Impact of Cardiac Amyloidosis on Patients With Severe Aortic Stenosis Who Undergo Transcatheter Aortic
Conditions: Valve Stenoses, Aortic, Amyloidosis Cardiac
Sponsor: Javier López Pais

Prognostic impact of cardiac amyloidosis in patients with severe aortic stenosis who 
underwent percutaneous implantation of an aortic prosthesis 
 
 
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Project AMY-TAVI (Protocol 29.09.2018) 
 
 
 
 
Title: Prognostic impact of cardiac 
amyloidosis in patients with severe aortic 
stenosis who underwent percutaneous 
implantation of aortic valve prosthesis  
 
Promoter: Hemodynamics Unit, Department of 
Cardiology in Santiago de Compostela Hospital 
Main Researcher: Diego López Otero 
Protocol Code: AMY-TAVI 
Project Type: An analytical and observational study of 
prospective cohorts at a multicenter and international 
level.  
 
 
 
 
 
 

Prognostic impact of cardiac amyloidosis in patients with severe aortic stenosis who 
underwent percutaneous implantation of an aortic prosthesis 
 
 
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Project AMY-TAVI (Protocol 29.09.2018) 
 
 
 
 
INDEX 
1. STUDY SUMMARY 
2. GLOSSARY AND DEFINITIONS 
3. LIST OF RESEARCHERS AND TASKS TO BE COMPLETED 
4. INTRODUCTION AND CURRENT STATUS OF THE MATTER 
5. BIBLIOGRAPHY 
6. RESEARCHERS GROUP EXPERIENCE ON THE MATTER 
7. HYPOTHESIS 
8. OBJECTIVE 
1. PRIMARY OBJECTIVE 
2. SECONDARY OBJECTIVES 
9. DESIGN 
1. POPULATION 
2. INCLUSION CRITERIA 
3. EXCLUSION CRITERIA 
4. AMYLOIDOSIS DIAGNOSES CRITERIA  
5. INFORMED CONSENT 
10. STATISTICAL  ANALYSIS PLAN AND SAMPLE SIZE  
1. STATISTICAL ANALYSIS 
2. SAMPLE SIZE  
11. PATIENTS FOLLOW-UPS 
1. INITIAL PATIENT ASSESSMENT  
2. CLINICAL FOLLOW-UP 
12. WORK PLAN 
13. DATA MANAGEMENT 
1. DATA COLLECTION 
2. DATA MANAGEMENT 
3. DATA CONFIDENTIALITY 
14. PRACTICAL USE OF THE RESULTS 
15. ETHICS 
16. ANNEX 
1. ECHOCARDIOGRAPHY PROTOCOL 
2. SCINTIGRAPHY PROTOCOL 
3. DECLARATION OF HELSINKI 
 
 

Prognostic impact of cardiac amyloidosis in patients with severe aortic stenosis who 
underwent percutaneous implantation of an aortic prosthesis 
 
 
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Project AMY-TAVI (Protocol 29.09.2018) 
 
 
1. STUDY SUMMARY 
 
Study title and code 
Prognostic impact of cardiac amyloidosis 
in patients with severe aortic stenosis 
who underwent percutaneous 
implantation of an aortic prosthesis 
(AMY-TAVI) 
Study type and design 
Observational analytical of multicenter 
prospective cohorts. 
Number  and characteristics of patients 
321 patients to whom a TAVI has been 
implanted by severe degenerative AoS. 
Study duration 
Recruitment period: All the consecutive 
patients to whom a non-invasive 
diagnostic study of amyloidosis 
(scintigraphy with tecnecio99 
pyrophosphate and electrophoresis of 
proteins in the blood) and to whom a 
percutaneous aortic prosthesis for severe 
aortic stenosis has been implanted. 
Follow-up period: clinical and 
echocardiographic monitoring will be 
done before discharge, in consultation at 
three, six, twelve, and twenty-four 
months. 
Inclusion: 1 year 
Follow-up: 2 years 
Data analysis:  6 months 
Total: 3 years and 6 months 
Events 
Main event: cardiovascular events 
(cardiovascular death, stroke, heart 
attack), death from any cause, 
readmission for cardiac insufficiency, 
pacemaker implantation, functional 
assessment 
Participating countries and centers  
Various Spanish and international centers 
will be invited to participate. 
 
 
 
 
 

Prognostic impact of cardiac amyloidosis in patients with severe aortic stenosis who 
underwent percutaneous implantation of an aortic prosthesis 
 
 
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Project AMY-TAVI (Protocol 29.09.2018) 
 
 
2. GLOSSARY AND DEFINITIONS 
AoS 
Aortic stenosis 
TAVI 
Percutaneous implant of aortic prosthesis 
GT99 
Scintigraphy with talio99 
PE 
Protein Electrophoresis 
CMR 
Cardiac Magnetic Resonance 
AVR 
Aortic valve replacement  
ECG 
Electrocardiogram 
6MT 
6-minute test 
HR 
Hazard ratio 
TTE 
Transthoracic echocardiography 
DCL 
Data Collection Logbook 
CREC 
Clinical research ethics committee 
IHC 
International harmonization conference 
 
3. LIST OF RESEARCHERS AND TASKS TO BE COMPLETED 
University Hospital Complex of Santiago de Compostela 
- 
Main researcher: 
o Diego López Otero 
 
- 
Collaborating researchers: 
o Ramiro Trillo Nouche, responsible for the Hemodynamic Department 
and the healthcare procedure for severe aortic stenosis and TAVI. 
o Javier López Pais, responsible for patient selection, coordination and 
data collection. 
o Virginia Pubul Núñez, responsible for the acquisition and interpretation 
of the GT99. 
o María Bastos Fernández, responsible for the acquisition and 
interpretation of the echocardiography.  
o Juan Carlos Sanmartín Pena, co-responsible for the healthcare 
procedure for severe aortic stenosis and TAVI. 
o Ana Belén Cid Álvarez, co-responsible for the healthcare procedure for 
severe aortic stenosis and TAVI. 
o Carmen Neiro Rey, co-responsible for the programming of tests, 
monitoring, and functional studies. 
o Montserrat Bello Seoane, co-responsible for the programming of tests, 
monitoring, and functional studies. 
o Mónica Gómez Fernández, co-responsible for the programming of tests, 
monitoring, and functional studies. 

Prognostic impact of cardiac amyloidosis in patients with severe aortic stenosis who 
underwent percutaneous implantation of an aortic prosthesis 
 
 
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Project AMY-TAVI (Protocol 29.09.2018) 
 
 
The intention is to increase the number of collaborating centers in order to carry out a 
study of an international multicentre by nature. (To begin with, other Sergas hospitals 
will not be taken into account). 
 
4. INTRODUCTION AND CURRENT STATUS OF THE MATTER 
The aortic stenosis. 
The degenerative aortic stenosis (AS) is a progressive disease that is related to 
atherosclerosis, inflammation, hemodynamic factors, and active calcification. Its 
prevalence is high and depends on age. More than 3% of people over 75 have aortic 
stenosis(1) and reaches 8% in people over 84(2). When symptoms appear, the disease 
is rapidly progressive, and a surgical or percutaneous aortic valve replacement is 
necessary. It is, therefore, a disease with a major impact on society. Without a doubt, 
we are facing a serious health problem that will increase in the future, giving the aging 
population. 
The percutaneous implantation of an aortic valve prosthesis. 
Given the nature of the degenerative aortic stenosis, most of the patients affected to a 
severe degree and requiring treatment have an advanced age. The fragility of aging 
combined with the comorbidities presented in many cases makes up one-third of the 
patients who are not candidates for a surgical valve and another third present a high 
risk. The percutaneous implantation of an aortic prosthesis (TAVI) emerges as a less 
invasive alternative for the treatment of aortic stenosis. The randomized study 
PARTNER demonstrated in cohort B a reduction in mortality with this technique 
compared with standard medical treatment in inoperable patients(3). In cohort A of 
the PARTNER study, no differences were found in the survival of patients with high-risk 
surgical treatment randomized to conventional surgery or percutaneous implant(4). 
The studies PARTNER-II and SURTAVI have more recently shown that those patients, 
with intermediate surgical risk, the percutaneous aortic prosthesis implant has similar 
results to those of conventional surgery (5)(6). This has increased the number of 
percutaneous aortic prosthesis implants exponentially. 
There are still many doubts about the long-term outcome after a TAVI implant. 
Avoiding the sternotomy and extracorporeal circulation by means of a percutaneous 
access has many advantages, but it also causes a series of technical and clinical 
problems: 1, cerebrovascular events, not only those due to the handling of large 

Prognostic impact of cardiac amyloidosis in patients with severe aortic stenosis who 
underwent percutaneous implantation of an aortic prosthesis 
 
 
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Project AMY-TAVI (Protocol 29.09.2018) 
 
 
vessels  during the procedure but also those occurred in the monitoring where there is 
still no solid evidence regarding antithrombotic handling. 2, Atrioventricular 
conduction disorders requiring permanent pacemaker implantation in 20-40% of the 
patients. 3, Vascular complications of the accesses. 4, Residual valve by the 
malpositioning of the prosthesis (creating a high residual gradient or insufficiency). 5, 
Valvular degeneration and durability of the prosthesis. 
Cardiac amyloidosis by transthyretin. 
The amyloid myocardial infiltration by transthyretin monomers and oligomers (TTR) 
with misfolding(7) is a serious and under-diagnosed disease which predominantly 
affects men over 60 years old and presents itself with symptoms of heart failure, 
orthostatic hypotension, and syncope (8). The infiltration of the cardiac conduction 
system can generate branch blocks, atrioventricular blocks, and atrial fibrillation.  
There are two types of amyloid that can infíltrate the myocardium: the amyloid light 
chain (AL or primary) within a monoclonal gammopathy, and the TTR amyloid. The 
latter in turn has two types of disease, familial, due to a mutation that causes 
transthyretin misfolding; and sporadic (formerly known as senile), which is due to 
incorrect folding of the common transthyretin. (8) 
The prevalence of the TTR amyloidosis in patients with AoS to whom a TAVI is 
performed is estimated at 14-16%(9)(10) and 6% in those over 65 years old to whom a 
surgical valve replacement is performed(11). It is especially associated with the AoS 
phenotype of low-flow and low gradient (9). Cardiac amyloid deposits were found in 
nearly one-third of autopsies of patients with TAVI implants who died before 30 
months(12), and the patients with AoS and amyloidosis have a prognosis similar to 
those with amyloidosis without AoS (13), It all states that the amyloidosis implies a 
worse prognosis (11). It is also suggested that amyloid infiltration may increase the 
requirements of permanent pacing (14). 
Recently the benefits of treatment with Tafamidis on patients with cardiac amyloidosis 
by TTR have been published. (15)  
Non-invasive diagnosis of cardiac amyloidosis. 
The echocardiogram is used to find interesting data of amyloidosis: increased 
thicknesses of the atrioventricular valves, interatrial septum valves or the free wall of 

Prognostic impact of cardiac amyloidosis in patients with severe aortic stenosis who 
underwent percutaneous implantation of an aortic prosthesis 
 
 
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Project AMY-TAVI (Protocol 29.09.2018) 
 
 
the right ventricle, the light pericardial effusions, high atrial dilation, a birefringence 
characteristic (sparkling) and a speed of S’ waveform mitral in the Doppler tissue lower 
than 6cm/s(9), but in no case is it diagnosed. The mechanical ventricular assessed by 
speckle tracking(tracking marks) may be useful in monitoring patients with amyloidosis 
as there seems to be a link between the longitudinal strain of the ventricular and the 
cardiac involvement (16). The longitudinal strain is decreased in the basal and middle 
segments and stored in the apical segments(17). This typical pattern helps obtain the 
differential diagnosis with other causes of hypertrophy ventricular, with high 
sensitivity and specificity (90-95% and 80-85%, respectively). The value of the strain is 
also an independent prognostic factor (18). 
 The maps on T1  MRI may have a role in the diagnosis (19), but it is still a technique in 
the early stage of development. 
Until recently, the diagnosis of amyloidosis required histological confirmation by 
biopsy and staining with Congo red of the sample. Recent studies have demonstrated 
the diagnostic capacity of the pyrophosphate tecnecio99 scintigraphy (20), with a 
sensitivity of more than 99% and a specificity of 86% for the TTR amyloidosis, being 
false positives due almost exclusively to the presence of AL amyloidosis. Therefore, the 
caption of radiotracer in grades 2 o 3 together with the absence of a monoclonal 
protein in the electrophoresis  has a specificity and a positive predictive value to 100% 
TTR amyloidosis. (21)  
 
5. BIBLIOGRAPHY 
1.  
Muszil D, Avelino A, Matesz K, Manuscript A, Asymmetric LPK, Schank JR, et al. 
NIH Public Access. Changes. 2009;34(6):1443–53.  
2.  
Lindroos M, Kupari M, Heikkilä J, Tilvis R. Prevalence of aortic valve 
abnormalities in the elderly: An echocardiographic study of a random 
population sample. J Am Coll Cardiol. 1993;21(5):1220–5.  
3.  
Leon MB, Smith CR, Mack M, Miller DC, Moses JW, Svensson LG, et al. 
Transcatheter Aortic-Valve Implantation for Aortic Stenosis in Patients Who 
Cannot Undergo Surgery. N Engl J Med. 2010 Sep 22;363(17):1597–607.  
4.  
Smith CR, Leon MB, Mack MJ, Miller DC, Moses JW, Svensson LG, et al. 
Transcatheter versus Surgical Aortic-Valve Replacement in High-Risk Patients. N 

Prognostic impact of cardiac amyloidosis in patients with severe aortic stenosis who 
underwent percutaneous implantation of an aortic prosthesis 
 
 
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Engl J Med. 2011 Jun 5;364(23):2187–98.  
5.  
Adams DH, Popma JJ, Reardon MJ, Yakubov SJ, Coselli JS, Deeb GM, et al. 
Transcatheter Aortic-Valve Replacement with a Self-Expanding Prosthesis. N 
Engl J Med. 2014;370(19):1790–8.  
6.  
Leon MB, Smith CR, Mack MJ, Makkar RR, Svensson LG, Kodali SK, et al. 
Transcatheter or Surgical Aortic-Valve Replacement in Intermediate-Risk 
Patients. N Engl J Med. 2016 Apr 2;374(17):1609–20.  
7.  
Rapezzi C, Quarta CC, Riva L, Longhi S, Gallelli I, Lorenzini M, et al. Transthyretin-
related amyloidoses and the heart: a clinical overview. Nat Rev Cardiol. 2010 
May 18;7:398.  
8.  
Ruberg FL, Berk JL. Transthyretin (TRR) Cardiac Amyloidosis. Circulation. 
2012;126(10):617–38.  
9.  
Castano A, Narotsky DL, Hamid N, Khalique OK, Morgenstern R, DeLuca A, et al. 
Unveiling transthyretin cardiac amyloidosis and its predictors among elderly 
patients with severe aortic stenosis undergoing transcatheter aortic valve 
replacement. Eur Heart J. 2017;38(38):2879–87.  
10.  
Scully PR, Treibel TA, Fontana M, Lloyd G, Mullen M, Pugliese F, et al. Prevalence 
of Cardiac Amyloidosis in Patients Referred for Transcatheter Aortic Valve 
Replacement. J Am Coll Cardiol. 2018;71(4):463–4.  
11.  
Treibel TA, Fontana M, Gilbertson JA, Castelletti S, White SK, Scully PR, et al. 
Occult transthyretin cardiac amyloid in severe calcific aortic stenosis. Circ 
Cardiovasc Imaging. 2016;9(8):1–10.  
12.  
Nietlispach F, Webb JG, Ye J, Cheung A, Lichtenstein S V., Carere RG, et al. 
Pathology of transcatheter valve therapy. JACC Cardiovasc Interv. 
2012;5(5):582–90.  
13.  
Sperry BW, Jones BM, Vranian MN, Hanna M, Jaber WA. Recognizing 
Transthyretin Cardiac Amyloidosis in Patients With Aortic Stenosis: Impact on 
Prognosis. JACC Cardiovasc Imaging. 2016;9(7):904–6.  
14.  
Adam Castaño M, Sabahat Bokhari M, Mathew S. Maurer M. Reply: Could late 
enhancement and need for permanent pacemaker implantation in patients 
undergoing TAVR be explained by undiagnosed transthyretin cardiac 
amyloidosis? J Am Coll Cardiol. 2015;65(3):311.  
15.  
Maurer MS, Schwartz JH, Gundapaneni B, Elliott PM, Merlini G, Waddington-
Cruz M, et al. Tafamidis Treatment for Patients with Transthyretin Amyloid 
Cardiomyopathy. N Engl J Med. 2018;NEJMoa1805689.  

Prognostic impact of cardiac amyloidosis in patients with severe aortic stenosis who 
underwent percutaneous implantation of an aortic prosthesis 
 
 
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16.  
Urbano-Moral JA, Gangadharamurthy D, Comenzo RL, Pandian NG, Patel AR.  
Speckle tracking three-dimensional echocardiography(tracking marks) in the 
cardiac amyloidosis of light chains: study of the parameters of the mechanical 
myocardial left and right ventricular. Rev Española Cardiol. 2015;68(08):657–64.  
17.  
Phelan D, Collier P, Thavendiranathan P, Popović ZB, Hanna M, Plana JC, et al. 
Relative apical sparing of longitudinal strain using two-dimensional speckle-
tracking echocardiography is both sensitive and specific for the diagnosis of 
cardiac amyloidosis. Heart. 2012 Oct 1;98(19):1442 LP-1448.  
18.  
Tendler A, Helmke S, Teruya S, Alvarez J, Maurer MS. The myocardial 
contraction fraction is superior to ejection fraction in predicting survival in 
patients with AL cardiac amyloidosis. Amyloid. 2015 Jan 2;22(1):61–6.  
19.  
Karamitsos TD, Piechnik SK, Banypersad SM, Fontana M, Ntusi NB, Ferreira VM, 
et al. Noncontrast T1 mapping for the diagnosis of cardiac amyloidosis. JACC 
Cardiovasc Imaging. 2013;6(4):488–97.  
20.  
Bokhari S, Castaño A, Pozniakoff T, Deslisle S, Latif F, Maurer MS. (99m)Tc-
Pyrophosphate scintigraphy for differentiating light-chain cardiac amyloidosis 
from the transthyretin-related familial and senile cardiac amyloidoses. Circ 
Cardiovasc Imaging. 2013 Mar 1;6(2):195–201.  
21.  
Gillmore JD, Maurer MS, Falk RH, Merlini G, Damy T, Dispenzieri A, et al. 
Nonbiopsy diagnosis of cardiac transthyretin amyloidosis. Circulation. 
2016;133(24):2404–12.  
 
7. HYPOTHESIS 
Amyloidosis is a disease which coexists in a high percentage of patients referred for 
TAVI by severe aortic stenosis; the pathophysiological mechanisms of this relationship 
have a negative impact on prognosis. 
 
8. OBJECTIVES 
8.1 PRIMARY OBJECTIVE 
A multicentre prospective study of cohorts consisting of two cohorts from patients 
who have been implanted with percutaneous aortic prosthesis as treatment for the 
severe degenerative aortic stenosis and compare the overall mortality and that for 

Prognostic impact of cardiac amyloidosis in patients with severe aortic stenosis who 
underwent percutaneous implantation of an aortic prosthesis 
 
 
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Project AMY-TAVI (Protocol 29.09.2018) 
 
 
cardiovascular and admissions for heart failure in the 2-year follow-up of patients 
diagnosed with amyloidosis (by GT99 and EFP) versus those that do not have it. 
8.1 SECONDARY OBJECTIVES 
1. As part of the hypothesis, we will analyze separately the impact of the presence of 
amyloidosis on the requirements of permanent pacing (permanent pacemaker 
implantation, pacemaker dependence, changes in the conduction system) 
2. Determine the impact on the functional class (measured by using the 6-minute test) 
of the coexistence of TTR amyloidosis in patients with a TAVI implant. 
3. Validate echocardiographic patterns of cardiac amyloidosis diagnosed by ventricular 
strain in a population with EAS undergoing a TAVI implant, as well as study the 
evolution of these patterns mid-term. 
 
9. DESIGN 
A multicentre, analytical, observational study of two cohorts. One cohort will be 
constituted by the patients to whom a TAVI will be implanted, and that meet non-
invasive diagnostic criteria of TTR amyloidosis (capting of radiotracer in grades 2 and 3 
and absence of monoclonal gammopathy the PE of urine or serum). The other cohort 
will be constituted by patients whose TAVI implants do not meet the diagnostic criteria 
for TTR amyloidosis. 
A clinical and echocardiographic follow-up will be done in all patients for two years. 
9.1. POPULATION 
Elderly patients, enlisted in the hospitalization area or from outpatients clinics that 
meet the following requirements will be included in the study: diagnosis of severe 
aortic stenosis by conventional echocardiographic criteria (maximum speed >4m/s, 
medium transvalvular gradient >40mmHg, area <1cm2). 
9.2. INCLUSION CRITERIA 
- Replacement of the aortic valve with a percutaneous prosthesis by severe aortic 
stenosis after evaluation by the Heart Team. 

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- Ability to understand and sign informed consent. 
- Do not meet any of the exclusion criteria. 
9.3. EXCLUSION CRITERIA 
- Death as a result of a complication of the procedure during hospitalization. 
- An associated mitral valve requiring intervention 
- TAVI implant valve-in-valve. 
- Does not sign informed consent. 
9.4. INFORMED CONSENT 
If the patient decides to participate, the patient information sheet will be given to the 
patient, and after reading the document and clarifying any doubts, if they agree to 
participate in the study, the patient will sign the informed consent. 
 
10. STATISTICAL ANALYSIS PLAN AND SAMPLE SIZE 
10.1 STATISTICAL ANALYSIS 
A descriptive analysis of both cohorts defining their clinical and angiographic profiles 
and a comparative study of the two groups (cohorts) will be performed. The 
categorical variables are expressed in percentages and the continuous as medium 
value  ± standard deviation.  The relative risk and the “Hazard Ratio” will be used as 
measures of effect when variables are analyzed with a temporal component, always 
with a confidence interval of 95%. For the comparison of categorical variables, the Chi-
square or the Fisher test will be used. To compare continuous variables, the “t“ of 
student test or analysis of variance will be used. The variables of time to event, such as 
the analysis of event-free survival will be analyzed by using the Kaplan- Meier 
technique and will be supplemented by the log-rank test. The multivariate analysis of 
the adverse events predictors during the follow-up will be done by means of the Cox 
regression model, determining the Odds Ratio for each variable. 
10.1 SAMPLE SIZE 

Prognostic impact of cardiac amyloidosis in patients with severe aortic stenosis who 
underwent percutaneous implantation of an aortic prosthesis 
 
 
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Project AMY-TAVI (Protocol 29.09.2018) 
 
 
Based on the information available, around 15% of the patients to whom a TAVI has 
been implanted show signs of amyloidosis. Mortality at two years after a TAVI is about 
17% and in patients with amyloidosis, it is estimated to be 40%. Based on this data, for 
a significance level of 5% (error α = 0,05) and a power of 80% (error β= 0,20), a sample 
size of 306 patients is estimated, assuming 5% of follow-up loses, it was decided to 
include 321 patients. The results in which the calculations are based are given at 2 
years, if not significant; monitoring would continue up to 3 years.  
 
11. PATIENTS FOLLOW-UPS 
11.1 INITIAL PATIENT ASSESSMENT 
An initial inclusion visit will assess whether the patient is a candidate who meets the 
inclusion criteria and none of the exclusion ones. After being informed, if they accept, 
the informed consent must be signed. The following data must be recorded: 
- Clinical parameters: age, sex, height, weight, body mass index, body surface, 
cardiovascular risk factors, medication intake, background interests, history of heart 
disease, presence of ischemic heart disease, cerebrovascular accident, peripheral 
artery disease, chronic obstructive pulmonary disease, symptoms (angina, syncope, 
dyspnea according to grades of the NYHA, other symptoms), blood pressure, heart 
rate, evidence of heart failure, assessment of functional class by means of 6MT and 
fragility assessment. 
- Analytical parameters: hematocrit, NT-proBNP, Troponin I, electrolytes, C-
reactive protein, creatinine, glomerular filtrate. 
-Protein electrophoresis: Rating presence of the monoclonal component 
- Scintigraphy with TC99: Assessment of values of tracer uptake. 
- Device parameters: Type of prosthesis, dimensions, expandable/ self-
expanding ball, post-dilatation. 
- ECG parameters: Rhythm, PR duration, pacemakers, implantable cardioverter 
defibrillator, QRS duration. 

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- The 6 minute-test: All patients in the study will have a test of 6 minutes 
before implantation and 6 months follow-up, according to the protocol after a TAVI, in 
order to carry out an objective functional assessment of the functional capacity. The 
distance covered will be determined, the variation of oxygen saturation by means of 
non-invasive pulse oximetry before and after the test and the appearance of 
symptoms with the effort according to the Borg scale. 
11.2. CLINICAL AND IMAGE FOLLOW-UP 
This will be done before discharge, in consultation at three, six, and 12 months and 
annually thereafter. All the visits have a window period to be carried out with 
additional tests within this protocol, one week before and one week after the due 
date. 
 
First visit (Basal Visit). 
- Assessment of patient history. 
- Detailed physical assessment of the patient. 
- Electrocardiogram. 
- Analysis. 
- Echocardiographic basal study, with ventricular strain measurements. 
- Perform 6-minute test. 
 
Follow-up visit at 3 months (more/less one week) 
- Clinical assessment. 
- Analysis. 
- Events assessment. 
 
Follow-up visit at 6 months (more/less one week) 
- Clinical assessment. 
- Analysis. 
- Perform 6-minute test. 
- Events assessment. 
 
Follow-up visit at 12 months (more/less one week) 
- Clinical assessment. 
- Electrocardiogram.  

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- Analysis. 
- Echocardiographic study with ventricular strain measurements. 
- Events assessment. 
 
Follow-up visit at 24 months (more/less one week) 
- Clinical assessment. 
- Analysis. 
- Echocardiographic study. 
- Events assessment. 
 
11.3 WITHDRAWAL FROM THE STUDY 
A) Withdrawal of a patient from the study because they withdraw the consent. 
B) Withdrawal of the study due to transfer to another health area 
 
12. WORK PLAN 
 
Basal 
3 
months 
6 
months 
1 year 
2 years 
Clinical history 
X 
 
 
 
 
Informed Consent 
X 
 
 
 
 
Clinical assessment 
X 
X 
X 
X 
X 
ECG 
X 
 
X 
X 
X 
Analysis1 
X 
 
X 
X 
X 
Protein electrophoresis 
X 
 
 
 
 
Scintigraphy with tecnecio99 
X 
 
 
 
 
Echocardiogram with strain 
X 
 
X 
X 
X 
The 6-minute test 
X 
 
X 
 
 
Events assessment  
 
X 
X 
X 
X 
 
 
1: includes: hemoglobin, hematocrit, lipid profile, NT-proBNP, TnT, electrolytes, C-reactive 
protein, creatinine, and glomerular filtrate. 
 
13. DATA MANAGEMENT 
13.1. DATA COLLECTION: 

Prognostic impact of cardiac amyloidosis in patients with severe aortic stenosis who 
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All the patients will receive standard treatment according to clinical practice guidelines 
and consensus in force. The criteria for inclusion/ exclusion will be assessed in all of 
them to confirm the feasibility of entering the study, which will be offered, explained, 
and then the appropriate informed consent collected. Subsequently, following the 
predefined protocol, a number of epidemiological variables, clinical, laboratory, 
electrocardiographic, echocardiographic, functionals, scintigraphy with Tc99basals of 
the patient will be collected before discharge. All previously detailed. All the data will 
be collected in the Data Collection Logbook (DCL), which is designed to register all the 
data required in the protocol and collected by the researcher. The data on the DCL will 
be coded. The data entry will be done by the researcher or the person assigned by 
them, and then that data will be transferred to the database of the study. The 
researcher or the appointed person in their team commits to completing the DCL on 
each visit made by the participant. All data corrections in the DCL will be made by the 
researcher or the person assigned in their team in accordance with the instructions 
given. After the last visit from the participant and the closure of the DCL, the 
researcher must attest, by signing the DCL, the authenticity of the data collected in the 
DCL, the coherence between the data in the DCL and the data contained in the source 
document.  
13.2DATA CONFIDENTIALITY: 
All documents and information provided to the researchers by the promoter regarding 
the study are confidential. Both the researcher and their collaborators undertake to 
use the information provided only within the framework of this study to carry out this 
protocol. This commitment remains as long as the confidential information has not 
been made public by the promoter. The protocol delivered to the researcher can be 
used by them or their collaborators to obtain informed consent from the participants 
entering the study. It should not be disclosed to third parties without the written 
consent of the promoter. This information will be treated in accordance with the rules 
of professional secrecy. The screening list must be completed when the researcher 
verifies that the participant can enter the study after confirming their record and 
medical history.  
14. PRACTICAL USE OF THE RESULTS 
The progressive aging of our population carries an increasing prevalence of 
degenerative aortic valve stenosis in our country, constituting a public health problem 

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of crucial importance. There is scientific evidence that amyloidosis is an 
underdiagnosed disease in clinical practice, which coexists simultaneously in a large 
number of patients suffering from aortic stenosis, and that could overshadow the 
prognosis. To determine the prevalence and prognostic impact of amyloidosis referred 
for percutaneous implantation of an aortic prosthesis is important in order to 
understand the interrelationship of both diseases. This information will guide us to 
take the appropriate measures in need of a screening and the early treatment that will 
allow optimizing the resources available. 
15. ETHICAL CONSIDERATIONS 
The project will be made in compliance with the Declaration of Helsinki (Annex III) of 
the 1964 World Medical Association 1964 and ratifications of the following assemblies 
(Tokyo 75, Venice 83, Hong Kong 89, Somerset West 96, Scotland 00, Seoul 08 and 
Fortaleza 13) on ethical principles for medical research on human beings, the RD 
1090/2015, of 24 December, clinical trials, specifically that stipulated in Article 38 on 
good clinical practice and the Agreement related to human rights and biomedicine), 
made in Oviedo the 4 of April 1997 and subsequent updates. 
The participating researchers in this study undertake that all clinical data collected 
from the study subjects are separated from the personal identification data so that the 
anonymity of the patient is ensured; respecting the law on  Personal Data Protection 
(Organic Law 15/1999, 13 of December), the RD 1720/2007 of 21 of December, 
approving  the Regulations implementing the Organic Law 15/1999, the Law 41/2002, 
of 14 of November (regulating the autonomy of the patient, rights and obligations 
regarding clinical information and documentation), as well as Law 3/2001, of 28 of 
May, (regulating the informed consent and medical history of the patients), the Law 
3/2005, of 7 of March, modification of  Law 3/2001 and the 29/2009 Decree of 5 of 
February , by which electronic access to medical histories is regulated. 
The clinical data of patients will be collected by the researcher in the Data Collection 
Logbook (DCL) specific to the study. Each DCL will be coded, protecting the identity of 
the patient. Only the research team and health authorities, who have a duty to 
maintain confidentiality, have access to all the data collected for the study. Third 
parties may only be provided with information that is not identified. Once the study is 
completed, the data will be destroyed. 
Sample handling will follow what is established in Law 14/2007 of 3 of July for 
Biomedical Research, in the RD 1301/2006, of 10 of November, establishing the 

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regulations for quality and safety for the donation, procurement, evaluation, 
processing, preservation, storage and distribution of human cells and tissues and 
abiding  the regulations for coordination and functioning for its use on humans and in 
the RD 1716/2011, of 18 of November, establishing the basic requirements for 
authorisation and functioning of biobanks for biomedical research and treatment of 
biological samples of human origin, and the functioning  and National Registry of 
Biobanks for the biomedical research is regulated. 
The treatment, communication, and transfer of data will be done according to what is 
stated in the General Data Protection Regulation (Regulation (UE) 2016/679 of the 
European Parliament and the Council, of 27 of April 2016). The center, where the 
information is obtained, is responsible for processing the data. The data collected will 
only be used for the purposes of the research study described in the protocol and 
maintained for the time needed to achieve the study objectives and in accordance 
with the applicable law.  
 
 
 

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ANNEX I: ECHOCARDIOGRAPHIC STUDY 
 
SUGGESTIVE ECHOCARDIOGRAPHIC FINDINGS:  
 
Cardiac 
involvement 
is 
generally 
characterized 
by 
the 
presence 
of 
an 
infiltrative/restrictive myocardiopathy with conventional echocardiographic findings 
that have been well described, but also with new echocardiographic findings identified 
using echocardiographic strain techniques. The main suggestive findings are: 
 
• Increased wall thickness ≥ 12 mm with a myocardium of a shiny appearance, 
refractile, granular or mottled (“sparkling”), resulting from the deposits of amyloid 
fibrils which are more echogenic than a normal myocardium; wall thickening VI 
medium > 15 mm was an independent factor associated with a worse prognosis.  
• Normal or small left ventricular cavity. 
• Ejection fraction of VI (FEVI) preserved or borderline > 50% (at least in the early 
stages of the disease; depressed in advanced stages) 
• Tissue Doppler: waves S´ and E´ reduced to basal level of the septal or lateral 
myocardium, reflecting a depressed longitudinal and an altered relaxation of VI. 
• Abnormal mitral filling pattern: from diastolic dysfunction of VI mild or moderate 
(transmitral pattern type I or II); and in advanced stages, a typical restrictive 
transmitral pattern, with a ratio E/A >2, E/E´ increased, and small wave A due to 
the atrial dysfunction. 
• The high filling pressure of VI may gradually lead to enlargement of the left atrium 
(diameter > 23 mm/m2; area > 20 cm2 or maximum volume > 28 ml/m2), which is 
useful for the differential diagnosis and it is also an independent factor associated 
with worse prognosis in AL. 
• Enlarged right atrium and dilated inferior vena cava, reflecting the filling pressure 
of the right ventricle (RV). 
• Increased thickness of the interatrial septum (this variable is much easier to 
measure in cardiac magnetic resonance (MRI) than in echocardiography). 
• Wall thickness free of increased RV (> 7 mm), with systolic and diastolic 
dysfunction of RV, associated with worse survival.  
• Thickening of the left and right valves, generally responsible for light 
insufficiencies. 

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• Reduced aortic ejection time (< 273 ms), described as a prognostic factor. 
• Mild pericardial effusion in 50% of the cases. The presence of effusion is an 
independent factor associated with worse survival;  
• Atrial thrombus can be found despite maintaining a sinus rhythm;  
 
 
ACQUISITION PROTOCOL 
. 
Acquire 3 beats in sinus rhythm and 5 beats in Atrial Fibrillation. 
Ensure good ECG signal. 
For Doppler curves, use adjusted scales to the corresponding valve gradients, scan 
speed 75 mm/s. 
Optimize sector width, gain, and compression to ensure maximum image quality. 
Do not perform any measurement in the study to be sent to the Core Lab (only the 
collection of images and imaging loops) 
 
Parasternal long axis (PLAX): 
2D 
Colour in mitral valve and aortic valve 
2D Zoom in aortic valve 
If aortic insufficiency: zoom color in the aortic valve. 
Mode M in aortic valve 
Modo M in VI 
 
Parasternal short axis (PSAX). 
2D 
Zoom 2D in aortic valve 
Colour in the aortic valve. 
Zoom 2D in pulmonary valve 
Colour in the pulmonary valve 
Pulsed Doppler in the pulmonary valve 
Colour in the pulmonary valve 
Colour in tricuspid valve 
Continuous Doppler in tricuspid valve  
Mode M in aortic valve and in VI (if PLAX has not been performed) 
 

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Parasternal in Right Cavities. 
2D 
Colour Doppler in the tricuspid valve. 
Continuous Doppler in the tricuspid valve. 
 
Apical 4C: 
2D 
2D with extended left Ventricle 
Colour Doppler in Mitral Valve 
Pulsed Doppler in Mitral Valve 
Pulsed Tissue Doppler in septal and lateral Mitral annulus. 
Continuous Doppler in Mitral Valve, if significant insufficiency  
Zoom color with PISA scale in Mitral Valve, if significant insufficiency. 
Colour Doppler in Tricuspid Valve 
Continuous Doppler in Tricuspid Valve 
Mode M in tricuspid annulus. 
Pulsed Tissue Doppler in the tricuspid annulus. 
3D: Acquisition of complete volume of the left ventricle (minimize the presence of 
artifacts and optimize profit adjustment) 
3D: Acquisition of complete volume centered in the right ventricle. 
 
Apical 5C 
2D 
Colour Doppler in aortic valve 
Pulsed Doppler in TSVI 
Continuous Doppler in aortic valve 
 
APICAL 2C 
2D 
Colour Doppler in Mitral Valve. 
Zoom color with PISA scale in the mitral valve, if significant insufficiency. 
 
APICAL 3C 
2D 
Zoom in TSVI 
Colour Doppler in aortic valve. 

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Pulsed Doppler in TSVI 
Continuous Doppler in aortic valve 
Colour Doppler in Mitral valve. 
If significant mitral insufficiency: continuous in Mitral and zoom color with PISA scale in 
Mitral. 
 
Subcostal plan 
2D 4C. 
Zoom  VD free wall (for measuring the thickness of the right ventricle) 
2D in VCI 
Mode M in VCI, from basal to deep inspiration.  
 
 
ACQUISITION PROTOCOL FOR ECHOCARDIOGRAPHY IMAGES FOR THE STUDY OF THE 
STRAIN. 
 
Aspects of images ACQUISITION: 
Configure the study to gain 3 consecutive beats in apnea. If the patient is in atrial 
fibrillation, acquire 5 beats, avoid extrasystole. 
Use a temporary resolution /frame rate of > 50 (ideally 60-80 frames/s). 
 
Plans for measuring of the STRAIN 
Acquire imaging of left ventricle expanded in the plans apical 4C, Apical 2C, and Apical 
3C, focussed on the left ventricle, ensuring a correct visualisation of the endocardial 
borders. 
Acquire imaging of the left atrium in the plans apical 4C, apical 2C, and apical 3C, 
focused on the atrium, ensuring correct visualization of all the atrium borders, avoiding 
an auricle and pulmonary veins.  
Acquire imaging centered in the right ventricle from the plan apical 4C, ensuring 
adequate visualization of the endocardial border of the free wall.  
 
 
REFERENCES FOR ANNEX II 
 
1. Lang RM, Badano LP, Mor-Avi V, el al. Recommendations for cardiac chamber 
quantification by echocardiography in adults: an update from the American 

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Society of Echocardiography and the European Association of Cardiovascular 
Imaging. Eur Heart J Cardiovasc Imaging. 2015; 16:233–270. doi: 
10.1093/ehjci/jev014.  
 
2. Pagourelias ED, et al. Echo Parameters for Differential Diagnosis in Cardiac 
Amyloidosis. 
Circ 
Cardiovasc 
Imaging. 
2017; 
10:e005588. 
DOI: 
10.1161/CIRCIMAGING.116.005588.)  
 
3. González-Lopez E, López-Sainz A, García-Pavia P. Diagnosis and treatment in 
cardiac amyloidosis by transthyretin. Progress and hope. Rev Esp Cardiol. 
2017;70(11):991–1004  
 
4. White JA, Fine NM. Recent Advances in Cardiovascular Imaging Relevant to the 
Management of Patients with Suspected Cardiac Amyloidosis. Curr Cardiol Rep 
(2016) 18: 77 DOI 10.1007/s11886-016-0752-7  
 
5. Phelan D, et al. Relative apical sparing of longitudinal strain using two-
dimensional speckle- tracking echocardiography are both sensitive and specific 
for the diagnosis of cardiac amyloidosis .Heart 2012; 98:1442e1448. 
doi:10.1136/heartjnl-2012-302353  
 
6. Bhogal S, et al. Cardiac Amyloidosis: An Updated Review With Emphasis on 
Diagnosis and Future Directions .Curr Probl Cardiol 2018; 43:10–34.  
 
7. Tendler A, Helmke S, Teruya S, Alvarez J, Maurer MS. The myocardial 
contraction fraction is superior to ejection fraction in predicting survival in 
patients with AL cardiac amyloidosis. Amyloid 2015;22:61-6  
 
8. Rubin, 
Jonah 
et 
al. 
Myocardial 
Contraction 
Fraction 
by 
M-Mode 
Echocardiography Is Superior to Ejection Fraction in Predicting Mortality in 
Transthyretin Amyloidosis. Journal of Cardiac Failure , Volume 24 , Issue 8 , 504 
- 511 
 
 

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ANNEX II: SCINTIGRAPHIC STUDY 
BASIC INFORMATION 
        Amyloidosis is a heterogeneous group of diseases in which there are abnormal 
extracellular insoluble fibril deposits, which are given the anatomopathological name 
of amyloid deposits. The amyloidosis can be classified into systemic or localized ways 
in an organ. Some types of this disease can infiltrate the heart, although only some 
types of these twenty known amyloidotic proteins produce cardiac affectation. These 
correspond to the varieties: senile, the secondary type AA, the primary type AL, and 
some inherited ways like the one related to transthyretin (ATTR), among others 
(AAPoA-I  and Afib). Of these, the most frequently related to the heart are the ATTR 
and AC-AL.  
        The 
99mTc-DPD (99mTc acid 3,3-diphosphono-1,2-propanedicarboxylic) is a 
bisphosphonate conventionally used in nuclear medicine for assessing bone pathology, 
but it has also demonstrated utility as an important technique to distinguish between 
the cardiac deposits in AC-TTR versus those in AC-AL, as they pose evolution, 
treatment and different prognoses as in the case of amyloidosis due to genetic defect 
with great transcendence for the family.  
        The cardiac scintigraphy with 99mTc-DPD is a tool that has proven useful for 
differentiating between the amyloidosis related with the transthyretin and the deposit 
by light chains of monoclonal immunoglobulin.  
 
PATIENT PREPARATION 
Previous preparation is not necessary. 
Avoid caffeine and toxins like tobacco 48 hours before the study. 
RADIOPHARMACEUTICALS, DOSAGE, AND ADMINISTRATION 
Compound: The radiopharmaceutical used is the 99mTc-DPD. The 99mTc-MPD can also 
be used. 
Dosage:  
- Adults: 20 mCi (740 Mbq). 
- For children, the dosage is calculated according to the weight and the clinical 
guidelines in the European Society of Nuclear Medicine for the 99mTc. 
 
Administration: intravenous. 
 
Dosimetry: The effective dosage (E) resulting from the administration of a quantity of 
700 MBq of acid 3,3-diphosphono-1,2-propanedicarboxylic labeled with technetium 
(99mTc) for an adult of 70 kg in weight is of 4 mSv. For an administered activity of 700 
MBq, the dosage of radiation absorbed by the target organ (bone) is 44,1 mGy, and  
33,6 mGy by the critical organ (bladder). 

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Dosage absorbed to organs (mGy/MBq): Bone 0,063; Heart 0,0012; Bladder 0,050; 
Kidneys 0,007; Bone marrow 0,0096,  Ovaries 0,0035, testicles 0,0024 and Kidneys 
0,007.  
INSTRUMENTATION 
 
Collimator: LEHR 
 
Window: 20% centered in 140 KeV  
 
Mode: selective cardiac images. 
 
Matrix: 256 x 256, 1.000 Kc.  
 
Zoom: 1,3.  
 
IMAGES ACQUISITION 
Patient supine with the head of the gamma camera focused on the area of interest 
(heart region). 
Planar Study and selective SPECT of the cardiac area. 
Projection:  
Full body: front and rear  
Selective cardiac areas: Anterior, 45º anterior left oblique, and lateral left.  
Images acquisition: 
Beginning of exploration: at 3 hours post-injection of the tracer. 
Study duration: - Accumulate 1000 kcts./ previous and subsequent projection. 
             - Accumulate 250 kcts./oblique projection. 
 
INTERPRETATION 
During the examination, on the one hand, the existence and intensity of the deposit in 
the myocardium will be assessed, and on the other hand, its distribution. Furthermore, 
the relation in the distribution of the radiotracer among the bone structures and soft 
parts as an indication of its proper clearance and image quality. 
          The intensity of the deposit is graduated according to a visual rating scale of 0 to 
3, in which the absence of capture is valued with a score of 0; the capture lower than 
bone (referred to as the adjacent rib), with: 1; similar to bone: 2 and higher capture 
than bone: 3.  
          The distribution of the deposit in the myocardium is assessed as focal deposit, 
diffuse deposit, deposit in a segment of the ventricular wall, diffuse ventricular deposit 
or diffuse biventricular deposit. The captured relation between the bone and the 

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background when this is very similar is graded 0, and when the bone is more or much 
more highlighted than the background, it is graded 1. 
 
REFERENCES TO ANNEX II 
 
Sanjay M. Banypersad, MRCP; James C. Moon, MD, MRCP; Carol Whelan, MD, MRCP; 
Philip N. Hawkins, PhD, FmedSci; Ashutosh D. Wechalekar, DM, MRCP, FRCPath. 
Updates in Cardiac Amyloidosis: A Review. December 20, 2016.  
 
Suzane Garcia Ferreira a,b,⁎, Alexandre Marins Rocha a, Osvaldo José Moreira do 
Nascimento a, Claudio Tinoco Mesquita. Role of 99mTc-DPD scintigraphy on 
discrimination of familial cardiac amyloidosis. November 2015.  
 
Brett W. Sperry, MD; Michael N. Vranian, MD; Rory Hachamovitch, MD; Hariom Joshi, 
MD; Asad Ikram, MBBS; Dermot Phelan, MD; Mazen Hanna, MD. Subtype-Specific 
Interactions and Prognosis in Cardiac Amyloidosis. December, 2016. 
 
Enrica Perugini, MD,* Pier Luigi Guidalotti, MD,† Fabrizio Salvi, MD,‡ Robin M. T. 
Cooke, MA,*Cinzia Pettinato, MD,† Letizia Riva, MD. Noninvasive Etiologic Diagnosis of 
Cardiac Amyloidosis Using 99mTc-3,3-Diphosphono-1,2-Propanodicarboxylic Acid 
Scintigraphy. Journal of American College of Cardiology 2005.  
 
Bjo¨ rn Pilebroa, Ole B. Suhrb, Ulf Na¨slunda, Per Westermarkc, Per Lindqvistd, and 
Torbjo¨ rn Sundstro¨me. 99mTc-DPD uptake reflects amyloid fibril composition in 
hereditary transthyretin amyloidosis. UPSALA JOURNAL OF MEDICAL SCIENCES, 2016. 
 
Wael A. AlJaroudi, MD, FACC, FAHA, FESC, FASE, FASNC,a Milind Y. Desai, MD,b, W. H. 
Wilson Tang, MD,b Dermot Phelan, MB, BCH, PhD,b Manuel D. Cerqueira, MD,b and 
Wael A. Jaber, MDb. Role of imaging in the diagnosis and management of patients with 
cardiac amyloidosis: State of the art review and focus on emerging nuclear techniques. 
2013 American Society of Nuclear Cardiology. 
 
F. Javier de Haro-del Moral, Alicia Sánchez-Lajusticia. Role of cardiac scintigraphy with 
99Tc-DPD in the discrimination of the cardiac amyloidosis subtype. Rev Esp Cardiol. 
2012; 65 (5); 440-446.  
 
J. Mallol. Radiopharmaceutical Manual. Iberoamerican Academy of Pharmacy; 
M.5.797- 2008.  
 
 

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ANNEX III: DECLARATION OF HELSINKI  
 
Initially submitted in 1964 
Original: English 
 
DECLARATION OF HELSINKI IN THE WORLD MEDICAL ASSOCIATION 
Ethical principles for medical research in human beings 
 
Adopted by the 18th General Assembly in the World Medical Association (WMA) 
Helsinki,Finland, June 1964 and amended by the 
29th General Assembly of the WMA, Tokyo, Japan, October 1975 
35th General Assembly of the WMA, Venice, Italy, October 1983 
41st General Assembly of the WMA, Hong Kong, September 1989 
48th General Assembly of the WMA, Somerset West, South Africa, October 1996 
and the 52nd General Assembly of the WMA, Edinburgh, Scotland, October 2000 and 
Explanatory note to paragraph 29, added by the General Assembly of the WMA, 
Washington 2002 
Explanatory note to paragraph 30, added by the General Assembly of the WMA, Tokyo 
2004 
 
A. INTRODUCTION 
1. The World Medical Association has passed the Declaration of Helsinki as a statement 
of ethical principles to provide guidance to doctors and other people who carry out 
medical research in humans. Medical research in humans includes research on human 
material or identifiable information. 
2. The duty of the doctor is to promote and safeguard the health of people. The 
knowledge and the awareness of the doctor are dedicated to the fulfillment of the 
duty.  
3. The Declaration of Geneva in the World Medical Association binds the doctor with 
the words "the health of my patient will be my first consideration", and the 
International Code of Medical Ethics declares: "The doctor must act only in the interest 
of the participant when providing medical care which might have the effect of 
weakening the physical and mental condition of the participant". 
4. Medical progress is based on research, which ultimately must rest in part on 
experimentation on humans. 
5. Medical research on human beings, concern for the welfare of human beings must 
take precedence over the interests of science and society.  
6. The main purpose of medical research involving humans is to improve preventive, 
diagnoses and therapeutic procedures, and also the understanding of the etiology and 
pathogenesis of the diseases. Even the best proven preventive, diagnoses and 
therapeutic methods must continuously be challenged through research to be 
efficient, effective, accessible and good quality. 

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7. In the present practice of medicine and medical research, most of the preventive, 
diagnoses and therapeutic procedures involve risks and costs. 
8. Medical research is subject to ethical standards that promote respect to all human 
beings and protect their health and individual rights. Some populations subject to the 
research are vulnerable and need special protection. The particular needs must be 
recognised of those that have economic or medical disadvantages. Attention must also 
be paid to those who cannot give or refuse consent for themselves, those who can give 
consent under pressure, those who will not personally benefit from the research and 
those that have the research combined with medical care. 
9. Researchers must know the ethical, lawful and legal requirements for the research 
on human beings in their own countries as well as the applicable international 
requirements. There is no allowance for an ethical or legal requirement to reduce or 
eliminate any measure of protection for human beings set in this Declaration. 
 
B. BASIC PRINCIPLES FOR ALL MEDICAL RESEARCH  
10. In a medical investigation it is the duty of the doctor to protect the life, health, 
privacy and dignity of the human being. 
11. The medical research in human beings must conform to the generally accepted 
scientific principles and must be based on thorough knowledge of scientific literature, 
other relevant sources of information, as well as laboratory experiments correctly 
carried out, and on animals, when appropriate. 
12. When researching, adequate attention must be paid to the factors that may harm 
the environment. The welfare of the animals used in the experiments must also be 
looked after. 
13. The project and method of all experimental procedure in human beings must be 
clearly formulated in an experimental protocol. This must be submitted for 
consideration, comment, advice, and when appropriate, approval, to an ethical 
assessment committee specially appointed, that must be independent of the 
researcher, promoter or any other kind of undue influence. It is understood that this 
independent committee should be in accordance with the applicable laws and 
regulations in the country where the experimental research takes place. The 
committee has the right to control ongoing trials. The researcher has the obligation to 
provide monitoring information to the committee, especially any serious adverse 
events. The researcher must also submit to the committee the information on funding, 
sponsorships, institutional affiliations, other possible conflicts of interest and 
incentives for the people in the study.   
14. The Project and method of all experimental procedures in human beings must be 
clearly formulated in an experimental protocol. This must be sent, for consideration, 
comment, advice, and when appropriate, approval, to a specially assigned committee 
of ethical assessment that must be independent of the researcher, promoter or any 
other kind of undue influence. It is understood that this independent committee 
should be in accordance with the applicable laws and regulations in the country where 
the experimental research takes place. The committee has the right to control ongoing 
trails. The researcher has an obligation to provide monitoring information to the 
committee, especially any serious adverse events.  The researcher should also submit 

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to the committee the information on funding, sponsorships, institutional affiliations, 
other possible conflicts of interest and incentives for the people in the study.   
 
15. The research protocol should always make reference to the ethical considerations 
that belong to the case and must indicate that the enunciated principles in this 
Declaration have been complied with. 
16. The medical investigation in human beings must be carried out only by qualified 
scientific people and under the supervision of a clinically competent doctor. The 
responsibility of human beings must always rest with a person with medical training 
and never on the participants of the research, even though they have given consent.  
17. Every medical research project in human beings must be preceded by a careful 
comparison of the expected risks with the foreseeable benefits for the individual or 
others. This does not preclude the participation of healthy volunteers in medical 
research. The design of all studies must be available to the public. 
18. Doctors must abstain from engaging in research projects on human beings unless 
they are confident that the risks involved have been adequately assessed and that it is 
possible to face them in a satisfactory manner. The experiment taking place must be 
suspended if the risks involved are more important than the expected benefits and if 
there is conclusive proof of positive or beneficial results. 
19. The medical research on human beings must only be carried out if the importance 
of the objective is greater than the inherent risk and the costs to the individual. This is 
especially important when humans are healthy volunteers. 
20. The medical research is only justified if there is a reasonable likelihood that the 
population on which the research is done, may benefit from the results. 
21. To take part in a research project, the individuals must be voluntary and informed 
participants. 
22. Always respect the right of participants in the research to protect their integrity. 
Every caution must be taken to protect the privacy of individuals, the confidentiality of 
the information on the participant and to minimise the consequences of the research 
on their physical and mental integrity and personality. 
23. In any research on human beings, each potential individual must receive adequate 
information about the objectives, methods, financial sources, possible conflicts of 
interest, institutional affiliations of the researcher, expected benefits, foreseeable risks 
and discomforts from the experiment. The person must be informed of the right to 
participate or not in the research and to withdraw their consent at any time without 
reprisal. After ensuring that the individual has understood the information, the doctor 
must then obtain, preferably in writing, the informed and voluntary consent of the 
person. If the consent cannot be obtained in writing, the process to obtain it must be 
formally documented and witnessed. 
24. When obtaining informed consent for the research project, the doctor must be 
particularly cautious when the individual is linked with them due to dependency or 
providing consent under pressure. In such a case, the informed consent must be 
obtained by a well informed doctor who does not participate in the research and is 
independent from this relationship. 

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25. When a person is deemed legally incompetent, physically or mentally unfit to 
obtain an informed consent or a minor, the researcher must obtain the informed 
consent from the legal representative and in accordance with the applicable law. 
These groups must not be included in the research unless it is necessary to promote 
the health of the population represented and this research cannot be carried out on 
legally competent people. 
26. If a person is deemed legally incompetent, as is the case of a minor, they are able 
to give their assent to participate or not in the research, the researcher must obtain it 
apart from the consent from the legal representative. 
27. Research on individuals from whom consent cannot be obtained even by 
representative or in advance, must only be done if the physical/mental condition that 
prevents obtaining the informed consent is a necessary characteristic of the 
population being researched. The specific reasons why research participants are used 
which cannot provide their informed consent must be stated in the experimental 
protocol for consideration and approval by the assessment committee. The protocol 
must state that the consent of the individual or legal representative to remain in the 
research must be obtained as soon as possible. 
28. Both authors and publishers have ethical obligations. When publishing the results 
of their research, the doctor is obliged to maintain the accuracy of the data and 
results. Both negative and positive results must be published or otherwise available to 
the public. The publication must cite the source of funding, institutional affiliations and 
any possible conflicts of interest. Reports on research that do not adhere to the 
principles described in the Declaration must not be accepted for publication. 
 
C. APPLICABLE PRINCIPLES WHEN MEDICAL RESEARCH IS COMBINED WITH MEDICAL 
CARE  
29. The doctor may combine medical research with medical care only to the extent 
that such research is justified by its preventive, diagnosis and therapeutic potential. 
When medical research is combined with medical care, the additional standards apply 
to protect the participants participating in the research. 
30. The possible benefits, risks, costs and effectiveness of new procedures must be 
assessed by its comparison with the best preventive, diagnoses and therapeutic 
methods available. This does not exclude a placebo, or any treatment, in studies where 
preventive, diagnoses and therapeutic procedures tested are not available.(See 
explanatory note*). 
31. At the end of the research, all the participants that participate in the study must be 
assured of the best preventive, diagnoses and therapeutic methods available, 
identified by the study (See explanatory note*). 
32. The doctor should fully inform the participants about the aspects of care related to 
the study. The refusal from the participant to participate in a research must never 
interfere with the doctor-participant relationship. 
33. When the preventive, diagnoses and therapeutic methods do not exist or result 
ineffective in the attention of a patient, the doctor, with the informed consent of the 
participant, may use preventive, diagnoses and therapeutic procedures which are new 
or not tested, if, in their opinion, this offers hope of saving life, restoring health or 

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alleviating the suffering. Whenever possible, those measures must be researched to 
evaluate its safety and effectiveness. In all cases, that new information must be 
registered and, when appropriate, published. All the relevant guidelines of this 
Declaration must be followed. 
 
*Explanatory note to paragraph 29 in the declaration of Helsinki in the WMA  
The WMA reaffirms that there must be great care when using trials with placebo, and 
in general, this methodology can only be applied if not counting on a tested and 
existing therapy. However, the trials with placebo are ethically acceptable in some 
cases, even if there is a tested therapy and the following conditions are met: 
-when for methodological, scientific and compelling reasons, its use is necessary to 
determine the effectiveness and safety of a preventive, diagnosis and therapeutic 
method; or 
- when a preventive, diagnosis or therapeutic method is tested for a disease of less 
importance which does not imply an additional risk, serious adverse effects or 
irreversible harm for the participants who receive the placebo. 
All the other provisions of the Declaration of Helsinki, especially the need for 
appropriate ethical and scientific reviews must be followed. 
 
*Explanatory note to paragraph 30 in the declaration of Helsinki in the WMA 
Hereby, the WMA reaffirms its position that it is necessary, during the process of the 
study plan, to identify the access after the testing of the participants in the study to 
preventive, diagnoses and therapeutic procedures that have resulted beneficial in the 
study or the access to another appropriate attention. The provisions for the access 
after testing or other attention must be described in the protocol of the study so that 
the ethical review committee can consider such arrangements during its review. 
 
9.10.2004 
The declaration de Helsinki (Document 17.C) is an official document of the World 
Medical Association, representing doctors in the world. Adapted in 1964 in Helsinki 
(Finland), was revised in 1975 in Tokyo (Japan), in 1983 in Venice (Italy), in 1989 in 
Hong Kong, in 1996 in Somerset West (South Africa), in 2000 in Edinburgh (Scotland), 
by the General Assembly of the WMA, Washington 2002 (adding an explanatory note 
for paragraph 29) and by the General Assembly of the WMA, Tokyo 2004 (adding an 
explanatory note for paragraph 30).