1. APROTININ an old kid back on the block
Stefan De Hert, MD, PhD
Department of Anaesthesiology and Perioperative Medicine
Ghent University Hospital
Ghent University
Belgium

2. disclosures ?

3. disclosures ?

4. the rise and fall of aprotinin
Actually, the title of this presentation could be the rise and fall of aprotinin.
Indeed, aprotinin is the example of a drug that became very popular at one time in history and then got critized and banned from clinical practice based on a number of controversial studies.
Let me give you a short overview of the history of aprotinin and explain you why the drug is now getting new attention and which the modalities are for its re-introduction in clinical practice.

5. aprotinin bovine pancreatic trypsin inhibitor (BPTI) or
basic trypsin inhibitor of bovine pancreas
derived from: cow parotid glands (1930)
bovine pancreas (1936)
bovine lung (1964)
The drug aprotinin (Trasylol, previously Bayer and now Nordic Group pharmaceuticals), is the small protein bovine pancreatic trypsin inhibitor (BPTI), or basic trypsin inhibitor of bovine pancreas 
It is an antifibrinolytic molecule that inhibits trypsin and related proteolytic enzymes.
As it inhibits pancreatic enzymes, it was initially used in the treatment for acute pancreatitis, in which destruction of the gland by its own enzymes is thought to be part of the pathogenesis.[
Initially named "kallikrein inactivator", aprotinin was first isolated from cow parotid glands in 1930 and independently as a trypsin inhibitor from bovine pancreas in It was purified from bovine lung in 1964.
C284H432N84O79S7

6. aprotinin = aspecific inhibitor of serine proteases trypsin  pancreas
plasmin  fibrinolytic system
(50 KIU / mL)
kallikrein  inflammatory chain
(200 KIU / mL) ….
1. The reason why the drug became very popular is related to its hemostatic effect because of the inhibition of the fibrinolytic enzyme plasmin.

7. Tice D, Worth MH Jr, Clauss RH, Reed GH
The inhibition of trasylol of fibrinolytic activity associated with cardiovascular operations
Surg Gynecol Obstet 1964; 119:
Indeed, from the early 60’s of last century the drug was used in order to reduce perioperative blood loss …

8. n = 2 x 11 pts blood loss: control: 1509 ± 388 mL
aprotinin: ± 48 mL
RBC transfusion: control: 1975 ± 520 g
aprotinin: ± 48 g
p < 0.001
n = 2 x 11 pts
But the real breakthrough of aprotinin started with the landmark publication of Royston in 1987 in the Lancet where he demonstrated that aprotinin dramatically reduced blood loss and the need for transfusion in redo open-heart surgery. 1.
Results = mean ± SEM
2. Of note, this study only had 11 pts in each group

9. In a systematic review, published in 2005, Ferguson and coll found that by 2002 a further 63 papers had been published since 1987
+ 63 papers

10. Cumulative meta-analysis of these RCTs found it was clear that aprotinin greatly decreased the need for perioperative transfusion
with an OR stablizing at 0.25 by the time of the 12th study in 1992.
Yet, over the next decade, a further 52 RCTs were undertaken. Fergusson’s explanation was not nice suggesting that the designers of these studies failed to do their homework by failing to cite previous publications and to perform adequate literature searches.

11. 1993 subsequent use in other major surgery  liver surgery
FDA approval for reducing blood loss during CABG surgery
subsequent use in other major surgery
 liver surgery
 orthopedic surgery
 …
1993  FDA approval; soon also in several European countries for CABG
1. Used also in other indications …

12. safety of aprotinin ?

13. observational, prospective cohort study n = 4374 adult CABG pts
The first observation study to cast doubt on the safety of aprotinin was this one from the group of Mangano ….
1, The study found that aprotinin was associated with an increased incidence of cardiac events, stroke, encephalopathy and mortality, where neither aminocaproic acid or tranexamic acid were associated with an increased risk.
2, And the same phenomenon was observed for renal dysfunction …
observational, prospective cohort study
n = 4374 adult CABG pts
aprotinin / aminocaproic acid / tranexamic acid vs no agent

14. n = 2 x 449 cardiac surgery pts
A second observational study published by Karkouti et al that same year found that aprotinin may be associated with renal dysfunction.

15. 2006  anaphylaxis  graft occlusion  stroke  + renal dysfunction
FDA safety concerns for aprotinin
 anaphylaxis
 graft occlusion
 stroke
 + renal dysfunction
As a consequence, in 2006, the FDA listed renal dysfunction along with anaphylaxis, graft occlusion and stroke amog the drug’s safety concerns.

16. However, the association of aprotinin and renal failure was disputed by Furnary and colleagues in 2007who reported that this was a confounding variable and that renal impairment was related to an increased packed red cell transfusion in the setting of cardiac surgery.

17. In 2007, Mangano and colleagues published a further analysis of the same data set that was used in their 2006 paper and the authors reported that aprotinin was independently predictive of 5-year mortality.

19. In 2008, the next important study, blood conservation using antifibrinolytics in a randomized trial – the BART study – was published.
The BART study was very influential because, unlike previous observational studies, it was a blinded RCT comparing aprotinin, tranexamic acid and aminocaproic acid in patients undergoing high-risk cardiac surgery.
The primary outcome was massive postoperaative bleeding and 30-day mortality was a secondary outcome.
The trial was terminated early by the safety committee because of a non-significant increased mortality in patients receiving aprotinin.
Indeed the BART study reported a higher 30-day mortality with aprotinin than with the other lysine analgues although there was less massive bleeding in the aprotinin group.

20. (p = 0.06)
(p = 0.05)

21. Following these publications, the manufacturer temporarily suspended marketing in November 2007.
In December 2007, the UK license of aprotinin was suspended on advice from the commission on human medicines.
In February 2008, the license was also suspended by the European Commission and finally aprotinin was permanently withdrawn from the market in 2008.
Of note, in some countries, aprotinin remained available on the market through a limited market plan.

22. the revival of aprotinin

23. news_and_events/news/2012/02/news_detail_
Mangano D et al.
N Engl J Med 2006; 354:
JAMA 2007; 297:
BART study
N Engl J Med; 2008; 358:
rappel-avis/
hc-sc/2011/13544a-eng.php
observational
not blinded, without randomization
= level 2- evidence; risk of bias and
non-causal relation
propensity-adjusted
= aprotinin in highest-risk pts
propensity scores missing for > 400 pts
choice of agents UNCLEAR
outcomes NOT well-defined
details of surgery lacking …
S3-01-Bayer.pdf
unexplained exclusion of 137 pts from analysis
reclassifications of outcomes
mortality trend different in excluded pts
heparin strategy inconsistent / inappropriate
lack of anticoagulation monitoring
no replication of findings …
First the Mangano studies:
Limitations in these studies strongly suggest that their conclusions may not be reliable.
First, the study ws observational, non-blinded, without randomization, and used unmatched groups.
The National Institute for Clinical Excellence (NICE) classifies the quality of such evidence as level 2-; associated with a high risk of confounding or bias and a significant risk that the relationship is NOT causal
Propensity-adjusted, multivariable logistic regression analyses were used to control for between-group differences at baseline. However, it is not clear whether such analysis has overcome the likely bias that aprotinin was administered to the sickest patients.
Cited risk factors for adverse outcomes were sometimes inappropriate such as for example, duration of education.
Moreover values for propensity scores for over 400 patients were missing and the reasons why patients received any specific agent or no agent remain unclear.
There were likely to be variations in practices between countries and centers, and outcomes were not clearly defined.
In addition, the authors failed to report details of the surgery, such as whether surgery was on- or off-pump, time on-pump, and number of vessels grafted.
These variables are all likely not only to influence the choice of the antifrinolytic agent but also outcome, and they are therefore a source of bias.
These multiple sources of confounding raised concerns regarding the validity of both Mangano papers.
The cardiovascular and renal drug advisory committee of the FDA reviewed the evidence from the study but could not endorse it because of questions about the methodologies used and because the data had not been independently reviewed by the FDA.
Second: whilst the BART study had the benefit of being a blinded RCT, a number of serious limitations were identified in this study.
First there is the unexplained exclusion of 137 pts from the analysis after randomization. A re-analysis which included these initially excluded pts reversed the statistical significance of the mortality signal for aprotinin.
There was an unusually large number of reclassifications of outcomes from the originally reported data, with a large (~75%) change rate in primary outcome (massive postoperative bleeding). Reclassifications were in opposite directions for aprotinin vs tranexamic acid and aminocaproic acid, favouring the latter, and these changes increased with the duration of the study.
The trend in mortality for the excluded patients in the study was opposite that for the included patients.
The way in which heparin was used for anticoagulation during CPB was inconsistent and sometimes inappropriate.
There was a lack of appropriate monitoring of patients’ anticoagulation with the ACT in that aprotinin could have influenced the reagent.
And finally, the findings of the BART study could not be reproduced because when data from other RCTs were analyzed together without data from the BART study, aprotinin was NOT associated with a higher risk of death in the aprotinin-treated patients
These re-analyses casted serious doubts about the validity of the results from the BART study come from health Canada
And the EMA
news_and_events/news/2012/02/news_detail_
jsp&mid=WC0b01ac058004d5c1

24. 2012/02/news_detail_001447.jsp&mid=WC0b01ac058004d5c1
2012/02/news_detail_ jsp&mid=WC0b01ac058004d5c1
Based on all these data and findings about all these weaknesses and pitfalls, experts, first from Health Canada concluded that the BART study could not be reliably used to assess the benefit-risk balance of aprotinin and that the benefits of aprotinin outweighed its risks.
1. Then the EMA also critically analyzed the available data and came to the same conclusion
After which they released a statement in February 2012, indicating that the benefits of aprotinin outweigh its risks in appropriately managed patients but only for the indication isolated CABG

25.  request to have a registry of use of aprotinin
NAPaR (Nordic Aprotinin Patient Registry)
= a non-interventional post-authorization safety study
of pattern of use of Nordic aprotinin
So, after this, market authorization was re-instituted and aprotinin became again available for clinical use in Europe and the current indication is: ….
However there are a number of conditions related to its use
They requested to have a registry of use of aprotinin. This is the NAPaR
Apart from the pattern of use, they were specifically interested in anti-coagulation strategy during CPB, the potential causal relationship with the use of aprotinin and renal dysfunction and finally, the potential safety issue of hypersensitivity reactions.
The registry specifically looks in detail to these items.
pattern of use
anti-coagulation strategy
~ renal dysfunction
hypersensitivity reactions

26. So, this is the current status for aprotinin.
For those who want some more background information; here are a number of recent articles on the topic.

27. NAPaR = Nordic Aprotinin Patient Registry
What is the NAPaR ?
NAPaR = Nordic Aprotinin Patient Registry
The NAPaR is a non-interventional Post-Authorisation Safety Study (PASS)
A registry is to be conducted to monitor the pattern of use of aprotinin
Mandatory Pharmacovigilance Risk Assessment Committee (PRAC)* endorsed protocol in March 2015
Record on patients information exposed to aprotinin at cardiac surgery centres in participating countries
Easy-to-use web-based database (“eCRF”)
Duration of study: at least three years or 12,000 patients… and the decision to terminate the Registry will be taken based on agreement with the PRAC
* The Pharmacovigilance Risk Assessment Committee (PRAC) is the European Medicines Agency's (EMA) committee responsible for assessing and monitoring the safety of human medicines.

28. Data Safety Monitoring Committee (DSMC)
Organisation
5 members meet every 6 months
Review of interim statistical analysis & yearly Post-Authorisation Safety Study report
Stefan De Hert (BE), Chair
AlexandreOuattara (FR)
David Royston (UK)
Jan Van Der Linden (SE)
Kai Zacharowski (DE)
Role
Safeguarding the interests of patients involved in the study and the safety of the interventions
Monitoring the overall conduct of the NAPaR study
Make recommendations to Nordic accordingly

29. First included patient
Status on September 6, 2018:
countries:
Active
Achieve regulatory acceptance
Pending regulatoy validation
Countries
Active sites
First included patient
Nbre of patients SE 4
02/2016
122 UK 31
03/2016
1,858 DE 6
02/2017
259 AT 1
09/2017 FI
05/2018
2,247 patients

30. procedures (pattern of use)
adult patients (N= 824)
Off-label procedures 1
N = 662 N
535
Missing Data (md)
127
Single procedure including aortic surgery
115 (21.50%)
56%
Single procedure excluding aortic surgery
185 (34.58%)
2 procedures including aortic surgery
99 (18.50%)
33%
2 procedures excluding aortic surgery
78 (14.58%)
3 procedures including aortic surgery
39 (7.29%)
11%
3 procedures excluding aortic surgery
19 (3.55%)
Majority of patients treated off-label (80%)
The off-label procedures have been stratified following the Eroscore II classification
Countries
Isolated on-pump CABG
Other procedures
Europe
162
662 UK
120 (16,42%)
611 (83,58%)
Sweden
9 (15%)
51 (85%)
Germany
32 (100%)
0 (0.00%)
Austria
1 (100%)
Most common off-label surgery consisted of single non-CABG procedure (300/535, 56%)
47% (235/535) of off-label procedures included ‘aortic surgery’

31. mortality rates in on label indications
mortality: isolated CABG
study
mortality definition
mortality rate
NAPaR
in-hospital mortality
1.41% (4/142)
Royston et al. Eur J Cardiothorac Surg 2009; 36:
in-hospital death or death within 30 days of the initial surgical hospitalisation
2.4% (21/862)
Wang et al. Ann Thorac Surg 2010; 89: 1489 – 95
1.2% (51/4,122)
Deloge et al. Eur J Anaesthesiol 2017; 34:
2.4%
mortality rate in patients treated within the official indication was comparable to or even lower than that reported in other studies