or opt-out of those procedures. Subjects who are non-CPAP 
users will have Holter monitoring done at Visits 3 and 6, but will not have Holter monitoring 
performed at Visits 2 and 5.  The Holter monitors will be connected by UW Sleep Laboratory 
Technicians or study team personnel. Subjects may disconnect the Holter monitor on their own, 
or they may come back to UW-Madison to have the Holter monitor removed.  A postage-paid 
envelope will be provided to subjects to return to Holter monitor by mail. 
3.2 Primary Study Endpoints 
The primary outcome variable is the difference in the individual slope of the MSNA – SaO2 
response curve at 6 weeks and baseline between treatment groups.  Basal normoxic MSNA 
and MSNA responses during hypoxia will be compared between the three treatment groups.  For 
purposes of quantification, MSNA is expressed as burst frequency (bursts/min), burst amplitude 
(arbitrary units) and total minute activity (burst frequency x mean burst amplitude).  Total minute 
activity of mSNA is an effective way to capture the change in both burst frequency and burst 
amplitude.  Both amplitude and burst frequency are influenced by disease states such as OSA 
and by environmental conditions such as hypoxia.  Postganglionic mSNA in the right peroneal 
nerve will be recorded directly using the microneurography technique (95). The neural signals are 
passed to a differential preamplifier, an amplifier, a band-pass filter, and an integrator (time 
constant=100 ms). Placement of the recording electrode within a muscle nerve fascicle is 
confirmed by the presence of muscle twitches in response to electrical stimulation, the 
appearance of afferent activity in response to tapping or stretching of muscle, but not gentle 
stroking of skin, in the appropriate receptive fields, and other criteria.  Once an acceptable neural 
recording (signal-to-noise ratio >3:1) is obtained, the subject is instructed to maintain the leg in a 
relaxed position for the duration of the study. Sympathetic bursts will be identified by computer-
assisted inspection of the mean voltage neurogram.   
 
3.3 Secondary Study Endpoints 
Secondary outcome variables will include assessment measures of cerebral and forearm blood 
flow during basal conditions and during graded hypoxia before and after study drug treatment.  
Plasma catecholamine concentrations before and after study drug and change in plasma 
catecholamines during hypoxia will be assessed as a secondary indicator of sympathetic 
activation.  Other secondary outcome variables will be assessed before and after study drug 
treatment, including: ventilation, aortic pulse wave velocity (a measurement of vascular stiffness), 
aortic augmentation index, % vasodilation during flow-mediated vasodilation (measurement of 
vascular endothelial function), apnea threshold, apnea-hypopnea index, % time spent below 90% 
oxygen saturation, heart rate variability, holter electrocardiogram measurements, and mean 24-


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 8 
Version: 23, 7-25-17 
CONFIDENTIAL 
hour blood pressure, mean nighttime blood pressure, mean daytime blood pressure, blood 
pressure load, and night/day blood pressure ratio.    
   
4 Subject Selection and Withdrawal 
4.1 Inclusion Criteria 
 
Inclusion Criteria 
• Males and females between ages of 21 and 65 years inclusive 
• Apnea hypopnea index or respiratory disturbance index (RDI) greater than or equal to 25 events/hour 
(diagnosed by Polysomnography or home study) 
• Subjects eligible for CPAP or BiPAP therapy. If subject is on CPAP or BiPAP, he/she must be 
on treatment for a minimum of 3 months prior to Visit 2. 
• Hypertension by clinical history of high blood pressure readings or diagnosis of hypertension 
(may be controlled with non-exclusionary medications) or average blood pressure ≥ 140/90 
mm Hg (only require systolic OR diastolic to be above threshold, not both for 
inclusion).  Blood pressures used for inclusion based on average blood pressure ≥ 140/90 
mm Hg  will be (using two measurements in prior 12 months – in PHI or other records OR 1 
prior elevated blood pressure and 1 blood pressure at screening) 
 
4.2 Exclusion Criteria 
 
Exclusion Criteria 
• If subject not using CPAP, having AHI > 60 events/hour or oxygen saturation ≤ 65% during 
sleep 
• Presence of clinical CV disease (coronary artery disease, angina, arrhythmias (subjects with 
sinus arrhythmias will be reviewed by PI for enrollment), stroke, TIA, cor pulmonale, etc.), 
heart failure, bruits, or diabetes mellitus by clinical diagnosis/history 
• Presence of pulmonary disease that results in significant hypoxemia (resting SaO2 < 88%) 
• Hypertriglyceridemia (triglycerides >300 mg/dL).   
• Diabetes (fasting plasma glucose > 125 mg/dL) 
• Subjects taking angiotensin converting enzyme inhibitors, angiotensin receptor antagonists, 
renin inhibitors, potassium-sparing diuretics (without accompanying loop/thiazide diuretic), 
allopurinol, oxypurinol, febuxostat, amoxicillin, ampicillin, azathioprine or mercaptopurine. 
*(see below for examples) 
• Subjects with chronic kidney disease (Serum creatinine >1.5 mg/dL) or history of significant 
hyperkalemia (Serum potassium > 5.2 mEq/L) with ARB therapy 
• Subjects with history of angioedema 
• Subjects with bilateral, modified radical or radical mastectomies 
• Subjects who have a Serum potassium > 5.0 mEq/L at the screening visit 
• Female subjects who are pregnant (determined by urine pregnancy test) or breastfeeding 
• Subjects with active MRSA or VRE (vancomycin resistant enterococcus) infection 
• History of adverse reaction to allopurinol,losartan, or zolpidem** 
• Subjects who are currently receiving cancer treatment or have received cancer treatment in 
the past 1 year. 
• Subjects who cannot swallow oral capsules 
• Subjects who are hospitalized or who have been recently hospitalized (last 2 weeks) 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 9 
Version: 23, 7-25-17 
CONFIDENTIAL 
• Inability to comply with or complete the protocol or other reasons at the discretion of the 
investigators 
 
*angiotensin converting enzyme inhibitors (lisinopril, enalapril, etc.) 
Angiotensin receptor antagonists (losartan, valsartan, etc.) 
Renin inhibitors (aliskiren) 
Potassium sparing diuretics (spironolactone, eplerenone, triamterene without hydrochlorothiazide, etc.) 
 
**If adverse reaction to zolpidem, may opt out of control of breathing study 
4.3 Subject Recruitment and Screening 
 
Screening Visit (Visit 1):  Subjects will come to the CRU or their local recruitment site for a brief 
screening visit.  Subjects should be instructed to be fasting for at least 12 hours prior to 
their visit and patients should hold any blood pressure medications on the morning of 
VISIT 1 (they may take at end of VISIT or after).  Investigators will explain the study, answer 
questions, and complete the informed consent and HIPAA process.  Potential subjects identified 
through sleep databases and registries will be contacted via telephone to obtain verbal consent 
or declination for participation in this study. A waiver of written documentation of informed consent 
has been approved by the IRB to obtain consent via telephone. At the in person screening visit, 
the subject will then sign a consent form witnessed by the study coordinator or investigator. The 
subject will then receive a signed copy of this consent form. All copies of the consent forms will 
be maintained in the subject’s binder. The subject  will then undergo: 
• 
a history and physical with the CRU nurse practitioner 
• 
undergo a 12-lead electrocardiogram (if the electrocardiogram has abnormalities of clear 
clinical significance, then a copy of the EKG with a letter explaining those abnormalities 
will be sent to the subject with instructions to share with his/her physician). 
• 
take a urine pregnancy test (for females of child-bearing age),  
• 
have a fasting blood sample taken to measure serum cholesterol, potassium, creatinine, 
uric acid, and glucose concentrations 
If the subject declines participating in the study, any personal and health related information will 
not be retained and will be destroyed upon completion of the telephone call. 
 
Subjects will not receive study medications until the end of Visit 3 and will be instructed to bring 
their study medication bottles with them to Visit 4 (two weeks after starting medication) and Visit 
6 (6 weeks after starting medication).  Study drug diary evaluation will be used to evaluate study 
drug adherence.  Subjects will receive an auto-CPAP machine or CPAP compliance card to use 
during the study. Subjects will not receive payment for Visit 6 if they do not return the CPAP 
machined or CPAP compliance card. Subjects will be instructed to maintain the baseline CPAP 
use throughout the study.  Subjects will complete baseline 24-hour ambulatory blood pressure 
monitoring (Visit 2) prior to receiving either study drug.  This will take place prior to their overnight 
visit to Madison (Visit 3).    
    
Study subjects will be recruited from the Wisconsin Sleep Center in Madison, Marshfield Clinic in 
the greater Marshfield, WI area, Aurora Bay Care in Green Bay, and Gundersen Health System 
in La Crosse, WI.  Wisconsin Sleep initiated a research subject registry and database in August, 
2008, and the subject registry invites all patients referred to clinic or laboratory to consider being 
contacted about research projects.  Data that is maintained in the subject database includes 
patient name, date of birth, gender, contact information, sleep study data, and diagnoses.  Drs. 
Dopp and Teodorescu will also recruit subjects at Wisconsin Sleep in their own clinics.  Subjects 
will be recruited in clinics at Wisconsin Sleep and after their sleep study at Wisconsin Sleep or 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 10 
Version: 23, 7-25-17 
CONFIDENTIAL 
after a home study done by Wisconsin Sleep.  Additional advertisements will be made available 
to subjects at other Sleep laboratories and clinics in the area, as well as at other clinics and 
medical centers in the greater Madison area (including, but not limited to:  Baraboo sleep 
lab/clinic, Sauk prairie hospital sleep lab/clinic, St. Mary's sleep lab/clinic, Portage sleep lab/clinic, 
Reedsburg sleep lab/clinic, etc. Permission will be obtained prior to posting flyers or 
advertisements.  An opt-out letter will be sent to qualifying subjects and the subjects will have 7 
days to opt out before we will contact them to provide further information about the study and 
inquire as to their interest.  Additional subjects who have participated in previous studies by 
investigators or other research groups who have expressed interest in future sleep apnea studies 
will be contacted about this study. Recruitment will also be completed with the use of social media 
and the internet. We will advertise on the UW Job Board, Craigslist, and Wisconsin Sleep’s 
website. We will implement an email blast to the UW Madison students, faculty and staff as well 
as use campus and local newspapers and other media. 
 
Recruitment at other sites: 
 
We will also recruit from the sleep laboratories and sleep clinics at Marshfield Clinic, Marshfield, 
WI, Gundersen Health System, La Crosse, WI, and Aurora Bay Care, Green Bay, WI.  Subjects 
will be recruited by the responsible investigators at the individual sites with the help of study 
coordinators.  Subjects will be identified following sleep studies in the sleep laboratories and 
during visits to clinic.  Subjects will be contacted and approached about study participation by 
individuals involved in his/her clinical care.  Subjects may also be identified using advertisements 
in the community and at the local medical centers if necessary.   
 
4.4 Early Withdrawal of Subjects 
4.4.1 When and How to Withdraw Subjects 
Allopurinol  
If a subject experiences an adverse effect that is thought to be significant or serious (e.g. renal 
impairment, maculopapular rash, etc.) subjects’ participation in the study will be ended.   
 
Losartan 
If a subject experiences clinically significant/serious adverse effects (angioedema, myalgias, 
acute renal failure), their participation will be ended.  If the subject experiences increases in serum 
creatinine (> 0.5 mg/dL) or serum potassium (above 5.0 mEq/L) the subject’s participation in the 
study will be ended.  Blood pressure will also be measured at the two week  monitoring visit (Visit 
4).  The dose of losartan will increase to 100mg daily if systolic blood pressure is > 120 mm Hg.  
For a systolic blood pressure between 100-120 mm Hg the subject may have dose increased if 
approved by study physician.  If the subject’s blood pressure is < 100/60 mm Hg and if the patient 
is dizzy and symptomatic during the study, the subject will discontinue study medication and go 
off study.  This can be done via a phone encounter or via an additional (unexpected) visit.  It is 
possible the subject’s participation will be terminated if the study physicians feel that is 
appropriate.   
 
Subjects may also be withdrawn from the study if they are diagnosed with a new and significant 
medical problem (e.g. they are hospitalized for an acute illness or major medical condition).  
Subjects may also be withdrawn from the study if they consistently fail to adhere to the protocol 
requirements.  Study physicians will communicate with subjects’ regular physician about care 
plans upon their withdrawal from the study.   
 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 11 
Version: 23, 7-25-17 
CONFIDENTIAL 
Additional (Unexpected) Visit: 
If a subject experiences an adverse event that requires them to be seen by study staff, they should 
be scheduled for a visit to their local center as soon as is feasible.  The PIs should be notified 
immediately and the physician at the local site should see the subject during the visit.  Depending 
on the complaint, the AE should be evaluated with appropriate objective measures (blood tests, 
blood pressure/heart rate measurements, etc.) and the case report forms for “Additional Visit” 
should be filled out (in subject binder).  These forms include: 
 
AE form 
Vitals form 
Additional visit form 
AE log 
 
The study physician will determine if the subject should be discharged from the study (if subject 
is discharged, then complete the “subject off study” form).  If the subject is discharged from the 
study, then he/she should have the 30-day post study evaluation performed 30 days after their 
study discharge (see below).    
4.4.2 Data Collection and Follow-up for Withdrawn Subjects 
We do not expect mortality or significant morbidity from the study.  However, all subjects will be 
instructed to contact investigators if any problems arise after study participation is discontinued.  
Investigators will contact subjects who are withdrawn from the study 30 days after study 
discontinuation and periodically thereafter per study physician discretion if necessary.  Data will 
be obtained and kept from any follow-up communications. 
5 Study Drug  
5.1 Description 
Allopurinol and losartan are both FDA approved medications.  Losartan is FDA approved for the 
treatment of hypertension.  Allopurinol is approved for the treatment of gout.  Study medications 
will be over encapsulated with lactose and all subjects will receive placebo with their active study 
medication in a double dummy design.  Zolpidem will also be given to subjects (upon discharge 
from the CRU at Visits 3 and 6) to help them sleep during the control of breathing studies.   
 
5.2 
Treatment Regimen 
Losartan 50 mg daily for two weeks, then increased to 100mg daily for 4 weeks 
 
OR 
 
Allopurinol 300 mg daily for 6 weeks  
 
OR 
 
Placebo capsule daily for 6 weeks 
 
All three medications will be taken orally (in capsules) by subjects.  The treatment regimen is 
summarized in Table 1. 
 
Table 1.  Treatment Regimen 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 12 
Version: 23, 7-25-17 
CONFIDENTIAL 
Arm 
Treatment 
Administered as: 
PRODUCT TYPE 
Active Losartan 
50mg Capsule  
Bottle A 
Active Losartan 
100mg Capsule 
Bottle B 
Placebo  
Allopurinol 
300mg 
Capsule 
Group I 
Losartan 
1 capsule  daily 
on Days 1-14 
 
1 Capsule daily 
on Days 15-42 
None 
None 
Group II 
Allopurinol 
None 
None 
None 
1 
Capsule 
Daily 
on 
Days 1-42 
Group III 
Placebo 
None 
None 
1 Capsule 
Daily 
on 
Days 1-42 
None 
Losartan subjects will take capsules from Bottle A for days 1-14, and capsules from Bottle B for days 15-
42.  If the subject’s systolic blood pressure is > 120 mm Hg, the subject will continue to take capsules daily 
from Bottle A (days 15-42).  If SBP is between 100-120 mm Hg, the subject may switch to bottle B with 
physician approval.   
 
Allopurinol and Placebo will also have Bottle A and B for each treatment and will contain identical drug in 
Bottle A and Bottle B.  Subjects will transition from Bottle A to Bottle B after day 14 in the allopurinol and 
placebo group.  This is done to maintain study blinding. 
5.3 Method for Assigning Subjects to Treatment Groups 
Randomization will be performed by the Pharmaceutical Research Center (PRC) using a scheme 
designed by the PRC and ICTR statisticians.  Subjects will be randomized and stratified based 
on their baseline use of continuous positive airway pressure (permuted block randomization).  
Subjects will be classified into the following three categories of baseline CPAP use:  1) < 1 hour 
per night; 2) between 1 and 4 hours per night; and 3) > 4 hours per night.  Subjects not using 
CPAP will be classified in the < 1 hour per night group.  Subjects will be instructed to continue the 
same CPAP use during the study as they were using at study entry. 
5.4 Preparation and Administration of Study Drug 
Study drug will be over encapsulated by the University of Iowa Pharmaceuticals (UIP) and stored 
and distributed by the University of Wisconsin Pharmaceutical Research Center and blinded 
according to a pre-designed scheme.  Subjects will be randomized at the end of visit 1 (screening 
visit) and will receive study drug at their baseline physiology study visit (visit 3).  UIP is an FDA-
registered facility compliant with 21 CFR parts 11, 210, and 211, licensed by the DEA to handle 
controlled substances (Schedules I-V), and capable of handling potent/cytotoxic materials.  UIP 
has been a cGMP facility for over 30 years. 
5.5 Subject Adherence Monitoring 
All subjects will receive instructions on how to take and record medication adherence in a diary.  
Subjects will not receive study medications until the end of the Physiology Visit in Madison (Visit 
3) and will be instructed to bring their study medications with them to Visit 4 (two weeks after 
starting medication at local study site) and Visit 6 (Six weeks after starting study medication – in 
Madison). Serum uric acid concentrations and plasma renin activity will be measured at baseline 
and after 6 weeks of study medication and will provide an indirect measure of drug adherence in 
subjects receiving allopurinol and losartan.   Adherence to CPAP therapy will be monitored at Visit 
4 (two weeks after starting medication) and Visit 6 (Six weeks after starting study medication).  
CPAP adherence can be determined from the electronic data card in the CPAP machine and 
downloaded into a printable report.  Each site will have a card reader and software for 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 13 
Version: 23, 7-25-17 
CONFIDENTIAL 
downloading the information from subjects’ CPAP machines at Visit 4. Subjects on BiPAP will use 
their machines instead of ours, and we will attempt to get their adherence data from them. 
 
5.5.1 CPAP Before and During Study 
Subjects who are using CPAP or BiPAP prior to the study will be asked to provide adherence data 
for the time period prior to them entering the study. Subjects who require CPAP or BiPAP must 
be on treatment for a minimum of 3 months prior to Visit 2. This information will be obtained from 
their CPAP/BiPAP machine or from their home health provider that services their CPAP/BiPAP 
machine. We will also ask for their permission to contact their sleep physician to obtain a copy of 
their sleep study report. This will help us to determine how severe their sleep apnea is. Consent 
to obtain this information from home health providers and sleep physician will be obtained in the 
informed consent document.  Subjects who qualify for the study will be instructed to continue to 
use CPAP/BiPAP the same amount during the study as they were using prior to the study.  If 
subjects are not using CPAP/BiPAP prior to study entry, they will be instructed to continue not 
using CPAP/BiPAP during the study.   
 
At Visit 3, subjects will be given a CPAP machine or CPAP compliance card to use during the 
study.  Subjects using BiPAP will continue to use their own BiPAP machine. We will set the 
pressure on this machine to the same pressure the subject was using on their machine at home.  
Subjects will bring their CPAP/BiPAP machine or compliance card with them to Visit 4 (at local 
site) and Visit 6 (in Madison) to have the adherence data checked.  Investigators should remove 
the compliance card from the CPAP machine and download the data at Visits 4 and 6.  After the 
download, the cards should be returned to the subjects’ CPAP machines.  Subjects will use the 
mask they had previously been using with the CPAP/BiPAP machine issued to them.  The 
Respironics CPAP machines given to subjects during the study will be capable of measuring 
residual breathing events to determine how effectively the machine is treating a subject’s sleep 
apnea. 
 
 
5.6 Prior and Concomitant Therapy 
Information about all medications (prescription, OTC and supplements) being taken will be 
collected at Study visits 1, 1A, 2, (if more than 90 days after Visit 1), 3, 4, 5, 6, and 7. Subjects 
are instructed to let the study coordinator know if they start any new medication while they are 
enrolled in the trial.  Other antihypertensive agents (diuretics, calcium channel blockers – see 
Appendix 3 for allowable blood pressure medications) will be allowed but doses should remain 
constant during the study period. Subjects taking non-hypnotic benzodiazepines, narcotic 
analgesics, quetiapine, or barbituates, on a daily basis or have taken them within 5 days (when 
using them p.r.n. (pro re nata or as needed) before the control of breathing study will not be 
allowed to participate in the control of breathing study (night 2, visits 3 and 6).  However, if a 
subject is taking a medication for sleep (e.g. hypnotic benzodiazepines, zaleplon, eszopiclone, 
etc.), they may participate in the control of breathing study if zolpidem may be taken instead of 
their usual sedative hypnotic for the control of breathing study. Subjects should not take PDE-5 
inhibitors (ie. sildenafil (Viagra), vardenafil (Levitra), or tadalafil (Cialis) within 72 hours (3 days) 
before or after taking nitroglycerin.  If a subject has taken any of these medications within that 
timeframe he/she will not be able to participate in the Stein lab portion of Visit 3. 
 
The following medications are not allowed to be taken: 
Angiotensin converting enzyme inhibitors, angiotensin receptor antagonists, renin-inhibitors, 
potassium-sparing diuretics, allopurinol, oxypurinol, febuxostat, amoxicillin, ampicillin, 
azathioprine or mercaptopurine. 
 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 14 
Version: 23, 7-25-17 
CONFIDENTIAL 
5.6.1 Pregnancy Testing and Birth Control 
Women of child-bearing age are eligible subjects for this study.  However, since study drugs are 
pregnancy rate “D” (losartan) and “C” allopurinol, female subjects will take a urine pregnancy test 
at Visit 1 to ensure they are not pregnant.  A urine pregnancy test will be repeated at Visit 1A if 
the study visit occurs more than 90 days after Visit 1. Furthermore, a urine pregnancy test will 
also be given at Visit 3 prior to study drug dispensing.  The Pregnancy and birth control case 
report form should be completed at Visit 1, Visit 1A (if applicable) and Visit 3.  On this case report 
form the method of birth control used should be documented and subjects are instructed to keep 
their birth control consistent throughout the study. Methods of birth control that are acceptable 
while participating in this study include oral contraceptives, hormonal contraceptives, hormonal 
implants, contraceptive patches, NuvaRing, intrauterine device hormonal, intrauterine device non-
hormonal, barrier method, spermicide, post-menopausal for greater than or equal to two (2) years, 
tubal ligation, bilateral oophorectomy, or hysterectomy, vasectomy, or abstinence. If a female is 
practicing abstinence, she should confirm this, including duration, and initial and date the case 
report form. 
 
5.7 Blinding of Study Drug 
Allopurinol and Losartan tablets will be over encapsulated and dispensed to subjects with a 
placebo using a double-dummy design.  Both subjects and investigators will be blinded to study 
drug assignment.  The PRC will have unblinding information should the need arise to unblind a 
subject’s study drug assignment. To obtain unblinding information, please call the University of 
Wisconsin Pharmaceutical Research Center at (608) 263-8863. 
 
5.8 Receiving, Storage, Dispensing and Return 
5.8.1 Receipt of Drug Supplies 
The PRC will order, receive and store all study medications for the trial.  The PRC complies with 
all Federal and State regulations and current good manufacturing practices. 
5.8.2 Dispensing of Study Drug 
The PRC performs all required study drug reconciliation and inventory procedures on a regular 
basis.  PRC drug record logs record the movement of study drugs out of PRC storage and all 
medications are checked by a PRC pharmacist prior to dispensing to the subject and study staff.  
Amount of study drug on hand and dispensed is constantly tracked in PRC records.  All study 
drugs will be dispensed by the PRC to subjects during Visit 3 in Madison.  Subjects will receive 
all study drugs for the 6-week treatment period at Visit 3 and will receive instructions from 
investigators prior to starting therapy.  Zolpidem (supplied by the PRC) will be given to subjects 
upon discharge from the CRU at VISITS 3 and 6 to be taken immediately before (9-10pm) the 
control of breathing study on night two.   
 
6 Study Procedures 
 
See Appendix 2 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 15 
Version: 23, 7-25-17 
CONFIDENTIAL 
6.1 Study Personnel 
Study subjects will be recruited from the Wisconsin Sleep Center in Madison (Dr. John Dopp, Dr. 
Barbara Morgan, and the study coordinator(s)); Marshfield Clinic in the greater Marshfield, WI 
area (Dr. Boero and study coordinator(s); Aurora Bay Care in Green Bay (Dr. Gapinski and study 
coordinator(s)); and Gundersen Health System in La Crosse, WI (Dr. Tobert and study 
coordinator(s)).  Dr. Dopp and Dr. Morgan will lead the overall project as coinvestigators at the 
University of WIsconsin-Madison.  WiNHR Central Administration Staff (Deb Kruser, Director, MJ 
Washburn, Regional Research Networks Coordinator, and Kelly Miller, Senior Research 
Coordinator) in collaboration with the UW Site Study Coordinator will also assist with data 
management, site communication and oversight of study conduct.  As described in section 8.7, 
CRIS Study Monitoring Personnel will perform monitoring throughout the project. 
6.2 Summary of Visits 
Subjects may complete some, but not all procedures without a protocol deviation occurring or 
affecting their study eligibility.  For logistical or other reasons, subjects may complete some of the 
procedures (but not all procedures) (i.e. in cases of non-exclusionary subject disabilities that 
preclude them from having certain procedures done or body habitus that does not allow 
completion of certain procedures.).  Reimbursement for participation will be pro-rated for Visits 3 
and 6 as in protocol section 11.2. 
 
Visit 1: Screen Visit (will be performed at each respective recruitment site)  
• 
informed consent, height and weight, heart rate, blood pressure, temperature, and 
electrocardiogram recorded, blood samples, urine pregnancy test, brief history (performed 
by study coordinator and local PI) 
• 
physical examination (local PI and UWHC medical staff (i.e., physician, physician’s 
assistant, or nurse practitioner)  for Madison subjects)    
Visit 1A: If visit 2 will occur more than 90 days after visit 1, subjects will undergo repeat 
screening procedures in a separate visit to the local study site (Visit 1A) including:  
 
Medical history, medication history, birth control assessment, blood samples, urine 
pregnancy test, AE assessment, electrocardiogram recorded (performed by the local PI, 
study coordinator, or medical staff (i.e., physician, physician’s assistant, or nurse 
practitioner)  
 
Visits 2 and 5:  Home Blood Pressure Monitoring (will be performed at each local site 
where subjects were originally enrolled)  
 
• 
Receive blood pressure monitor and Holter/heart rate variability monitor* to wear for 24 
hours along with the diary recording of activity each time they engage in activity and each 
time the blood pressure cuff inflates; subject returns to the study site the following day (all 
activities performed by coordinator--local PI consulted if necessary).  Subjects at UW-
Madison will come to the School of Pharmacy Clinical research space of the Office of 
Clinical Trials Examination Space (H6/136 CSC) to be set up with their 24-hour blood 
pressure cuff and Holter/heart rate variability monitor.  Adverse events will be assessed 
at visits 2 and 5. 
 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 16 
Version: 23, 7-25-17 
CONFIDENTIAL 
*Holter and Heart rate variability measurements are optional and will only be offered to UW subjects.  
They can choose to opt-in or opt-out of those procedures. Subjects who are non-CPAP users will have 
Holter monitoring done at Visits 3 and 6, but will not have Holter monitoring performed at Visits 2 and 5.”  
The Holter monitors will be connected by UW Sleep Laboratory Technicians or study team personnel. 
Subjects may disconnect the Holter monitor on their own, or they may come back to UW-Madison to have 
the Holter monitor removed.  A postage-paid envelope will be provided to subjects to return to Holter 
monitor by mail. 
 
Visits 3 and 6: (each visit requiring a one or two night stay in Madison at the CRU at UW 
and the VA Hospital for all subjects enrolled in the study) 
• 
Day 1:  Vital signs, Heart Rate; Forearm Blood Flow; Blood Samples; Cerebral blood 
flow; Nerve 
activity; Reduced 
oxygen 
exposure 
and 
increased 
oxygen 
breathing; overnight sleep study and heart rhythm measurements at the CRU to measure 
brain waves, respiration, blood oxygen level, snoring, leg movements, heart rhythm, and 
heart rate variability* (performed by PIs and sleep technician). A physical exam will also 
be obtained. If bruit are noted on physical exam the subject will no longer be able to 
participate in the study. The presence of a bruit would potentially indicate previously 
unrecognized cardiovascular disease.  
• 
Day two: Ultrasound of blood vessels and heart; non-invasive tonometry; assessment of 
CPAP use; assessment of daytime sleepiness by completing the Epworth sleepiness 
scale; Study Medication dispensation (V3) and collection (V6);  Control of Breathing 
Experiments during sleep stay at VA hospital.  Subjects will receive zolpidem (sleeping 
medication) upon discharge from the CRU to be taken around 10pm at the start of the 
control of breathing study.   (Ultrasounds performed in Dr. Stein’s laboratory; control of 
breathing studies performed by sleep technician).  Subjects who do not have someone to 
drive them home, do not want to take a complimentary cab home or do not want to go 
back to the CRU to sleep and have breakfast prior to having a driver/cab take them home 
from Visits 3 and 6 may opt out of the control of breathing study on the second night.  
When a subject arrives for Visits 3 and 6, he/she will be asked if they have a driver to take 
them home, need a cab ride home or if they agree to go back to the CRU to sleep and 
have breakfast prior to having a driver/cab take them home after the control of breathing 
study.   
• 
Visit 6 will also include AE Assessment and diary review in addition to the procedures 
mentioned above. 
*Holter and Heart rate variability measurements are optional and will only be offered to UW 
subjects.  They can choose to opt-in or opt-out of those procedures. Subjects who are non-CPAP 
users will have Holter monitoring done at Visits 3 and 6, but will not have Holter monitoring 
performed at Visits 2 and 5.”  The Holter monitors will be connected by UW Sleep Laboratory 
Technicians or study team personnel. Subjects may disconnect the Holter monitor on their own, 
or they may come back to UW-Madison to have the Holter monitor removed.  A postage-paid 
envelope will be provided to subjects to return to Holter monitor by mail. 
 
 
 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 17 
Version: 23, 7-25-17 
CONFIDENTIAL 
Visit 4:  Monitoring visit (performed at each local site where subject was enrolled) 
• 
measurement of blood pressure, heart rate, temperature, and height and weight, blood 
sample collection, Study drug compliance and diary review, AE assessment (all performed 
by local site coordinator or CRU nurses for Madison subjects)  
Visit 7: One month Follow-up (performed at each local site where subject was enrolled) 
• 
Subjects will be contacted by phone for completion of a final AE assessment (performed 
by local site coordinator)  
6.3 Sample collection 
Blood samples collected during Visits 1 and 4 will be processed for the specific laboratory 
parameter at the local site and assayed in the clinical labs (or couriered from LaCrosse to 
Marshfield to be assayed).  Samples will be collected and sent to the clinical laboratory for 
measurement of serum creatinine, serum potassium, serum glucose, lipid panels, serum uric acid, 
etc.  At Visits 3 and 6 at UW-Madison, blood samples will be drawn at rest while the subject is 
breathing room air.  Uric acid will be collected and assayed in the UW Hospital and Clinics Clinical 
Laboratory and other samples will be collected for research use.  These samples collected for 
research use will be centrifuged, aliquoted, and stored in the locked freezer space at the 
University of Wisconsin School of Pharmacy for future assay.  Stored blood samples will be 
stripped of identifiers and only labeled with study subject number and Visit number.  Plasma will 
be sent to the TTR Analytical Laboratory Core at the University of Iowa College of Pharmacy for 
plasma catecholamine measurement.  Other plasma will be assayed at the UW School of 
Pharmacy for oxidative stress markers.  
 
6.4 Study communication and oversight 
At the start of the trial, study binders will be set up for each study site with the current protocol 
version (will include protocol number and date).  At any time during the study when the 
protocol changes, a new protocol will be sent to each site to be included in the study binder 
and to be kept in a relevant study folder electronically with the study coordinator.  The new 
protocol will be sent along with a summary of protocol changes denoted separately to the 
study coordinator, the IRB and co-Investigators.  Discussion of protocol changes will also be 
an item of discussion at the bi-weekly WiNHR coordinator conference calls. 
 
When the UW HS IRB approves amendments to the protocol, copy of the approval will 
accompany the summary of changes approved and the new protocol version sent to the study 
sites.  Electronic copies of the approval documents will be forwarded to each site with the 
other electronic documents and stored electronically and in paper form in an IRB folder at 
each study site.  At each biweekly WiNHR coordinator conference call, reconciliation of all 
current approvals will be performed by all the investigators and their study binders and 
electronic files. 
 
Reporting of adverse events and unanticipated problems will be performed at all centers 
through the Principal Investigators to comply with the UW HS IRB requirements.  Investigators 
at all study sites will be instructed to notify the Principal Investigators of all adverse events 
immediately upon awareness of their occurrence.  The Principal investigators and the study 
coordinator in Madison will provide the information to the DMC and the IRB in the appropriate 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 18 
Version: 23, 7-25-17 
CONFIDENTIAL 
time frame.  Unanticipated problems will be discussed at each WiNHR biweekly coordinator 
conference call and at quarterly investigator meetings. 
 
For events that are unexpected, immediately life threatening or severely debilitating to other 
current subjects, and caused by or probably related to the study drug, the DMC and IRB will 
be notified immediately.   
 
Any deviations from the protocol at any of the research sites will be discouraged and if occur, 
will be reported to the UW HS IRB if the deviation or exception presents a significant safety 
risk to the subject or potential subject.  As per Good Clinical Practices, protocol deviations will 
be documented and reviewed at the quarterly investigator meetings.  If changes are necessary 
for the project, protocol modifications will be submitted to the HS IRB to change the protocol 
to fit the required changes. 
6.5 Unexpected Findings 
Whenever tests like the electrocardiogram (Visit 1), cardiac output (Visits 3 and 6), or serum 
laboratory tests are done, there is the chance of an unexpected finding. Subjects will be 
informed of all findings of clear clinical significance which may be revealed during the 
electrocardiogram, cardiac output procedure (limited echocardiography), and serum 
laboratory tests. Findings of clear clinical significance are those for which we generally know 
the risks of non-treatment and for which treatment may be available. If unexpected findings 
for the electrocardiogram, cardiac output, or serum laboratory tests are discovered, a letter 
describing the findings will be sent to the subject and to his/her physician with permission from 
the subject.  
 
 
7 Statistical Plan 
7.1 Sample Size Determination 
The primary outcome will be the change in the slope response of MSNA during progressive 
hypoxia from baseline to 6 weeks.  From previous data from our lab we have seen an approximate 
standard deviation for slope in OSA patients under similar progressive hypoxia of 8-10.   
 
Based on interim data, a power calculation for the differences and standard deviations in primary 
outcome variable estimated that we would have 90% power (this is increased power from our 
original sample size calculation that used 80%) to determine a difference of 3.07 (1.13 vs -1.94) 
at a significance level of 0.017 (Bonferroni adjusted alpha for 3 tests) if we had complete MSNA 
slope response data from 20 subjects per group.   
 
We had originally proposed to enroll 50 subjects per group.  Our interim analysis determined that 
if we obtain complete data in 20 subjects per group, we are likely to find a significant difference.  
We will enroll as many subjects as we can in the remaining time frame to give us the best possible 
chance at finding a difference.  Consistent with DMC recommendations, we will reduce our 
sample size to 100 subjects.  To account for drop-outs, we will ask to enroll 120 subjects to get 
100 subjects who proceed to randomization.     
 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 19 
Version: 23, 7-25-17 
CONFIDENTIAL 
7.2 Statistical Methods 
Data will be analyzed by a statistician blinded to drug assignment. Demographic comparisons to 
assess effectiveness of randomization will be examined between treatment arms by ANOVA and 
chi-square tests where appropriate.  Differences in baseline values for all variables between 
treatment arms will be assessed using ANOVA.  The paired differences in these same variables 
will be compared after the 6-week study period.  These values include all measured physiologic 
variables, such as resting heart rate, blood pressure, plasma catecholamine concentrations, 
MSNA, cerebral blood flow, forearm blood flow, flow-mediated vasodilation, pulse wave velocity, 
augmentation index, polysomnographic measures, Pcrit, apnea threshold, and others.   
 
The primary outcome variable is the difference in the individual slope of the MSNA – SaO2 
response curve at 6 weeks and baseline between treatment groups.  The MSNA – SaO2 response 
curve will be graphed with MSNA as the dependent variable and SaO2 as the independent 
variable during progressive hypoxia.  Slope response curves will also be calculated for heart rate, 
ventilation, forearm blood flow, cerebral blood flow, and blood pressure during hypoxia, hyperoxia 
and controlled breathing periods.  MSNA will be expressed as burst frequency (bursts/min), burst 
amplitude (arbitrary units) and total minute activity (burst frequency x mean burst amplitude). 
MSNA will be averaged over the 5-min measurement periods at baseline, 90%, 85%, 80% 
hypoxia, and during controlled breathing. During hyperoxic trials, 15-sec averages of MSNA will 
be obtained, and an average of the 4 trials will be computed. The slope response of MSNA during 
progressive hypoxia will be calculated and will be analyzed using ANOVA.   Changes in 
polysomnographic measures from baseline to the end of the 6-week treatment period between 
the treatment groups will be compared using ANOVA.  If significant ANOVA p-values are found, 
then post-hoc multiple comparison Tukey HSD tests will examine differences between each 
arms.  Multiple regression will be used to determine the influence of subject characteristics and 
hours of CPAP use on the primary outcome, and on the effect of treatment on the primary 
outcome. 
 
7.3 Subject Population(s) for Analysis 
This study will follow a Intent to Treat (ITT) philosophy.  Loss to follow-up is a possibility and can 
cause issues with final analysis because the “missingness” of the data may not be random.  We 
will try to determine if there is any cause for drop out and summarize the reasons accordingly.  
We have accounted for drop outs in our projected sample sizes to assure sufficient power to 
remain in our tests.  We will test the sensitivity of the analysis results by comparing the analysis 
between dropping those with missing data and utilizing last observation carried forward.  In regard 
to the measurements taken at each time period, if a subject is unwilling or unable to complete the 
full progressive hypoxia we will calculate the slope for that individual based on the data we were 
able to attain and use that in the final analysis. 
8 Safety and Adverse Events 
8.1 Definitions 
Unanticipated Problems Involving Risk to Subjects or Others 
Any incident, experience, or outcome that meets all of the following criteria:  
• Unexpected in nature, severity, or frequency  (i.e. not described in study-related documents 
such as the IRB-approved protocol or consent form, the investigators brochure, etc) 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 20 
Version: 23, 7-25-17 
CONFIDENTIAL 
• Related or possibly related to participation in the research (i.e. possibly related means 
there is a reasonable possibility that the incident experience, or outcome may have been 
caused by the procedures involved in the research) 
• Suggests that the research places subjects or others at greater risk of harm (including 
physical, psychological, economic, or social harm). 
 
Adverse Event 
An adverse event (AE) is any symptom, sign, illness or experience that develops or worsens in 
severity during the course of the study.  Intercurrent illnesses or injuries should be regarded as 
adverse events.  Abnormal results of diagnostic procedures are considered to be adverse events 
if the abnormality: 
• results in study withdrawal 
• is associated with a serious adverse event 
• is associated with clinical signs or symptoms 
• leads to additional treatment or to further diagnostic tests 
• is considered by the investigator to be of clinical significance 
 
Serious Adverse Event 
Adverse events are classified as serious or non-serious.  A serious adverse event is any AE 
that fulfills at least one of the following criteria:  
• is fatal 
• is life-threatening 
• requires or prolongs hospital stay 
• results in persistent or significant disability or incapacity 
• is a congenital anomaly or birth defect 
• is an important medical event 
 
Important medical events are those that may not be immediately life threatening, but are clearly 
of major clinical significance.   They may jeopardize the subject, and may require intervention to 
prevent one of the other serious outcomes noted above.  For example, drug overdose or abuse, 
a seizure that did not result in in-patient hospitalization, or intensive treatment of bronchospasm 
in an emergency department would typically be considered serious.  
 
All adverse events that do not meet any of the criteria for serious should be regarded as non-
serious adverse events.  
 
Adverse Event Reporting Period 
Adverse events must be reported once the subject undergoes any study procedures and adverse 
events must be reported during the entire active study period (6 weeks) and for 30 days following 
the last administration of study treatment.  
 
General Physical Examination Findings 
At screening, any clinically significant abnormality shall be recorded as a preexisting condition.  
Throughout the study, any new clinically significant findings/abnormalities that meet the definition 
of an adverse event must also be recorded and documented as an adverse event.  
 
Post-study Adverse Event 
All unresolved adverse events will be followed by the investigator until the events are resolved, 
the subject is lost to follow-up, or the adverse event is otherwise explained.  At the last scheduled 
visit, the investigator will instruct each subject to report any subsequent event(s) that the subject, 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 21 
Version: 23, 7-25-17 
CONFIDENTIAL 
or the subject’s personal physician, believes might be related to participation in this study.  The 
investigator should notify the study sponsor and IRB of any serious adverse event or death 
occurring up to 30 days after the subject has discontinued or terminated study participation that 
may be related to this study.  Study staff will contact the subject 30 days after study participation 
has ended to ask about possible adverse effects after study drug discontinuation. 
 
Abnormal Laboratory Values 
A clinical laboratory abnormality should be documented as an adverse event if any one of the 
following conditions is met:  
• The laboratory abnormality is not otherwise refuted by a repeat test to confirm the 
abnormality 
• The abnormality suggests a disease and/or organ toxicity (e.g. serum creatinine increases 
more than 0.4 mg/dL) 
• The abnormality is of a degree that requires active management; e.g. change of dose, 
discontinuation of the drug, more frequent follow-up assessments, further diagnostic 
investigation, etc. 
 
Hospitalization, Prolonged Hospitalization or Surgery 
Any adverse event that results in hospitalization or prolonged hospitalization will be documented 
and reported as a serious adverse event unless specifically instructed otherwise in this protocol.  
Any condition responsible for surgery should be documented as an adverse event if the condition 
meets the criteria for an adverse event.  
 
Neither the condition, hospitalization, prolonged hospitalization, nor surgery are reported as an 
adverse event in the following circumstance: 
• Hospitalization or prolonged hospitalization for diagnostic or elective surgical procedures 
for a preexisting condition.  Surgery should not be reported as an outcome of an adverse 
event if the purpose of the surgery was elective or diagnostic and the outcome was 
uneventful. 
8.2 Recording of Adverse Events 
At each contact with the subject, the investigator will seek information on adverse events by 
specific questioning and, as appropriate, by examination.  Information on all adverse events will 
be recorded immediately in the source document (AE Assessment form), and also in the 
appropriate ongoing subject AE log.  All clearly related signs, symptoms, and abnormal diagnostic 
procedures results will be recorded in the source document. 
 
All adverse events occurring during the study period will be recorded.  The clinical course of each 
event will be followed until resolution, stabilization, or until it has been determined that the study 
treatment or participation is not the cause.  Serious adverse events that are still ongoing at the 
end of the study period will be followed up to determine the final outcome.  Any serious adverse 
event that occurs up to 30 days after Visit 6 and is considered to be possibly related to the study 
treatment or study participation will be recorded and reported immediately. 
8.3 Reporting of Serious Adverse Events and Unanticipated Problems 
All serious adverse events (AE) will be reported to the appropriate authorities, including the UW 
IRB and DMC in the required timeframe.  Adverse events and SAEs will be reported to the DMC 
and study sponsor (NHLBI) within the same time frame as required for the IRB (see 8.3.1 and 
8.3.2 below). 
 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 22 
Version: 23, 7-25-17 
CONFIDENTIAL 
Investigators must conform to the adverse event reporting timelines, formats and requirements of 
the various entities to which they are responsible, but at a minimum those events that must be 
reported are those that are: 
• related to study participation, 
• unexpected, and  
• serious or involve risks to subjects or others  
• Multiple occurrences of an AE that, based on an aggregate analysis, is determined to 
be an unanticipated problem. There should be a determination that the series of AEs 
represents a signal that the AEs were not just isolated occurrences and involve risk to 
human subjects. 
• An AE that is described or addressed in the investigator’s brochure, protocol, or 
informed consent documents that occurs at a specificity or severity that is inconsistent 
with prior observations OR for which the rate of occurrence in the study represents a 
clinically significant increase in the expected rate of occurrence.  
• Any AE or safety finding (e.g., based on animal or epidemiologic data) that would cause 
the sponsor to modify the investigator’s brochure, study protocol, or informed consent 
documents, or would prompt other action by the IRB to ensure the protection of human 
subjects.  
 
(see definitions, section 8.1).   
 
If the report is supplied as a narrative, the minimum necessary information to be provided at the 
time of the initial report includes: 
• 
Study identifier 
• 
Study Center 
• 
Subject number 
• 
A description of the event 
• 
Date of onset 
• 
Study AE Assessment form 
• 
Current status 
• 
Whether study treatment was discontinued 
• 
The reason why the event is classified as 
serious 
• 
Investigator assessment of the association 
between the event and study treatment 
8.3.1 Investigator reporting: notifying the IRB 
All adverse events and serious adverse events will be reported to the IRB at the appropriate and 
required time intervals.  This will include acutely and yearly at the annual protocol renewal.  
 
Will be reported immediately:  
• Any serious adverse event that occurs any time during or up to 30 days after the research 
study, which in the opinion of the principal investigator meets all 3 of these criteria: 
1. Is Unexpected (An event is “unexpected” when its specificity and severity are not 
accurately reflected in the protocol-related documents, such as the IRB-approved 
research protocol, any applicable investigator brochure, and the current IRB-approved 
informed consent document and other relevant sources of information, such as product 
labeling and package inserts.) 
2.  Is Related to the research procedures (An event is “related to the research procedures” 
if in the opinion of the principal investigator or sponsor, the event was more likely than 
not to be caused by the research procedures.) 
3. Is Immediately life-threatening or severely debilitating to other current subjects 
Will be reported within 14 days: 
• 
Any other adverse events listed below will be reported within 14 days 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 23 
Version: 23, 7-25-17 
CONFIDENTIAL 
1. A single occurrence of a serious, unexpected event that is uncommon and strongly 
associated with drug exposure (such as angioedema); 
2. A single occurrence, or more often a small number of occurrences, of a serious, 
unexpected event not commonly associated with drug exposure, but uncommon in the 
study population. 
3. Multiple occurrences of an AE that, based on an aggregate analysis, is determined to be 
an unanticipated problem. There should be a determination that the series of AEs 
represents a signal that the AEs were not just isolated occurrences and involve risk to 
human subjects (e.g. a comparison of rates across treatment groups reveals higher rate 
in the drug treatment arm versus a control). 
4. An AE that is described or addressed in the protocol or informed consent documents that 
occurs at a specificity or severity that is inconsistent with prior observations OR for which 
the rate of occurrence in the study represents a clinically significant increase in the 
expected rate of occurrence. 
5. Any AE or safety finding (e.g., based on animal or epidemiologic data) that would cause 
the sponsor to modify the investigator’s brochure, study protocol, or informed consent 
documents, or would prompt other action by the IRB to ensure the protection of human 
subjects. 
Will be reported annually at continuing progress review: 
• Occurred locally (or at any WiNHR study sites); 
• Are related to the research study but unexpected;  
• Were not assessed as placing subjects or others at increased risks (including 
physical, psychological, economic, or social harm) than was previously known 
or recognized;  
• Were not assessed as resulting in new information that needed to be 
disseminated to participants; and  
8.3.2 Sponsor reporting:  
Notifying NHLBI 
The investigators will appropriately notify NHLBI using expedited reporting to the NHLBI Program 
Officer for serious adverse events and unexpected problems. The reporting of adverse events 
and SAEs will be reported to NHLBI within the same timeframe as required by the IRB (see 8.3.1 
above).  Expedited safety reports will be sent concurrently to the DMC and NHLBI and the DMC 
response will be forwarded to NHLBI within 14 days. 
8.3.3 Sponsor reporting: Notifying the FDA 
 
The investigators will appropriately notify the FDA in safety reports. The following describes the 
safety reporting requirements by timeline for reporting and associated type of event: 
 
• Within 7 calendar days 
Any study event that is: 
– associated with the use of the study drug 
– unexpected,  
– fatal or life-threatening, and  
• Within 15 calendar days 
Any study event that is: 
– associated with the use of the study drug, 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 24 
Version: 23, 7-25-17 
CONFIDENTIAL 
– unexpected, and 
– serious, but not fatal or life-threatening 
-or- 
– a previous adverse event that was not initially deemed reportable but is later found to 
fit the criteria for reporting (reporting within 15 calendar days from when event was 
deemed reportable). 
 
8.4 Unblinding Procedures 
Unblinding of the study medication will be done in emergent circumstances where the identity of 
the study medication needs to be known.  All efforts will be made to maintain blinding except in 
the case of urgent medical necessity.  If a subject needs to be unblinded, the study staff should 
contact the PIs and the PRC and the DMC for unblinding (using contact information below).  The 
PI will work with the PRC to unblind the situation without compromising the blinding of remaining 
subjects.  The investigator will follow all NIH and institutional rules for the timely and appropriate 
notification of the governing IRB.  In most cases, the unblinding will be part of managing an SAE, 
and will be reported with the SAE, however, in cases where unblinding was not associated with 
an SAE, such actions should be reported in a timely manner.   
 
Principal Investigators: 
 
John Dopp:  
 
(608) 265-9352 (office phone); (608) 628-5756 (mobile) 
Barbara Morgan: 
(608) 265-2087 (office phone) 
 
Pharmaceutical Research Center (PRC):  (608) 263-8863 
 
Data Monitoring Committee (DMC):  DMC contact Amy Siedschlag (608) 263-9508 
8.5 Stopping Rules  
 
In case there is a significant beneficial or harmful effect due to one or both of the treatment arms, 
which would require early termination of the study, we plan on having three interim data analyses.  
Interim analysis will be performed after approximately 33%, 66%, and 100% of the subjects 
complete the 6-week study period.  Stopping criteria for benefit/harm will be based on the primary 
outcome variable analysis with O'Brien-Fleming alpha spending thresholds for three interim 
analyses.  These thresholds are: p=0.0005 at look 1, p=0.01413 at look 2, and p=0.04507 at final 
analysis.  The DMC, concurrent with and additional to the interim analyses, will meet on a 
quarterly basis to evaluate the protocol, subject accrual rates, subject retention, adverse events, 
and other aspects of the study.  In those meetings which correspond to the times determined for 
interim analyses the DMC will also evaluate the benefit or harm of the treatments. 
8.6 Independent Data and Safety Monitoring Board 
The Data and Safety Monitoring Committee used for this trial will be the University of Wisconsin 
Institute for Clinical and Translational Research DMC.  The project’s statistician, Dr. DeMets sits 
on the committee along with other ICTR personnel to oversee the project. Mr. Hetzel (our study 
statistician) will make the data presentations to the ICTR DMC.  Dr. DeMets will recuse himself 
from discussion of this project during DMC meetings to avoid conflict of interest for discussion of 
this protocol.  In case there is a significant beneficial or harmful effect due to one or both of the 
treatment arms, which would require early termination of the study, we plan on having three 
interim data analyses.  Interim analysis will be performed after approximately 33%, 66%, and 
100% of the subjects complete the 6-week study period.  Stopping criteria for benefit/harm will be 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 25 
Version: 23, 7-25-17 
CONFIDENTIAL 
based on the primary outcome variable analysis with O'Brien-Fleming alpha spending thresholds 
for three interim analyses.  These thresholds are: p=0.0005 at look 1, p=0.01413 at look 2, and 
p=0.04507 at final analysis.  The DMC, concurrent with and additional to the interim analyses, will 
meet on a quarterly basis to evaluate the protocol, subject accrual rates, subject retention, 
adverse events, and other aspects of the study.  In those meetings which correspond to the times 
determined for interim analyses the DMC will also evaluate the benefit or harm of the treatments. 
 
Data will be summarized and provided directly to the DMC through the OnCore CRM clinical trial 
software used for data collection during the study.  The DMC will keep minutes of its meetings 
and the PIs and all study staff will receive verbal and written summaries of their reviews and 
recommendations. 
 
8.7  Study Coordination and Data Monitoring 
The WiNHR Central Administrative Office personnel based at the University of Wisconsin, along 
with UW ICTR Clinical Research Infrastructure System (CRIS) Study Monitoring Service 
personnel, will complete Site Initiation Visit procedures at each performance site after IRB 
approval is confirmed and before enrollment of any subjects into the study.  The Site Initiation 
Visit will include a summary of all monitoring activities and expectations during the study.  The 
WiNHR Central Administrative Office personnel will monitor study activities at each site through 
biweekly study personnel conference calls and through weekly assessment of OnCore study 
data.  In addition, the study will be formally monitored using the the (CRIS) Study Monitoring 
Service.  Beyond the Site Initiation Visit activity described above, CRIS Monitoring personnel will 
perform study monitoring at each performance site following enrollment of their first 
subject, ongoing interim study monitoring every 3-6 months throughout the study, along with a 
closeout study monitoring visit upon completion of the study.  Finally, this study will utilize the UW 
ICTR Data and Safety Monitoring Committee.  The project’s statisticians, Dr. DeMets and Dr. 
Hetzel will sit on the committee along with other ICTR personnel to oversee the project.  Data will 
be summarized and provided directly to the DMC through the OnCore CRM clinical trial software 
used for data collection during the study.  The DMC will keep minutes of its meetings and the PIs 
and all study staff will receive verbal and written summaries of their reviews and 
recommendations. 
 
The University of Wisconsin will be the lead site of the study.  Specific responsibilities for 
coordination inherent to being a lead site are as follows: 1) WiNHR will facilitate biweekly study 
coordinator conference calls for discussion of current accrual and study-specific questions; 2) 
WiNHR Central Administration personnel (Deb Kruser, WiNHR Director, MJ Washburn, WiNHR 
Regional Research Networks Coordinator, Kelly Miller, WiNHR Senior Research Coordinator and 
Greg Guilfoil, WiNHR Senior Research Coordinator.) will serve as first-line communication conduit 
for questions from outlying sites and will refer questions, as necessary, to the UW-Madison 
Investigators; 3) WiNHR Central Administration Personnel will provide regular updates to the 
WiNHR Advisory Committee regarding study progress and pertinent updates; 4) UW-Madison 
and other site Investigators will participate on WiNHR biweekly study coordinator calls, as needed; 
5) UW-Madison Investigators, working with WiNHR Central Administration, will facilitate ongoing 
site Investigator teleconferences and/or  live meetings for discussion of study progress; 6) UW 
Madison Investigators will participate with WiNHR Central Administration personnel in Site 
Initiation Visits which will be completed prior to the enrollment of subjects at any performance 
sites; 7) WiNHR Central Administration personnel will work with ICTR Study Monitoring Service 
personnel to coordinate their site visits; 8) the UW-Madison study coordinator and sleep tech will 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 26 
Version: 23, 7-25-17 
CONFIDENTIAL 
perform study-specific activities with enrolled subjects and will communicate regularly with 
WiNHR Central Administration personnel and the Study Investigators. 
9 Data Handling and Record Keeping 
9.1 Confidentiality 
Information about study subjects will be kept confidential and managed according to the 
requirements of the Health Insurance Portability and Accountability Act of 1996 (HIPAA).  All 
subjects will sign an informed consent document and HIPAA authorization that names specific 
privacy and confidentiality rights.  Study data will be maintained per Federal and State data 
policies.  Subject data that is entered and stored in OnCore CRM database is coded with a study 
number generated by the OnCore system.  The ICTR data management team will work with 
investigators to collect only relevant, applicable study data, which will be entered in a secure web-
based system with restricted access. 
 
OnCore is an electronic data management system that will be used to capture, edit, manage, and 
export study data for analysis. 
 
The research team will manage study data through OnCore.  Access to the OnCore system is 
restricted to those that have been granted access by the software administrators. User access 
requires supervisor approval, completion of HIPAA and Human Subjects Protection training, and 
completion of role-based training in the OnCore system.  In addition, users’ access is limited to 
protocols for which they have some responsibility of protocol, subject, or data management. 
Within those protocols, the ability to view and modify data is restricted based on their role in the 
conduct of the research project (e.g. regulatory staff do not have the privilege to view subject 
identifiable information). 
 
In addition, the technical components of this software are managed by the UW-Madison’s 
Bioinformatics Computing Group (for server maintenance, software upgrades, etc.), and security 
and software support is provided by ICTR administrative staff.  
All communication between the clients and the OnCore application takes place via Hypertext 
Transfer Protocol over Secure Socket Layer or HTTPS. HTTPS provide the ability for normal web 
based communication over an encrypted Secure Socket Layer (SSL) connection. This ensures 
that data passing between the client and OnCore is protected from unauthorized attempts to 
access the data. 
 
The OnCore system is web-based software, with data stored on secure servers. Data exported 
from the OnCore System, is exported with indirect identifiers (i.e. with study ID number per 
subject) for statistical and data monitoring purposes in an MS Excel or SAS format. Upon 
exportation, the clinical research management system has no control over how it is manipulated 
or managed. 
9.2 
Source Documents 
All source data will be kept and merged and/or entered into the electronic clinical trial software 
(OnCore CRM).  Source data is all information, original records of clinical findings, observations, 
or other activities in a clinical trial necessary for the reconstruction and evaluation of the trial.  
Source data are contained in source documents.  Examples of these original documents, and 
data records include: hospital records, clinical and office charts, laboratory notes, memoranda, 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 27 
Version: 23, 7-25-17 
CONFIDENTIAL 
subjects’ diaries or evaluation checklists, pharmacy dispensing records, recorded data from 
automated instruments, copies or transcriptions certified after verification as being accurate and 
complete, microfiches, photographic negatives, microfilm or magnetic media, x-rays, subject files, 
and records kept at the pharmacy, at the laboratories, and at medico-technical departments 
involved in the clinical trial. 
9.3 
Case Report Forms 
The study case report form (CRF) is the primary data collection instrument for the study.  All data 
requested on the CRF must be recorded.  All missing data must be explained.  If a section of the 
CRF is left blank because the procedure was not done or the question was not asked, please 
document reason on the source document.  If the item is not applicable to the individual case, 
document “N/A”. 
 
9.4 
Data Collection at Sites   
Data will be collected at Marshfield Clinic, Gundersen Health System, and Aurora Bay Care at 
the screening visit (Visit 1) and subject monitoring visit (Visit 4).  In addition, 24-ambulatory blood 
pressure monitoring will be performed at each site (Visits 2 and 5).  Data from these visits 
including subject demographic information, vital signs, laboratory test values, and other subject 
information will be uploaded into OnCore CRM clinical trial software.  As a result all investigators 
will be able to access subject data from the screening visit and monitoring visit.  It is vital that all 
investigators can access data from the monitoring visit for safety reasons and monitoring of the 
study medications.  Data for the primary outcome variable will be uploaded into OnCore from Visit 
3 and Visit 6 and will be made available for the data and safety monitoring committee when they 
meet quarterly and for interim/final analyses after 33%, 66%, and 100% of subjects have been 
enrolled.   
 
9.5 Data analysis at Brigham and Women’s Hospital 
 
Dr. Scott Sands, at Brigham and Women’s Hospital will analyze some study data by isolating 
individual waveforms obtained from sleep studies.   Data will be transmitted to Dr. Sands by 
using a restricted Box folder within the password protected UW Box program.  The data that will 
be shared with Dr. Sands consists of individual waveforms from sleep studies.  The waveform 
files will be labeled with the subject’s unique study ID number and will not contain any other 
identifiers.  Coded sleep study files will be uploaded for Dr. Sands to access.  He will extract 
data from individual waveforms, and then summarize and score the data, and make the scored 
files available on Box.  Dr. Sands will not have access to the key that links the code to 
identifiable data.  Brigham and Women’s Hospital (Partners IRB) is deferring IRB oversight to 
the University of Wisconsin-Madison.  UW-Madison IRB will serve as the IRB of record for this 
study.   
 
 
 
 
9.6 Data sharing with 2016-0988 study (PI: Claudia Korcarz) 
 
Study data collected in this study will be shared with the 2016-0988 study (PI: Claudia Korcarz).  
The rationale for using this data in the 2016-0988 study is that this data contains information 
about PAP compliance, risk factors, demographics, etc.  IRB approval has already been 
obtained for the use of this data in the 2016-0988 protocol.   


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 28 
Version: 23, 7-25-17 
CONFIDENTIAL 
 
10 Ethical Considerations 
This study is to be conducted according to US and international standards of Good Clinical 
Practice (FDA Title 21 part 312 and International Conference on Harmonization guidelines), 
applicable government regulations and Institutional research policies and procedures. 
 
This protocol and any amendments will be submitted to a properly constituted independent 
Institutional Review Board (IRB), in agreement with local legal prescriptions, for formal approval 
of the study conduct.  The decision of the IRB concerning the conduct of the study will be made 
in writing to the investigator and a copy of this decision will be provided to the sponsor before 
commencement of this study.   
 
All subjects for this study will be provided a consent form describing this study and providing 
sufficient information for subjects to make an informed decision about their participation in this 
study.  This consent form will be submitted with the protocol for review and approval by the IRB 
for the study.  The formal consent of a subject, using the IRB-approved consent form, must be 
obtained before that subject undergoes any study procedure.  The consent form must be signed 
by the subject and the investigator-designated research professional obtaining the consent.  
11 Study Finances 
11.1 Funding Source 
The trial has been funded by the NIH – National Heart Lung and Blood Institute  
11.2 Subject Stipends or Payments 
Subjects will be reimbursed $50 for completing the screening visit (Visit 1), $25 for subjects 
undergoing re-screening procedures, if necessary (Visit 1A), and $50 for the 2-week monitoring 
visit (Visit 4).  Subjects who choose to wear the Holter/Heart rate variability equipment will also 
be paid $25 for each night the equipment is worn at home. Subjects will receive up to $400 for 
each study visit they complete in Madison (2 night visit) (Visits 3 and 6).  This amount may be 
prorated if the subject is only able to complete certain portions of the procedures. The prorated 
amounts are $100 for completion of the Physiologic Studies for day 1 of Visit 3 and 6, $100 for 
the completion of the sleep studies for night 1 of Visit 3 and Visit 6, $100 for the completion of the 
Vascular Studies in Dr. Stein’s Lab for Visit 3 and Visit 6, and $100 for the completion of the 
Control of Breathing Studies on night 2 for Visit 3 and Visit 6. If the subject completes all 
procedures, they will receive $400 for each Visit 3 and Visit 6. In addition, subjects will receive 
$100 for completing the study and the post-study follow-up questions with study staff (Visit 7).  
Subjects would receive $1075 if they complete the entire study, including re-screening Visit 1A 
and the optional Holter procedures. 
 
If subjects do not return the loaned CPAP machine or CPAP compliance card at Visit 6, they will 
not receive the payment for Visit 6 or the 30-day follow-up phone call until they return the loaned 
CPAP machine or compliance card. 
 
Subjects who travel more than 40 miles one way to Madison for Visits 3 and 6 will be reimbursed 
for their travel.  Subjects will be reimbursed $150 for traveling to visit 3 and visit 6 ($150 for each 
visit, $300 total).  In addition, hotel accommodations at the Best Western Inn Towner (two blocks 
from UW Hospital) are available for a spouse/significant other and or family of subjects enrolled 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 29 
Version: 23, 7-25-17 
CONFIDENTIAL 
at Aurora Bay Care, Gundersen Health System, and Marshfield Clinic for the 2- night study visit 
in Madison.  This is provided at no cost to the subject. 
 
12 Publication Plan 
Study data and results will be published and shared as mandated by NIH and Federal data sharing 
regulations. 
 
13 References 
 
1. Das AM and Khayat R. Hypertension in obstructive sleep apnea: risk and therapy. 
Expert Rev Cardiovasc Ther 2009;7:619-626. 
2. Narkiewicz K, van de Borne PJ, Pesek CA, Dyken ME, Montano N and Somers VK. 
Selective potentiation of peripheral chemoreflex sensitivity in obstructive sleep apnea. 
Circulation 99: 1183-1189, 1999. 
3. Imadojemu VA, Mawji Z, Kunselman A, Gray KS, Hogeman CS and Leuenberger UA. 
Sympathetic chemoreflex responses in obstructive sleep apnea and effects of 
continuous positive airway pressure therapy. Chest 2007;131:1406-1413. 
4. Marcus N, Li Y, Bird C, Olson E, Smith S, Sorenson K, Schultz H and Morgan B. Chronic 
intermittent hypoxia alters chemoreflex control of lumbar sympathetic nerve activity and 
carotid body protein expression. FASEB J 2009;23: 1008.1. 
5. Peng YJ, Overholt JL, Kline D, Kumar GK and Prabhakar NR. Induction of sensory long-
term facilitation in the carotid body by intermittent hypoxia: implications for recurrent 
apneas. Proc Natl Acad Sci U S A 2003;100:10073-10078. 
6. Li Y-L, Gao L, Zucker IH, Schultz HD.  NADPH oxidase-derived superoxide anion 
mediates angiotensin II-enhanced carotid body chemoreceptor sensitivity in heart failure 
rabbits.  Cardiovascular Res 2007;75:546-554. 
7. Warner G, Skatrud JB and Dempsey JA. Effect of hypoxia-induced periodic breathing on 
upper airway obstruction during sleep. J Appl Physiol 62: 2201-2211, 1987. 
8. Wellman A, Jordan AS, Malhotra A, Fogel RB, Katz ES, Schory K, Edwards JK and 
White DP. Ventilatory control and airway anatomy in obstructive sleep apnea. Am J 
Respir Crit Care Med 170: 1225-1232, 2004. 
9. Kelley EE, Hock T, Khoo NK, Richardson GR, Johnson KK, Powell PC, Giles GI, 
Agarwal A, Lancaster JR Jr, Tarpey MM.  Moderate hypoxia induces xanthine 
oxidoreductase activity in arterial endothelial cells.  Free Rad Biol Med 2006;40:952-959. 
10. McCord JM.  Oxygen-derived free radicals in post-ischaemic tissue injury.  N Engl J Med 
1985;312:159-163. 
11. Landmesser U, Spiekermann S, Preuss C, Sorrentino S, Fischer D, Manes C, Mueller M 
and Drexler H. Angiotensin II induces endothelial xanthine oxidase activation: role for 
endothelial dysfunction in patients with coronary disease. Arterioscler Thromb Vasc Biol 
2007;27:943-948. 
12. Laursen JB, Rajagopalan S, Galis Z, Tarpey M, Freeman BA and Harrison DG. Role of 
superoxide in angiotensin II-induced but not catecholamine-induced hypertension. 
Circulation 1997;95:588-593. 
13. Moller DS, Lind P, Strunge B and Pedersen EB. Abnormal vasoactive hormones and 24-
hour blood pressure in obstructive sleep apnea. Am J Hypertens 2003;16:274-280. 
14. Hassoun PM, Feng-Sheng Y, Cote CG et al.  Upregulation of xanthine oxidase by 
lipopolysaccharide, interleukin-1 and hypoxia.  Am J Respir Crit Care Med.  158:299-
305. 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 30 
Version: 23, 7-25-17 
CONFIDENTIAL 
15. Kayyali US, Donaldson C, Huang H et al.  Phosphorylation of xanthine 
dehydrogenase/oxidase in hypoxia.  J Biol Chem.  2001; 276:14359-14365. 
16. El Solh AA, Saliba R, Bosinski T et al. Allopurinol improves endothelial function in sleep 
apnoea: a randomised controlled study. Eur Respir J. 2006;27:997-1002. 
17. Engleman HM, Wild MR.  Improving CPAP use by patients with the sleep 
apnea/hypopnea syndrome.  Sleep Med Rev.  2003;7:81-99. 
18. Barone-Kribbs N, Pack A, Kline L  et al.  Effects of one night without CPAP treatment on 
sleep and sleepiness in patients with obstructive sleep apnea.  Am Rev Respir Dis.  
1993;47:1162-1168. 
19. Weaver T, Kribbs N, Pack A, et al.  Night-to-night variability in CPAP use over the first 
three months of treatment.  Sleep.  1997;20:278-283. 
20. Fitzpatrick MF, Alloway CE, Wakeford TM, MacLean AW, Munt PW, Day AG. Can 
patients with obstructive sleep apnea titrate their own continuous positive airway 
pressure? Am J Respir Crit Care Med 2003;167:716-722. 
21. Grote L, Hedner J, Grunstein R, Kraiczi H.  Therapy with CPAP: incomplete elimination 
of sleep related breathing disorder.  Eur Respir J. 2000;16:921-927. 
22. National Commission on Sleep Disorders Research.  Wake up America: a national sleep 
alert.  Washington, DC: Government Printing Office; 1993. 
23. Strohl KP, Redline S.  Recognition of obstructive sleep apnea.  Am J Respir Crit Care 
Med.  1996;154:279-289. 
24. Levinson PD, McGarvey ST, Carlisle CC et al.  Adiposity and cardiovascular risk factors 
in men with obstructive sleep apnea.  Chest.  1993;103:1336-1342. 
25. Phillips B, Cook Y, Schmitt F et al.  Sleep apnea:  prevalence of risk factors in the 
general population.  South Med J. 1989;82:1090-1092. 
26. Peppard PE, Young, T, Palta, M et al.  Prospective study of the association between 
sleep-disordered breathing and hypertension.  N Engl J Med.  2000; 342:1378-1384.  
27. Dyken ME, Somers VK, Yamada T et al.  Investigating the relationship between stroke 
and obstructive sleep apnea.  Stroke.  1996; 27:401-407. 
28. Partinen M. Guilleminault C. Daytime sleepiness and vascular morbidity at seven-year 
follow-up in obstructive sleep apnea patients.  Chest. 1990; 97:27-32. 
29. He J, Kryger MH, Zorick FJ et al.  Mortality and apnea index in obstructive sleep apnea: 
experience in 385 male patients.  Chest.  1988; 94:373-376. 
30. Marin JM, Carrizo SJ, Vicente E et al. Long-term cardiovascular outcomes in men with 
obstructive sleep apnoea-hypopnoea with or without treatment with continuous positive 
airway pressure: an observational study. Lancet. 2005;365:1046-1053. 
31. Narkiewicz K, Kato M, Phillips BG et al. Nocturnal continuous positive airway pressure 
decreases daytime sympathetic traffic in obstructive sleep apnea. Circulation. 
1999;100:2332-2335. 
32. Ip MS, Tse HF, Lam B, Tsang KW, Lam WK.  Endothelial function in obstructive sleep 
apnea and response to treatment.  Am J Respir Crit Care Med. 2004;169:348-353.  
33. Becker HF, Jerrentrup A, Ploch T, Grote L, Penzel T, Sullivan CE, Peter JH.  Effect of 
nasal continuous positive airway pressure treatment on blood pressure in patients with 
obstructive sleep apnea.  Circulation. 2003;107:68-73. 
34. Fletcher EC, Lesske J, Behm R, Miller CC 3rd, Stauss H, Unger T.  Carotid 
chemoreceptors, systemic blood pressure, and chronic episodic hypoxia mimicking 
sleep apnea.  J Appl Physiol. 1992;72:1978-1784. 
35. Fletcher EC, Lesske J, Culman J, Miller CC and Unger T. Sympathetic denervation 
blocks blood pressure elevation in episodic hypoxia. Hypertension 1992;20:612-619. 
36. Fletcher EC, Lesske J, Qian W, Miller CC, III and Unger T. Repetitive, episodic hypoxia 
causes diurnal elevation of blood pressure in rats. Hypertension 1992;19:555-561.   


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 31 
Version: 23, 7-25-17 
CONFIDENTIAL 
37. Carlson JT, Hedner J, Elam M et al. Augmented resting sympathetic activity in awake 
patients with obstructive sleep apnea. Chest. 1993;103:1763-1768. 
38. Dimsdale JE, Coy T, Ziegler MG, Ancoli-Israel S, Clausen J. The effect of sleep apnea 
on plasma and urinary catecholamines. Sleep. 1995;18:377–381. 
39. Xie A, Skatrud JB, Puleo DS, Morgan BJ.  Exposure to hypoxia produces long-lasting 
sympathetic activation in humans.  J Appl Physiol.  2001;91:1555-1562. 
40. Peng YJ, Overholt JL, Kline D et al. Induction of sensory long-term facilitation in the 
carotid body by intermittent hypoxia: implications for recurrent apneas. Proc Natl Acad 
Sci U S A. 2003;100:10073-10078. 
41. Greenberg HE, Sica A, Batson D et al. Chronic intermittent hypoxia increases 
sympathetic responsiveness to hypoxia and hypercapnia. J Appl Physiol. 1999;86:298-
305. 
42. Sica AL, Greenberg HE, Ruggiero DA et al. Chronic-intermittent hypoxia: a model of 
sympathetic activation in the rat. Respir Physiol. 2000;121:173-184. 
43. Prabhakar NR, Peng YJ, Jacono FJ et al. Cardiovascular alterations by chronic 
intermittent hypoxia: importance of carotid body chemoreflexes. Clin Exp Pharmacol 
Physiol. 2005;32:447-449. 
44. Lam SY, Fung ML, Leung PS. Regulation of the angiotensin-converting enzyme activity 
by a time-course hypoxia in the carotid body. J Appl Physiol. 2004;96:809-813. 
45. Allen AM.  Angiotensin AT1 receptor-mediated excitation of rat carotid body 
chemoreceptor afferent activity.  J Physiol. 1998;510:773-781. 
46. Leung PS, Lam SY, Fung ML.  Chronic hypoxia upregulates the expression and function 
of AT(1) receptor in rat carotid body.  J Endocrinol 2000;167:517-524. 
47. Li Y-L, Gao L, Zucker IH, Schultz HD.  NADPH oxidase-derived superoxide anion 
mediates angiotensin II-enhanced carotid body chemoreceptor sensitivity in heart failure 
rabbits.  Cardiovascular Res. 2007;75:546-554. 
48. Hassoun PM, Feng-Sheng Y, Cote CG et al.  Upregulation of xanthine oxidase by 
lipopolysaccharide, interleukin-1 and hypoxia.  Am J Respir Crit Care Med. 
1998;158:299-305. 
49. Kayyali US, Donaldson C, Huang H et al.  Phosphorylation of xanthine 
dehydrogenase/oxidase in hypoxia.  J Biol Chem.  2001; 276:14359-14365. 
50. Valdes V, Mosqueira M, Rey S, Del Rio R, Iturriaga R.  Inhibitory effects of NO on 
carotid body:  contribution of neural and endothelial nitric oxide synthase isoforms.  Am J 
Physiol.  2003;284:L57-L68. 
51. Prabhakar NR.  Neurotransmitters in the carotid body.  Adv Exp Med Biol.  1994;360:57-
69. 
52. Weiss JW, Liu MD, Huang J. Sleep Apnoea & Hypertension: Physiological basis for a 
causal relationship of obstructive sleep apnoea to hypertension. Exp Physiol. 
2007;92:21-26. 
53. El Solh AA, Saliba R, Bosinski T et al. Allopurinol improves endothelial function in sleep 
apnoea: a randomised controlled study. Eur Respir J. 2006;27:997-1002. 
54. Ip MS, Lam B, Chan LY, Zheng L, Tsang KW, Fung PC, Lam WK.  Circulating nitric 
oxide is suppressed in obstructive sleep apnea and is reversed by nasal continuous 
positive airway pressure.  Am J Respir Crit Care Med. 2000 Dec;162(6):2166-71.  
55. Kato M, Roberts-Thomson P, Phillips BG et al. Impairment of endothelium-dependent 
vasodilation of resistance vessels in patients with obstructive sleep apnea. Circulation. 
2000;102:2607-2610. 
56. Kraiczi H, Caidahl K, Samuelsson A et al. Impairment of vascular endothelial function 
and left ventricular filling : association with the severity of apnea-induced hypoxemia 
during sleep. Chest. 2001;119:1085-1091. 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 32 
Version: 23, 7-25-17 
CONFIDENTIAL 
57. Nieto FJ, Herrington DM, Redline S et al. Sleep apnea and markers of vascular 
endothelial function in a large community sample of older adults. Am J Respir Crit Care 
Med. 2004;169:354-360. 
58. Phillips SA, Olson EB, Morgan BJ et al. Chronic intermittent hypoxia impairs 
endothelium-dependent dilation in rat cerebral and skeletal muscle resistance arteries. 
Am J Physiol Heart Circ Physiol. 2004;286:H388-H393. 
59. Milstein S and Katusic Z.  Oxidation of tetrahydrobiopterin by peroxynitrite: implications 
for vascular endothelial function.  Biochem Biophys Res Comm.  1999;263:681-684. 
60. Schulz R, Mahmoudi S, Hattar K et al. Enhanced release of superoxide from 
polymorphonuclear neutrophils in obstructive sleep apnea. Impact of continuous positive 
airway pressure therapy. Am J Respir Crit Care Med. 2000;162:566-570. 
61. Barcelo A, Miralles C, Barbe F et al. Abnormal lipid peroxidation in patients with sleep 
apnoea. Eur Respir J. 2000;16:644-647. 
62. McNally JS, Davis ME, Giddens DP, et al.  Role of xanthine oxidoreductase and 
NAD(P)H oxidase in endothelial superoxide production in response to oscillatory shear 
stress.  Am J Physiol. 2003;285:H2290-H2297. 
63. Minoguchi K, Yokoe T, Tazaki T, Minoguchi H, Tanaka A, Oda N, Okada S, Ohta S, 
Naito H and Adachi M. Increased carotid intima-media thickness and serum 
inflammatory markers in obstructive sleep apnea. Am J Respir Crit Care Med 
2005;172:625-630. 
64. Tanriverdi H, Evrengul H, Kara CO, Kuru O, Tanriverdi S, Ozkurt S, Kaftan A and Kilic 
M. Aortic stiffness, flow-mediated dilatation and carotid intima-media thickness in 
obstructive sleep apnea: non-invasive indicators of atherosclerosis. Respiration 
2006;73:741-750. 
65. Baguet JP, Hammer L, Levy P, Pierre H, Launois S, Mallion JM and Pepin JL. The 
severity of oxygen desaturation is predictive of carotid wall thickening and plaque 
occurrence. Chest 2005;128:3407-3412. 
66. Drager LF, Bortolotto LA, Figueiredo AC, Silva BC, Krieger EM and Lorenzi-Filho G. 
Obstructive sleep apnea, hypertension, and their interaction on arterial stiffness and 
heart remodeling. Chest 2007;131:1379-1386. 
67. Kohler M, Craig S, Nicoll D, Leeson P, Davies RJ and Stradling JR. Endothelial function 
and arterial stiffness in minimally symptomatic obstructive sleep apnea. Am J Respir Crit 
Care Med 2008;178:984-988. 
68. Phillips C, Hedner J, Berend N and Grunstein R. Diurnal and obstructive sleep apnea 
influences on arterial stiffness and central blood pressure in men. Sleep 2005;28:604-
609. 
69. Tsioufis C, Thomopoulos K, Dimitriadis K, Amfilochiou A, Tousoulis D, Alchanatis M, 
Stefanadis C and Kallikazaros I. The incremental effect of obstructive sleep apnoea 
syndrome on arterial stiffness in newly diagnosed essential hypertensive subjects. J 
Hypertens 2007;25:141-146. 
70. McEniery CM, Qasem A, Schmitt M, Avolio AP, Cockcroft JR and Wilkinson IB. 
Endothelin-1 regulates arterial pulse wave velocity in vivo. J Am Coll Cardiol 
2003;42:1975-1981. 
71. Wilkinson IB, Qasem A, McEniery CM, Webb DJ, Avolio AP and Cockcroft JR. Nitric 
oxide regulates local arterial distensibility in vivo. Circulation 2002;105:213-217. 
72. Phillips BG, Narkiewicz K, Pesek CA, Haynes WG, Dyken ME and Somers VK. Effects 
of obstructive sleep apnea on endothelin-1 and blood pressure. J Hypertens 1999;17:61-
66. 
73. Phillips SA, Olson EB, Lombard JH and Morgan BJ. Chronic intermittent hypoxia alters 
NE reactivity and mechanics of skeletal muscle resistance arteries. J Appl Physiol 
2006;100:1117-1123. 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 33 
Version: 23, 7-25-17 
CONFIDENTIAL 
74. Carmeliet P and Jain RK. Angiogenesis in cancer and other diseases. Nature 
2000;407:249-257. 
75. Duyndam MC, Hulscher TM, Fontijn D, Pinedo HM and Boven E. Induction of vascular 
endothelial growth factor expression and hypoxia-inducible factor 1alpha protein by the 
oxidative stressor arsenite. J Biol Chem 2001;276:48066-48076. 
76. Brown NJ. Aldosterone and vascular inflammation. Hypertension 2008;51:161-167. 
77. Takahashi S, Nakamura Y, Nishijima T, Sakurai S and Inoue H. Essential roles of 
angiotensin II in vascular endothelial growth factor expression in sleep apnea syndrome. 
Respir Med 2005;99:1125-1131. 
78. Bleeke T, Zhang H, Madamanchi N, Patterson C and Faber JE. Catecholamine-induced 
vascular wall growth is dependent on generation of reactive oxygen species. Circ Res 
2004;94:37-45. 
79. Gerasimovskaya EV, Ahmad S, White CW, Jones PL, Carpenter TC and Stenmark KR. 
Extracellular ATP is an autocrine/paracrine regulator of hypoxia-induced adventitial 
fibroblast growth. Signaling through extracellular signal-regulated kinase-1/2 and the 
Egr-1 transcription factor. J Biol Chem 2002;277:44638-44650. 
80. Stenmark KR, Gerasimovskaya E, Nemenoff RA and Das M. Hypoxic activation of 
adventitial fibroblasts: role in vascular remodeling. Chest 2002;122:326S-334S. 
81. Tulis DA and Prewitt RL. Medial and endothelial platelet-derived growth factor A chain 
expression is regulated by in vivo exposure to elevated flow. J Vasc Res 1998;35:413-
420. 
82. Wilson E, Mai Q, Sudhir K, Weiss RH and Ives HE. Mechanical strain induces growth of 
vascular smooth muscle cells via autocrine action of PDGF. J Cell Biol 1993;123:741-
747. 
83. Hudgel DW, Gordon EA, Thanakitcharu S and Bruce EN. Instability of ventilatory control 
in patients with obstructive sleep apnea. Am J Respir Crit Care Med 1998;158:1142-
1149. 
84. Onal E, Burrows DL, Hart RH and Lopata M. Induction of periodic breathing during sleep 
causes upper airway obstruction in humans. J Appl Physiol 1986;61:1438-1443. 
85. Younes M, Ostrowski M, Atkar R, Laprairie J, Siemens A and Hanly P. Mechanisms of 
breathing instability in patients with obstructive sleep apnea. J Appl Physiol 
2007;103:1929-1941. 
86. Prabhakar NR, Peng YJ, Jacono FJ, Kumar GK and Dick TE. Cardiovascular alterations 
by chronic intermittent hypoxia: importance of carotid body chemoreflexes. Clin Exp 
Pharmacol Physiol 2005;32:447-449. 
87. Reichmuth KJ, Dopp JM, Barczi SR, Skatrud JB, Wojdyla P, Hayes JD and Morgan BJ. 
Impaired Vascular Regulation in Patients with Obstructive Sleep Apnea: Effects of CPAP 
Treatment. Am J Respir Crit Care Med 2009;180:1143-1150. 
88. Brian JE, Jr. Carbon dioxide and the cerebral circulation. Anesthesiology 1998;88:1365-
1386. 
89. Xie A, Skatrud JB, Morgan B, Chenuel B, Khayat R, Reichmuth K, Lin J and Dempsey 
JA. Influence of cerebrovascular function on the hypercapnic ventilatory response in 
healthy humans. J Physiol 2006;577:319-329. 
90. Xie A, Skatrud JB, Barczi SR, Reichmuth K, Morgan BJ, Mont S and Dempsey JA. 
Influence of cerebral blood flow on breathing stability. J Appl Physiol 2009;106:850-856. 
91. Foster GE, Hanly PJ, Ostrowski M and Poulin MJ. Ventilatory and cerebrovascular 
responses to hypercapnia in patients with obstructive sleep apnoea: Effect of CPAP 
therapy. Respir Physiol Neurobiol 165: 73-81, 2009. 
92. Marcus N, Li Y, Bird C, Olson E, Smith S, Sorenson K, Schultz H and Morgan B. Chronic 
intermittent hypoxia alters chemoreflex control of lumbar sympathetic nerve activity and 
carotid body protein expression. FASEB J 2009;23:1008.1. 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 34 
Version: 23, 7-25-17 
CONFIDENTIAL 
93. Spicuzza L, Bernardi L, Balsamo R, Ciancio N, Polosa R and Di Maria G. Effect of 
treatment with nasal continuous positive airway pressure on ventilatory response to 
hypoxia and hypercapnia in patients with sleep apnea syndrome. Chest 2006;130:774-
779. 
94. Wang D, Grunstein RR and Teichtahl H. Association between ventilatory response to 
hypercapnia and obstructive sleep apnea-hypopnea index in asymptomatic subjects. 
Sleep Breath 2007;11:103-108. 
95. Vallbo AB, Hagbarth KE, Torebjork HE & Wallin BG. Somatosensory, proprioceptive, 
and sympathetic activity in human peripheral nerves. Physiol Rev 1979;59:919-957. 
96. Dopp JM, Wiegert NA, Moran JJ, Muller D, Weber SM, Hayney MS.  Humoral immune 
responses to influenza vaccinationin patients with obstructive sleep apnea.  
Pharmacotherapy 2007;27:1483-1489. 
97. Hoffman RP, Sinkey CA, Dopp JM, Phillips BG.  Systemic and local adrenergic 
regulation of muscle glucose utilization during hypoglycemia in healthy subjects.  
Diabetes  2002; 51:734-42. 
98. Agapitov A, Correia ML, Sinkey CA, Dopp JM, Haynes WG.  Impaired skeletal muscle 
and skin microcirculatory function in human obesity.  J Hypertens 2002;20:1401-1405. 
99. Hoffman RP, Sinkey CA, Dopp JM, Phillips BG. Lack of effect of alpha- and beta-
adrenergic inhibition on forearm glucose uptake despite differences in forearm blood 
flow in healthy humans.  Metabolism 2002; 51:1506-1513. 
100. 
Ainslie PN, Ashmead JC, Ide K, Morgan BJ, Poulin MJ.  Differential responses to 
CO2 and sympathetic stimulation in the cerebral and femoral circulations in humans.  J 
Physiol. 2005;566:613-24. 
101. 
Khayat RN, Przybylowski T, Meyer KC, Skatrud JB, Morgan BJ.  Role of sensory 
input from the lungs in control of muscle sympathetic nerve activity during and after 
apnea in humans.  J Appl Physiol. 2004;97:635-40. 
102. 
Halliwill JR, Morgan BJ, Charkoudian N.  Peripheral chemoreflex and baroreflex 
interactions in cardiovascular regulation in humans.  J Physiol. 2003;552:295-302.  
103. 
Przybyłowski T, Bangash MF, Reichmuth K, Morgan BJ, Skatrud JB, Dempsey 
JA.  Mechanisms of the cerebrovascular response to apnoea in humans.  J Physiol. 
2003;548:323-32. 
104. 
Derchak PA, Sheel AW, Morgan BJ, Dempsey JA.  Effects of expiratory muscle 
work on muscle sympathetic nerve activity.  J Appl Physiol. 2002;92:1539-1552. 
105. 
Katragadda S, Xie A, Puleo D, Skatrud JB, Morgan BJ.  Neural mechanism of the 
pressor response to obstructive and nonobstructive apnea.  J Appl Physiol. 
1997;83:2048-2054. 
106. 
Morgan BJ, Denahan T, Ebert TJ.  Neurocirculatory consequences of negative 
intrathoracic pressure vs. asphyxia during voluntary apnea.  J Appl Physiol. 
1993;74:2969-2975. 
107. 
Morgan BJ, Crabtree DC, Palta M, Skatrud JB.  Combined hypoxia and 
hypercapnia evokes long-lasting sympathetic activation in humans.  J Appl Physiol. 
1995;79:205-213. 
108. 
St Croix CM, Morgan BJ, Wetter TJ, Dempsey JA.  Fatiguing inspiratory muscle 
work causes reflex sympathetic activation in humans.  J Physiol. 2000;529:493-504. 
109. 
Aaslid R, Markwalder T, Nornes H.  Noninvasive transcranial Doppler ultrasound 
recording of flow velocity in basal cerebral arteries.  J Neurosurg. 1982;57:769-774. 
110. 
Otis SM, Ringelstein EB.  The transcranial Doppler examination: principles and 
applications of transcranial Doppler sonography.  In: Tegeler CH, Babikian VL, Gomez 
CR, Neurosonology, Mosby, St. Louis, MO, 1996, pp. 113-128. 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 35 
Version: 23, 7-25-17 
CONFIDENTIAL 
111. 
Giller CA, Bowman G, Dyer H, Mootz L, Krippner W.  Cerebral arterial diameters 
during changes in blood pressure and carbon dioxide during craniotomy.  Neurosurgery. 
1993;32:737-742. 
112. 
Serrador JM, Picot PA, Rutt BK, Shoemaker JK, Bondar RL.  MRI measures of 
middle cerebral artery diameter in conscious humans during simulated orthostasis.  
Stroke. 2000;31:1672-1678. 
113. 
Bishop CCR, Powell S, Rutt D, Browse NL.  Transcranial Doppler measurement 
of middle cerebral artery blood flow velocity:  a validation study.  Stroke. 1986;17:913-
915. 
114. 
Kirkham FJ, Padayachee TS, Parsons S, Seargeant LS, House FR, Gosling RG.  
Transcranial measurement of blood velocities in basal cerebral arteries using pulsed 
Doppler ultrasound: velocity as an index of flow.  Ultrasound Med  Biol. 1986;12:15-21. 
115. 
Pyke KE, Hartnett JA and Tschakovsky ME. Are the dynamic response 
characteristics of brachial artery flow-mediated dilation sensitive to the magnitude of 
increase in shear stimulus? J Appl Physiol 2008;105:282-292. 
116. 
Aeschlimann SE, Mitchell CK and Korcarz CE. Ultrasound brachial artery 
reactivity testing: technical considerations. J Am Soc Echocardiogr 2004;17:697-699. 
117. 
Laurent S, Cockcroft J, Van BL, Boutouyrie P, Giannattasio C, Hayoz D, Pannier 
B, Vlachopoulos C, Wilkinson I and Struijker-Boudier H. Expert consensus document on 
arterial stiffness: methodological issues and clinical applications. Eur Heart J 
2006;27:2588-2605. 
118. 
Karamanoglu M, O'Rourke MF, Avolio AP and Kelly RP. An analysis of the 
relationship between central aortic and peripheral upper limb pressure waves in man. 
Eur Heart J 1993;14:160-167. 
119. 
Sweitzer NK, Shenoy M, Stein JH, Keles S, Palta M, LeCaire T and Mitchell GF. 
Increases in central aortic impedance precede alterations in arterial stiffness measures 
in type 1 diabetes. Diabetes Care 2007;30:2886-2891. 
120. 
Rechtschaffen A and Kales A. A manual of standardized terminology, 
techniques, and scoring system for sleep stages of human subjects. Washington, DC: 
US Government Printing Office, 1968. 
121. 
Young T, Finn L, Peppard PE, Szklo-Coxe M, Austin D, Nieto FJ, Stubbs R and 
Hla KM. Sleep disordered breathing and mortality: eighteen-year follow-up of the 
Wisconsin sleep cohort. Sleep 2008;31:1071-1078. 
122. 
Young T, Palta M, Dempsey J, Skatrud J, Weber S and Badr S. The occurrence 
of sleep-disordered breathing among middle-aged adults. N Engl J Med 1993;328:1230-
1235. 
123. 
Gold AR, Marcus CL, Dipalo F and Gold MS. Upper airway collapsibility during 
sleep in upper airway resistance syndrome. Chest 2002;121:1531-1540. 
124. 
Kirkness JP, Schwartz AR, Schneider H, Punjabi NM, Maly JJ, Laffan AM, 
McGinley BM, Magnuson T, Schweitzer M, Smith PL and Patil SP. Contribution of male 
sex, age, and obesity to mechanical instability of the upper airway during sleep. J Appl 
Physiol 2008;104:1618-1624. 
125. 
Patil SP, Punjabi NM, Schneider H, O'Donnell CP, Smith PL and Schwartz AR. A 
simplified method for measuring critical pressures during sleep in the clinical setting. Am 
J Respir Crit Care Med 2004;170:86-93. 
126. 
Xie A, Skatrud JB and Dempsey JA. Effect of hypoxia on the hypopnoeic and 
apnoeic threshold for CO(2) in sleeping humans. J Physiol 2001;535:269-278. 
127. 
Nakayama H, Smith CA, Rodman JR, Skatrud JB and Dempsey JA. Carotid body 
denervation eliminates apnea in response to transient hypocapnia. J Appl Physiol 
2003;94:155-164. 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 36 
Version: 23, 7-25-17 
CONFIDENTIAL 
128. 
Wilson CR, Satoh M, Skatrud JB and Dempsey JA. Non-chemical inhibition of 
respiratory motor output during mechanical ventilation in sleeping humans. J Physiol 
1999;518 ( Pt 2):605-618. 
129. 
Johns MW. Reliability and factor analysis of the Epworth Sleepiness Scale. Sleep 
1992;15:376-381. 
130. 
Pickering TG, Hall JE, Appel LJ, Falkner BE, Graves J, Hill MN, Jones DW, Kurtz 
T, Sheps SG and Roccella EJ. Recommendations for blood pressure measurement in 
humans and experimental animals: Part 1: blood pressure measurement in humans: a 
statement for professionals from the Subcommittee of Professional and Public Education 
of the American Heart Association Council on High Blood Pressure Research. 
Hypertension 2005;45:142-161. 
131. 
Hla KM, Young T, Finn L, Peppard PE, Szklo-Coxe M and Stubbs M. 
Longitudinal association of sleep-disordered breathing and nondipping of nocturnal 
blood pressure in the Wisconsin Sleep Cohort Study. Sleep 2008;31: 795-800. 
132. 
Hla KM, Young TB, Bidwell T, Palta M, Skatrud JB and Dempsey J. Sleep apnea 
and hypertension. A population-based study. Ann Intern Med 1994;120: 382-388. 
133. 
Stickland MK, Morgan BJ, Dempsey JA.  Carotid chemoreceptor modulation of 
sympathetic vasoconstrictor outflow during exercise in healthy humans. J Physiol. 
2008;586:1743-1754. 
134. 
Product Information: Alloprim™, allopurinol. Nabi, Boca Raton, FL, 2003. 
135. 
Murrell GAC, Rapeport WG. Clinical pharmacokinetics of allopurinol. Clin Pharm. 
1986;11:343–353. 
136. 
Day RO, Graham S, Wong H, et al. Pharmacodynamics of oxypurinol after 
administration of allopurinol to healthy subjects. Br J Clin Pharmacol. 1996;41:299–304. 
137. 
Day RO, Miners J, Birkett DJ, et al. Relationship between plasma oxipurinol 
concentrations and xanthine oxidase activity in volunteers dosed with allopurinol. Br J 
Clin Pharmacol. 1988;26:429–434. 
138. 
Breithaupt H, Tittel M. Kinetics of allopurinol after single intravenous and oral 
doses. Eur J Clin Pharmacol. 1982;22:77–84. 
139. 
Sasaki M, Fujimura A, Harada K, et al: Clinical pharmacology of multiple-dose 
losartan, and angiotensin II receptor antagonist, in patients with essential hypertension. 
J Clin Pharmacol 1996;36:403-408. 
140. 
Munafo A, Christen Y, Nussberger J, Shum LY, Borland RB, Lee RJ, Waeber B, 
Biollaz J, Brunner HR.  Drug concentration response relationships in normal volunteers 
after oral administration of losartan, an angiotensin II receptor antagonist.  Clin 
Pharmacol Ther 1992;51:513-521. 
141. 
Ohtawa M, Takayama F, Saitoh K, et al: Pharmacokinetics and biochemical 
efficacy after single and multiple oral administration of losartan, an orally active 
nonpeptide angiotensin II receptor antagonist, in humans. Br J Clin Pharmacol  
1993;35:290-297. 
142. 
Ebert TJ, Morgan BJ, Barney JA, Denahan T, and Smith JJ. Effects of aging on 
baroreflex regulation of sympathetic activity in humans. Am J Physiol 1992;263: H798-
803. 
143. 
Jacobsen TN, Morgan BJ, Scherrer U, Vissing SF, Lange RA, Johnson N, Ring 
WS, Rahko PS, Hanson P, and Victor RG. Relative contributions of cardiopulmonary 
and sinoaortic baroreflexes in causing sympathetic activation in the human skeletal 
muscle circulation during orthostatic stress. Circ Res 1993;73:367-378. 
144. 
St Croix CM, Satoh M, Morgan BJ, Skatrud JB, and Dempsey JA. Role of 
respiratory motor output in within-breath modulation of muscle sympathetic nerve activity 
in humans. Circ Res 1999;85:457-469. 
 


Pharmacologic Interventions for Cardiovascular Disease in Obstructive Sleep Apnea 
 
 
Page 37 
Version: 23, 7-25-17 
CONFIDENTIAL 
14 Appendices 
• Appendix 1 - Descriptions of Study Procedures 
• Appendix 2 - Table of Study Procedures and Study Visits 
• Appendix 3 – Exclusionary Medication List