Risk of Microbial Translocation in Patients Undergoing Per-Oral Endoscopic Myotomy (POEM) for Achalasia: Antibiotic Prophylaxis or Short-therapy
Conditions: Achalasia
Sponsor: Istituto Clinico Humanitas

 
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Risk of Microbial Translocation in patients undergoing Per-Oral Endoscopic 
Myotomy (POEM) for Achalasia: Antibiotic Prophylaxis or Short-therapy? 
 
Promotore della sperimentazione: Dr. Roberta Maselli 
 
Codice dello studio: POEM-MT01 
 
Version N° 1- 15 Oct 2016 
 
 
  
 
 
 
 

 
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Contact addresses 
 
 
 
 
 
 
 
Principal investigator: 
Dr. Roberta Maselli 
Phone: 02-82247570 
Fax: 02-82242292 
e-mail: roberta.maselli@humanitas.it  
Study Coordinator: 
 
Dr. Roberta Maselli 
Phone: 02-82247570 
Fax: 02-82242292 
e-mail: roberta.maselli@humanitas.it 
Institutional Address 
IRCCS Humanitas Research Hospital 
Via A.Manzoni 56 
20089 Rozzano, Milano 
Collaborating center 
Università degli Studi di Roma 
“La Sapienza” 
Piazzale Aldo Moro 5  
00185 Roma 

 
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Signatures Page 
 
 
 
 
 
STUDY COORDINATOR SIGNATURE (where applicable) 
 
 
Printed name 
 
 
Role & Department 
 
 
 
 
 
Signature 
 
Date 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
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CENTRE SIGNATURE – PRINCIPAL INVESTIGATOR 
 
I have read this Protocol Amendment relevant to the study entitled “Risk of Microbial 
Translocation in patients undergoing Per-Oral Endoscopic Myotomy (POEM) for Achalasia: 
Antibiotic Prophylaxis or Short-therapy?” and I agree to conduct the study as detailed herein and in 
compliance with guidelines for Good Clinical Practice and applicable regulatory requirements. I 
will provide all study personnel under my supervision with all information provided by the Sponsor 
and I will inform them about their responsibilities and obligations. 
 
 
 
Printed name 
 
 
Role & Department 
 
 
 
Address 
 
 
 
 
Signature 
 
Date 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
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INDEX 
SUMMARY .......................................................................................................................... 7 
1 
BACKGROUND AND INTRODUCTION .................................................................... 10 
2 
RATIONALE OF THE STUDY ................................................................................... 11 
3 
OBJECTIVES OF THE STUDY .................................................................................. 11 
4 
PATIENT SELECTION CRITERIA ............................................................................. 11 
4.1 
Inclusion criteria .................................................................................................................. 12 
4.2 
Exclusion criteria ................................................................................................................. 12 
5 
STUDY DESIGN ......................................................................................................... 12 
6 
STUDY TREATMENT ................................................................................................ 13 
7 
CLINICAL EVALUATION, LABORATORY TESTS AND FOLLOW-UP .................... 14 
8 
STATISTICAL CONSIDERATIONS ........................................................................... 14 
9 
FORMS AND PROCEDURES FOR COLLECTING DATA AND DATA MANAGING 14 
10 
ETHICAL CONSIDERATIONS ............................................................................... 15 
10.1 
Patient protection ................................................................................................................. 15 
10.2 
Subject identification – Personal Data protection ............................................................ 15 
10.3 
Informed consent.................................................................................................................. 16 
11 
CONFLICT OF INTEREST ..................................................................................... 16 
12 
DATA OWNERSHIP ............................................................................................... 16 
13 
PUBLICATION POLICY ......................................................................................... 17 
14 
STUDY TIME TABLE ............................................................................................. 17 
15 
REFERENCES........................................................................................................ 18 

 
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Summary   
 
Title 
Risk of Microbial Translocation in patients undergoing Per-Oral 
Endoscopic Myotomy (POEM) for Achalasia: Antibiotic 
Prophylaxis or Short-therapy 
Study coordinator 
Dr. Roberta Maselli 
Protocol identifying code 
POEM-MT01 
Protocol version date 
Version N° 1. 15 Oct 2016 
Background and rationale 
Achalasia is a primary rare esophageal motor disorder of the 
esophagus (annual incidence of 1:100,000 persons). Recently, a 
new endoscopic technique, Per-Oral Endoscopic Myotomy 
(POEM), has been introduced with excellent success rates. 
Several studies have evaluated complications of POEM but there 
is a lack of knowledge on the potential risk of bacteremia or 
microbial translocation during the endoscopic intervention and, 
also, there aren’t evidences regarding the use of antibiotics 
before/after POEM. Microbial translocation (MT) is the passage 
of both viable and nonviable microbes across the anatomically 
intact GI barrier to the mesenteric lymph nodes, and possibly 
other 
tissues. 
Gram-negative 
bacteria 
contain 
lipopolysaccharides (LPSs) coating their thin peptidoglycan cell 
wall. The presence of LPS, an endotoxin, in the plasma has been 
correlated to sepsis and septic shock through the activation of the 
inflammatory host defence via binding to soluble CD14 (sCD14) 
which initiates downstream cytokines (like IL-6, IL-8 and tumor 
necrosis factor (TNF-α)) and, also, through the production of 
sCD14 and LPS-binding protein (LBP) by the innate immune 
system. 
Considering POEM a clean-contaminated procedure, it should be 
assessed whether the post-POEM fever or systemic inflammation 
is a cytokinin-mediated or an infection-related fever. Thus, aim 
of the study is to evaluate the presence of inflammation 
mediators, bacteremia and microbial translocation post POEM, 
to guide future antibiotic prophylaxis/therapy in patients 
undergoing this procedure. 
Population and patient 
selection criteria 
Study population: patients with esophageal achalasia (diagnosed 
accordingly to the international guidelines) scheduled for POEM, who 
are willing to partecipate in a RCT and who are able to give informed 
consent.  
Inclusion citeria: 
1. Patients who had a diagnosis of achalasia defined by symptoms, HR 
manometry and X-ray accordingly to guidelines, who have no absolute 
contraindications to undergo interventions under general anesthesia  
2. Age over 18 years 
3. Ability to provide and to give informed consent 
4. Body temperature under 37 °C on the day before and just before 
POEM 
Exclusion criteria: 
1. Antibiotic use within 1 week before the procedure 

 
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2. Patients who had chronic inflammatory diseases (such as 
rheumatic arthritis or inflammatory bowel diseases) and/or 
known neoplasia 
3. Inability to obtain written informed consent 
4. Patient unwilling to participate to the study 
5. Impossibility to be subjected to the invasive endoscopic 
procedure or general anesthesia for the presence of comorbidities 
Study design and study 
duration 
In this study we will enroll all consecutive patients with 
esophageal 
achalasia, 
diagnosed 
by 
clinical 
symptoms, 
esophageal HR manometry and X-ray barium swallow 
(accordingly to the international guidelines) evaluated during a 
gastroenterological outpatient visit. These patients will undergo 
POEM at our institution from June 2017 to June 2019. 
Duration of the study (patient enrolment; treatment duration, 
follow up, etc.): we expected six months for the approval of the 
study by the Ethics Committee of our Institute, personnel 
training, study protocol registration; twenty-four months of 
enrollment, interventions and follow-up; six months for 
processing data and for the drafting of the final manuscript. 
Description of study 
treatment/product/intervention 
All patients who will undergo POEM at Humanitas Research 
Hospital from June 2017 to June 2019 will be enrolled in a 
prospective, interventional randomized clinical trial (RCT). 
Patients will be randomized in two groups. The Group A, 
prophylaxis group, will receive antibiotics (Cefazoline 2 gr i.v.) 
only before procedure whereas Group B, short therapy group, 
will receive antibiotics before POEM (Cefazoline 2 gr i.v.), 
continued 
for 
the 
first 
24 
hours 
and 
then 
per 
os 
(Amoxicilline/Clavulanic Acid 3 gr/die) for 3 days. For each 
patient we will be evaluated: plasma dosage of IL-6, IL-1β, 
TNF-α, sCD4, LPB, LPS and blood cultures.  
Objectives 
Considering POEM a clean-contaminated procedure, it should be 
assessed whether the post-POEM fever or systemic inflammation 
is a cytokinin-mediated or an infection-related fever. Thus, aim 
of the study is to evaluate the presence of inflammation 
mediators, bacteremia and microbial translocation post POEM, 
to guide future antibiotic prophylaxis/therapy in patients 
undergoing this procedure.  
Statistical methods, data 
analysis 
From a retrospective internal analysis on POEMs performed 
from October 2015 to October 2016 in our istitution we 
estimated that, in patients submitted to short antibiotic therapy, 
the prevalence of fever and/or systemic inflammation is about 
30%. We want to demonstrate that giving only prophylaxis 
antibiotics, the prevalence of fever and/or systemic inflammation 
will be not higher than 10% difference with and alpha error of 
5% and a power of 80%: we need 62 patients each group, for a 
total of 124 patients.  
Data will be described as number, percentage and confidence 
interval, or mean, standard deviation and confidence interval, or 

 
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median 
and 
interquartile 
interval 
where 
appropriated. 
Differences between groups will be explored with chi square 
test, with fisher correction if appropriated with qualitative data, 
or with wilcoxon test or t student test if quantitative. 
No interim analysis is planned. 
Study time table 
- duration of the study (patient enrolment; treatment duration, 
follow up, etc.): we expected six months for the approval of the 
study by the Ethics Committee of our Institute, personnel 
training, study protocol registration; twenty-four months of 
enrollment, interventions and follow-up; six months for 
processing data and for the drafting of the final manuscript. 
 
- when the final report will be provided: December 2019 
 
- overall study duration from the First Patient First Visit (FPFV): 
twenty-four months plus five days 
 
 
 
 

 
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1 Background and introduction 
Achalasia is a primary rare esophageal motor disorder of the esophagus with an estimated annual 
incidence of 1:100,000. It is characterized by the absence of esophageal peristalsis and by failure of 
the lower esophageal sphincter to relax upon swallowing. Recently, a new endoscopic technique, 
POEM has been introduced as an alternative treatment to surgery with excellent success rates.  
Several studies have evaluated complications of POEM such as bleeding, perioperative perforation, 
pneumothorax, pneumoperitoneum, but there is a lack of knowledge on the potential risk of 
bacteremia or microbial translocation (MT) during the endoscopic intervention.  
Moreover, several reviews reported the routine use of pre-procedure antibiotic prophylaxis for 
POEM with no specifying regarding the type and dosage of antibiotic . The latest ASGE (American 
Society od Gastrointestinal Endoscopy) guidelines on antibiotic prophylaxis for GI (gastro-
intestinal) endoscopy do not illustrate the management of antibiotic for POEM. On the contrary, in 
clinical practice most centers use to administer antibiotics just after the procedure; no evidences 
have been published regarding the use of antibiotics before/after POEM.  
The reason for the development of post-POEM fever or systemic inflammation has not been 
investigated yet. It is possible that the long procedure time and contact time of electric knives to the 
submucosal layer could potentially promote GI bacteria translocation. Moreover, the extensive 
submucosal defect caused by POEM exposes deep GI layers to the indigenous bacterial flora which 
may theoretically cause microbial translocation and/or bacteremia.  
The presence of bacteremia during gastrointestinal endoscopy has been established in several 
studies. MT is the passage of both viable and nonviable microbes across through the anatomically 
intact GI barrier to the mesenteric lymph nodes, and possibly other tissues. All bacteria including 
those colonizing the gut can be divided into two groups, Gram-negative and Gram-positive bacteria, 
depending on their cell wall structure and outer membrane. Gram-negative bacteria contain LPSs 
coating their thin peptidoglycan cell wall. Over the past decades, the presence of LPS, commonly 
referred to as endotoxin, in the plasma has been correlated to multiple diseases including sepsis and 
septic shock. LPSs presence in the peripheral circulatory system could thus signify potential 
bacterial invasion, requiring rapid host response. Potent pyrogens, endotoxins activate the 
inflammatory host defence via binding to sCD14 (sCD14), which initiates downstream cytokines 
important for the clearance of bacterial infections. 
Additionally to the circulating IgM, IgA, and IgG antibodies directed against the LPS core antigen 
that neutralize its activity, the innate immune system produces soluble factors, such as sCD14, and 
LBP. CD14 is an LPS coreceptor expressed by peripheral blood monocytes and tissue macrophages. 
Following LPS stimulation, CD14+ monocytes/macrophages secrete sCD14 and shed surface 
CD14, which binds to LPS. 
Taken together, it is clear that several circulating factors, including sCD14 and LBP act as 
fundamental lines of defence against systemic stimulation of the immune system by translocated 
microbial antigens. 
When CD14 transfers LPS to monocytes two specific signalling pathways are activated both 
leading to the recruitment of numerous downstream molecules, which finally results in the 
transcription of proinflammatory cytokines such as interleukin IL-6, IL-1β and TNF-α. 
Since the manifestation of febrile state is dependent upon the pro-inflammatory response towards 
endotoxins, sCD14, LPS and LBP could be used as a suitable biomarker associated with microbial 
translocation, and IL-6, IL-1β and TNF-α as a biomarkers of inflammatory pattern. 
 

 
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2 Rationale of the study 
Literature does not describe a unique management toward antibiotic prophylaxis of patients that 
must undergo POEM. The same guidelines on antibiotic prophylaxis for GI endoscopy do not 
illustrate the management of antibiotic for POEM. Several reviews reported the routine use of pre-
procedure antibiotic prophylaxis for POEM (without specifying the type of antibiotic and dosage) 
but, on the contrary, in clinical practice most centers use to administer antibiotics just after the 
procedure.  
Our study, by evaluating the presence of inflammation mediators, bacteremia and microbial 
translocation post POEM, would like to investigate the origin of the post-POEM fever or post-
POEM systemic inflammation (if these are cytokinin-mediated or infection-related).  
 
In this way we could: 
- standardize the management of antibiotic therapy for patients undergoing POEM 
- allow adequate treatment selection and avoid unnecessary antibiotic usage 
- reduce the risk of antibiotic resistance 
- optimize the length of hospitalization 
 
3 Objectives of the study 
The primary objective of our study is determine whether fever (TC > 38°C) and systemic 
inflammation (increased in white blood cells (WBC) and/or protein C-reactive (PCR)) that may 
occur following POEM, are cytokine-mediated or infection-related and whether they are affected by 
prophylaxis or antibiotic short therapy. 
The secondary objectives of our study are: 
- assess the presence of post-POEM bacteremia though the result of blood cultures  
- assess the presence and quantify the degree of post-POEM microbial translocation through the 
dosage sCD4, LPB and LPS 
- quantify the degree of post-POEM systemic inflammation through the dosage of IL-6, IL-1β and 
TNF-α 
Statistical hypotesis: the aim of the study is to evaluate the presence of inflammation mediators, 
bacteremia and microbial translocation post-POEM, to guide future antibiotic prophylaxis/therapy 
in patients submitted to POEM. The study wants to assess the non-inferiority of antibiotics 
prophylaxis versus antibiotics short therapy in terms of fever and/or systemic inflammation control. 
 
4 Patient selection criteria 
Study population: patients older than 18, with esophageal achalasia (diagnosed accordingly to the 
international guidelines) scheduled for POEM, who are willing to partecipate in a RCT and who are 
able able to give informed consent.  
Clinical setting: the study will be conducted at the institute IRCCS Humanitas Research Hospital 
(Rozzano, Milano) at the operating unit of Gastroenterology and Digestive Endoscopy and at the 
Institute Sapienza University of Rome, Policlinico Umberto I° at the Department of Public Health 
and Infectious Diseases.  
Enrollment procedure: the investigator or his/her representative who will perform the procedure will 
explain the nature of the study to the subject and answer all questions regarding the study, in 

 
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particular he/she will illustrate the purpose of the study, the absence of risk for the patient, making 
it clear that the current procedure of POEM will not be changed. The subjects review, signed and 
dated the informed consent (a copy of the IC form will be given to the subject and the original 
placed in subject medical form).  
Accrual time: from June 2017 to June 2019 (patients enrollment) 
In our study we will not be involved vulnerable populations. 
 
4.1 Inclusion criteria  
1. Patients who had a diagnosis of achalasia defined by symptoms, HR manometry and X-ray 
accordingly to guidelines, who have no absolute contraindications to undergo interventions under 
general anesthesia  
2. Age over 18 years 
3. Ability to provide and to give informed consent 
4. Body temperature under 37 °C on the day before and just before POEM 
 
4.2 Exclusion criteria 
1. Antibiotic use within 1 week before the procedure 
2. Patients who had chronic inflammatory diseases (such as rheumatic arthritis or inflammatory 
bowel diseases) and/or known neoplasia 
3. Inability to obtain written informed consent 
4. Patient unwilling to participate to the study 
5. Impossibility to be subjected to the invasive endoscopic procedure or general anesthesia for the 
presence of comorbidities 
 
5 Study Design  
In this study we will enroll all consecutive patients with esophageal achalasia, diagnosed by clinical 
symptoms, esophageal HR manometry and X-ray barium swallow (accordingly to the international 
guidelines) evaluated during a gastroenterological outpatient visit. These patients will undergo 
POEM at our institution from June 2017 to June 2019. 
In our center, these patients usually undertake a preoperative routine (blood tests, cardiological 
evaluation, chest x-rays and anesthesia assessment) some weeks earlier, in order to test the 
feasibility of general anesthesia for the endoscopic invasive procedure. Patients are admitted to the 
gastroenterology department the morning of the procedure and, therefore, they are accompanied in 
the operating room.  
The POEM procedure will be carried out according to the current clinical practice. After an EGDS a 
solution is endoscopically injected in the mid esophagus to produce a large cushion, thereby 
promoting a safe mucosal incision. After submucosal injection a longitudinal large incision is 
performed to allow the entrance of the endoscope into the submucosal layer. Then, the submucosal 
layer is longitudinally dissected to create a tunnel from the esophageal submucosal incision to the 
gastric side. The created tunnel will be the working space to perform the myotomy. During the 
tunnelling any possible bleeding is promptly endoscopically treated. The endoscopic myotomy 
(ether selective of the circular muscle layer or both circular and longitudinal muscle layer) is then 
performed. Finally the mucosal incision is clipped by standard endoscopic clips. 
After the procedure, patients are moved to the ward where they received the appropriate supportive 
care (pain reliever and antiemetic as needed, intravenous hydration and IPP) and they remain 

 
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fasting. The next day blood tests are performed to rule out the occurrence of major complications; 
esophageal X-Ray are performed where needed. Then patients resume to eat with liquid diet and 
then with a soft diet. Finally, on the same day, in the absence of complications patients is 
discharged. Our current routine provides antibiotic coverage with Cefazolin ev during the entire 
duration of stay.  
When performing the clinical laboratoristic work-up, subjects will be evaluated on the basis of the 
inclusion and exclusion criteria. They will be proposed for the study and, if they agree to 
participate, they will sign the informed consent after adequate explanations. All patients who agree 
to participate in the study will be randomized, by a 1:1 randomization when arrived in the operating 
room (T0). At the same time, first blood samples will be collected for plasma cytokine dosage (IL-
6, IL-1β, TNF-α, sCD4, LPB and LPS) and bacteremia. Patients randomized to the group A, 
prophylaxis group, will receive a dose of antibiotics (Cefazoline 2 gr iv) 30 minutes before the 
procedure and patients randomized to the group B, short therapy group, will receive a dose of 
antibiotics (Cefazoline 2 gr iv); in this latter group the antibiotic will be also continued during 
hospitalization and at home (Cefazoline 6gr/day for the first 24 hours then orally 
Amoxicilline/Clavulanic Acid 3 gr/day for 3 days). For patients allergic to beta-lactams, a different 
class of antimicrobials will be prescribed according to the Institution’s clinical practice and 
patient’s characteristics. 
At the end of the procedure (T1) and at 24 hours after procedure (T2) the second and the third blood 
samples, respectively, will be carried out (for bacteremia, IL-6, IL-1β, TNF-α,  sCD4, LPB and 
LPS). In addition, blood samples will be performed to evaluate white blood count and PCR, as 
routinely done 24 hours after POEM. Additional blood cultures will be performed in case of fever 
(body temperature> 38 °C). It is finally scheduled a phone call five days after the procedure to 
complete the clinical follow-up and to ensure the compliance of recruitment of therapy for patients 
enrolled in group B. 
 
6 Study treatment  
Patients randomized to group A (prophylaxis group) will undergo a prophylactic antibiotic therapy 
with Cefazoline 2 gr administered by intravenously, 30 minutes before the procedure. Instead, 
patients randomized in the group B (short therapy group) will be subjected to a broader antibiotic 
coverage characterized by Cefazoline 2 gr i.v. 30 minutes before the procedure and then continued 
for the next 24 hours (Cefazoline 6 gr/ day i.v.) and, thereafter, orally for the next 3 days 
(Amoxicilline / Clavulanic Acid 3 gr / day). The dosage of antibiotics will be modified according to 
creatinine clearance. For patients allergic to beta-lactams, a different class of antimicrobials will be 
prescribed according to the Institution’s clinical practice and patient’s characteristics. 
In case the POEM will last for more than 4 hours, an additional administration of Cefazoline (2 gr 
i.v.) will be given.  
 
The object of our study is not to investigate the efficacy and safety of a new drugs. The drugs that 
we will use (Cefazoline, Amoxicilline/Clavulanic Acid) are commonly used in clinical practice and 
in the management of patients who undergo POEM, and part of GCP (Good Clinical Practice). 
 
Patient compliance will be monitored by means of a telephone survey. 
 
Stopping rules/discontinuation criteria: in our study will be not used experimental drugs, the patient 
will leave the study only in case of occurrence of allergic reaction to the drug used. 
 

 
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7 Clinical evaluation, laboratory tests and follow-up  
For each subject, blood samples will be collected at T0, T1, T2 for the following analyses: 
- 5 ml of whole blood will be inoculated on blood cultures bottles and will be transferred to the 
Humanitas Microbiology Laboratory within 3h from collection and processed as daily practice to 
evaluate the presence of bacteria in the blood; 
- 5 ml of whole blood sample for plasma collection. Briefly, plasma will be collected from EDTA 
whole blood samples within 6 hours of collection by centrifugal force of 1000g for 20 mins, will be 
then removed and stored at -80°C until the enzyme-linked immunosorbent assays (ELISA) for IL-6, 
IL-1β, TNF-α, sCD4, LPB and LPS will be performed. Experiments will be performed in duplicate. 
 
In the event that the patients develop fever accompanied by clinical signs of sepsis or develop a 
complication to the procedure (eg, aspiration pneumonia) specific changes of antibiotic therapy will 
be administered according to the clinical judgment of the physician. In case of positive blood 
culture to a specific bacterium, a specific therapy will, therefore, guided by antibiogram. 
 
There aren't drugs not allowed. 
 
8 Statistical considerations  
Randomization: patients will be randomized by a 1:1 randomization.  
Blinding: not applicable. 
 
From a retrospective internal analysis on POEMs performed from October 2015 to October 2016 in 
our institution we estimated that, in patients submitted to shot antibiotic therapy, the prevalence of 
fever and/or systemic inflammation is about 30%. We want to demonstrate that giving only 
prophylaxis antibiotics, the prevalence of fever and/or systemic inflammation will be not higher 
than 10% difference with and alpha error of 5% and a power of 80%: we need 62 patients each 
group, for a total of 124 patients.  
Data will be described as number, percentage and confidence interval, or mean, standard deviation 
and confidence interval, or median and interquartile interval where appropriated. Differences 
between groups will be explored with chi square test, with fisher correction if appropriated with 
qualitative data, or with wilcoxon test or t student test if quantitative. 
No interim analysis is planned. 
 
9 Forms and procedures for collecting data and data managing 
Data that will be gathered: indicators of systemic inflammation and of microbial translocation, 
presence of bacteremia, procedure's complications.  
The forms/tools used for the retrieval of information and their validity and reliability: all the tools 
used to obtain the data have been validated and certified annually by clinical engineering of our 
institute and the calibration, adequacy and validation certificates are preserved by them and can be 
consulted in case of need. 
The measures/indicators used: plasma dosage of sCD14, LPS, LBP, IL-6, IL-1β and TNF-α, results 
of blood cultures, presence of fever (TC > 38°C), increase in inflammatory markers (white blood 
cells and PCR). 
The potential sources of biases in the retrieval of information regarding study subjects and 
interventions/treatments: any possible missing data, because for example some of the participants 
could drop out before the end of the study, because participants could not follow the protocol either 

 
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deliberately or accidentally, or because some outcomes could be not measured correctly or cannot 
be measured at all at one or more time points. 
Duration and frequency of follow up: follow-up is performed with a phone call five days after the 
procedure. 
Expected estimate of subjects lost to follow up and possible implications for the findings of the 
study: 5% estimated lost to follow up; this will not affect the result of the study because the aim of 
the study is mainly focalized on the first 24 hours after POEM, when the patients is still 
hospitalized. 
Potential confounding factors and methods for taking into account their effect: with possible 
underlying co-infections could confound the results of our study, for this reason will not be enrolled 
patients with fever or patient who had taken antibiotics until 1 week before the procedures. Other 
potential confounders: patients with biochemical signs of inflammations/infections (increased PCR 
and/or WBC count), for this reason patients with any known systemic chronic inflammation disease 
or neoplasia will not be enrolled. 
Identification of source data definition (electronic clinical reporting form (e-CRF)): N/A because 
we will not use e-CRF but a paper CRF for records. Data will be finally collected in a specific 
electronic database and validated by an external monitor. 
 
10 Ethical considerations 
10.1 Patient protection 
The responsible investigator will ensure that this study is conducted in agreement with either the 
Declaration of Helsinki (Tokyo, Venice, Hong Kong and Somerset West amendments) or the laws 
and regulations of the country, whichever provides the greatest protection of the patient. 
The protocol has been written, and the study will be conducted according to the ICH Guideline for 
Good Clinical Practice  
The protocol and its annexes are subject to review and approval by the competent Independent 
Ethics Committee(s) (“IEC”). 
10.2 Subject identification – Personal Data protection 
All records identifying the subject must be kept confidential and, to the extent permitted by the 
applicable laws and/or regulations, not be made publicly available. The name of the patient will not 
be asked for nor recorded at the Data Center. A sequential identification number will be 
automatically attributed to each patient registered in the study. This number will identify the patient 
and must be included on all case report forms. In order to avoid identification errors, patient initials 
and date of birth will also be reported on the case report forms. 
Any and all patient information or documentation pertaining to a clinical trial, to the extent 
permitting, through a “key” kept anywhere, regardless of whether such key is supplied along with 
the information or documentation or not, must be considered as containing sensitive personal data 
of the patient, and is therefore subjected to the provisions of applicable data protection (“privacy”) 
regulations. Breach of such regulations may result in administrative or even criminal sanctions. 
Particularly, an information sheet prepared according to such regulations and a form to evidence the 
consent of patients to the processing of such data must therefore accompany the informed consent 
administered to the patient (see paragraph 14.3 below). Such information must (i) identify the roles 
of the holder (“titolare”) and processor (“responsabile”, appointed by the holder) of the patient 
personal data (also if not directly identifying the patient), as well as the purposes of the personal 

 
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data collection and processing (medical treatment and related/unrelated scientific research), (ii) 
adequately describe the flows of communication involving them, particularly if third parties should 
become involved, and (iii) seek the patient’s prior and specific consent to such processing. 
Patient information or documentation may be considered “anonymous”, and as such not subject to 
privacy regulations, only when no key whatsoever, permitting the identification of the patient, is 
any longer available. 
Particular attention should therefore be paid (and information/consent materials adapted 
accordingly) whenever patient data are supplied to third parties and may be autonomously 
processed, or biological samples/materials are taken and kept for future research purposes, 
associated or not with the pathology considered in the study.  
10.3 Informed consent 
All patients will be informed of the aims of the study, the possible adverse events, the procedures 
and possible hazards to which he/she will be exposed, and the mechanism of treatment allocation. 
They will be informed as to the strict confidentiality of their patient data, but that their medical 
records may be reviewed for study purposes by authorized individuals other than their treating 
physician. An example of a patient informed consent statement is given as an appendix to this 
protocol. 
It will be emphasized that the participation is voluntary and that the patient is allowed to refuse 
further participation in the protocol whenever he/she wants. This will not prejudice the patient’s 
subsequent care. Documented informed consent must be obtained for all patients included in the 
study before they are registered or randomized at the Data Center. This must be done in accordance 
with the national and local regulatory requirements. 
For European Union member states, the informed consent procedure must conform to the ICH 
guidelines on Good Clinical Practice. This implies that “the written informed consent form should 
be signed and personally dated by the patient or by the patient’s legally acceptable representative”. 
A copy of  Informed consent should be attached to this Protocol Template. 
 
11 Conflict of Interest 
Any investigator and/or research staff member who has a conflict of interest with this study (such as 
patent ownership, royalties, or financial gain greater than the minimum allowable by their 
institution) must fully disclose the nature of the conflict of interest.   
 
12 Data ownership 
According to the ICH Guidelines on Good Clinical Practice the sponsor of a study (the Institution, 
should the investigator or study coordinator act as sponsor in the performance of her/his 
institutional duties under the employment or collaboration agreement with Humanitas) is the owner 
of the data resulting therefrom. All centers and investigators participating in the study should be 
made aware of such circumstance and invited not to disseminate information or data without the 
Institution’s prior express consent. 
  

 
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13 Publication Policy 
After completion of the study, the project coordinator will prepare a draft manuscript containing 
final results of the study on the basis of the statistical analysis. The manuscript will be derived to 
the co-authors for comments and after revision will be sent to a major scientific journal. 
All publications, abstracts, presentations, manuscripts and slides including data from the present 
study will be submitted to and reviewed by the Study Coordinator for coordination and 
homogeneity purposes: specific advance periods  for submission and review may be specified in the 
protocol. The timing of publications (in the event several Centers should be participating in the 
Study) may be coordinated, and publication delayed if patentable inventions should be involved (for 
the time required in order to file the relevant patent applications); otherwise, according to the 
MoH’s Decree of May 12, 2006, investigators cannot be precluded from or limited in publishing the 
results of their studies (IECs must verify that no excessive restriction is contained in the protocols 
submitted to their review and approval).  
14 Study time table 
Duration of the study (patient enrolment; treatment duration, follow up, etc.): we expected six 
months for the approval of the study by the Ethics Committee of our Institute, personnel training, 
study protocol registration; twenty-four months of enrollment, interventions and follow-up; six 
months for processing data and for the drafting of the final manuscript. 
 
Final report will be provided: December 2019 
 
Overall study duration from the First Patient First Visit (FPFV): twenty-four months plus five days 
 
The study is designed in accordance with GCP and for this reason we do not foresee any additional 
risks for the patients rather than known antibiotics- and procedures-related risks. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
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