TRALICases and Chapter Session

Tanya Petraszko MD FRCPCCBS BC & YukonJanuary 2010

What is Transfusion-related Acute Lung Injury?

Sudden onset of “Acute Lung Injury” occurring within 6 hours of a transfusion

Acute Lung Injury Hypoxemia New bilateral chest X-ray infiltrates No evidence of volume overload

Canadian Consensus Conference Definition. Kleinman et al. Transfusion 2004;44:1774-89

Features of TRALI

SOB and abnormal CXR Fever, hypotension

onset is usually within the first 1-2 hours of transfusion, but may be delayed up to 6 hours

Normal CXR Patient’s CXR

TRALI Mortality

TRALI appears to be the most common cause of transfusion-related mortality

Reports to the FDA of transfusion-related deaths: TRALI > bacterial sepsis > acute hemolysis

Estimated rate of fatalities is 5 – 10%

Transfusion Related Fatalities Reported to FDA 2003-2005

Cause #cases (% of fatalities) TRALI 72 (32%) Bacterial sepsis 25 (11%) AHTR 23 (10%) Other 71 (32%) Not clearly tx related 34 (15%) Total 225 (100%)

Leading Cause of Adverse Transfusion Events reported to Health Canada

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118 113 112

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1 / 105

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Evolution of Transfusion RisksHIV and HCV vs TRALI

Estimated TRALI fatality (?)

Estimated TRALI risk

Adapted from Goodnough L and Aubuchon J

WBC Antibody Hypothesis Antigen-Antibody reaction triggers TRALI

Activation

Lodged in pulmonary capillaries

Recipient WBC- Neutrophils- Lymphocytes- Monocytes

Donor-Anti-HNA-Anti-HLA I-Anti-HLA II

Release of substances causing pulmonary endothelial damage and capillary leak

Courtesy of Y. Lin

Clinical Correlation

Right, single lung transplant

10 weeks after transplant, transfused PRBCs for anemia

HLA B44 antibody in PRBC donor

HLA B44 antigen present in lung donor tissue only

Elegant clinical example of targeted endothelial injury from a blood transfusion

Dykes et al. Br J Haematol 2000.

What evidence is there for this

Many studies demonstrate HLA or leukoagglutinating alloantibodies in donors

Several examples of donor-recipient antigen/antibody concordance

Anti HLA class I and II implicated Anti-neutrophil antibodies implicated

Evidence for antibody theory ex vivo isolated rabbit lung model severe lung vascular leakage was

reproduced using a human anti-neutrophil antibody in the presence of cognate, human 5b-positive neutrophils

In contrast, no vascular leakage was noted in lungs perfused in the absence of either antibody, neutrophils, or a complement source.

No permeability increase occurred with the use of 5b-negative neutrophils

Seeger Blood. 76:1990.

Antibody theory animal models isolated perfused rat lungs perfused them with plasma containing anti HNA-2a mAb and

human neutrophils. If neutrophil expression of HNA-2a antigen was >70%, ALI

was manifest; if antigen expression was <30%, ALI was not manifest. However, if the lungs were primed with fMLP then lung

injury was induced in the group with < 30% antigen expression.

Thus, if there is no relevant first hit (priming with fMLP), TRALI may not be manifest even with cognate antibody unless the cognate antigen expression meets a certain threshold.

also demonstrated that anti-HNA 2a directly activates neutrophils in the absence of complement

Sachs Blood. 107(3): 2006.

Anti-neutrophil Antibodies (HNA-2a)

Sachs et al, Blood 2006.

+FMLP

Slide from Looney MD, UofC, SF

Non antibody mediated TRALI

15% of typical TRALI cases fail to demonstrate antibody

alternate hypothesis that some cases may be non-antibody mediated

Biologically active molecules contained in stored blood products lysophosphatidylcholines interleukins

What evidence is there for this Rats pre-treated with LPS developed acute lung injury

(ALI) when the lungs were perfused with plasma Rats did not get ALI with LPS alone

Rats pretreated also got ALI with lipids extracted from stored 42 day old human RBCs or platelets

Did NOT get ALI if given lipids from 5 day product thus plasma or lipids from fresh, human RBCs or

platelets did not cause lung injury.

Silliman CC J. Clin.Invest 101(7): 1998

Silliman CC Transfusion. 43: 2003

Two Event Model

Recipients of blood containing anti WBC antibodies don’t all get TRALI

perhaps two events required to manifest TRALI (as in ARDS) Predisposing clinical condition Infusion of biologically active mediators

First event results in activation of pulmonary endothelium with neutrophil priming

Second event results in activation of the neutrophils adherent to pulmonary endothelium and subsequent lung injury

Two event hypothesis

Silliman et al. Blood 2003;101:454-62Kleinman et al. Transfusion 2004;44:1774-89

1st event: Underlying clinical condition of patient (inflammation, infection, surgery)

Activation of pulmonary endothelium with increased adhesion molecules

2nd event: Transfusion of biologically active lipids or antibodies

Activation

Release of substances causing pulmonary endothelial damage and capillary leak

Evidence from animal models 2-event in vivo rat model first event

saline (NS) or endotoxin (LPS)

second event plasma from stored, human packed red blood cells

(PRBCs) or antibodies (OX18 and OX27) against rat MHC Class I

antigens NS treated rats did not develop TRALI

With plasma nor with antibody LPS treated rats developed ALI

With plasma and with antibody

Kelher Blood. 113(9):2009.

Neutrophil depleted rats had experiment

repeated and all failed to demonstrate ALI Confirmed that ALI is neutrophil dependent

also demonstrated that anti-MHC Class I localized to the neutrophil surface

They concluded that TRALI is the result of 2 events with the examples of the second event being plasma from stored blood and antibodies that prime neutrophils.

Case of recurrent TRALI52 year old female 4 months post allo BMT for myelofibrosis; transfusion dependent

2007-02-10 received 2 units red cells uneventfully as an outpatient. Discharged but became acutely SOB on way home,

returned to ER CXR showed bilateral new infiltrates Decreased sats, intubated and ventilated for 30 hours then

extubated and recovered.

Results of Investigation Recipient results: post- sample: Negative

Donor 1st unit negative Donor 2nd unit positive for anti HLA antibody

Crossmatch donor 2 with recipient positive

Case of recurrent TRALI52 year old female 4 months post allo BMT for myelofibrosis; transfusion dependent

2007-03-19 Had been discharged home receiving regular outpatient

transfusions 2x/week after transfusion of 2 units of RCC developed new onset

SOB, new CXR infiltrates and PO2<60. Required mechanical ventilation.

Results of Investigation Recipient results: pre-samples: positive anti HLA post-sample: positive anti HLA No crossmatch done

Donor 1st unit positive anti HLA antibody Donor 2nd unit positive anti HLA antibody

Two event model

In first TRALI event, implicated donor associated platelet unit did not cause TRALI in the recipient it was given to

That recipient was not primed or did not have cognate antigen….

Evidence from animal models in vivo mouse model BALB/c wild-type mice positive for MHC

Class I antigen were injected with MHC Class I mAb via the jugular vein and sacrificed at 2 hours.

Control mice were knock out mice negative for the antigen

Injection of MHC Class I mAb produced severe ALI in wild type mice with 50% mortality compared to controls negative for the cognate antigen.

Looney MRJ. Clin. Invest. 116:(2006)

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MHC I mAb

MHC I antibody produces severe ALI in mice

Evidence from animal models

• The MHC Class I mAb binds throughout the body and prominently in the lung microvasculature.

• The investigators demonstrated that this antibody also binds to neutrophils, but does not cause direct neutrophil activation.

Looney MRJ. Clin. Invest. 116:(2006)

Evidence from animal models Mice were also neutrophil depleted with GR-1 antibody

these mice were protected from lung injury. Thus the absence of circulating neutrophils renders the bound

antibody on lung tissue inert and unable to cause vascular permeability.

Using a knockout mouse it was further demonstrated that it was the Fc gamma receptor which was essential in responding to the antibody challenge.

The Fc gamma receptor knock out mice were fully protected from lung injury, however when wild type neutrophils were transfused back, the lung injury was restored.

Thus in this model, MHC Class I antibody binds to class I antigen on the endothelium then presents the Fc portion of the antibody to the neutrophil Neutrophil is then activated thru Fc gamma receptor engagement.

demonstrated a critical role for neutrophil Fc gamma receptors in mediating ALI.

Looney MRJ. Clin. Invest. 116:(2006)

Summary so far Clinical features of TRALI Leading cause of TR mortality Majority appear to be antibody

mediated Animal models support this

So….why don’t we always see TRALI when antibody and cognate ag are present?

Effect of environment on priming Mice were housed in specific-pathogen free

barrier conditions vs. non-barrier housing Repeated TRALI model

Barrier mice were protected from TRALI non barrier mice did develop ALI.

Barrier mice demonstrated lower levels of circulating neutrophils suggesting that this is the reason why they did not

respond to the second hit since this model is neutrophil dependent.

if barrier mice were primed with LPS prior to receiving anti-MHC Class I mAb, then they did develop ALI in a dose-dependent manner.

Looney. J Clin Invest. 119(11): 2009

Anti-neutrophil Antibodies (HNA-2a)

Sachs et al, Blood 2006.

+FMLP

Slide from Looney MD, UofC, SF

Role of neutrophils in ALI microfluidics model of the lung microcirculation. MHC Class I mAb is immunoadsorbed to the artificial

capillary bed, and the unbound antibody is removed by washing.

Next, Fc gamma receptor knockout neutrophils or wild-type neutrophils are perfused.

video demonstrates that Fc gamma receptor knockout neutrophils zip right through the capillaries as they cannot engage the bound antibody

wild-type neutrophils all get stuck as they engage the bound antibody through their Fc gamma receptor.

Effect of hematopoetic cells in priming hematopoietic chimeras TLR4 mice, which

are LPS unresponsive TLR4 mice reconstituted with TLR4 bone

marrow were protected in the two-event model of TRALI (LPS + MHC Class I mAb).

However, when TLR4 mice were reconstituted with wild-type bone marrow, lung injury was manifest

thus demonstrating that hematopoietic cells are the critical population that must be primed to produce lung injury.

Looney. J Clin Invest. 119(11): 2009

Conclusions from animal models Well developed animal models support the

two event hypothesis of TRALI. effective ‘second hits’ in TRALI

Neutrophil or MHC Class I antibodies or plasma or lipids from stored, human RBCs

Priming of hematopoietic cells is necessary in experimental TRALI and may be the ‘first hit’.

Common to all experimental models, neutrophils are the key effector cell in TRALI.

How can we prevent TRALI?

We have evidence that antibodies are responsible for TRALI in a primed host

How can we identify a primed host? Fresher product for the primed host?

How can we prevent TRALI? TRALI reduction strategies have focused on

donors with antibodies and specifically plasma containing products

Secondary – Measures taken to identify the cause of a reaction that has occurred and prevent it from happening again

Primary – Preventive measures taken to eliminate the risk before it happens in the first place

How can we prevent TRALI Secondary…defer implicated donors

Donors associated with a reported TRALI are investigated for anti HLA and anti neutrophil antibody and deferred if positive

Pilot 2001 – 2006, SOP 2006 Primary…avoid donors with anti HLA or anti

neutrophil antibody what have blood suppliers done and what

has the effect been Donor loss Reduction of TRALI

Which donors develop WBC antibodies?

WBC alloimmunization may occur following previous exposure to WBCs through pregnancy or transfusion 332 female plateletpheresis donors 17% had detectable anti-HLA antibody Frequency of HLA antibodies increased with

pregnancy:

0 pregnancies: 7.8%1-2 pregnancies: 14.6% 3 or more: 26.3%

Densmore et al. Transfusion 1999;39:103-6

Primary Prevention 2003 – UK changed component production

policy moving to predominantly male plasma for transfusion

2003 2004 2005

TRALI 36 23 23

Highly likely/probable

22 13 6

Chapman et al. Vox Sang 2006;91(Suppl 3):227

SHOT data 10 years hemovigilance in UK and impact of

preferential use of male donor plasma Risk of highly likely/probable TRALI due to

FFP decreased 15.5 per million units 1999-2004 3.2 per million units issued 2005-2006 (p=

0.0079) Risk of highly likely/probable TRALI due to

platelets decreased 14 per million to 5.8 per million

Chapman.Transfusion 2009;49:440-452

TRALI reduction measures CBS-predominantly male plasma

CBS moved to predominantly male plasma for transfusion Oct 2008

Female plasma diverted fractionation

Year 2001 2002 2003 2004 2005 2006 2007 2008 2009 Q1

Total 6 6 6 16 14 21 35 19* 1

Tor 4 6 6 10 6 10 26 5 1

Edm 1 0 0 3 3 1 0 0* 0

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TRALI the CBS Experience – Definite and Possible

BC B

C &

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SMH: GIFT, LIFT, LCT, X-Match when possible CBS:Luminex HLA

TRALI Reduction Measures – Platelet Apheresis

On July 20, 2009 CBS started asking all female platelet donors if they’d ever been pregnant

Any responding positively redirected to whole blood

Donor Loss YTD

June 2009 there were 2394 female apheresis platelet donors

By Nov 2009 only 300 active female apheresis platelet donors

32% of the remaining donors converted to whole blood or plasma (approx 765 donors)

Net loss of approx 1329 donors

BC&Y Platelet Imports and Issues (Doses)

0.0 K

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2.5%

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17.5%

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Imports 650 1,335 2,344 3,642 2,559

Issues 12,829 14,318 15,599 17,412 17,719

% Imports/Issue 5% 9% 15% 21% 14%

2004/05 2005/06 2006/07 2007/08 2008/09

F2008/09 reversed the trend of large rises in the number of platelet imports.

TRALI – Platelet Apheresis In BC, LVPs allowed CBS to continue to meet

platelet demand with fewer imports

The number of platelet apheresis units collected since implementation of the TRALI reduction measures has not decreased, in fact increased with 38% growth

BC Yukon data National growth: 6% National data: LVP target 34.5% actual YTD

23.4%

Summary Primary Prevention

Reports of TRALI have decreased since shift to male plasma

Consistent with experience elsewhere Donor loss due to deferral of

previously pregnant females largely mitigated by collection of double platelets

Will there be enough snow for the Olympics??

Questions?