Treatment of Crohn's Complex Perianal Fistulas With Tissue Transplantation by Local Injection of Micro-fragmented Autologous Adipose Tissue. Multicenter Randomized Controlled Prospective Study
Phase: NA
Conditions: Crohn Disease
Sponsor: Lipogems International spa

 
 
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Protocol synopsis 
Protocol title 
Treatment of complex perianal fistulas of Crohn’s disease with tissue 
transplantation by local injection of microfragmented autologous 
adipose tissue. Prospective, multicenter, randomized controlled trial 
Short title 
ATTIC 
Version no. (date) 
2 (11 January 2023) 
Study design 
Interventional, non-pharmacologic, prospective, randomized, 
controlled, double blind, parallel groups, profit, multicentric 
EudraCT 
-- 
Principal 
investigator 
Prof. Silvio Laureti 
Sponsor 
Lipogems International S.p.A. 
Experimental center IRCCS Azienda Ospedaliero-Universitaria di Bologna, Policlinico San 
Orsola-Malpighi, Bologna, Italy 
1. Rationale 
Crohn’s disease is a chronic inflammatory process that can affect the 
entire digestive tract and, in about one-third of cases, leads to the 
development of perianal fistulas or abscesses. Perianal Crohn’s 
disease causes pain, release of purulent or fecal material, and anal 
sphincter destruction, which all have deleterious effects on patients’ 
quality of life. 
Perianal fistulas rarely heal spontaneously or after medical or 
surgical treatment. The role of surgery is to sanitize sepsis, improve 
quality of life, and defer the need for ileostomy or colostomy. The 
availability of anti-TNF antibodies (e.g. infliximab), however, has 
made fistula healing a realistic therapeutic goal. While infliximab is a 
powerful drug, if not used correctly patients may have septic 
complications. Thus, surgical drainage is a fundamental step in the 
treatment algorithm. The preferred procedure is cone-like 
fistulectomy, which creates a surgical wound that heals from the 
inner opening outwards, avoiding abscess development. Combined 
therapy (surgery and a biological drug) is the gold standard for 
perianal Crohn’s disease. Still, it is ineffective in about one-third of 
patients, necessitating the need for new therapies. 
A new approach to promote healing of perianal fistulas is the local 
injection of mesenchymal stem cells (MSC), which have 
immunomodulatory activity. Some studies tested the efficacy of 
allogenic MSCs that had been expanded in vitro under laboratory 
conditions of good manufacturing practice. An alternative is to use 

 
 
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autologous preparations that are rich in MSC-like cells but have not 
been expanded in vitro. These minimally manipulated materials can 
be prepared from adipose tissue using a Lipogems medical device, 
which mechanically fragments lipoaspirates into submillimeter 
particles suitable for transplantation. We previously tested 
microfragmented autologous adipose tissue for efficacy against 
perianal Crohn’s fistulas in a non-controlled study of 15 patients and 
obtained healing in 10 patients at 24 weeks. This new randomized 
controlled trial will test the efficacy and safety of transplantation of 
microfragmented autologous adipose tissue in patients with perianal 
Crohn’s disease refractory to standard therapy. 
2. Study objectives Objective 
Outcome 
 
 
Primary 
To determine the efficacy of 
circumferential, submucosal 
infiltration of 
microfragmented autologous 
adipose tissue around the 
internal fistula opening and 
in the perianal adipose tissue 
around the fistula: combined 
remission 
Combined remission: both (i) absence 
of drainage at gentle finger 
compression of all treated external 
openings that were draining at 
baseline and (ii) absence of pelvic 
MRI evidence of abscesses > 3 mm 
near the treated fistulas, 24 weeks 
after the intervention 
Study hypothesis: combined 
remission rates of 10 % in the control 
group and 40 % in the treatment 
group 
 
 
Secondary To determine the efficacy of 
circumferential, submucosal 
infiltration of 
microfragmented autologous 
adipose tissue around the 
internal fistula opening and 
in the perianal adipose tissue 
around the fistula: clinical 
remission, clinical response, 
failure, time to response, 
safety, and improvement in 
quality of life 
Clinical remission: absence of 
drainage at gentle finger compression 
of all treated external openings that 
were draining at baseline*1 
Clinical response: closure of ≥ 50 % of 
all treated external orifices* 
Time to clinical remission in the 
subgroups of patients with combined 
remission: time from intervention to 
combined remission 
Quality of life: PDAI, IBDQ and WPAI 
scores* 
C-reactive protein* 
Safety: adverse events, serious 
 
1After the study was over, we discovered that the definition of this outcome had been written incorrectly in the 
original protocol in Italian. The correct definition is reported here and has been communicated to 
ClinicalTrials.gov. 

 
 
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adverse events, and suspected 
unexpected serious adverse reactions 
(SUSARs)* 
* Evaluated at 2 weeks (± 2 d), 4 
weeks (± 2 d), 8 weeks (± 2 d), 12 
weeks (± 3 d) and 24 weeks (± 5 d) 
3. Study design 
Interventional, non-pharmacologic, prospective, randomized, 
controlled, double blind, parallel groups, profit, multicentric 
4. Patient selection 
  
Target 
population 
Patients with Crohn’s disease and complex perianal fistulas 
refractory to standard treatment (surgical sanitization and local or 
systemic biological therapy). Complex fistulas will be defined 
according to the American Gastroenterological Association and will 
include transsphincteric fistulas involving over 30% of the external 
anal sphincter, suprasphincteric fistulas, extrasphincteric fistulas, 
horseshoe fistulas, and fistulas with multiple tracts. 
 
Inclusion criteria – Age > 18 years at surgery 
– Crohn’s disease confirmed by histology and diagnostic imaging 
– Complex perianal, fistulizing disease refractory to standard 
treatment (surgical drainage and local or systemic administration 
of anti-TNF for at least one year) 
– Written informed consent 
– Ability to understand the study and participate for the entire 
duration 
 
Exclusion criteria – More than one internal opening or three external openings 
– Ileostomy or colostomy 
– Anovaginal or rectovaginal fistulas 
– Active infection with HIV, HCV, HBV or tuberculosis 
– Abdominal Crohn’s disease that could require general surgery 
during the study 
– Active oncologic or lymphoproliferative disease that could make it 
difficult to obtain at least 60 cc lipoaspirate 
– Clinical conditions that could compromise the outcome of surgery 
or follow-up 
– Pregnant or breastfeeding women 
 
Concomitant 
treatments 
Use of biological drugs, low-availability steroids or mesalamine in the 
4 weeks before enrollment and azathioprine or mercaptopurine in 

 
 
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the 6 months before enrollment will be allowed in a stable dosage 
for the entire study. Concomitant treatments will be reported. 
 
Study groups 
Treatment group (A): surgical sanitization with cone-like fistulectomy 
and infiltration of microfragmented autologous adipose tissue at the 
internal orifice (40 patients) 
Control group (B): surgical sanitization with cone-like fistulectomy, 
infiltration of physiological saline, and placement of a suture at the 
internal orifice (40 patients) 
5. Study 
procedures 
Patients will be enrolled after protocol approval by the ethics 
committee of each center. Enrollment will be competitive, i.e. each 
center will enroll at least 8 patients until the total number (80) is 
reached. 
 
Randomization 
Randomization will be centralized (i.e. not done at any study center). 
A computer-generated randomization list will be made, and patients 
will be assigned to the two groups in a 1:1 ratio. At each center, an 
investigator will identify eligible patients according to the inclusion 
and exclusion criteria and will present to them the scope of the 
study, the protocol and the procedures. After a patient provides 
consent, another investigator (not involved in enrollment, surgery or 
postoperative assessment) will contact the randomization center to 
obtain this patient’s assignment (i.e. the first place available on the 
randomization list). 
 
Data collection 
Each patient will be assigned an alphanumeric identification code 
generated by Lipogems International S.p.A. using the blockrand 
package v1.5 in R Statistical Software v4.2.0. The code will be 
compliant with rules on the privacy of subjects of clinical trials. All 
the data from the study will be registered by an investigator 
uninvolved in study procedures (including randomization and 
surgery). Radiological data will be saved in digital form on each 
institute’s computer system. The other data will initially be recorded 
on case report forms (paper). For each patient, the study center will 
keep a folder with all the completed forms, including the signed 
informed consent form and privacy agreement. Information such as 
the identification code, personal data, clinical reports, informed 
consent, and privacy agreement will be kept in the patient’s file and 
made available only in the case of audit or inspection. During the 
study, data from the paper forms will be progressively inserted into a 
database (regularly backed up). 
 
Enrollment and 
randomization 
Potentially eligible patients will undergo a screening evaluation by an 
investigator, uninvolved in the surgical procedures, according to the 
inclusion and exclusion criteria. Eligible patients will be told of the 
study’s goals and procedures and asked to provide informed consent 

 
 
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and authorize the use of their data. 
Once a patient has been enrolled, an investigator will collect 
baseline clinical data, and the patient will undergo basic blood 
testing, measurement of serum C-reactive protein, and pelvic MRI. 
After enrollment, participants were randomly assigned in a 1:1 ratio 
to either the intervention group or control group through an 
Interactive Web Response System (IWRS). 
Patients in the treatment and control groups will undergo the same 
pre- and postoperative procedures and will be operated on by a 
surgeon uninvolved in their assessments.   
The study will be double blind, as follows: randomization and 
statistical analyses will be done by an investigator not involved in 
assessments or surgery. Clinical assessments will be done by 
investigators uninvolved in surgery and blinded to group assignment. 
The radiologist who evaluates the MRI scans will be blind to patients’ 
assignment. The surgeons will not be allowed to share information 
on the treatments and will not participate in clinical assessments. 
In case of emergency, the principal investigator can decide to 
unblind a patient, who will then be excluded from the study. In these 
cases, the study sponsor will be notified as soon as possible. The 
date and reason for unblinding will be reported in the patient’s 
clinical file. 
 
Preoperative 
assessment (T0) 
On the day of surgery, the following data will be collected or 
measured: 
– Any use of biological drugs, low-availability steroids or 
mesalamine in the 4 weeks before enrollment and azathioprine or 
6-mercaptopurine in the 6 months before enrollment 
– Serum C-reactive protein level 
– Pelvic MRI scan 
– PDAI score 
– IBDQ score 
– WPAI scores 
 
Preparation of 
microfragmente
d adipose tissue 
– Lipogems 
system 
Lipogems is a class II A sterile medical device for the fragmentation 
of adipose tissue (with CE mark subsequent to assessment by 
notified body 0476). The US Food and Drug Administration approved 
the use of Lipogems devices for liposuction, microfragmentation 
without additional manipulation, and lipofilling in plastic surgery, 
reconstructive surgery and other surgical applications (22 December 
2014; class II device under 21 CFR 878.5040, Section 510(k) 
premarket notification). 
The single-use Lipogems device reduces lipoaspirates to 

 
 
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submillimeter clusters enriched in pericytes, with intact vascular 
structure, free of oil or blood. Because microfragmentation is 
achieved by mechanical means, without centrifugation or additives 
(e.g. enzymes), the resulting tissue product is considered minimally 
manipulated. With the Lipogems 60 cc kit, approximately 60 mL 
lipoaspirate is obtained in less than 20 min. 
 
Surgical 
procedures 
Liposuction and tissue transplantation will be done in a single 
surgical procedure. 
 
 
Treatment 
group (A) 
Phase 1. 
Liposuction 
Liposuction will be done under general or spinal 
anesthesia. A plastic surgeon will choose a donor 
site (lateral or lower abdomen or thigh) in which 
100 cc modified Klein solution (500 cc physiological 
saline, 1 cc epinephrine 1/1000 IU) will be injected 
using a 17G cannula on a 60 cc Luer-lock syringe. 
Adipose tissue will be aspirated (50-100 cc) using a 
13G cannula on a 20 mL VacLok syringes. 
 
 
 
Phase 2. Tissue 
processing 
The lipoaspirate will be immediately processed in a 
Lipogems device. The microfragmented tissue will 
be collected in a 60 cc syringe and allowed to settle 
for the removal of excess physiological saline. The 
remaining product will be transferred to 10 cc 
syringes. 
 
 
 
Phase 3. 
Surgery and 
tissue 
transplantation 
Each patient will undergo a proctological exam 
under anesthesia for the identification of fistula 
tracts and any abscesses. Purulent material will be 
drained, and the tracts will be curetted. Necrotic or 
inflamed tissue will be removed by cone-like 
fistulectomy. Then, 15 cc microfragmented 
autologous adipose tissue will be injected using a 
21G needle into the submucosa around the 
internal fistula opening and along the remaining 
fistula tract. 
 
 
Control group 
(B) 
Phase 1. Sham 
liposuction 
A 6-0 absorbable suture will be placed on the skin 
of the thigh to simulate liposuction. 
 
 
 
Phase 2. 
Surgery 
Each patient will undergo a proctological exam 
under anesthesia for the identification of fistula 
tracts and any abscesses. Purulent material will be 
drained, and the tracts will be curetted. Necrotic or 
inflamed tissue will be removed by cone-like 
fistulectomy. Then, 10 cc physiological saline will be 
injected using a 21G needle around the internal 
fistula opening and along the remaining fistula 

 
 
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tract. The internal fistula orifice will be closed using 
a 2-0 absorbable polyglactin suture. 
 
Postoperative assessments 
 
 
2 weeks (T2) 
C-reactive protein, clinical exam (assessment of drainage at gentile 
finger compression), PDAI, IBDQ and WPAI scoring; registration of 
any adverse events 
 
 
4 weeks (T4) 
C-reactive protein, clinical exam (assessment of drainage at gentile 
finger compression), PDAI, IBDQ and WPAI scoring; registration of 
any adverse events 
 
 
8 weeks (T8) 
C-reactive protein, clinical exam (assessment of drainage at gentile 
finger compression), PDAI, IBDQ and WPAI scoring; registration of 
any adverse events 
 
 
12 weeks 
(T12) 
C-reactive protein, clinical exam (assessment of drainage at gentile 
finger compression), PDAI, IBDQ and WPAI scoring; registration of 
any adverse events 
 
 
24 weeks 
(T24) 
C-reactive protein, clinical exam (assessment of drainage at gentile 
finger compression), PDAI, IBDQ and WPAI scoring; registration of 
any adverse events. Pelvic MRI 
 
 
 
Control patients who do not achieve combined remission will be 
offered treatment with microfragmented autologous adipose tissue. 
 
Pelvic MRI 
MRI scanning will be done on a 1.5-3T MR scanner (Signa HDxt, GE 
Medical Systems) with an 8-channel phased array body coil (without 
endoanal coil). Axial T1- and T2-weighted fast spin-echo sequences 
with 5 mm slices will be used to assess sphincter anatomy and fistula 
extent. Short tau inversion recovery sequences, in axial, sagittal and 
coronal planes, and 3 mm slices will be used to distinguish fistulas 
from adjacent tissue and to assess soft tissue edema and 
inflammation in the perianal and perirectal areas. 
Radiological fistula healing will be defined as a lack of T2 
hyperintensity and lack of abscesses > 3 mm. 
 
Safety 
At every follow-up visit, an investigator will record the occurrence of 
adverse events, serious adverse events and SUSARs between the 
date when the patient gave written informed consent and the end of 
the study. 
 
 
Adverse 
events 
An adverse event is any sign, symptom or disease, unfavorable and 
unwanted, that has a temporal association with use of the 
experimental product, even in the absence of a causal relation. The 
clinical conditions and pathologies that patients had before the 

 
 
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beginning of the study are considered adverse events only if they 
worsen during the study. 
Every adverse event that occurs during the study will be registered. 
For every adverse event, the investigators will promptly provide 
appropriate standard care. If necessary, the patient will be 
withdrawn from the study. 
 
 
Serious 
adverse 
events 
A serious adverse event is an undesired sign, symptom or clinical 
condition that: 
1. Is fatal, 
2. Is life threatening, 
3. Requires or prolongs hospitalization, 
4. Results in disability or serious or prolonged incapacity, 
5. Causes a congenital anomaly or birth defect. 
Hospitalization for a preexisting condition will be considered a 
serious adverse event, unless it had been planned prior to the start 
of the study. 
 
 
Classification 
of causality 
The causal relation between an adverse event and the treatments 
will be classified as: 
– Unknown 
– Unrelated 
– Impossible 
– Possible 
– Probable 
– Certain 
The investigators will rank the severity of each adverse event to one 
of the following categories: 
– Mild: the symptom is evident but tolerable. 
– Moderate: it negatively affects normal activity. 
– Severe: a severe effect, incapacity to work, need to withdraw 
from the study. 
 
 
Reporting of 
adverse 
events 
All adverse events will be documented in the case report forms. The 
investigators are responsible for correctly registering the events and 
for notifying healthcare authorities of all undesired events during 
the study. For each adverse event, the following will be recorded: 
– Identification code and initials of the patient 
– Randomization code 

 
 
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– Description of the symptom or event 
– Classification of the event and its severity 
– Dates of onset and outcome 
– Frequency (once, occasionally, frequently, continually) 
– Treatment (none, pharmacological, non-pharmacologic, dose 
reduction, temporary suspension, hospitalization) 
– Causal relation with the study product 
– Outcome (resolved, not yet resolved, sequelae of disability or 
incapacity, death, unknown) 
Any serious adverse event or any adverse event that can affect the 
safety of the patient or affect the conduct of the study, whether 
correlated or not to the treatments, will be notified immediately 
(within 24 h) to Lipogems International Spa. Lipogems will forward 
the information to healthcare authorities as required by law. 
 
 
Withdrawal 
Patients can withdraw from the study for the following reasons: 
a. personal decision (even without motivation) 
b. as decided by an investigator 
c. in case of an adverse event. 
The reason for withdrawal will be reported in the case report form. 
6. Sample size and 
statistical 
analyses 
Sample size was calculated assuming the use of Fisher’s exact test to 
assess the difference between groups in combined remission at T24. 
Assuming combined remission rates of 10 % in the control group and 
40 % in the intervention group, a two-tailed test at a significance 
level of 0.05 would require a sample of 72 patients (36 per group) to 
achieve 80 % power. Assuming a drop-out rate of 10 %, the sample 
size was set to 80 (40 per group).   
Qualitative variables will be reported as counts and range. 
Quantitative variables will be reported as mean, standard deviation 
and range. Differences between groups in categorical variables, 
including the primary outcome, will be tested for significance using 
Fisher’s exact test or chi-squared test. The distribution of continuous 
variables will be assessed using the Shapiro-Wilk test. Normally 
distributed variables will be assessed for significance using Student’s 
t test; otherwise, the Mann-Whitney test will be used. Longitudinal 
data will be analyzed using two-way ANOVA for repeated measures 
and multiple regression. 
7. Study duration 
Total, 24 months: enrollment, 12 months; follow-up, 6 months; data 
analysis, 3 months; writing, 3 months 

 
 
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8. Publication 
The results of the study will be written up in a manuscript, which will 
be sent to a peer-reviewed medical journal for publication. 
9. Ethical and 
regulatory issues 
The study will adhere to the ethical principles of the Declaration of 
Helsinki (64th WMA General Assembly 2013), to ICH-GCP guidelines 
and ISO 14155 - Clinical Investigation of medical devices for human 
subjects, and to Italian laws on clinical trials. 
 
Ethical approval The study protocol will be approved by each center’s ethics 
committee before the study begins. 
 
Protocol changes Any change to this protocol will be presented by the sponsor to the 
ethics committees for approval. 
 
Informed 
consent 
Written informed consent will be obtained from each patient before 
entering the study. The informed consent form will be approved by 
the ethics committees.   
 
Privacy 
The privacy of patients in the study will be protected in accordance 
with ICH/GCP and local legislation. Patients’ identities will be known 
only to the investigators and the Clinical Monitor (in accordance with 
UE regulation 2017/745). 
 
Data access 
The investigators will allow study monitoring by the sponsor. They 
will also allow healthcare authorities and ethics committee members 
direct access to study documentation during audits.   
 
Data ownership 
Study data will be the property of Lipogems International S.p.A. 
 
Funding 
The study is funded by Lipogems International S.p.A. without costs 
to the Italian healthcare system. 
 
Ethical 
guidelines 
The study will adhere to the Declaration of Helsinki (Fortaleza 2013) 
and to the principles of good clinical practice as expressed in Italian 
law (D.leg. 24 giugno 2003, n. 211 alla G.U. n. 184 del 09/08/2003). 
 
Insurance 
Lipogems International S.p.A will provide insurance for all study 
subjects. 
10 Premature study 
termination 
The investigators have the right to stop the study at any moment for 
medical or administrative considerations. Reasons for the 
interruption will be documented, and the ethics committee of the 
Italian Ministry of Health will be informed.