Cryospray Therapy for Benign Airway Stenosis: a Randomized Pilot Study
Phase: NA
Conditions: Pulmonary Disease
Sponsor: Vanderbilt University Medical Center

 
 
 
 
 
Cryospray Therapy Versus Standard of 
Care for Benign Airway Stenosis 
(CryoStasis) 
 
NCT04996173 
 
Protocol 
Version Date: 27May2025 

 
 
Cryospray Therapy for Benign Airway Stenosis: a pilot 
study 
(pilot-CRYOSTASIS) 
 
 
Protocol 
Principal investigators: 
Fabien Maldonado, MD 
Industry Sponsor: 
US Endoscopy/STERIS 
 
 
Protocol Version Date  
27MAY2025

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PROTOCOL SYNOPSIS 
 
 
Title 
Cryospray Therapy for Benign Airway Stenosis: a 
pilot study 
Short title 
(pilot-CRYOSTASIS) 
Study objective 
1. To assess the effectiveness of cryospray 
therapy used in addition to current 
standard of care endoscopic therapies in 
preventing short term recurrent airway 
stenosis. 
2.  
Study device 
truFreeze Spray Cryotherapy System (US 
Endoscopy) 
Design 
, Multicenter, investigator-initiated, industry-
funded, randomized 1:1, superiority study. The 
protocol allows for inclusion of an additional 
external site if recruitment is insufficient after 6 
months for study. 
Study Centers 
Multicenter at academic and community hospitals 
Number of subjects 
40 
Timeline 
12-month recruitment, 6 months follow up 
Inclusion criteria 
1. Referral to interventional pulmonology or 
otorhinolaryngology for endoscopic 
management of suspected benign tracheal 
stenosis. 
2. Significant tracheal stenosis defined by 
stenosis ≥ 50% of tracheal lumen assessed 
on chest CT or clinical symptomatology 
warranting evaluation. 
3. Able to provide informed consent. 
4. Age > 18 

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Exclusion criteria 
1. Inability to provide informed consent. 
2. Pregnancy 
3. Known or suspected malignant central 
airway stenosis 
4. Patient has already been enrolled in this 
study. 
5. Study subject has any disease or condition 
that interferes with safe completion of the 
study including: 
a. Hypoxemia with need for 
supplemental oxygen ≥ 2L/min 
by nasal canula 
b. Pneumothorax in the previous 12 
months 
c. Severe COPD (defined as a 
FEV1/FVC < 70% and FEV1 < 
30% predicted) and/or severe 
persistent asthma. 
d. Hemodynamic instability with 
systolic blood pressure <90 
mmHg or heart rate > 120 
beats/min, unless deemed to be 
stable with these values by the 
attending physicians. 
6. Prior complications with SCT (Spray 
cryotherapy) 
7. Contraindication to rigid bronchoscopy 
8. Significant tracheomalacia or alterations 
in cartilage integrity on CT (Computed 
Tomography) that would require stent 
placement or surgical referral. 
9. Greater then 1 BCAS intervention, 
excluding cricotracheal resection, within 
6 months before enrollment. 
Primary endpoint 
The degree of re-stenosis at 6 months, expressed 
as the percentage of airway lumen volume within 

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the stenotic segment, at 6 months compared to 
personal best patency volume on CT scan 
performed within two weeks of the study 
intervention. If patients require repeat intervention 
before six months based on clinical course, a CT 
chest before this re-intervention will be analyzed 
for the primary outcome. 
Secondary endpoint 
• 
Change in PEF (Secondary) 
• 
Change in QOL questionnaire 
(Secondary) 
• 
Incidence of complications between 
groups (Secondary) 
 
Statistical methodology 
We hypothesize the patency assessed at 6 months 
will be 90% of baseline in the intervention group 
and 70% in the control group, with SD 20% in the 
control group. A sample size of 16 subjects in 
control and 16 subjects in intervention group will 
have 80% power to detect an increase in stenotic 
volume of 20% from control arm with type I error 
of 0.05. Eight additional patients will be recruited 
to account for attrition and crossover. Study will 
be done using the intention-to-treat principle. 
Continuous and categorical variables will be 
analyzed using the appropriate methodology 
accounting for non-normal distributions. Patients 
will be blinded to intervention including those 
evaluating the volumetric CT’s and those 
performing statistical analysis. 

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Table of Contents 
1. Abbreviations ................................................................................................................................. 7 
2. General Study Information ......................................................................................................... 8 
2.1 Investigate team and research environment .............................................................................. 8 
3. Statement of compliance. .............................................................................................................. 8 
4. Purpose of the study ...................................................................................................................... 8 
4.1 Overview. ................................................................................................................................. 8 
4.2 Hypothesis ................................................................................................................................ 9 
4.3 Aims and Objectives................................................................................................................. 9 
5. Study Design. .............................................................................................................................. 10 
5.1 General Study Design ............................................................................................................. 10 
5.1.2 Intervention group ............................................................................................................... 10 
5.1.3 Control group ...................................................................................................................... 11 
5.1.4 Both groups ......................................................................................................................... 11 
5.1.5 Design Schematic. ............................................................................................................... 11 
6. Subject Information. .................................................................................................................. 12 
7. Outcomes .................................................................................................................................... 12 
8. Statistical Design ........................................................................................................................ 13 
8.1 Expected outcomes ................................................................................................................. 13 
8.2 Randomization ........................................................................................................................ 13 
8.3 Power Calculation and target accrual...................................................................................... 13 
8.4 Analytical Plan ....................................................................................................................... 14 
8.5 Measurements ......................................................................................................................... 14 
9. Outcome Measure Synopsis....................................................................................................... 15 
9.1 Quantitative imaging of stenotic volume. ............................................................................... 16 
10. Clinical COPD Questionnaire ................................................................................................... 17 
11. Safety. .......................................................................................................................................... 17 
12. Data Handling ............................................................................................................................ 18 
12.1 Data Collection ...................................................................................................................... 18 
12.2 Data and Safety Monitoring. .................................................................................................. 19 
12.3 Data Handling and Record Keeping. ..................................................................................... 19 

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13. Regulatory Considerations ....................................................................................................... 19 
13.1 Protocol Review and Amendments ...................................................................................... 20 
13.2 Informed Consent ................................................................................................................ 20 
13.3 Ethics and Good Clinical Practice. ...................................................................................... 20 
13.4 Confidentiality. ................................................................................................................... 20 
13.5 Study Termination. ............................................................................................................. 21 
14. Costs ............................................................................................................................................ 21 
15. Study Coordination .................................................................................................................... 21 
15.1 Trial Compliance ................................................................................................................ 21 
15.2 Changes to Protocol and Informed Consent Document ....................................................... 21 
15.3 Protocol Deviations ............................................................................................................. 22 
15.4 Monitoring and Quality Assurance. ..................................................................................... 22 
15.5 Data Verification. ................................................................................................................ 22 
15.6 Study Documentation .......................................................................................................... 22 
15.7 Closure of the Study. ........................................................................................................... 22 
15.8 Records Retention................................................................................................................. 23 
16. Publications................................................................................................................................. 23 
17. Citations ...................................................................................................................................... 23 
18. Consort Diagram ........................................................................................................................ 25 
19. Timeline ...................................................................................................................................... 26 
20. Study Assessment Table ............................................................................................................ 26 
21. COPD Clinical Questionnaire ................................................................................................... 27 

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NoAAC: North American airway collaborative 
VICTR: Vanderbilt Institute for Clinical and Translational Research 
Clinical and Translational Science Award (CTSA) 
REDCap: Research Electronic Data Capture 
FDA: Food and Drug Administration 
BCAS: Benign central airway stenosis 
COVID19: Cornavirus-19 
LN2: Liquid nitrogen 
SOC: Standard of care 
LMA: Laryngeal mask airway 
iSGS: Idiopathic subglottic stenosis 
QOL: Quality of life 
PEF: Peak expiratory flow 
FEV1: Forced expiratory volume in one second 
FVC: Forced vital capacity 
6MWD: 6-minute walk distance 
COPD: Chronic Obstructive Pulmonary Disease 
CCQ: COPD Clinical Questionnaire 
VUMC: Vanderbilt University Medical Center 
ITS: Idiopathic tracheal stenosis 
BMI: Body mass index 
ROI: Region of interest 
GSE: General safety events 
SAE: Serious adverse events 
eCRF: Electronic case report form 
IRB: Institutional review board 
HIPPAA: Health Information Portability and Accountability Act 
GCP: Good clinical practice 
1. Abbreviations 

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Principal Investigators: Fabien Maldonado, MD. 
Statistician: Heidi Chen, PhD 
Industry Sponsor: US Endoscopy/STERIS 
Title: Cryospray Therapy for Benign Airway Stenosis: a pilot study 
Protocol Version Date:  27, May, 2025 
2.1 Investigate team and research environment 
Vanderbilt has a complex airway center including interventional pulmonology, otorhinolaryngology, and 
thoracic surgery.. The Vanderbilt University Institute of Imaging Science is a trans-institutional initiative 
within Vanderbilt University serving physicians, scientists, students, and corporate affiliates. Vanderbilt 
Institute for Clinical and Translational Research (VICTR) VICTR is Vanderbilt’s virtual home for 
clinical and translational research. Supported by the Vanderbilt University Medical Center’s Office of 
Research and the NIH sponsored Clinical and Translational Science Award (CTSA), the mission of the 
institute is to transform the way ideas and research discoveries make their way from origin to patient 
care. REDCap (Research Electronic Data Capture) is a secure web application for building and managing 
online surveys and databases. While REDCap can be used to collect virtually any type of data (FISMA 
(Federal Information Security Management Act), and HIPAA- compliant environments), it is specifically 
geared to support online or offline data capture for research studies and operations. 
 
 
This human subject study will comply with all applicable federal, state, and local laws and regulations, 
including accepted standards of good clinical practice as adopted by current Food Drug Administration 
(FDA) regulations and statutes. Participating study sites shall only allow individuals who are 
appropriately trained and qualified to assist in the conduct of the study. 
 
 
4.1 : Overview 
Benign central airway stenoses (BCAS) are an important cause of pulmonary morbidity and mortality. 
2. General Study Information 
3. Statement of compliance 
4. Purpose of the study 

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1. Because of the Cornavirus-19 (COVID19) pandemic we anticipate a substantial increase in BCAS in 
the coming years primarily due to post intubation tracheal stenosis. There are two general options 
available for management of BCAS: surgical resection or endoscopic management. Surgery is the 
preferred definitive option; however, the first therapeutic attempt is typically endoscopic with surgery 
reserved for recurrence2. In addition, many patients are not candidates for surgical resection due to 
comorbid conditions, anatomical characteristic such as length or complexity of stenosis. Symptomatic 
stenosis frequently recurs resulting in numerous airway procedures3. For example, at our institution, we 
have been doing 3-4 balloon dilation procedures a week for BCAS. Procedures such as dilation, thermal 
ablative therapies, and stenting may result in additional airway injury due to disruption of extracellular 
matrix4. Spray cryotherapy (SCT) is a novel FDA cleared technique which allows for Liquid nitrogen 
(LN2) to be delivered in a metered fashion via a 7F catheter used and studied extensively in the 
esophagus5. The proposed advantage of SCT in the treatment of BCAS is the possibility to decellularize 
the tracheal mucosa, leaving the extracellular matrix intact, which may then serve as a scaffold for 
appropriate wound repair6, shifting the abnormal wound response after conventional endoscopic 
management to a more normal healing process7. Thus, SCT has the potential for long-lasting endoscopic 
management of BCAS and is used as part of routine clinical care by many physicians, but there is a 
relative paucity of peer-reviewed data, and no randomized controlled trial has been performed. 
Management decisions require adequate assessment of the position, extent, and severity of the obstructing 
or stenotic segment. This has historically been done with endoscopic measurements which are subjective, 
recurrent symptoms or abnormal pulmonary function tests8 which are universally a late finding in airway 
stenosis, typically occurring when the stenosis exceeds 70%9. 
4.2 : Hypothesis 
We hypothesize that the addition of SCT to standard endoscopic treatment of benign airway stenosis will 
result in decreased stenosis recurrence at 6 months as estimated by quantitative radiologic assessment of 
the stenotic volume. 
4.3 : Aims and Objectives 
1. To assess the effectiveness of cryospray therapy used in addition to current standard of care 
endoscopic therapies in preventing short term recurrent airway stenosis with a multicentric 
outcome evaluation  

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5.1 : General Study Design 
Subjects will be randomly allocated in a 1:1 fashion to intervention standard of care (SOC) + SCT and 
control (SOC) groups. Participants will be blinded to the use of SCT to prevent knowledge of their group 
assignment from biasing their subjective assessments. Adjudicators in the primary outcome 
measurements (quantitative CT) will be blinded to group assignment as well. Patient will be excluded 
who have received greater then 1 BCAS intervention within 6 months of enrollment. All operators will be 
experienced trūFreeze users with proper passive venting training as defined by trūFreeze protocol. All 
operators will have a minimum experience of 5 prior procedures. 
5.1.2 : Intervention group 
Patients with BCAS randomized to the intervention group will undergo conventional endoscopic 
treatment and SCT. The airway will be managed with a Dedo laryngoscope, rigid bronchoscope, or a 
laryngeal mask airway (LMA) per operator preference and lesion location. Airway pictures will be 
documented through our propriety bronchoscopy software. The truFreeze spray cryotherapy system will 
be used to apply liquid nitrogen spray through a flexible catheter passed though the working channel of a 
therapeutic bronchoscope or rigid laryngoscope. Typical dosimetry with passive venting without suction 
starts with up to 2 cycles with standard endoscopy, with one cycle after. Spray applications will be 
adjusted as needed based on the size and region being ablated for up to a 10 second spray time. Standard 
of care endoscopic management may include radial mucosal cuts using a thermal modality, and 
mechanical dilation. 
5.1.3 : Control group 
5.  Study Design 

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Patients randomized to the control group will undergo the same airway management and standard of care 
interventions without use of SCT before and after intervention. 
5.1.4 : Both groups 
Peak expiratory flow (PEF) as well as a quality of life (QOL) questionnaires will be obtained within 2 
weeks before and 2 weeks after the procedure. A repeat thin-cut chest CT or neck to quantitatively 
estimate of the patency volume will be obtained within 5 weeks of the first procedure to establish 
personal best patency volume (baseline patency volume, see below). A final CT scan will be done at 6 
months after the index study procedure to evaluate the degree of restenosis. A QOL questionnaire and 
check-in survey will be done at the at the 3-month. Patients will have a final QOL questionnaire at the 6- 
month mark. Patients will be doing continuous PEF measurements weekly throughout the study with a 
mobile application as mentioned below. 
Throughout the follow up period if a patient meets one or both of the following criteria: 1) drop in peak 
flow to 30% of previous value 2) drop in dyspnea score by 30% then the investigators will review if the 
patient needs an early CT and possibly repeat bronchoscopy. If patients undergo a repeat procedure before 
the 6-month CT they will be analyzed in an early reintervention subgroup. These patients will be 
including in the final analysis as per intention-to-treat and thereafter followed as per follow-up procedures 
outlined in the protocol. Patients in the SCT group may get repeat SCT as per discretion of the 
proceduralist. Patients in the SOC will not be able to get SCT and may only have repeat dilation with 
laser radial cuts. 
 
 
   
6.  Subject Information 

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Inclusion criteria: 
1. Referral to interventional pulmonology or ENT for endoscopic management of suspected benign 
tracheal stenosis. 
2. Significant tracheal stenosis defined by stenosis ≥ 50% of tracheal lumen assessed on chest CT or 
clinical symptomatology warranting evaluation. 
3. Able to provide informed consent 
4. Age > 18 
 
Exclusion criteria: 
1. Inability to provide informed consent. 
2. Pregnancy 
3. Known or suspected malignant central airway stenosis 
4. Patient has already been enrolled in this study. 
5. Study subject has any disease or condition that interferes with safe completion of the study 
including: 
e. Hypoxemia with need for supplemental oxygen ≥ 2L/min by nasal canula 
f. Recent pneumothorax in the previous 12 months 
g. Severe COPD (defined as a FEV1/FVC < 70% and FEV1 < 30% predicted) and/or severe 
persistent asthma. 
h. Hemodynamic instability with systolic blood pressure <90 mmHg or heart rate > 120 
beats/min, unless deemed to be stable with these values by the attending physicians. 
6. Prior complications with SCT 
7. Contraindication to rigid bronchoscopy 
8. Significant tracheomalacia or alterations in cartilage integrity that would require stent placement 
or surgical referral as assessed by CT imaging. 
9. Greater then 1 BCAS intervention within 6 months before enrollment 
 
 
Primary endpoint: 
The primary endpoint will be the degree of re-stenosis at 6 months expressed as the percentage of airway 
lumen volume within the stenotic segment at 6 months compared to personal best patency volume on CT 
scan performed within five weeks of the study intervention. If patients require repeat intervention before 
six months based on clinical course, a CT chest before this re-intervention will be analyzed for the 
primary outcome. All analyses will be performed using OsiriX (Pixmeo, Geneva, Switzerland, see 
below). 
7.  Outcomes 

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Secondary and exploratory endpoints: 
• 
Change in PEF (Secondary) 
• 
Change in QOL questionnaire (Secondary) 
• 
Incidence of complications between groups (Secondary) 
 
 
 
8.1 : Expected Outcomes 
• 
We anticipate being able to enroll 40 patients over 1-year period at VUMC with the possibility of 
opening the study at a second site if enrollment is deemed insufficient at 6 months (15 patients 
enrolled or less) 
• 
We expect to observe a significant difference in percent change of stenotic volume between SCT- 
treated patients and controls suggesting decreased recurrence in SCT-treated patients. 
8.2 : Randomization 
Patients will be randomly allocated in a 1:1 ratio in permuted blocks of 4 and 6 to intervention or control 
arms via randomization module in REDCap. Randomization tables will be set up by a study statistician 
with no role in recruitment, enrollment, or subsequent randomization and will not be available to study 
personnel performing these tasks. 
8.3 : Power Calculation and target accrual 
A search of the literature in idiopathic tracheal stenosis (ITS) shows that clinical recurrence after first 
intervention can happen as early as 6 months. A study by Perotin et al out of 9 institutions of 23 patients 
with ITS found recurrence of stenosis to be 30% at 6 months, 59% at 2 years, and 87% at 5 years with a 
delay of 14 to 16 months 10. Further a study by Gelbard et al evaluated 603 patients who underwent 
endobronchial dilation in ITS and found the time to stenosis reoccurrence averaged 1.3 years, which was 
considerably shorter than cricotracheal resection 11. We hypothesize that the earliest signs of loss of 
airway patency will be evident at 6 months by radiologic CT-based volumetric assessment, which will 
detect early (as opposed to clinical) recurrence. Our results will inform the design of future larger studies. 
We hypothesize the patency assessed at 6 months will be 90% of baseline in the intervention group and 
70% in the control group, with SD 20% in the control group. The sample size estimation was completed 
using two-sample t-test. For the pilot study, a sample size of 16 subjects in control and 16 subjects in 
intervention group will have 80% power to detect an increase in stenotic volume of 20% from control arm 
with type I error of 0.05. An additional 8 patients will be enrolled (attrition), for a total target accrual of 
40 patients. Likert scale will be used after treatment to determine the minimal clinically important 
difference for future studies using this endpoint. 
8.4 : Analytical Plan 
8.  Statistical Design 

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Descriptive statistics including means, standard deviations, and ranges for continuous parameters, as well 
as percentages and frequencies for categorical parameters will be presented. Investigations for outliers 
and assumptions for statistical analysis, e.g., normality and homoscedasticity will be made. If necessary, 
data will be transformed using Box-Cox power transformation. Comparisons between groups, i.e. 
intervention vs control, will be made using either the t-test or Wilcoxon Rank Sum test for continuous 
variables (such as patency volume, visual analog scales) and Chi-square test for categorical variables 
(such as incidences of pneumothorax). Other than the primary end point of percentage change of patency 
volume at 6 month, we will apply linear regression to assess the 6-month patency volume difference 
between control and intervention groups with the adjustment of baseline patency volume. Patients will be 
analyzed according to the per protocol and intention to treat principle. For peak flows, COPD 
questionnaire, intermediatory time points will be analyzed using mixed model analysis for repeated 
continuous variables starting from 2 weeks pre-procedure, 3 months and then to 6 months post procedure. 
8.5  Measurements: 
PEF, quality of life using the Clinical COPD CCQ. Incidence of complications (hypoxemia, bleeding, 
pneumothorax rate, barotrauma). Baseline characteristics (age, sex, body mass index (BMI), smoking 
status, underlying disease process). CT Chest without contrast with 3D quantitative reconstruction. 
Peak expiratory flow monitoring: Evidence supports the use of PEF as a simple, efficient, and 
accessible way of monitoring progression of tracheal stenosis. PEF has previously been captured in 
clinical encounters as component of the decision-making algorithm for surgical intervention in tracheal 
stenosis patients. Mobile Device Software ‘App’ can robustly capture longitudinal patient reported 
outcome measures.. 
CQS mobile computing group: Established expert programmer and data scientist support 
dedicated to mobile application development and support. Prior work with the CQS team 
successfully created and deployed a mobile app (Airflo™) for tracheal stenosis patient PEF 
reporting. Leveraging a Research Electronic Data Capture (REDCap) foundation, mobile apps are 
built on open-source mobile computing "MyCap" infrastructure (https://projectmycap.org/). 
MyCap leverages REDCap, ResearchKit (Apple: iOS), and ResearchStack (Android) to capture 
patient reported outcome measures, and physiologic tasks (such as maximum phonation time, 
voice samples, and peak expiratory flow rate) using iOS and Android mobile devices. REDCap is 
used to define tasks/instruments/surveys to be completed by participants. MyCap translates 
REDCap task metadata into a structure compatible with ResearchKit and ResearchStack. When a 
project participant completes a task, MyCap converts the results into a format compatible with 
REDCap before synchronizing back to the REDCap project. 
 
 
9.1 : Quantitative imaging of stenotic volume 
Airway segmentation and measurement of the patency volume will be performed as follows by two 
experienced operators (FM and R). DICOM images of CT scans will be transferred to the LungCloud™ 
platform. 
9.  Outcome Measure Synopsis 

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The LungCloud™ is provided by Neurotargeting LLC, a spin-off of Vanderbilt University. The 
LungCloud™ is a health IT clinically approved system for the collection of complete data sets in the 
clinical flow, including but not limited to clinical outcome, radiology notes and scans. The LungCloud™ 
is set up as a network of accounts that can be interconnected around a clinical study. The managing 
institution creates a Clinical Trial inside the LungCloud™ and ties it to each of the institutions. Each 
institution is then able to collect and share data for the clinical study. Data is collected in the clinical flow 
and from PACS and stays at all time in the LungCloud™ instance of the institution owning the data. 
Image data is encrypted and sent to the institution’s LungCloud™. Automatic uploading of images is 
achieved through what is referred to as the LungDrive. This is a process that runs on the end user’s 
machine and works in a way similar to Box or Dropbox, i.e., any new image volume placed in a specific 
folder is automatically encrypted and uploaded to the account repository. The non-imaging data can be 
entered into the system and associated with the imaging data through web-based forms customized to the 
end users. Images are reconstructed in 3D and can be reviewed in any view (sagittal, coronal or axial). 
The system offers a series of web-based user interfaces that permit inspecting and visualizing both 
imaging and non-imaging data. 
Deanonymized DICOM images will be transferred and analyzed using the OsiriX (Pixmeo, Geneva, 
Switzerland) threshold-based region growing algorithm function for volume segmentation. Briefly, the 
process is as follows: The operator identifies proximal and distal ends of the tracheal stenotic and use the 
region of interest (ROI) function to segment the trachea in 3 to 4 different areas spanning the length of the 
stenotic segment using the closed polygonal function (Figure 5). The ROI volume function will then be 
used to generate missing intervening ROIs and the repulsor function will be used to manually adjust the 
boundaries of the ROI. The volume of the final airway will then be computed and a three-dimensional 
image saved for documentation (Figure 6). 
To classify the degree of tracheal stenosis, we will manually annotate the trachea (wall, stenosis, and 
patency) on all scans. Expert manual review will be performed by one of two experienced investigators 
(FM, AR) In an exploratory analysis to enable routine automated analysis of stenotic volumes, we will 
perform transfer learning using our abdomen/thoracic model using leave-one-out cross-validation. 
Mixture modeling with image intensity classification will be used to separate the tracheal wall, stenosis 
region, and patency following our work in the abdomen. 
 
Figure 5. Closed polygonal function-derived 2D-tracheal segmentation 

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Figure 6. 3D reconstruction and computation of the patency volume 
 
 
Nouraei and colleagues successfully validated the Clinical Chronic Obstructive Pulmonary Disease 
Questionnaire in a group of patients with laryngotracheal stenosis. Total and domain-specific scores also 
corrected with FEV1 and PEFR. The authors found statistically significant differences between total and 
domain specific CCQ scores across populations of patients with different preoperative stenosis severities 
(P <0.001 in all cases; one‐way analysis of variance). Furthermore, there were statistically significant 
differences between preoperative and postoperative total and domain‐specific CCQ scores (P <0.001 in 
all cases; Student’s t‐test)18. 
We plan to use this questionnaire at carefully selected timepoints throughout the study timeline to assess 
symptom improvement pre- and post-intervention. 
 
 
 
General Safety Events (GSE): 
General safety events will be reported within our data collection protocol which includes safety events 
associated with the procedure (Spray Cryotherapy or Ballon Dilation): bleeding, pneumothorax, infection, 
airway trauma, air embolism. 
Serious adverse event (SAE) 
A serious adverse event (SAE) is an undesirable sign, symptom, or medical condition. 
which: 
• 
is fatal or life-threatening; 
10. Clinical COPD Questionnaire 
11. Safety 

17 of 27 
 
 
• 
requires or prolongs inpatient hospitalization; 
• 
results in persistent or significant disability/incapacity; 
• 
constitutes a congenital anomaly or birth defect; or 
• 
jeopardizes the participant and requires medical or surgical intervention to 
prevent one of the outcomes listed above. 
 
Events not considered to be serious adverse events are hospitalizations for: 
 
 
• 
routine treatment or monitoring of the studied indication, not associated with 
any deterioration in condition, or for elective procedures 
• 
elective or pre-planned treatment for a pre-existing condition that did not 
worsen 
• 
emergency outpatient treatment for an event not fulfilling the serious criteria 
outlined above cannot resulting in inpatient admission. 
• 
respite care 
 
 
General Instructions for Reporting Serious Adverse Events 
The Institutional Review Board and Industry Sponsor be notified of all SAEs, within 7 business days after 
the treating institution becomes aware of the event. Only SAEs related to research procedures will be 
reported to the IRB. 
 
 
12.1 Data Collection 
The Vanderbilt University Office of Research will be used as a central location for data processing and 
management. Vanderbilt University, with collaboration from a consortium of institutional partners, has 
developed a software toolset and workflow methodology for electronic collection and management of 
research and clinical trial data. REDCap (Research Electronic Data Capture) is a secure, web-based 
application that is flexible enough to be used for a variety of types of research. REDCap provides an 
intuitive user interface that streamlines project development and improves data entry through real-time 
validation rules (with automated data type and range checks). 
REDCap also provides easy data manipulation (with audit trails for reporting, monitoring and querying 
patient records) and an automated export mechanism to common statistical packages (SPSS, SAS, Stata, 
R/S-Plus). In addition to traditional data capture functionality, REDCap’s survey capabilities are a powerful 
12. Data Handling 

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tool for building and managing online surveys. The research team can create and design surveys in a web 
browser and engage potential respondents using a variety of notification methods. All data collection 
projects rely on a thorough, study-specific data dictionary, defined by all members of the research team in 
an iterative, self-documenting process. This iterative development and testing process results in a well- 
planned and individualized data collection strategy. 
REDCap servers are housed in a local data center at Vanderbilt, and all web-based information transmission 
is encrypted. REDCap was developed specifically around HIPAA-Security guidelines and is recommended 
to Vanderbilt researchers by both our Privacy Office and Institutional Review Board. REDCap has been 
disseminated for local use at more than 940 other academic/non-profit consortium partners in 75 countries. 
Vanderbilt leads the REDCap Consortium, which currently supports more than 99,000 projects and 128,000 
users. 
12.2 Data and Safety Monitoring 
 
The Independent Data Monitoring Committee (iDMC) will be comprised of two members with 
experience in airway stenosis, thoracic surgery and interventional pulmonary. Dr. Samira 
Shojaee, and Dr. Eric Grogan both from Vanderbilt University Medical Center (VUMC). Both 
members will meet periodically to discuss data and any SAEs. The group maintains authority to 
intervene in the conduct of studies as necessary to ensure clinical-research is performed at the 
highest quality standards. 
 
Both members have a clinical expertise include interventional pulmonology, minimal invasive 
diagnostics, airway stenosis and lung cancer. They also both have extensive experience in the 
minimally invasive management of malignant airway obstruction, complex airway management 
and difficulty airways. In addition, Dr. Shojaee specializes in the diagnosis and management of 
pleural diseases and symptom relief with minimally invasive procedures such as indwelling 
pleural catheters and pleurodesis. Dr. Eric Groan is also NIH R01 funded and co-chair of the 
MAS Lung Cancer Lab 
 
 
12.3 Data Handling and Record Keeping 
An electronic case report form (eCRF) is required and must be completed for each included participant. 
The completed dataset should not be made available in any form to third parties, except for authorized 
representatives of proper Health/Regulatory Authorities, without written permission from Vanderbilt. 
To enable evaluations and/or audits from health authorities and Vanderbilt, the site investigator agrees to 
keep records including: The identity of all participants (sufficient information to link records; e.g., hospital 
records), all original signed informed consent forms, copies of all source documents, and detailed records 
of any study drug or device disposition. To comply with international regulations, the records should be 
retained by the investigator in compliance with regulations. 
Queries resulting from review of the eCRFs will be generated for the site and corrections will be made by 
the study site personnel. This will be done on an ongoing basis. 
 
 
13. Regulatory Considerations 

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13.1 Protocol Review and Amendments 
Information regarding study conduct and progress will be reported to the Institutional Review Board 
(IRB) per current institutional standards. 
The trial will not be initiated until there is approval by the IRB of the protocol, informed consent 
document and any other material used to inform the patient about the nature of the trial. The IRB should 
be duly constituted according to regulatory requirements. The investigator will inform the IRB of the 
progress of the trial at least yearly. 
Any changes to the protocol will be made in the form of a written amendment and must be approved by 
the sponsor-investigator and the IRB prior to implementation. Protocol changes to eliminate an immediate 
hazard to a trial patient may be implemented by the investigator immediately. The investigator must then 
immediately inform any applicable local IRB and the sponsor-investigator (or designee). 
The sponsor-investigator (or designee) is responsible for the coordination and development of all protocol 
amendments. Once approved by the sponsor-investigator, Vanderbilt will disseminate this information to 
the participating study sites. 
13.2 Informed Consent 
The investigator (or his/her designee) will explain to each participant the nature of the study, its purpose, 
the procedures involved, the expected duration, the potential risks and benefits involved and any 
discomfort it may entail. Each participant will be informed that participation in the study is voluntary, that 
s/he may withdraw from the study at any time, and that withdrawal of consent will not affect subsequent 
medical treatment or relationship with the treating physician(s) or institution. 
The informed consent will be given by means of a standard written or in electronic format, written in non- 
technical language, which will be IRB approved. The participant should read and consider the statement 
before signing and dating it and will be given a copy of the document. No patient will enter the study or 
have study-specific procedures done before his/her informed consent has been obtained. 
In accordance with the Health Information Portability and Accountability Act (HIPAA), the written 
informed consent document (or a separate document to be given in conjunction with the consent 
document) will include a subject authorization to release medical information to the study sponsor and 
supporting agencies and/or allow these bodies, a regulatory authority, or Institutional Review Board 
access to subjects’ medical information that includes all hospital records relevant to the study, including 
subjects’ medical history. 
13.3 Ethics and Good Clinical Practice 
This study will be carried out in compliance with the protocol and Good Clinical Practice (GCP), as 
described within: 
1. ICH Harmonized Tripartite Guidelines for Good Clinical Practice 1996. 
2. Declaration of Helsinki, concerning medical research in humans (Recommendations Guiding 
Physicians in Biomedical Research Involving Human Subjects, Helsinki 1964, amended Tokyo 1975, 
Venice 1983, Hong Kong 1989, Somerset West 1996). 
The investigator agrees to adhere to the instructions and procedures described within the above and 
thereby to adhere to the principles of Good Clinical Practice with which the above conform. 

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13.4 Confidentiality 
It is the responsibility of the investigator to ensure the confidentiality of patients participating in the trial 
and all of their medical information is maintained. CRFs and other documents submitted to regulatory 
authorities must not contain the name of a trial patient. All patients in the trial will be identified by a 
unique identifier which will be used on all CRFs and any other material submitted to regulatory 
authorities. All case report forms and any identifying information must be kept in a secure location with 
access limited to the study staff directly assisting with the trial. 
13.5 Study Termination 
The sponsor-investigator reserves the right to terminate the study at any site and at any time. Reasons for 
study termination may include, but are not limited to, the following: 
• Investigator non-compliance with the protocol, GCP or regulatory requirements. 
• Insufficient enrollment if not supplemented adequately by additional site as defined by 15 patients 
enrolled or less at 6 months. 
• Safety concerns. 
• Decision by suppliers to modify or discontinue the availability, development or manufacture of 
protocol-indicated treatment or device. 
• A request to discontinue the study by the IRB or a recognized regulatory authority. 
 
 
Subjects will not be paid to participate in the study. 
There will be no additional costs to subjects for participating in this study. Subjects and/or their insurance 
companies will be responsible for all care provided as part of the standard of care stated above. 
Full study budget can get found in the separate budget allocation sheet. 
 
 
 
15.1 Trial Compliance 
This is an investigator-industry sponsored study. The Principal Investigators, Fabien Maldonado, M.D. 
are conducting the study with the sponsor US Endoscopy with the device truFreeze Spray Cryotherapy. 
Vanderbilt is the Lead site for this study. All aspects of the study will be carefully monitored by the Lead 
site for compliance with applicable government regulations with respect to current GCP and standard 
operating procedures. 
15.2 Changes to Protocol and Informed Consent Document 
14. Cost 
15. Study Coordination 

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Any change to the protocol and informed consent document must be reviewed and approved by the Lead 
site before being submitted to the Institutional Review Board/Independent Ethics Committee (IRB/IEC) at 
participating institutions and the industry sponsor. Amendments should not be implemented until all 
necessary approvals have been obtained, except when necessary to eliminate an immediate hazard to study 
subjects. 
15.3 Protocol Deviations 
The Lead site is responsible for implementing and maintaining quality assurance and quality control to 
ensure that studies are conducted according to the protocol, GCP, and all applicable regulatory 
requirements. A protocol deviation is any noncompliance with the protocol. Noncompliance can be on the 
part of the study participant, the investigator, or the study site staff. Deviations to the protocol are not 
permitted except when necessary to eliminate an immediate hazard to study subjects. 
15.4 Monitoring and Quality Assurance 
As the Lead site, Vanderbilt has responsibilities to health authorities to take all reasonable steps to ensure 
the proper conduct of the study with regard to ethics, protocol adherence, integrity, validity of the data 
recorded on the CRFs, and adherence to regulations regarding GCP and the protection of human subjects. 
15.5 Data Verification 
Data will be collected via eCRFs and entered into the database per Lead site guidelines. The Lead site will 
check data accuracy by performing source data verification. Source data verification is a direct comparison 
of the entries made on the CRFs against the appropriate source documentation. This will be conducted 
remotely, with the possibility of on-site verification periodically. Discrepancies in the data will be brought 
to the attention of the investigator and/or the investigator’s staff. Any necessary corrections will be made 
directly to the eCRFs or via queries by the investigator and/or the investigator’s staff. 
15.6 Study Documentation 
Each participating site is responsible for submitting copies of all relevant regulatory documentation to the 
Lead site. The required documents include but are not limited to the following: IRB approvals (i.e., 
protocol, consent form, amendments, patient brochures, recruitment material, etc.), each participant’s 
informed consent, enrollment form, eligibility checklist, summary of unanticipated problems or protocol 
deviations, and documentation of expertise of the investigators. The Lead site will provide each 
participating site with a comprehensive list of the necessary documents. Specified members at each 
participating site will submit all pertinent regulatory documents to the Lead site, for storage in a secure 
location. It is the responsibility of the participating sites to maintain copies of all documentation 
submitted to the Lead site. 
15.7 Closure of the Study 
The Lead site reserves the right to discontinue a site at any time during the study for medical or 
administrative reasons such as: 
· Unsatisfactory enrollment; 
· GCP noncompliance; 

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· Inaccurate or incomplete data collection; 
· Falsification of records; 
· Failure to adhere to the study protocol. 
15.8 Records Retention 
FDA regulations (21 CFR §312.62[c]) require that records and documents pertaining to the conduct of 
this study, including CRFs, consent forms, laboratory test results and physical exam, imaging and medical 
procedure records, must be retained by each site’s Principal Investigator for 2 years after marketing 
application approval. If no application is filed, these records must be kept 2 years after the study is 
discontinued and the applicable national and local health authorities are notified. 
Following closure of the study, each participating site will maintain a copy of all site study records in a 
safe and secure location. The Lead site will inform the investigator at each site at such time that the 
records may be destroyed. 
 
 
Any manuscript or releases resulting from the collaborative research must be approved by the investigator 
and will be circulated to applicable participating sites/investigators prior to submission for publication or 
presentation. A publication plan consistent with the international Committee of Medical Journal Editors 
(ICMJE) will be created prior to analysis ad publication of any data. All data will be made available to 
authors as required. The publication of sub-studies, post-hoc analyses, or single-center experiences will 
not precede the primary multicenter publication. Publication of results will be determined by the 
investigators, without limitations from the funder. Input (non-binding) will be obtained from the funder 
who will be given access to the data after closure of the study. All authors are expected to disclose 
financial or affiliations that could be considered conflicts of interest per journal or medical society 
requirements. 
 
 
1. Barros Casas, D., Fernández-Bussy, S., Folch, E., Flandes Aldeyturriaga, J. & Majid, A. Non- 
malignant central airway obstruction. Arch. Bronconeumol. 50, 345–354 (2014) 
2. Brigger, M. T. & Boseley, M. E. Management of tracheal stenosis. Curr. Opin. Otolaryngol. 
Head Neck Surg. 20, 491–496 (2012) 
3. Qiu, X.-J., Zhang, J., Wang, T., Pei, Y.-H. & Xu, M. Nonstent Combination Interventional 
Therapy for Treatment of Benign Cicatricial Airway Stenosis. Chin. Med. J. 128, 2154–2161 
(2015) 
4. Shapshay, S. M., Beamis, J. F., Jr, Hybels, R. L. & Bohigian, R. K. Endoscopic treatment of 
subglottic and tracheal stenosis by radial laser incision and dilation. Ann. Otol. Rhinol. Laryngol. 
96, 661–664 (1987) 
5. Gosain, S., Mercer, K., Twaddell, W. S., Uradomo, L. & Greenwald, B. D. Liquid nitrogen spray 
cryotherapy in Barrett’s esophagus with high-grade dysplasia: long-term results. Gastrointest. 
Endosc. 78, 260–265 (2013) 
16. Publications 
17. Citations 

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6. Au, J. T., Carson, J., Monette, S. & Finley, D. J. Spray cryotherapy is effective for 
bronchoscopic, endoscopic and open ablation of thoracic tissues. Interact. Cardiovasc. Thorac. 
Surg. 15, 580–584 (2012) 
7. Krimsky, W. S., Broussard, J. N., Sarkar, S. A. & Harley, D. P. Bronchoscopic spray 
cryotherapy: assessment of safety and depth of airway injury. J. Thorac. Cardiovasc. Surg. 139, 
781–782 (2010) 
8. Sackner, M. A. Physiologic features of upper airway obstruction. Chest 62, 414–417 (1972) 
9. Simonsson, B. G. & Malmberg, R. DIFFERENTIATION BETWEEN LOCALIZED AND 
GENERALIZED AIRWAY OBSTRUCTION. Thorax 19, 416–419 (1964) 
10. Perotin, J.-M. et al. Endoscopic management of idiopathic tracheal stenosis. Ann. Thorac. Surg. 
92, 297–301 (2011) 
11. Gelbard, A. et al. Comparative Treatment Outcomes for Patients With Idiopathic Subglottic 
Stenosis. JAMA Otolaryngol. Head Neck Surg. 146, 20–29 (2020) 
12. Nouraei, S. A. R. et al. Validation of the Clinical COPD Questionnaire as a psychophysical 
outcome measure in adult laryngotracheal stenosis. Clin. Otolaryngol. 34, 343–348 (2009) 

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18. Consort Diagram 

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20. Study Assessment Table 
Study Procedures 
(Subject level) 
Pre-Procedure 
Post Procedure 
3 months 
6 months 
Pathology review/Cryobiopsy 
Non-SOC (1) 
CT scan (Chest) 
Non-SOC (1) 
SOC (1) 
Physician’s Time/Exam 
SOC (1) 
SOC (1) 
Bronchoscopy 
SOC (1) 
Questionnaire Delivery 
Non-SOC (1) 
Non-SOC (1) 
Non-SOC (1) 
Non-SOC (1) 
PEF 
Non-SOC (1) 
Non-SOC (1) 
Non-SOC (1) 
Non-SOC (1) 
 
Study Procedures 
(Site level) 
Site start up 
Non-SOC (1) 
IRB Cost 
INV 
Radiology Review setup 
Non-SOC (1) 

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Serious adverse events 
INV 
Archiving/Document 
storage 
Non-SOC (1) 
Statistician 
Non-SOC (1) 
Segmentation and 
Volumetric Analysis 
Non-SOC (1) 
RedCAP Development 
Non-SOC (1) 
 
 
 
 
21. COPD Questionnaire (CCQ) 

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