Pathogeninaktivierung
Wolfgang R. Mayr
Blutspendewesen
Blutsicherheit
Spenderauswahl
Produktion
Spendertestung
PI
Klinische Indikation QM
Hämovigilanz
www.roteskreuz.at
Blutspendewesen
2
Infektionsrisken
Diagnostisches Fenster
Bekannte Viren ohne Testung
Unbekannte (emerging) Viren
Bakterien
Protozoen
Jeder Erreger, der während der
asymptomatischen Phase der Infektion, im
Blut vorkommt, kann via Transfusion
übertragen werden
www.roteskreuz.at
Blutspendewesen
3
Testung (BSZ W,N,B)
1950
01. 1973
02. 1979
07. 1985
01. 1989
07. 1990
04. 1995
02. 1999
06. 2003
10. 2003
AB0, RH, Ery-Ak, Lues
AuSH Ag
HBs Ag
HIV Ak
ALT
HCV Ak
Neopterin
NAT – HBV, HCV, HIV
NAT – Parvo B19
NAT - HAV
www.roteskreuz.at
Blutspendewesen
4
Restrisiko 2009
HCV, HIV
1 : 5.000.000
HBV
1 : 500.000
(okkulte HBV Infektionen)
www.roteskreuz.at
Blutspendewesen
5
www.roteskreuz.at
Blutspendewesen
6
www.roteskreuz.at
Blutspendewesen
7
www.roteskreuz.at
Blutspendewesen
8
Pathogeninaktivierung - PI
Fensterperiode – Viren mit Test
Unbekannte Viren
Bakterien
Protozoen
Leukozyten (TA GvHD)
www.roteskreuz.at
Blutspendewesen
9
Leukozyten
Zell-assoziierte Viren (CMV, EBV, ...)
Febrile Transfusionsreaktionen NHFTR
HLA / HNA Alloimmunisierung
Transfusions-assoziierte Immunmodulation
TA GvHD
www.roteskreuz.at
Blutspendewesen
10
Pathogeninaktivierung
Wirksamkeit
Klinik
Pharmakokinetik
Toxizität
Toxizität
Akut
Mehrfachanwendung
Subchronisch
Reproduktiv
Genotoxizität
Carcinogenität
Phototoxizität
Venenirritation
www.roteskreuz.at
Blutspendewesen
11
www.roteskreuz.at
www.roteskreuz.at
Blutspendewesen
13
Schroeder DD &
Mozen MM,
Science 1970; 168:
1462-1464
www.roteskreuz.at
Blutspendewesen
14
Prion-removal capacity ...
(Thyer J et al, Vox Sang 2006; 91: 292-300)
www.roteskreuz.at
Blutspendewesen
15
PI – stabile Blutprodukte
Pasteurisierung (60°C, 10 h)
Erhitzen (trocken, feucht)
S/D (solvent-detergent)
Nanofiltration
www.roteskreuz.at
Blutspendewesen
16
Burnouf T,
Transf Med Rev 2007;
21, 101-117
www.roteskreuz.at
Blutspendewesen
17
PI - FFP
Pool
S/D inaktiviert „Octaplas“
Einzelspender
Quarantäne
PI durch Amotosalen
PI durch Methylenblau
PI durch Riboflavin
www.roteskreuz.at
Blutspendewesen
18
S/D FFP versus EinzelspenderFFP
FFP
S/D FFP
Quarantä
Quarantäne FFP
PI Einzelspender
FFP
Standardisierung
(Volumen, Proteine,
Zellgehalt, ...)
Hoch
Gering
Gering
Umhüllte Viren
Sicher
Okkulte HBV
Sicher
Nicht-umh. Viren
B19, HAV
NAT
Nicht getestet
Relativ sicher
„Emerging“ Viren
Pool
Einzelspender
Relativ sicher
www.roteskreuz.at
Blutspendewesen
19
PI - labile Produkte
Thrombocyten
Hauptproblem: bakterielle Infektion
www.roteskreuz.at
Blutspendewesen
20
Blajchman MA et al, Transfus Med Rev 2005; 19: 259-272
Division of
Blood Group
REDUCTION OF SEPTIC TRANSFUSION
REACTIONS WITHOUT BACTERIA DETECTION
(Andreu et al, ISBT Science Series 2008; 3: 124-132)
•
Reduction of initial bacterial contamination
–
–
–
–
•
Hygiene, skin desinfection
Diversion of the first 35 ml
Postdonation information
Donor selection
Reduction of bacterial growth
– Practice for delivery
– Universal leukodepletion
– Storage of whole blood before processing (2 – 20 h)
•
Improvement of knowledge and practice on bacterial
contamination
– Management of possible bacterial infection
– Research on bacterial contamination
Division of
Blood Group
Bacterial screening of apheresis platelets
(ARC 2004-2006)
Eder AE et al,
Transfusion
2007; 47: 113411341142
Division of
Blood Group
te Boekhorst PAW et al, Transfusion 2005; 45: 514-519
Division of
Blood Group
Screening platelet concentrates for bacterial
contamination: low numbers of bacteria and
slow growth in contaminated units mandate
an alternative approach to product safety
Murphy WG et al, Vox Sanguinis 2008; 95: 13-19
Conclusions:
Screening platelet concentrates for bacterial
contamination using the most sensitive method
available has a sensitivity of less than 40% because
of the low numbers of bacteria in the initial
contamination. Effective resolution of this problem will
require a pathogen-inactivation technique.
Division of
Blood Group
PI – labile Blutprodukte:
Thrombocyten
Verhinderung der DNA Replikation:
Photosensitizer + Licht
Verhinderung der DNA Replikation:
UVC (254nm) Behandlung
www.roteskreuz.at
Blutspendewesen
26
Nucleic Acids Must “UnZip”
During Pathogen Replication
DNA / RNA
Department for
Blood Group
Strand Separation
Replication of
Nucleic Acids
and Pathogen
Blood Donation Center for Vienna, Lower Austria and
Burgenland,
Austrian Red Cross
Amotosalen
Mechanism of Action
UVA Illumination
Amotosalen
DNA or RNA
of pathogen
Docking
Department for
Blood Group
Permanent Crosslinking
Blood Donation Center for Vienna, Lower Austria and
Burgenland,
Austrian Red Cross
INTERCEPT Blood System for Platelets Buffy
Coat Preparation
Buffy Coat
Platelets
1
InterSol
Solution
Pooling
Container
2
Whole Blood
Separation
Department for
Leukodepletion
Filter
3
Pooling and Addition
of InterSol Solution
Centrifugation
to Pellet Red
Blood Cell
4
Separation
of Platelets
Blood Donation Center for Vienna, Lower Austria and
Burgenland,
Austrian Red Cross
INTERCEPT Blood System
Amicus Platelet Preparation
Amicus enables the
automatic collection of
one or more therapeutic dose(s)
of SDP in 35% plasma
and 65% InterSol Solution.
Department for
Blood Group
Blood Donation Center for Vienna, Lower Austria and
Burgenland,
Austrian Red Cross
INTERCEPT Blood System for Platelets Process Timeline
Start Processing
Within
one day
of collection
1
Department for
Blood Group
Illuminate
Removal & Storage
Incubate platelets
3
on the CAD for a minimum of
for
4 hours: SDP
~3-6 minutes 6 hours: buffy coat platelets
J/cm2
2
3
Blood Donation Center for Vienna, Lower Austria and
Burgenland,
Austrian Red Cross
Mirasol PRT System Primary Modes of Action
The Mirasol PRT System inactivates disease-causing agents by
altering their nucleic acids in two primary ways:1
1. UV light only: reversible inactivation
UV light alone breaks chemical bonds in the
nucleic acids of pathogens
2. UV light + riboflavin: irreversible inactivation
Riboflavin molecules form complexes with nucleic acids
UV light from the Mirasol Illuminator activates the
riboflavin molecule in the complex
Photoactivated riboflavin induces a chemical alteration to
the functional groups (such as guanine bases) of nucleic
acids making pathogens unable to replicate
1
Kumar et al 2004
Mirasol PRT System Performance
Reduction of active
pathogen load
Typical
performance
Viruses
~2–6 log
(enveloped, nonnon-enveloped;
intracellular, extracellular)
(99.0–99.9999%)
Bacteria
~2–5 log
(Gram +, Gram -)
(99.0–99.999%)
Parasites
Inactivation of white
blood cells
White blood cell inactivation
≥ 3.0 to ≥ 5.0
(≥ 99.9% to ≥ 99.999%)
Typical
performance
≥ 6.0
(>99.9999%)
References
Ruane, et al., 2004; Goodrich, et al., 2006a,b;
CaridianBCT data on file.
Ruane, et al., 2004; Goodrich, et al., 2006b; CaridianBCT
data on file.
Cardo, et al., 2006; Cardo, et al., 2007;
Rentas, et al., 2007; Tonnetti, et al., 2007;
Sullivan, et al., 2008.
References
Fast, et al., 2006a. (in-vitro)
Cytokine production and
expression
prevented
Fast, et al., 2006a. (in-vitro)
GraftGraft-versusversus-Host Disease
prevented
Fast, et al., 2006b. (animal model)
Alloimmunization &
transplant rejection
prevented
Asano, et al., 2007. (animal study)
UVC Behandlung
(Mohr H et al, Transfusion 2009; 49: 1956-1963)
Beutel 19 x 38 cm
Thrombocyten: in 350 ml (109 / ml)
Bestrahlung mit UVC (254 nm)
Agitation (100 rpm, horizontal)
www.roteskreuz.at
Blutspendewesen
34
UVC Behandlung
Mohr et al, Transfusion 2009; 49: 1956-1963
www.roteskreuz.at
Blutspendewesen
35
PI bei Thrombocyten
Stramer et al, Transfusion 2009; 49: August Suppl 1S – 29S
www.roteskreuz.at
Blutspendewesen
36
Amotosalen: Apherese- und BCThrombocyten
CCI nach 1 und nach 24h geringer als bei
nicht behandelten Thrombocyten
Hämostaseologische Funktion nicht verändert
Keine unerwünschte Nebenwirkungen
www.roteskreuz.at
Blutspendewesen
37
Amotosalen: Hämovigilanz
www.roteskreuz.at
Blutspendewesen
38
Red Cell Processing
S-303
CAD
Collected Red
Cells with
Additive Solution
Department for
Blood Group
Mixing
Container
Reaction
Container
Final
Storage
Container
Blood Donation Center for Vienna, Lower Austria and
Burgenland,
Copyright 2001 – Cerus Corporation
Austrian Red Cross
INTERCEPT Blood System for
Red Blood Cells
Proposed Process Timeline
Start Processing
Treatment
Removal and
Storage
S-303
CAD
Collected red blood
cells with additive
solution;
Leukodepleted
Incubate
Remove
Reconstitute
red blood cells
residual
S-303 and mix
and S-303
compound
with red blood cells at 19-25°C for at 19-25°C for
12 hours
8 hours
------- Development of an automated process is on-going -------
1
Department for
Blood Group
2
3
Blood Donation Center for Vienna, Lower Austria and
Burgenland,
Austrian Red Cross
PI of RBC (S-303, Inactine, ...)
Treated red cells demonstrated acceptable recovery and comparable
comparable
lifelife-span to conventional red cells
Repeated transfusions of autologous treated red cells did not provoke
provoke
autoauto-antibody formation
PostPost-transfusion recovery after repeated transfusions of autologous
treated red cells was comparable to that of untreated red cells and to
that of single exposures of treated red cells
Full unit transfusions of treated red cells were well tolerated
? Production of antibodies against treated RBC
www.roteskreuz.at
Blutspendewesen
41
Lublin DM, Transfusion 2000; 40: 1285-1289
www.roteskreuz.at
Blutspendewesen
42