Study of TB Lesions Obtained in Surgery: in Search of Best Biomarkers Correlating With TB Pathology, Clinical Features, MDR Cases and Prognostic
Conditions: Tuberculosis, Thoracic Surgery, Tuberculosis, Pulmonary, Tuberculosis, Multidrug-Resistant
Sponsor: Fundació Institut Germans Trias i Pujol

SH-TBL (Study of Human TB Lesions) 
 
 
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PROTOCOL & INFORMED CONSENT 
 
TITLE: “Study of TB lesions obtained in surgery: in search of best biomarkers correlating 
with TB pathology, clinical features, MDR cases and prognosis”  
Acronym: SH-TBL (Study of Human TB Lesions). 
NCT number: NCT02715271 
Version: v3 
Date: 20th April 2016 
 
 
 
 
 
 

SH-TBL (Study of Human TB Lesions) 
 
 
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TABLE OF CONTENTS 
 
1. 
STATE OF ART AND RATIONALE ........................................................................ 3 
2. 
AIM .......................................................................................................................... 5 
3. 
HYPOTHESIS .......................................................................................................... 5 
4. 
SPECIFIC OBJECTIVES .......................................................................................... 5 
5. 
M&M ........................................................................................................................ 5 
1. 
Study design........................................................................................................... 5 
2. 
Study population and eligibility ............................................................................. 6 
6. 
ETHICS: ...................................................................................................................... 7 
7. 
PARAMETERS AND MEASUREMENTS ............................................................................ 8 
8. 
RELEVANT BIBLIOGRAPHY .............................................................................. 13 
9. 
ANNEX 1. PATIENT’S TISSUE SAMPLES TO BE COLLECTED WITHIN THE 
PROSPECTIVE SUBSTUDY ......................................................................................... 14 
10. ANNEX 2. PROTOCOL TO PROCESS PATIENT’S BLOOD AND URINE 
SAMPLES ...................................................................................................................... 16 
11. ANNEX 3. INFORMED CONSENT FORM ............................................................ 18 

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1. 
STATE OF ART AND RATIONALE 
 
Nowadays, Tuberculosis (TB) is still an international public health problem, in spite of 
existing an effective treatment. Every year, 9 million people get sick and 1 million of them die. 
Recently, one of the major problems for health systems (in terms of both medical aspect and 
budget) of all countries are those cases due to Multi-Resistant to Drugs strains (MDR-TB), 
which represented up to half a million last year. The average length of hospital stay and 
duration of treatment are increasing due to MDR-TB cases, with an associated increasing 
burden on National TB Control Programmes. Moreover, these cases have poor treatment 
outcomes associated and  
In Spain, tuberculosis is not a big problem, and yet during 2013 we had 5539 notified cases 
and an incidence of 13 cases per 100.000 people is estimated.  However, due to our national 
history of fight against the disease, we have one of the most skilled scientific network at 
international level in TB research field, the Experimental Tuberculosis Unit (Unitat de 
Tuberculosi Experimental, UTE) being a renown reference in Europe.  
In this context, new approaches to ameliorate the response to TB treatment are a priority; 
especially those aiming to reduce morbidity, mortality and treatment duration, all taking into 
account the MDR-TB cases. Moreover, these new options are required to be worldwide 
available, to be used immediately and for any patient, implying them needed to be cheap.  
During the last 20 years, a lot of progress has been made in the design and development of 
new drug and vaccine candidates, but this is a hard, time and cost-consuming activity with low 
or none output at short-term and doubtful output at long-term either (only one out of 10.000 
new compounds will arrive to the market and only after 15 years of a rigorous pre-clinical and 
clinical development). 
Meanwhile, physicians deal with clinical TB management based on old premises, most of 
them strictly clinical and empiric approaches, and the TB drug pipeline stills remains sparse.  
Nowadays in TB, there is any protector correlator, diagnose or prognosis biomarker known. 
International scientific community dedicated to TB field is focused to find it, and for this reason 
many resources are dedicated.  During the last years, different genetic profiles has been found 
in peripheral blood related with TB disease, pointing that this genetic signatures involving more 
than one parameter could establish the key of futures therapeutic approaches. However, the 
same authors of these projects specify that it is necessary to check, compare and validate these 

SH-TBL (Study of Human TB Lesions) 
 
 
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results with the genetic profiles found in the TB lesions. In our opinion these data would be not 
necessarily equal but complementary which would explain the reason of the disease generation 
in some infected patients and not in others, the different prognostic for distinct patients, the 
role of inflammation in the disease and infection progression and the response to treatments 
and vaccines. Moreover, they will provide robustness and parameter restriction to the profiles 
found in blood. For a further commercial exploitation, it is better a profile with 3 common 
characters in blood and lesion than 10 parameters in blood. However, the reason why the 
researches use peripheral blood instead of samples of lesions in these studies is only due to the 
availability of samples. The majority of top countries in TB research works in collaboration 
with African countries which have high incidence of TB and also have a high rate of coinfection 
with HIV masking and difficult works like ours, in which the histopathology is very important.  
On the other hand, after an age when surgery was a common treatment in Europe and America 
this technique was no longer necessary after the emergence of anti-TB drugs in the 50’s and 
they fell into disuse. Nowadays, almost any neither European nor American physician has the 
experience in this type of practice. However the irruption of drug multiresistant strains has 
changed the landscape. Asia, and especially Eastern countries (with exURSS countries in lead) 
have a high number of MDR cases and still very high rates of TB. Georgia is one of these 
countries. It has an incidence 10 times higher than Spain, a low rate of HIV-coinfection and 
they are still obligated to operate their TB patients, because more than a half are MDR and fails 
to the standard treatment.  
 
NCTLD is the reference center in Georgia dedicated entirely to TB and with highly 
expert surgeons in clinics and surgery of TB lesions and yet, without any or few experience on 
research. Dr. Vilaplana and her team are experts of basic research of the disease; they have 
been involved in all aspects of TB research like developing and evaluating vaccines and new 
therapies; and also immunology in clinical trials and pathology of lesions in experimental 
animal models.  This created association is based in professional loyalty between these two 
teams ensures the creation of a unique international collection, that after being exploited by the 
two teams will generate an information that can be used in collaboration with other leading 
international groups to ensure a translation and exploitation of obtained results.   
 
 
 

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2. 
AIM 
To perform a genetic, protein and microbiological analysis on a unique collection of biological 
samples of TB patients (sensitive and MDR-cases). 
3. 
HYPOTHESIS 
The correlation of the morphologic, microbiological, genetic and histopathological 
characteristics of TB lesions obtained in therapeutical surgery with the clinical forms and 
features of the patients will provide essential information 
a) on the role of the host in the mechanisms associated to the generation and evolution 
of active TB and 
b) about future diagnostic and/or prognostic biomarkers of TB disease. 
All this information could be used for patients stratification and/or to design new therapeutic 
strategies. 
4. 
SPECIFIC OBJECTIVES  
To achieve the general aim of the project, we plan to achieve 2 specific objectives: 
3.1. Description study of the morpho-histopathological characteristics of TB lesions and 
its correlation with the clínico-pathological features of TB patients who underwent 
therapeutical surgery during 2014-2016. The aim of this objective is to conduct a 
descriptive study to know about the characteristics of TB patients which need therapeutical 
surgery, and to evaluate its impacto on patients’ morbility and mortality. 
3.2. Prospective study of the TB lesions obtained in therapeutical chest surgery. The aim 
of this objective is to obtain a genic-proteomic profile of in tissues which correlate to the 
histopathology of the granulomas and/or the clinic-pathological features of TB patients. 
 
5. 
M&M 
1. 
Study design  
The project is part of a collaboration agreement between the UTE and the NCTLD, and already 
obtained the Ethics Committee approval (IRB00007705 NCTLD Georgia #1, IORG0006411). 
The protocol approved includes lesions’ collection during therapeutical surgery, as well as the 
collection of demographic data, clinical features and analytical data of the patients included in 
the study, data on their evolution pre and post-surgery, complications and survival. The data 

SH-TBL (Study of Human TB Lesions) 
 
 
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collection is done in the Georgian center supervised by the NCTLD team and coordinated by 
the IP, Dr Vilaplana, and the samples and data will be shipped to the UTE for their analysis in 
collaboration with the Georgian team. 
The project is an observational study including 2 substudies: 
1. Description study of the morpho-histopathological characteristics of TB lesions and its 
correlation with the clínico-pathological features of TB” (Objective 1). Data on patients 
(all cases, both DS and MDR/XDR-TB) therapeutically operated an their lesions during 
an initial period 2014-2016 will be retrospectively recorded in a spreadsheet file from 
the surgical notebooks in an anonimized way.  
2. For the “Prospective study of the TB lesions obtained in therapeutical chest surgery” 
(Objective 2): Clinical and epidemiological data and tissue, blood and urine samples 
will be anonymously collected from patients submitted to therapeutical surgery.  
 
2. 
Study population and eligibility 
 
Study Population 
Patients undergoing therapeutic surgery for their Pulmonary Tuberculosis at NCTLD in Tbilisi, 
Georgia.  
• For Description study of the morpho-histopathological characteristics of TB lesions and its 
correlation with the clínico-pathological features of TB” (Objective 1): Data on patients (all 
cases, both DS and MDR/XDR-TB) therapeutically operated an their lesions during an 
initial period 2014-2016 will be retrospectively recorded in a spreadsheet file from the 
surgical notebooks in an anonimized way.  
• For the “Prospective study of the TB lesions obtained in therapeutical chest surgery” 
(Objective 2):  
Eligibility: 
Inclusion Criteria: 
• Patients of all ages and sexes undergoing therapeutical surgery for their Pulmonary 
Tuberculosis at NCTLD in Tbilisi, Georgia indicated as per clinical routine 
Exclusion Criteria: 
• Non consenting to donate samples and/or data for the study 

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6. 
Ethics:  
The project protocol and associated documents was reviewed and approved by both the ethics 
committee of the NCTLD (IRB00007705 NCTLD Georgia #1, IORG0006411) and the 
Germans Trias i Pujol Research Institute (IGTP) ethics committee (EC: PI-16-171).  
 
In the retrospective study, the patients included underwent surgery during the past 2-5 years 
time, and they already filled in the Informed Consent for the surgery. In this study, only the 
epidemiological and clinical data of the patients will be collected in an anonymized way 
together with the histopathological data from the lesions obtained. No extra samples or data are 
needed for this substudy, thus no extra Informed Consent will be needed. 
 
For the prospective substudy, an informed consent (attached as Annex 3) will be obtained for 
the collection of biological material and data from all study participants before they were 
enrolled.  
Each patient will be given a sequential number coded as SH-TBL XXX (XXX meaning the 
correspondent number of patient).  
For each patient, a non anonymized form created ad-hoc for this project will correlate him/her 
with his/her projects’ code. This form will be to be kept together with the Medical Record of 
each patient as the Hospital Records copy. A spreadsheet file will be filled-in with the following 
data of each patient included: name, the Hospital Record Number and the correspondent 
Project’s code (SH-TBL XXX). Another form created ad-hoc will be filled-in for each patient 
included. In order to comply with the patients’ rights of confidentiality, this form will be 
anonymized, without the name and surnames of the patient and where only the project’s code 
will be recorded, together with all the clinical data associated to the specific case. A spreadsheet 
file will be filled-in with the following data of each patient included: Project’s code (SH-TBL 
XXX), clinical characteristics, characteristics of the lesion to be removed, and monitoring data. 
In order to comply with the ethics requirements of samples’ collections, both spreadsheets will 
be kept as 2 different documents and stored in 2 different computers.  
All studies will be conducted blindly.  
 

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7. 
Parameters and measurements  
There are 2 cohorts/study groups, each corresponding to one of the substudies, with different 
outcome measures and parameters: 
4.2.1. FOR THE “DESCRIPTION STUDY OF THE MORPHO-HISTOPATHOLOGICAL 
CHARACTERISTICS OF TB LESIONS AND ITS CORRELATION WITH THE CLINICO-
PATHOLOGICAL FEATURES OF TB” (OBJECTIVE 1): 
Retrospective cohort.  
Primary Outcome Measures: 
• Description of Clinical and Epidemiological Data of the TB patients enrolled. Time Frame: 
at baseline. Descriptive analysis; data recorded in a spreadsheet created ad-hoc. 
•Description of the Histopathological characteristics of the TB lesions of TB patients enrolled. 
Time Frame: at baseline. Descriptive analysis; data recorded in a spreadsheet created ad-hoc 
 
From surgical notebooks of TB patients submitted to therapeutical surgery the following data 
will be recorded (anonymously) in a spreadsheet file: 
Patients’ data (epidemiological and clinical data): 
• 
Sex 
• 
Age 
• 
Smoking and alcohol intake habits 
• 
Comorbilities: diabetes, immunosuppression, HIV-coinfection, renal disease, COPD, 
others 
• 
TB form: pulmonary, extrapulmonary, pulmonary+extrapulmonary, disseminated 
• 
Type of TB according to drug sensitivity (WHO definitions) 
• 
History of previous treatment 
• 
Anatomical site of TB  
• 
Date of TB diagnosis 
• 
Date of Treatment initiation 
• 
number of lesions found in radiology assays 

SH-TBL (Study of Human TB Lesions) 
 
 
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• 
Bacteriology cured? (WHO definitions) 
• 
Time of negativization of culture (in months) 
• 
Indication for surgery, main reason 
• 
Number of lesions operated  
• 
Type of surgery done 
• 
Date of surgery 
• 
Post-surgery complications 
• 
Final outcome (WHO definitions) 
• 
Date of death if appropriate 
For each lesion operated: 
• 
Type of lesion 
• 
Presence of fibrosis/cirrhosis 
• 
Size (mm) 
• 
picture 
 
Work plan and estimated calendar: 
Data will be recorded during the first year (ideally during the first 6 months), and the analysis 
and reporting done during the second year of the project.  
Statistics 
Data obtained will be described (descriptive analysis) but also statistically analyzed by 
multivariant analysis with the aim to find correlations between the different variables included 
and to identify the factors which might have an impact on the final outcome.  
 
4.2. FOR THE “PROSPECTIVE STUDY OF THE TB LESIONS OBTAINED IN 
THERAPEUTICAL CHEST SURGERY” (OBJECTIVE 2): 
Prospective cohort: data and samples will be prospectively included from TB patients 
undergoing therapeutical surgery (DS- and MDR/XDR-TB). Data will be recorded in an 

SH-TBL (Study of Human TB Lesions) 
 
 
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electronic form which has been already created ad-hoc (ElectronicDataCapture, Clincapture, 
Clinovo, http://www.clinovo.com/products/clincapture). In this form, we have included: 
clinical and epidemiological data of patients; data on collection and process of biological 
samples; Health Quality of Life (HQoL) questionnaires data.  
Primary Outcome Measures: 
• Description of Clinical and Epidemiological Data of the TB patients enrolled. Time Frame: 
at baseline. Descriptive analysis; data recorded in a spreadsheet created ad-hoc. 
•Description of the Histopathological characteristics of the TB lesions of TB patients enrolled. 
Time Frame: at baseline. Descriptive analysis; data recorded in a spreadsheet created ad-hoc 
•Obtention of a genic-proteomic profile of TB lesions tissues which correlate to the 
histopathology of the granulomas and/or the clinic-pathological features of TB patients. Time 
Frame: at baseline. Descriptive analysis, correlation with histopathological characteristics of 
TB lesions and with clinical and epidemiological data of TB patients 
Secondary Outcome Measures 
•Change in Immunological responses. Time Frame: at baseline; and at the moment of 
discharge, an average of at day 15 post-enrollment. Immunological responses will be measured 
2.1.3. Identification of biomarkers in blood at protein level in blood and urine; validation in 
tissue and blood of genic biomarkers secreted (ELISA, qPCR, immunohistochemistry). 
•Change in Health Quality of Life Measurements. Time Frame: at baseline (before surgery) 
and through study completion, an average of 1 year. Measured with Health Quality of Life 
Questionnaires.  
 
Data will be recorded in an electronic form which has been already created ad-hoc 
(ElectronicDataCapture, Clincapture, Clinovo, http://www.clinovo.com/products/clincapture). 
In this form, the following data will be collected:  
• Epidemiological and clinical data will be collected from all patients at baseline (prior 
surgery).  
• Health Quality of Life (HQoL) questionnaires: SGUL’s Respiratory Questionnaire, 
Kessler-10 and BCN-Q will be used to evaluate the social and psychological factors 
associated to the patients included in the study. The patients will be reevaluated at the 
moment of discharge and during the follow-up if they are scheduled as per routine.  

SH-TBL (Study of Human TB Lesions) 
 
 
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The following samples will be prospectively collected from patients undergoing therapeutic 
surgery and enrolled in the prospective substudy: 
• Tissue samples from the TB lesions 
• Blood samples for plasma obtention and whole blood for RNA studies 
• urine 
During surgery, the following tissue samples from the following areas of the removed lesions 
will be collected:  
1. cavity center 
2. cavity internal wall 
3. cavity external wall 
4. nearby visually healthy tissue around the cavity 
If a TB lesion (nodulus) is detected, samples of it will also be collected.  
These samples will be used for 1) the genetic and proteomic studies; 2) histopathology; 3) 
microbiologic studies. For histopathological analysis a single piece including the 4 areas of the 
granulomas will be collected. For genetic and proteomic and microbiological studies, 
individual samples will be collected from each area.  
If there will be no possibility to collect 3 samples of each zone for these 3 purposes, then the 
priority will be 1st: samples for genetic and proteomic studies; 2nd: samples for microbiology 
and 3rd: samples for histopathology. 
A protocol for collecting and processing patient’s tissue samples has been attached as Annex 
1. 
Blood sample for plasma (in a 10mL EDTA tube) and whole blood for RNA studies (in a 2,5mL 
PaxGene RNA tube) plus a urine sample will be collected prior to surgery (baseline) and at the 
moment of clinical discharge. Extra-samples will be collected in next post-surgery follow-up 
visit if possible (as indicated by clinical management routine). A protocol for processing blood 
and urine samples has been attached as Annex 2. 
 
Samples collected will be stored to perform the planned studies and the remaining samples will 
be kept for further studies if considered appropriate by the PI and according to extra-funding 

SH-TBL (Study of Human TB Lesions) 
 
 
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possibilities. All samples collected will be recorded and labeled according to a specific code 
for the present project (SH-TBL plus the number correspondent to the patient’s number).  
 

SH-TBL (Study of Human TB Lesions) 
 
 
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8. 
RELEVANT BIBLIOGRAPHY 
1.  Who. Global tuberculosis report 2015 (WHO/HTM/TB/2015). 2014; Available from: 
http://apps.who.int/iris/bitstream/10665/191102/1/9789241565059_eng.pdf?ua=1 
2.  World Health Organization. WHO Tuberculosis country profiles [Internet]. 2015 [cited 
2015 Sep 30]. Available from: http://www.who.int/tb/country/data/profiles/en/ 
3.  Zumla A, Maeurer M. Rational development of adjunct immune-based therapies for drug-
resistant tuberculosis: Hypotheses and experimental designs. J Infect Dis. 
2012;205(Suppl 2):335–9.  
4.  Schito M, Migliori GB, Fletcher HA, Mcnerney R, Centis R, D ’ambrosio L, et al. 
Perspectives on Advances in Tuberculosis Diagnostics, Drugs, and Vaccines. Clin Infect 
Dis. 2015 Oct 15;61Suppl 3:S102-18. doi: 10.1093/cid/civ609. 
5.  Berry MPR, Graham CM, Mcnab FW, Xu Z, Bloch SAA, Oni T, et al. An Interferon-
Inducible Neutrophil-Driven Blood Transcriptional Signature in Human Tuberculosis.  
6.  Kaufmann SHE, Lange C, Rao M, Balaji KN, Lotze M, Schito M, et al. Progress in 
tuberculosis vaccine development and host-directed therapies-a state of the art review. 
Lancet Respir Med [Internet]. Elsevier Ltd; 2014;2(14):301–20. Available from: 
http://dx.doi.org/10.1016/S2213-2600(14)70033-5 
7.  Anderson ST, Kaforou M, Brent AJ, Wright VJ, Banwell CM, Chagaluka G, et al. Diagnosis 
of Childhood Tuberculosis and Host RNA Expression in Africa. N Engl J Med. 
2014;370:1712–23.  
8.  Maertzdorf J, Ota M, Repsilber D, Mollenkopf HJ, Weiner J, Hill PC, et al. Functional 
correlations of pathogenesis-driven gene expression signatures in tuberculosis. PLoS 
One. 2011;6(10):e26938. doi: 10.1371/journal.pone.0026938. Epub 2011 Oct 28. 
9.  Zumla A, Abubakar I, Raviglione M, Hoelscher M, Ditiu L, McHugh TD, et al. Drug-
Resistant tuberculosis-current dilemmas, unanswered questions, challenges, and priority 
needs. J Infect Dis. 2012;205(Suppl 2).  
10.  
Dara M, Sotgiu G, Zaleskis R, Migliori GB. Untreatable tuberculosis: is surgery the 
answer? Eur Respir J. 2015 Mar;45(3):577-82. doi: 10.1183/09031936.00229514. 
11.     Madansein R, Parida S, Padayatchi N, Singh N, Master I, Naidu K, Zumla A, Maeurer 
M. Surgical treatment of complications of pulmonary tuberculosis, including drug-
resistant 
tuberculosis. 
Int 
J 
Infect 
Dis. 2015 
Mar;32:61-7. 
doi: 
10.1016/j.ijid.2015.01.019. 
 

SH-TBL (Study of Human TB Lesions) 
 
 
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9. 
ANNEX 1. PATIENT’S TISSUE SAMPLES TO BE COLLECTED 
WITHIN THE PROSPECTIVE SUBSTUDY 
For each patient, and lesion removed, samples of the removed lesions should be obtained and 
labeled with the patient’s code, the data of collection, the zone of obtention and the purpose 
according to the following premises:  
 
ZONE OF OBTENTION 
CODE 
cavity center 
C 
cavity internal wall 
I 
cavity internal wall 
E 
visually 
healthy 
tissue 
around the cavity 
H 
nodulus 
N 
 
 
 
 
 
 
 
PURPOSE 
CODE 
NUMBER OF SAMPLES 
TO BE COLLECTED  
Samples for gene expression and protein 
profile (Host expression) 
HExp 
1 per each zone 
Microbiology 
Mic 
1 per each zone 
histopathology 
Path 
1 single piece with all zones 

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Samples for gene expression and protein profile  
1. Do not freeze tissues before immersion in RNAlater Solution. 
2. Cut large tissue samples to ≤0.5 cm in any single dimension. 
3. Place the fresh tissue in 5–10 volumes of RNAlater Solution in a properly labeled criotube. 
4. Do not freeze samples in RNAlater Solution immediately; store at 4°C overnight (to allow 
the solution to thoroughly penetrate the tissue), remove supernatant, then move to –80°C 
for long-term storage. 
Samples for histopathology 
Samples will be collected as one single piece including the different parts of granuloma, and 
should be fixed processed in 10% buffered formalin and sent to the UTE for it’s histopathology 
analysis.  
Samples for microbiology: 
The principal aim is to obtain M.tuberculosis isolates for further characterization (whole 
sequencing).  
 
If there is limited quantity of biological material, sampling for genetic and proteomic studies 
shall be prioritized and the volume of samples collected for microbiology and histopathology 
could be reduced (always expressing it as incidence in the samples collection form).  
 
 

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10. 
ANNEX 2. PROTOCOL TO PROCESS PATIENT’S BLOOD AND 
URINE SAMPLES 
For the samples’ quality, keep on ice or at the fridge (<4ºC) until they are processed, which 
should be done in less than 1 hour after their extraction. Labels: each microtube should be 
labeled with the patient’s code and the date it was obtained. 
For serum collection:  
1. 
Extract 10 ml of blood in tubes with EDTA (anticoagulant). It should be attempted to 
perform the blood extraction at the same time as a routine blood test. 
2. 
Invert the tube 10 times to mix the anticoagulant with blood (do not shake). 
3. 
Immediately after, centrifuge at 2000g and 4°C for 10 minutes (if it’s not possible to do 
it immediately after, maintain the sample at 4°C in ice or otherwise in 4°C water). 
4. 
Isolate the plasma (approx. 3 ml) right after the centrifugation (avoid over half an hour 
elapse) and aliquot in 2-3 properly labeled microtubes (patient’s code number, date, the 
letter “P” for plasma and the tube number).  
5. 
Store immediately the plasma at 80°C until its shipment. 
For whole blood collection for RNA:  
1. 
Extract 2,5 ml of blood into PAXgene Blood RNA Tubes properly labeled (patient’s 
code number, date, and the tube number). It should be attempted to perform the blood 
extraction at the same time as a routine blood test. 
2. 
Invert the tube 10 times (do not shake) and store immediately at -80ºC until its shipment. 
 
For urine collection:  
1.  Collect urine  samples in the collection tube and aliquote it in properly labeled 15mL 
tubes (maximum 10mL of urine per tube), properly labeled with patient’s code number, 
date, the letter “U” for urine and the tube number. Store immediately at -20ºC until its 
shipment.  
 
 
 

SH-TBL (Study of Human TB Lesions) 
 
 
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10 ml of blood
Invert tube 10 mes
Centrifuge 2000g, 4°C, 10 min
P1
P2
P3
Aliquot plasma in labelled tubes
Freeze immediately and store at
-80°C un l its shipment in dry ice
!
< 1 hour
Vacutainer® tube
with EDTA
PaxGene
RNA tube
2.5 ml of blood
Invert tube 10 mes
Fresh urine
Aliquot urine in 15mL labelled tubes
Freeze immediately and store at
-20°C un l its shipment in dry ice

SH-TBL (Study of Human TB Lesions) 
 
 
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11. 
ANNEX 3. INFORMED CONSENT FORM    
  
INFORMED CONSENT SHEET 
 
This data sheet may contain terms that you don´t understand.  Do not hesitate to ask your  
doctor or the attending personal any questions that you may have. 
 
Title:  “Study of TB lesions obtained in surgery: in search of best biomarkers correlating with 
TB pathology, clinical features, MDR cases and prognostic: SH-TBL (Study of Human TB 
Lesions).” 
 
 
We  are  writing  to  inform  you  about  a  research  study  we  would  like  you  to  
join.  The study was  approved  by  the  Clinical  Research  Ethics  Committee  of  the National 
Center for Tuberculosis and Lung Diseases (NCTLD) and is conducted according all the 
current European Laws on Ethical Research.  
 
 
Our  intention  is just  to provide  you  with  the correct  and  necessary  information  
to help  you consider  and decide  if you want to participate  or not. For this, we are asking 
you to read this information carefully.  The  study’s  doctor  will  clarify  any  doubts  you  
may  have after reading  the explanation.   
 
 
The aim of this document is to offer you information in order you can decide if you 
want to hand us your biological samples exclusively with research purposes, in the way we 
will explain you below. The biological samples consist in one/two blood sample(s) plus tissue 
samples of the extracted lesion for which you will be operated. To conduct the present Project 
we do expect to collect biological samples from approximately 100 patients having had 
surgery in the NCTLD within a year.  
 
Why is this study taking place? 
 
Tuberculosis is still an important problem around the world.  Every year 100 million 
people acquire this infection.  Its prevention is currently very difficult, since this is a disease 
caused by an airborne bacterium (Mycobacterium tuberculosis).  Of all people  getting  
infected,  only some  will  develop  the  disease  and,  to  this  date,  we  still  don´t  know  

SH-TBL (Study of Human TB Lesions) 
 
 
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why  some  people  get sick and why other’s  don´t. Georgia has a high burden of people ill 
from tuberculosis (TB), half of them being due to bacilli resistant to common therapies, and 
for this reason its physicians and surgeons are experts on TB management. A research team 
from Barcelona (Catalonia, Spain) leaded by Dr Vilaplana and in collaboration with the 
surgery team of the NCTLD headed by Dr. Vashakidze and the microbiology team of the 
same institute (Dr. Shubladze) want to study the TB lesions removed within routine surgery 
of this disease in order to 1) understand the mechanisms of disease generation; 2) find new 
therapies and 3) find new diagnostic methods to stratify TB patients. The final aim of the 
study is to better address and manage the patients’ disease.   
 
What will you have to do in this study? 
 
You have been approached because you will be surgically intervened for your 
condition.  
 
If you do accept to participate, we will use the lesion removed to perform our studies.  
 
This will not modify any procedure from the prescribed routine designed by your 
surgeon. This will not represent any added procedure to your clinical management plan but 
an extra blood sample collection.  
 
The study  consists  of using the tissue sample removed within routine surgery of your 
disease and 2 blood samples (one prior to surgery and another during follow-up visit).The  
samples  will  be  kept  in  the  Experimental  Tuberculosis  Unit  lab  (person  in  charge: Dr. 
Cristina  Vilaplana)  and  will  be  handled  according  to  the Biomedical  Research Law 
14/2007.  They will be labeled with the study’s research code (a numeric  one  that  does  not  
identify  you  directly),  and  they  will  only  be used  for  the study’s  purposes  and  they  
will be  kept  during  10  years  for future  research  related  to the disease  and previously  
approved  by an Ethics Committee. 
 
 
The results will be introduced in a database and will be used for scientific purposes, 
and the data will always be handled by specialized personnel. 
 
Which are the benefits, risks and alternatives to participate in this study? 
 
The participation in this study is voluntary, and if you are not willing to collaborate in 
it, this will not represent any harm for you.  
 
Your participation  in  this  study  won´t  change the  treatment  you are  receiving right  
now  for your condition.   

SH-TBL (Study of Human TB Lesions) 
 
 
20 
 
If you do not participate in this  study,  you will  still  receive  the  more  appropriate  
treatment  for  your condition,  as  if  you would  during  the participation. Your participation 
will not modify any procedure from the prescribed routine designed by your surgeon. This will 
not represent any added procedure to your clinical management plan but an extra blood sample 
collection.  
 
 
As no intervention will be given to you besides the routine schedule to manage your 
condition, no potential side effects are expected from this study. Blood collections can cause 
local discomfort or bruises in the puncture area.  
 
It is not  expected  that  you will have  a particular  benefit  from  this  study.  Your 
participation in this study will have no cost for you and you are not going to receive any 
economic reward or any other kind of reward for participating.   
 
 
In any case, your participation in this study may be of great help to understand 
your disease and may contribute to design better diagnostic and therapeutic approaches from 
which other patients might benefit in a short future.  
 
Do you have to remain in the study? 
 
Your participation is voluntary.  If you do not want to participate in the study, tell the 
study’s doctor.  At any time you can decide not to continue without giving any reason for it. 
If you don´t want to participate in the study or if you leave the study, you will be treated as 
usual.  
 
If you   decide  to  stop  participating  in  this  study,  please  contact  the  study’s 
physician. 
 
All data and samples collected before your withdrawal from the study will be used for 
the purposes of the study.   
 
 
Even if  you  are  not  interested  in  participating  in  the  study  anymore,  your  health 
information  is  very  important  for  this  study.  We will ask you to let  us  gather  information 
about  your health  till  the  completion  of  the  study.  For this reason,  and  with  your 
permission,  we  would  be  in contact  with  you  or,  if it was  not  possible,  with  your  
relatives, friends  or your physician.  We will ask for the relevant contact information in your 
case. 
What happens with your personal data? 
 
The data  will be coded to  preserve   your   identity,   according   to  the  Organic  

SH-TBL (Study of Human TB Lesions) 
 
 
21 
Spanish Law  15/1999  from  December 13th,  devoted   to  personal   data  protection.   
According   to  this  law  you  have  the  right  to access,  change,  oppose  and  cancel  your  
data.  To do any of these you must contact your study’s physician.  You can rest assured that 
your  clinical  history  information  will be  strictly  confidential  and  your  identity  will  be  
kept  anonymous.   
 
When you will accept to participate, a code will be generated and be used to match 
your samples with your clinical data without need to use your personal data. Only the people 
responsible of this study (principal investigator and investigator team) will have access to 
these data if necessary. Your identity will  not  be  revealed  to  anybody  except  when  
required  by  medical  emergencies   or legal requests.  
 
Your    identity will remain confidential even if some results of the study are 
published.   
 
This research study will not generate any data directly applicable to you regarding 
your condition, as the value of the results obtained is expected at long-term.  
 
Questions and Answers: 
If you have any question or doubt related to this study please contact your physician.

SH-TBL (Study of Human TB Lesions) 
 
 
22 
 
 
INFORMED CONSENT FORM FOR SAMPLES’ DONATION 
 
Systematic collection of blood and tissue samples for the study of Tuberculosis  
 
Patient’s code number: _____________________ 
 
 I 
________________________________________________________________ 
(complete 
name) 
 
…have read the information sheet that was given to me 
…have been able to make inquiries about the study  
…have received enough information about the study 
…understand that my participation is voluntary 
…understand that I can withdraw myself from the study: 
 
1. Whenever I want 
2. Without giving any explanations 
3. Without it affecting my medical care 
For which I hereby consent that the samples obtained during my attention care may be stored and 
used in medical research authorized by the ethics’ committee and preserving the anonymity. 
 
Date and place: ___________________ 
 
Signature: _______________________