PATHOGEN INACTIVATION

IN DONATED BLOOD

HEALTH TECHNOLOGY ASSESSMENT SECTION

MEDICAL DEVELOPMENT
i DIVISION
MINISTRY OF HEALTH MALAYSIA
018/2010
DISCLAIMER

Technology review is a brief report, prepared on an urgent basis, which draw on restricted
reviews from analysis of pertinent literature, on expert opinion and / or regulatory status where
appropriate. They are not subjected to an external review process. While effort has been made to
do so, this document may not fully reflect all scientific research available. Additionally, other
relevant scientific findings may have been reported since completion of the review.

Please contact: [email protected], if you would like further information.

Health Technology Assessment Section,

Medical Development Division
Ministry of Health Malaysia
Level 4, Block E1, Precinct 1
Federal Government Administrative Centre
62590 Putrajaya

Tel: 603 88831246

Fax: 603 8883 1230
Available at the following website: http://www.moh.gov.my

ii
Prepared by:

Ms Mariammah Krishnasamy
Assistant Director (Scientific officer - Microbiology)
Health Technology Assessment Section
Ministry of Health Malaysia

Reviewed by:
Datin Dr Rugayah bt Bakri
Deputy Director
Health Technology Assessment Unit
Ministry of Health Malaysia

DISCLOSURE
The authors of this report have no competing interest in this subject and the preparation of this
report is totally funded by the Ministry of Health, Malaysia

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EXECUTIVE SUMMARY
Introduction
Several pathogen inactivation methods are available such as Chemical treatment, Photochemical,
Leukocytes depletion, Ultraviolet B irradiation and others. Worldwide numerous infectious
agents, have been identified as potential threats to blood supply. Newly discovered agents
namely transfusion transmitted virus (TTV), SEN-V (SEN virus), Human herpes virus-8 (HHV-
8), Hepatitis G Virus (HGV), West Nile Virus and Prions present a unique challenge in assessing
the possible risk to safety of blood and plasma products. Current blood donor screening and
disease testing have shown to reduce the incidence of blood transfusion-transmitted diseases.
Pathogen inactivation or reduction technology represents a proactive approach to blood safety.
The ultimate goal of pathogen inactivation is to maximally reduce the transmission of potential
pathogens without significantly compromising the therapeutic efficacy of the cellular and protein
constituents of blood. This must be accomplished without introducing toxicities into the blood
supply, neoantigen formation and subsequent antibody production. Investigations into some
technologies are at an advanced preclinical or already in clinical stages. Evidently, the use of
such procedures raises a number of issues, including efficiency, the damage inflicted to labile
blood products, toxicity in patients, and cost-effectiveness.

This review was requested by a Senior Consultant Pathologist, Y. Bhg. Datin Dr. Nor Azian
Azlan A. Samad from Pathology Department of Melaka Hospital

Aims/objectives
To assess the safety, efficacy, effectiveness, and cost effectiveness of pathogen inactivation or
pathogen reduction technology used to treat blood products such as plasma, platelet and red
blood cells (RBC) for safe blood and blood products.

Results and conclusion

A. PLASMA
Safety
i) Photochemical Treatment (Amotosalen)
Fair level of evidence on the safety aspect of photochemical treatment (Amotosalen) used
for pathogen inactivation in plasma.
ii) Chemical treatment (Solvent/ detergent [SD-treated])
Evidence showed SD (treated) - plasma caused some deaths in liver transplant, severe liver
disease and known coagulapathies patients
iii) Methylene Blue
Poor evidence on the safety aspect of Methylene Blue technology used in plasma products.
There was no long term studies conducted on carcinogenicity and reproductive toxicity in
Methylene blue treated plasma.
Effectiveness
i) Psoralen light treatment (Amotosalen- HCL)
Fair evidence on the effectiveness of INTERCEPT blood system when used on Platelet-
plasma.
ii) Riboflavin light treated (RLT)
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 Poor evidence on the effectiveness of Riboflavin light treated (RLT) technology when used
in plasma. Clinical studies using Mirasol (riboflavin) pathogen inactivation technology is
still underway.
iii) Solvent Detergent (SD- plasma)
Fair evidence on the effectiveness of solvent detergent technology, when used for plasma
and fresh frozen plasma.
Cost Effectiveness
i) SD- FFP transfusion was cost effective in patient’s ≤ 48 years of age and in older patients
with good clinical prognosis.
ii) In another baseline study on cost effectiveness of transfusing virus-inactivated plasma
instead of standard plasma showed a cost-effectiveness ratio of $2,156,398 +/- $257,587 per
quality-adjusted life year gained. The transfusion of virus-inactivated plasma produces little
health benefit at a very high cost.
iii) There is no retrievable evidence on cost effectiveness for other pathogen inactivation
technologies used for plasma.
Recommendation
Based on the above review:
Photochemical treatment (Amotosalen) and Solvent detergent (SD- plasma) may be
recommended as pathogen inactivation technology for plasma.
However, SD – treated plasma is not recommended for patients after liver transplant, severe
liver disease and with known coagulopathies.
Insufficient evidence to recommend Methylene Blue and Riboflavin light treated
technologies as pathogen inactivation technologies for plasma. Hence, more clinical
research is warranted before considering the technologies for plasma or fresh frozen plasma.

B. PLATELETS
Safety
i) Amotosalen and Mirosal (Riboflavin)
Good evidence was obtained on the safety aspect of Amotosalen (photochemical treatment)
and Mirosal (Riboflavin) used as pathogen inactivation technology for platelets. There was
good patients tolerance for platelets treated with amotosalen. Both Amotosalen and Mirosol
technologies did not affect the quality of platelets concentrates.
Effectiveness
i) Amotosalen and Mirosal (Riboflavin)
Fair evidence was obtained with regards to the effectiveness of Amotosalen (photochemical
treatment) and Mirosal (Riboflavin) pathogen inacativation technology for platelets
ii) Thionine/UV Light
Low level of evidence on the effectiveness of Thionine/UV Light used as pathogen
inactivation technology for platelets.
Cost effectiveness
i) Amatosalen
Sufficient evidence to show that Amatosalen (photochemical technology) is cost effective
compared with non treated plaletets used for transfusions.
Recommendation
Based on the above review:-
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 Amotosalen and Mirasol (Riboflavin) pathogen inactivation technologies can be used for
platelet concentrates.
More research is warranted on Thionine /UV Light when used as pathogen inactivation on
platelet concentrates.

C. RED BLOOD CELLS (RBCs)

Safety
i) S 303
Fair level of evidence to show the safety of S303 when using as pathogen inactivation
technology for RBC.
Effectiveness
i) S 303
Fair evidence obtained on the effectiveness of S303 as a pathogen inactivation technology
when used on RBC.
ii) PEN110
Poor evidence for the effectiveness of PEN110 as a pathogen inactivation technology when
used on RBC.
Cost effectiveness
No retrievable evidence on the cost effectiveness of S303 and PEN110 pathogen
inactivation technologies when used for RBC.

Methods
This review was based on two Health Technology Assessment (HTA) reports, a World Health
Organisation guideline, seven Randomised control trials (RCT), two laboratory experiment study
design, two cost effectiveness studies, three narrative reviews and an abstract on pathogen
inactivation or pathogen reduction technologies.

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PATHOGEN INACTIVATION IN DONATED BLOOD
1. INTRODUCTION
The approach to reduce pathogens in the blood supply depends on a combination of activities
such as donor education; donor selection (volunteer and nonremunerated); donor screening,
direct questioning on risk behaviour; testing for selected known causative agents; consulting
lists of previously deferred donors; meticulous examination, and others. Blood is still not a
safe product for many reasons. Most of the screening techniques cannot detect the pathogens
in the blood sample during the “window period” of a disease. Pooling of blood fractions from
many asymptomatic infected blood donors, in order to raise product yield, can raise the
infection rates 1. There is also possibility of introducing a new emerging pathogen into the
blood supply, for which no contemporary test exists. A very good example to illustrate this is
the transmission of the Human Immunodeficiency Virus (HIV) in the late 1970's and early
80's, to thousands of haemophiliac patients (Corash, 1998). To further reduce the risks of
pathogen transmission via blood transfusion pathogen inactivation steps have been routinely
applied to labile blood product such as fresh frozen plasma (FFP) and blood cellular
components. Several pathogen inactivation methods are available such as solvent/ detergent
treatment of FFP, Methylene treatment of FFP, Leukocytes depletion, Ultraviolet B
irradiation and Gamma Irradiation. 1 Numerous infectious agents (found worldwide) have
been identified as potential threats to blood supply. These newly discovered agents namely
transfusion transmitted virus (TTV), SEN-V (SEN virus), Human Herpes virus-8 (HHV-8),
Hepatitis G Virus (HGV), West Nile Virus and Prions present a unique challenge in
assessing the possible risk on safety of blood and plasma products. This makes pathogen
inactivation even more important.2 Current blood donor screening and disease testing has
shown to reduce the incidence of transfusion-transmitted diseases. Pathogen inactivation of
blood products represents a proactive approach to blood safety.3 The ultimate goal of
pathogen inactivation is to maximally reduce the transmission of potential pathogens without
significantly compromising the therapeutic efficacy of the cellular and protein constituents of
blood.
This must be accomplished without introducing toxicities into the blood supply and without
causing neoantigen with subsequent antibody production. Investigations into some
technologies are at an advanced preclinical or already in clinical stages. Evidently, the use of
such procedures raises a number of issues, including efficiency, the damage inflicted to labile
blood products, the toxicity in patients, and the effectiveness.

This review was requested by a senior consultant pathologist, Y.Bhg. Datin Dr. Nor Azian
Azlan A. Samad from Pathology Department of Melaka Hospital. This review was requested
in order to obtain information regarding safety and effectiveness of pathogen reduction or
inactivation technologies used on donated blood.

2. OBJECTIVES

To assess the safety, effectiveness, efficacy, and cost effectiveness of pathogen inactivation
or pathogen reduction technology used to treat blood products such as plasma, platelet and
red blood cells (RBC) for safe blood and blood products.
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 3. TECHNICAL FEATURES
Pathogen reduction technologies on blood products (such as plasma, platelets and red blood
cells (RBCs) can be categorized into three categories namely Chemical treatments,
Photochemical treatment and photodynamic treatments.

There are several pathogen inactivation technologies used in Plasma, Platelets and RBCs.
Pathogen inactivation techniques used for plasma includes the Solvent detergent, Amotosalen
+ UV, Methlene blue + Light, and Riboflavin. As for platelets the pathogen inactivation
technologies used are the Amotosalen+ UV, Riboflavin + UV and UV (± thionine). Inactine,
S303, Riboflavin + UV, and thiopynilium +light are used as pathogen inactivation
technologies for red blood cells. The status of pathogen inactivation technologies for blood
products (such as plasma, platelet and red blood cells) are described in Table 1.

Table 1: Status of pathogen inactivation technologies

Blood Company Method US status EU status

Component
Plasma Octapharma Solvent detergent Licensed Licensed
Cerus Amotosalen +UV Phase III Licensed
Macopharma Metylene blue + Light - Licensed
Navigant Riboflavin +UV - Licensed

Platelet Cerus Amotosalen +UV Phase III Licensed

Navigant Riboflavin +UV Phase II Licensed
Macopharma UV (±thionine) Preclinical Preclinical

Red Cells Vitex Inactine Abondoned Abondoned

Cerus S-303 Phase II Licensed
Navigant Riboflavin +UV Preclinical Licensed
America red cross Thiopyrilium + light Preclinical Licensed

Note: Phase 1 volunteer safety studies, Phase II: Patient safety studies. Phase III:
Patient efficacy studies. Licensed: by FDA in USA and/or CE marked in Europe .4

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3.1 PLASMA
Photochemical treatment
3.1.1 Methylene Blue (MB)
First-generation MB systems involved the addition of an MB stock solution to each thawed
plasma unit for a final concentration of 1μM. Then the MB-treated plasma is subjected to a
white light luminescence of 45000 lux or more for 1 hour. A more advanced commercially
manufactured MB system by MacoPharma (Mouvaux, France) uses an inline system
consisting of a membrane filter, MB dye, illumination bag, elimination filter, and storage
bag. The 0.65-μm membrane filter removes platelets, leukocytes, and debris. The plasma
then passes through tubing containing an MB pill that dissolves as the plasma flows through
the tubing into the illumination bag. The MB-containing plasma is subjected to double-sided
illumination by sodium high-intensity low-pressure lamps emitting yellow light at a
wavelength of 590 nm for 15 to 20 minutes. Plasma is then passed through an MB
elimination filter that removes greater than 95% of the residual dye and photoderivative
byproducts (especially phenothiazide dyes azure A and B) that exhibit in vitro mutagenic
properties. The Macro pharma system is CE marked and implanted under good
manufacturing practices( GMP) in several europian countries. However, a question still
remains concerning the genotoxicity of MPB and may introduce long term side effects. 5

3.1.2 Psoralen light treatment (PLT- Plasma) or Amotosalen Ssystem

A photochemical treatment system using a novel psoralen material (S-59, developed by Cerus
and Baxter Corporations) and UV-A (long wavelength) ultraviolet light, has been developed
to inactivate viruses in single donor units of Fresh frozen Plasma (FFP). Upon exposure to
UVA, the S-59 forms non-reversible covalent monoadducts with DNA and RNA. This system
can be used to treat single 250-300 mL plasma units or jumbo units containing 500-600 mLs
of plasma. During the photochemical treatment with psoralen-UVA, individual units of
plasma are either separated from whole blood or collected by apheresis technology and
transferred into a plastic container. S-59 is added to a concentration of 150 μM through an
integral closed system into a UV-A transparent vessel containing the plasma. After five
minutes incubation, the S-59 treated plasma is illuminated with shaking for three minutes in a
high intensity UV-A-light box. After illumination process the plasma is transferred to another
container with an S-59 reduction reduction device (SRD), which is used to remove most of the
photoproducts caused by light activation of the S-59. After S-59 reduction device processing
for 1 hour, the plasma is transferred via a closed system into a final container for freezing at
18˚C and can be stored for up to 1 year.

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 INTERCEPT, THE S-59 AMOTOSALEN SYSTEM

3.1.3 Riboflavin light treated plasma (RLT-plasma)

Another Method applies essential natural vitamins (Vitamin B2 or riboflavin) that absorb both
visible lights (the Mirasol system). Riboflavin and UV light (280- 360) damage nucleic acids by
direct electron transfer, production of singlet oxygen and generation of hydrogen peroxide with
formation of hydroxyl radicals hence damage the DNA/RNA of pathogens and it can proceed in
the absence of oxygen. Currently, riboflavin is classified as a Generally Regarded As Safe
(GRAS) compound by US FDA. 5

3.1.4. Solvent Detergent (SD)

SD viral inactivation technologies were first licensed for treatment of clotting factor concentrates
in 1985. Before 1984, a combination of ethyl ether (20%) and Tween 80 (1%) were found
effective in inactivating HBV and NANBHV in factor VIII and IX concentrates with minimal
loss in activity. In 1986, TNBP and 0.2% sodium cholate were shown to be effective at
inactivating HBV, NANBHV, and HIV.
Solvent detergent (SD) treatment disrupts the membranes of enveloped virus, bacteria,
and eukaryotes. Infectivity is lost via this action. Many combinations of tri-(n-butyl)- phosphate
(TNBP) or ethyl ether (organic solvents) with Tween 80, sodium cholate, and Triton X-100 (non-
ionic detergents) have been used in the past. The SD combination most often used now is TNBP
with Triton X-100.This compound acts as an organic solvent and removes lipids during SD
processing by extracting and sequestering them into a separate micellar (colloidal) phase. This
non-ionic detergent disrupts lipid bilayers for easier extraction and stabilizes TNBP. 6

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3.2. PLATELET
Different techniques are available to inactivate pathogen in cellular blood components suc as for
platelets. The techniques include psorelans (Amotosalan) , methylene blue, porphyrins, UV light,
Gamma irradiation and riboflavin (RB)

3.2.1 Psorelan
Amotosalen hydrochloride is synthetic psorelan compound that intercalates into helical region of
DNA or RNA after illumination with ultraviolet ray light reacts with pyrimidine bases to form
inter nucleic and intra nucleic acid stands crosslink. This is a fix bond and thus the strands of
DNA cannot separate and replicate.

3.2.2 Riboflavin (RB)

UV light has been used as a pathogen inactivation method since it can damage nucleic acids.
However, UV light alone has not proven to be an effective approach to pathogen inactivation of
cellular components such as platelets. Therefore, RB in combination with UV light in the 265-
370nm wavelength is being developed as a clinical method for pathogen inactivation of several
blood components including platelets. It is believed that RB acts by intercalation with nucleic
acids and when exposed to light causes damage to DNA by production of singlet’s oxygen,
electron transfer, production of hydrogen peroxide and hydroxyl radicals that lead to nucleic acid
base damage and DNA strands breaks.7

3.2.3 PORPHYRINS
Porphyrins are “a large class of deeply coloured red or purple, fluorescent crystalline pigments,
with natural or synthetic origin, having in common a substituted aromatic macrocyclic ring
consisting of four pyrrole-type residues, linked together by four methine bridging groups”. All
porphyrin-like compounds have a strong absorption band around 400nm called band. Porphyrins
have applications in a variety of novel chemical and medicinal procedures including the use of
water-soluble porphyrins in photo dynamic theraphy. It was claimed that photosensitizers have
been used for detection and treatment of cancer. It is also an effective modality against antibiotic
resistant bacteria and cell free viruses.

Photodynamic interactions were described to take place wherever sensitizer, light and oxygen are
simultaneously present. Inflammatory tissue was described to manifest similarities in porphyrin
retention and therefore bacterial and viral infected tissues may become targets for photodynamic
treatment. It is independent of the antibiotic sensitivity spectrum of the treated pathogen and it
has an efficient and non-recovering anti-microbial killing effect upon illumination of Gram
positive bacteria. Bacterial PDT is affected by the use of various sensitizers, as a general rule
non-charged or positively charged molecules are effective in photoinactivation of
Staphylococcus sp. In order to photosensitive Gram (-) bacteria such as Pseudomonas
aeruginosa, it is necessary to introduce a small peptide polymyxin-B nona-peptide (PBNP) which
stimulate the translocation of porphyrin through the outer membrane of these bacteria and makes
PDT possible. Gram negative cell killing by the use of PBNP and DP broadens the antibacterial

11
spectrum of photodynamic inactivation and opens new horizons for this modality as a wide
spectrum drug when antibiotic resistance is the main concern8

3.3. Whole Blood and Red Blood Cells (RBC)

3.3.1 S- 303
Pathogen Inactivation techniques used for RBC includes the S-303. This method is developed by
the same manufacturer as amotosalen. S-303 is a part of class of compounds known as Frales
(Frangible Anchor Linker Effectors). These tripartite molecules are composed of a nucleic acid
anchor, an effector moiety, and a frangible linker. The effector moiety binds nucleic acids
covalently, and the frangible linker breaks down forming an inactive negatively charged species
(S-300). This prevents further binding to the nucleic acid, thus rendering the DNA or RNA non-
functional. The completely light-independent reaction proceeds due to a shift in pH, which
occurs as the S-303 is added to red blood cell concentrates. The process as developed requires
the addition of S-303 to red cells at a concentration of 150 mg/mL, which is then incubated for
12 hours at room temperature. The red cells are incubated with a compound absorption device
(CAD) for an additional 8 hours after transfer to a storage container to remove any residual S-
303 and S-300. The CAD remains in the red cells throughout storage. 6

3.3.1 PEN110
PEN110 is another pathogen inactivation compound that also disrupts nucleic acids. Like S-303,
light is not required for the reaction and it can therefore be used to treat red cells. PEN110 is a
compound chemically related to binary ethyleneimine and is known as an ethyleneimine
oligomer. The altered molecule is a cation, selective for nucleic acids, and highly watersoluble.
As the molecule is very small, it readily diffuses through cell membranes, which, the
manufacturer claims, makes it more effective against non-lipid enveloped viruses, such as
parvoviruses. PEN110 forms ionic bonds with nucleic acids, activating the molecule by
protonation of the aziridino nitrogen. The active form can then alkylate a proximal nucleophilic
center such as the N7 position of guanine in DNA. This results in the opening of the imidazole
ring structure of guanine, which creates a break in the strand and creates a stop message. The
nucleic acid is thus inactivated and becomes useless as a template. Binary ethyleneimines, like
psoralens, have been used in nucleic acid research since the 1980s. The development and use of
PEN110 for pathogen reduction of blood products has occurred only over the past 15 years. In
the process known as INACTINE treatment, PEN110 is added to create a 0.1% (vol/vol)
concentration and incubated at room temperature for 6 hours.

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PEN110 is then removed by washing with unbuffered saline to levels below the limit of
detection when using HPLC (30 ng/mL). The units are then ready for storage.6

4. METHODOLOGY

4.1 SEARCH METHODS

Literature were searched through electronic databases which included Medline, PubMed,
Cochrane Database for Systematic Review, EBM Review HTA Database, INAHTA database
and general databases such as Google and Yahoo.

The search strategy used the terms, which were either used singly or in various combinations:
“Pathogen Inactivation”, “white blood cells”, “Pathogen Reduction”, “Pathogen Inactivation
AND Blood” ,“Blood products” platelets, plasma and “Red Blood Cells”. The search was limited
to articles on human. There was no language limitation applied in the search.

4.1.1 SELECTION OF STUDIES INCLUDED /EXCLUDED

All the studies pertaining to safety, efficacy, effectiveness and cost effectiveness of pathogen
inactivation or pathogen reduction technology in donated blood were included in this review.
A critical appraisal of all relevant literature was performed using Critical Appraisal Skills
Program (CASP) checklists and the evidence graded according to the US/Canadian Preventive
Services Task Force (Appendix 1)
Data were extracted and summarized in evidence table as in Appendix 2. The data were not
pooled and only non-qualitative analysis was carried out.

5. RESULTS AND DISCUSSION

This review was based on two Health Technology Assessment (HTA) reports, a World Health
Organisation guideline, seven Randomised control trials (RCT), two laboratory experiment study
design, two cost effectiveness studies, three narrative reviews and an abstract on pathogen
inactivation or pathogen reduction technologies.

A. PLASMA
5.1 . A. SAFETY
Photochemical treatment (Amotosalen)
INAHTA briefs by Catalan Agency for Health Technology Assessment and Research-CAHTA,
Spain (CAHTA), a systematic review revealed that pathogen inactivation technique using
Amotosalen (photochemical treatment) is safe for plasma products.9

A phase III Randomised control trial (RCT) conducted by Mintz et al in 2006 was reviewed by
Health Technology Assessment by Canadian Agency for Drugs and Technologies in Health
(CADTH). A randomised, controlled, double-blind Phase III trial was conducted with
Photochemical treatment of fresh-frozen plasma (FFP) with amotosalen and ultraviolet (UV)
(PCT FFP) or control FFP for therapeutic plasma exchange (TPE) in patients with thrombotic

13
thrombocytopenic purpura (TTP). Thirty five patients were randomly assigned to receive plasma
(n=17) or control plasma (n=18). These patients received plasma until a remission (defined as a
platelet count of 150 x 109/L for two consecutive days) was achieved or for a maximum of 35
days. Those patients were observed for adverse events (AEs) for seven days after the last
transfusion. This study found that there were no significant difference between groups in terms
of the time to remission, relapse rate, time to relapse, total volume of plasma exchanged and
number of plasma unit exchanged. The author concluded that the safety profile of PCT- treated
FFP did not differ significantly from the control FFP. This study received funding from Cerus,
the manufactures of INTERCEPT. 10 level 1
Chemical Treatment (Solvent/detergent-treated)
World Health Organisation (WHO) guidelines reported that based on two small RCTs
(Williamson LM et al. 1999 and Lerner RG et al. 2000) and two clinical trials (Inbal A et al.
1993 and Horowitz MS et al. 1998) showed that solvent/detergent (SD)-treated-plasma can
replace FFP in all of its indications. These include the replacement of coagulation factors and in
the treatment of thrombotic thrombocytopenic purpura. However, several deaths were reported
in liver transplant patients who received a product provided by one manufacturer in the USA.
Although the link with this product or with reduced levels of some anticoagulant proteins in SD-
plasma is uncertain, the US manufacturer’s product label has been amended to indicate that this
product should not be used in patients undergoing liver transplant or in patients with severe liver
disease and known coagulopathies. 11 Level III
Methylene blue (MB)
A narrative review paper by Bryant BJ. et. al. 2007, indicated that millions of Methylene blue
(MB) single units of plasma have been transfused in Europe without unexpected adverse
outcomes. Methylene blue–treated plasma has replaced S-D plasma in Belgium and is the sole
plasma product used in the United Kingdom for patients born after January 1, 1996. However,
the long-term effects of exposure to even minute amounts of residual MB and its photo
derivatives have not been studied in larger cohorts. To date, no adverse effects have been
reported. Long-term studies of carcinogenicity and reproductive toxicity have not been
conducted. 12 Level III

5.2. A EFFECTIVENESS
PLASMA
Psorelan light treatment (Photochemical)
Baxter/ Cerus (INTERCEPT blood system) is a method applying amotosalen HCL (Formally
known as S-59) and long wave length ultraviolet (UVA) for plasma pathogen reduction. Only
two small randomised controlled trials were published as abstracts, have been performed
comparing PLT-Plasma and FFP. One of these is the patients with acquired coagulopathy,
including liver transplant and one involving exchange for thrombocytopenic purpura (TTP).
They demonstrated a similar effect for platelet –plasma and traditional FFP. In patients with
congenital coagulation deficiencies, recoveries were within the therapeutic range for FII, FVII,
FIX and FX after transfusion of 15-20mL. PLT-plasma per kg, while t1/2 was reduced for F1, FII
and FXII; both PT and APTT values showed significant correction after transfusion.5 Level III

Riboflavin light treated plasma (RLT-Plasma)

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Another method applies an essential natural vitamin (Vitamin B12 or riboflavin) that absorbs
both visible and UV light( the Mirasol system). In this narrative review paper by Solheim
BG, 2006, revealed that Riboflavin and UV light (280-360nm) could damage nucleic acids
by direct electron transfer. The production of singlet oxygen and generation of hydrogen
peroxide with formation of hydroxyl radicals hence damage to pathogen DNA/RNA. This
process has been proven effective against a wide range of pathogens, including bacteria,
intracellular HIV-1, West Nile virus (WNV) and porcine parvovirus in preclinical studies of
platelets and plasma. However the clinical studies using the Mirasol PRT for plasma are still
underway. 5 Level III

Pooled Solvent Detergent plasma (SD- plasma)

In vitro procedures have demonstrated that the SD method eliminates all lipid-envolped
viruses, including West Nile virus except for vaccinia virus which demonstrates a relative
resistance. The poxvirus family has unique structure which exhibits a general high
physiochemical resistance. The SD- plasma production process also eliminates bacteria,
protozoa cells and cellular fragments. It has no effect on nonlipid coated viruses, B19 and
Hepatitis A virus (HAV), have been dispelled by screening of the strating material with
Nuclear amplification technology. A reduction of the prion load which has been observed
with SD production process is still under further investigation. The same SD method has
been applied for all SD-plasma products. There are differences in in manufacturing processes
which resulted in some important differences in product conmposition. In Europe, the
production process patented by OctapharmaAG (Lanhen, Switzerland). In US, SD-plasma
was produced under the brand name PLAS+SD by Vitex(Watertown, MA). Nine studies
covered the indication for plasma examined the clinical efficacy and tolerance of SD-
plasma. It was administered to 376 patients. Eight of the studies were European SD- plasma,
while one observational study was with US SD-plasma. Four of the studies were prospective
randomised trials, three SD-plasma versus Methylene blue light treatement- plasma(MBLT-
plasma). It was concluded that SD-plasma was proved as effective as FFP or MBLT-plasma.
5 Level III

5.3. A COST EFFECTIVENESS

There was a cost effectiveness study to estimate the incremental cost/life year saved for SD-
FFP compared with untreated FFP, having controlled for post-transfusion mortality and
patient age. Various infective and non infective transfusion –related complication were
considered. The findings showed that the discounted cost/life year saved for SD-FFP use in
the UK was £22 728 [95%, CI: £22 604- 22853] for neonates and £98 465 (95%CI:£97 924-
99 005) for patient age 70. The cost effectiveness ratio was below £50 000/life year saved
for patients ≤ 48 years of age and below £30 000/life year saved for those ≤21 years of age.
In transfusion recipients with no significant morbidity, the cost –effectiveness ratio was £12
335 for neonates and £61 692 for 70-years old. The most important driver of cost-
effectiveness was transfusion-related acute lung injury (TRALI), on account of its relatively
high incidence and mortality rate. Inclusion of non- infectious complications suggests that
SD-FFP is cost effective in patients ≤48 years of age and in older patients with good clinical
prognosis. 13

15
 The cost effective study by Pereri et al. 2000, investigated the costs and utilities of
transfusing virus-inactivated plasma. Probability distributions for patients' age and sex and
for the number of blood components transfused were determined in 924 plasma recipients in
a tertiary-care hospital. Other values were obtained from the medical literature. Results of the
baseline and sensitivity analyses are the mean (+/- SD) of 10 simulations with 10(7) patients
per simulation.

In the baseline analysis, transfusing virus-inactivated plasma instead of standard plasma

prolonged the quality-adjusted survival by 1 hour and 11 minutes per patient, at a cost-
effectiveness ratio of $2,156,398 +/- $257,587 per quality-adjusted life year gained. Cost-
effectiveness was most sensitive to the patients' mean age, the incremental cost per unit of
virus-inactivated plasma, the HIV and hepatitis C virus transmission rates, and the short-term
mortality of plasma recipients due to their underlying diseases. The finding showed that
compared to most accepted medical procedures, the transfusion of virus-inactivated plasma
produces little health benefit at a very high cost. This poor cost-effectiveness ratio is due to
the low current risk of infection with transfusion-transmitted viruses and to the greater age
and poor short-term prognosis of most plasma recipients. 14

6.0. A CONCLUSION
PLASMA
Safety
i) Photochemical Treatment (Amotosalen)
Fair level of evidence on the safety aspect of photochemical treatment
(Amotosalen) used for pathogen inactivation in plasma.
ii) Chemical treatment (Solvent/ detergent [SD-treated])
Evidence showed SD (treated) - plasma caused some deaths in liver transplant, severe
liver disease and known coagulapathies patients
iii) Methylene Blue
Poor evidence on the safety aspect of Methylene Blue technology used in plasma
products. There was no long term studies conducted on carcinogenicity and
reproductive toxicity in Methylene blue treated plasma.
Effectiveness
i) Psoralen light treatment (Amotosalen- HCL)
Fair evidence on the effectiveness of INTERCEPT blood system when used on
platelet-plasma.

ii) Riboflavin light treated (RLT)

Poor evidence on the effectiveness of Riboflavin light treated (RLT) technology when
used in plasma. Clinical studies using Mirasol (riboflavin) pathogen inactivation
technology is still underway.
iii) Solvent Deteregent ( SD- plasma)
Fair evidence on the effectiveness of solvent detergent technology, when used for
plasma and fresh frozen plasma.

16
Cost Effectiveness
i) SD- FFP transfusion was cost effective in patient’s ≤ 48 years of age and in older
patients with good clinical prognosis.
ii) In another baseline study on cost effectiveness of transfusing virus-inactivated plasma
instead of standard plasma showed a cost-effectiveness ratio of $2,156,398 +/-
$257,587 per quality-adjusted life year gained. The transfusion of virus-inactivated
plasma produces little health benefit at a very high cost.
iii) There is no retrievable evidence on cost effectiveness for other pathogen inactivation
technologies used for plasma.

7.0. A RECOMMENDATION
Based on the above review:
Photochemical treatment (Amotosalen) and Solvent detergent (SD- plasma) may be
recommended as pathogen inactivation technology for plasma.
However, SD – treated plasma is not recommended for patients after liver transplant,
severe liver disease and with known coagulopathies.
Insufficient evidence to recommend Methylene Blue and Riboflavin light treated
technologies as pathogen inactivation technologies for plasma. Hence, more clinical
research is warranted before considering the technologies for plasma or fresh frozen
plasma.

B. PLATELET
5.1.B SAFETY
INAHTA briefs by Catalan Agency for Health Technology Assessment and Research-
CAHTA, Spain (CAHTA) also reported that pathogen inactivation treatment using
Amotosalen (photochemical treatment) is also safe to be used in platelets concentrates. The
results from active hemosurveillance subsequent to its commercialisation showed good
patient (adult and pediatric) tolerance in 14 493 transfusions of platelet concentrations treated
with amotosalen.9

Health Technology Assessment by Canadian Agency for Drugs and Technologies in Health
(CADTH), reviewed the pathogen reduction technologies (PRTs) of INTERCEPT Blood
System and Mirasol. INTERCEPT was licensed for use in Europe in 2006 and Mirasol
received European approval in 2008. These technologies are currently not licensed in North
America. Both systems used a photochemical treatment (PCT) approach which combines the
blood products with photoactive chemical (amotosalen) for INTERCEPT and [riboflavin
(vitamin B2)] for Mirasol. Both photochemical processes were claimed to inactivate
pathogens without adversely affecting the quality or safety of blood products. 10 level 1

The Randomised Controlled Trial (RCT) by Ambruso et al (2009) had compared Mirasol
treated platelet with control platelets (no treatment with PRT). Patients involved were all had
a confirmed diagnosis of thrombocytopenia due to chemotherapy or heamatopoetic stem cell
transplantation. The serum of 44 patients received Mirasol- treated platelets and 22 patients
received controlled platelets. Patients over 28-day study period was analysed for the
presence of neoantigens. No patients in either group had antibodies to platelet neoantigens.
17
Therefore the author concluded that neoantigen formation is not a side effect of the Mirasol
treatment. The immunereactivity of Mirasol-treated platelets was similar to that of control
platelets. However, it was indicated that this study was funded by Mirasol manufactures.10
level 1

CADTH also reviewed a prospective cohort study on the safety aspect of INTERCEPT-
treated platelet (amotosalen +UVA) components in a broad patient population. Blood
transfusion centres using the INTERCEPT Blood System for routine production of platelet
components were invited to participate in the study. A total of 5,106 transfusions with
INTERCEPT –treated platelets were performed in 651 patients over a period of
approximately two years. Patients with heamatological or oncological conditions accounted
for the majority (58.1%) of the participants. The mean number of transfusion per patients was
7.8 ±16.2 (SD) with a range of one to 156. Adverse events (AEs) were reported in 55
transfusions [1.1%; 95% CI; 0.81 to 1.40] corresponding to 42 patients (6.4%). AEs
classified as related to transfusion included chills, fever, uticaria (hives), dyspenea (shortness
of breath), nausea or vomiting, itching, flushing and hypotension. In this study, three AEs
were reported as serious. The author reported that AEs occurred after a single transfusion or
after multiple transfusions. Therefore, the authors suggested that repeated exposure to
INTERCEPT-treated platelets did not increase the risk of transfusion- related reaction.
INTERCEPT-treated platelets were well- tolerated and that the AEs profile of the patients
receiving INTERCEPT – treated platelets was consistent with other studies evaluating
conventional prepared platelets. However, this study did not have a control group receiving
untreated platelets. This represents a limitation of the study. Furthermore this study received
a funding from the manufactures (Cerus) and two of the authors were employed by Cerus. 10
level 1

Another RCT, double blind, intention to treat analysis conducted (in vivo phase II study) by
Van Rhenen et. al. 2003, between June 1998 to 2000 (EURO SPRITE trial) on
photochemical treatment (PCT) for platelets. One hundred three patients received either
standard control platelets prepared by buffy coat method of buffy coat prepared platelets
treated with amatosalen for pathogen inactivation. Fifty-two patients received 390
transfusions of amotosalen-treated platelets and 51 patients received 286 transfusions of
control platelets. All patients were evaluated following platelet transfusion for transfusion-
related symptoms and signs. Post transfusion events classified as acute transfusion reactions
included fever, chills, nausea, skin rash, urticaria, bronchospasm, tachycardia, hypotension,
hypertension, hemoglobinuria, hemolysis, and change in vital signs within six hours
following platelet transfusion. In cycle 1, 6% of test transfusions were associated with acute
reactions, compared with 5% of reference platelet transfusions (p ≥ 0.61). A total of 147
types of adverse events were reported in 5% or more of the patient population evaluated
during cycle 1 and cycle 2. Overall, there was no difference in the incidence of adverse
events by system organ class between the two treatment groups. Serious adverse events were
reported in 14 (27%) patients randomised to the test platelet treatment group and in 13 (25%)
patients randomised to the reference platelet group. Adverse events reported as related to
platelet transfusion were not statistically significantly different between the groups (p=0.20).
No study transfusions demonstrated transfusion-associated bacteremia. Four test patients and
five reference patients died while on study. No deaths were related to study transfusions

18
adverse events. Based on peritransfusion hemostatic assessments, 71% of test and 63% of
reference patients had at least one episode of bleeding before transfusion (p=0.36). A high
and equal proportion of patients had at least one hemorrhagic adverse event. The most
frequent hemorrhagic adverse events were epistaxis (42% test vs 41% reference), gingival
bleeding (17% test vs 12%reference), injection site hemorrhage (17% test vs 6% reference),
purpura (15% test vs 14% reference), petechiae (13% test vs 16%reference), and hematoma
(13% test vs 8% reference). Only 6% of patients in each group experienced severe
hemorrhagic adverse events. Bleeding also was assessed indirectly by comparing the number
of red blood cell units transfused during the transfusion periods. Based on this clinical trial,
pooled platelet components prepared with PCT and stored for up to 5 days offer the potential
to reduce transfusion-associated infections and inactivate residual contaminating leukocytes
using a processing system that is compatible with the current method of preparing therapeutic
doses of buffy coat platelets. 15 Level 1

PLATELETS
5.2 B. EFFECTIVENESS

PHOTOCHEMICAL TREATMENT
A) Amotosalen system, S-59 + UVA
HTA report by CADTH, included the SPRINT trial (randomised control phase III trial)
which evaluated the clinical effectiveness of PCT (amatosalen+ UVA) treated platelets in
patients with thrombocytopenia. In this RCT, the patients were randomised to receive either
PCT-treated platelets (n=318) or control platelets (n=327). There were no significant
differences in the baseline patient’s characteristics. The patients received platelets until they
became independent of transfusion (seven days without transfusions) or for a maximum of 28
days. Before receiving transfusion, platelet counts of the group did not differ significantly
(PCT group = 15.1 x 109/L, control group = 15.2 x 109/L). Patients receiving transfusions
with PCT- treated platelets had lower post- transfusion platelet counts than patients in the
control group (36.5 x 109/L versus 49.5 x 109/L). Besides that patients in the PCT group had
a shorter interval time between transfusions than the control group (8.4 versus 6.2). The
author noted the mean dose of platelets per transfusion was lower in PCT group compared
with the control group [3.7 x 1011 platelet per mililitre (mL) versus 4.0 x 1011 platelets/L].
Despite the differences in the secondary outcome the author concluded that the effectiveness
of PCT platelets was similar to that of control platelets. This study was funded by Cerus, the
manufactures of INTERCEPT. 10 Level 1
In another RCT double blind, intention to treat analysis conducted by Van Rhenen et al 2003
between June 1998 to 2000 (euroSPRITE trial.) The study population was patients with
thrombocytopenia or receiving therapy expected threshold of <20x109) gave informed
consent and were randomised to receive transfusion support. A total of 103 patients (52 test
and 51 references) received at least one study transfusion.The test platelets which was pooled
concentrates were leukoreduced by filtration followed by photochemical treatment (PCT)
with 150µM S-59 and 3 J/cm2, ultraviolet A (UVA) treatment (Helinx Technology; Cerus
Corperation, Concord CA) for pathogen and leukocyte inactivation. The targeted platelet
content of reference pooled components was 3.0 x 1011 platelets. During the cycle 1,
19
311(80%) of 390 test transfusion and 256(90%) of 286 reference transfusion were prepared
according to protocol. The mean total platelet dose of on-protocol transfusion for both
treatment groups and reference group was similar (test mean dose 21.3x1011 vs. reference
mean total dose 21.2 x10 11). PCT– treated and non- PCT – treated platelets gave comparable
24-hour post transfusion platelet counts for equal doses in cycle 1 and post transfusion
platelet counts for patients enrolled in cycle 2. Limited number of patients enrolled in cycle
2 (10 test and 2 reference patient). The treatment groups did not differ significantly with
respect to the 1- hour platelet count (p=0.47). The estimated effect of PCT was an increase in
the one- hour platelet count of 4x 109/L. The interaction terms for PCT and dose were not
significant (p=0.47). Similarly, the treatment groups did not differ significantly with respect
to the 24-hour platelet count in cycle 2 (P =0 .90). The estimated effect of PCT was an
increase in the platelet count of 1 x109/L (95% CI, -10 to 11 x 109/L). PCT treated and non-
PCT-treated platelets provided comparable 1- hour and 24-hour platelet counts for equal
doses in cycle 2. The data support the conclusion that pooled buffy-coat platelet components
treated with the S-59 pathogen inactivation device and stored for up to 5 days were
comparable to conventional platelet components used for transfusion in thrombocytopenic
patients. 15 Level 1

Kerikhoffs JL et. al. 2010, conducted a multicentre open label randomised and non-
inferiority trial. This trial compared the clinical effectiveness of buffy-coat derived
leucoreduced platelet concentrates (PC) stored for up to 7 days in plasma with platelets
stored in platelet additive solution III (PASIII) without and with amotosalen HCI/ ultraviolet-
A (UVA) photochemical pathogen reduction (PR-/PASIII).

Based on the abstract of this study, the primary endpoint evaluated was 1-h corrected count
increment (CCI), secondary endpoint was 24-h CCI, bleeding, transfusion requirement on red
cells and PC, platelet transfusion interval and adverse transfusion reactions. Compared to
plasma-PC in the intention to treat analysis of 278 evaluable patients the mean differences for
1-h CCI of PR-PASIII-PC and PASIII-PC was -31% (p<0.0001) and -9%(P= n.s)
respectively. Twenty-seven patients (32%) had bleeding events in the PR- PASIII arm, as
compared to 19(19%) in the plasma arm and 14(15%) in the PASIII arm (p=0.034). Despite
the potential advantages of pathogen (and leukocyte) inactivation of amotosalen-HCI/UVA-
treated platelet products, their clinical efficacy is inferior to platelets stored in plasma,
warranting a critical reappraisal of employing this technique for clinical use. 16 Level 1

In vitro studies have found that after processing through the INTERCEPT Blood System,
there is no insignificant drop (in the range of 8-10%) in the platelets present in the bag. This
was due to the manipulation of platelets through three different bags, and agitation needed to
mix the amotosalen with platelets and to expose amotosalen and photoproducts to the
compound absorption device. In the Moog et al study, single-donor platelet concentrates
contained mean platelet counts of 3.06±0.27x1011. On completion of the photochemical
treatment, the mean count was 2.62±0.27 x 1011, representing a loss of 9.7%. Also in the
Moog study, significant decrease in pH was identified after PCT, with a mean drop to 6.98±
0.08.

20
Compared to the control, however this pH level was well above the required 6.8 limit
required in Europe, and the pH remained above 6.8 until the end of the study, at 5 days of
storage. 6 level III

Another in Vitro study looked at PCT platelet survival following indium-III radiolabelling of
the treated autologous platelets. The normal span of platelets survival is broad ranging from 5
to 7.3 days within 33-66% recovery. Recovery in PCT treated was 43%±8.7% and life span
was 4.8±1.3 days. Although within the lower range of normal and significantly lower than
controls, the findings were consistent with other in vitro studies. The author found that, in
vitro studies, the label dissociation was higher in PCT platelets than in control platelets. 6 level
III

MIRASOL riboflavin (vitamin B2)

A multicenter, open-label, parallel-group noninferiority RCT was conducted in France by the
Etablissement Francais du Sang and university hospitals (n = 6). This study compared PRT-
PLTs (MIRASOL) and standard (reference) platelet products transfused to thrombocytopenic
hematology and/or oncology patients. The aim of this clinical trial was to determine whether
pathogen-reduced PLTs (PRT-PLTs) are as effective as standard untreated platelet products.
The corrected count increment 1 hour post transfusion (CCI1hour) was evaluated in patients
with chemotherapy-induced thrombocytopenia. Safety information of PRTPLTs was also
documented on all adverse events. The primary efficacy outcome was the CCI 1 hour
measured 30 to 90 minutes post transfusion for each of a maximum of eight on-protocol
platelet transfusions per patient occurring within the 28-day treatment period. Six centers
enrolled 118 patients into the study between December 2005 and September 2007: 60
patients received PRT-PLTs and 58 received reference PLTs.
The pre-specified primary outcome analysis for the CCI 1hour was based on a maximum of
eight PLT transfusions per patient occurring in the 28-day treatment period: 258 for PRT-
PLTs and 209 for the reference group (total 467). The test for homogeneity of treatment
effects between sites for the CCI 1hour was not significant (p = 0.1728), indicating that data
from all sites could be pooled to estimate the treatment effect. The Least square (LS) mean
CCI 1hour in the PRT-PLT group was 11,725 (Standard error [SE], 1140) and in the
reference group was 16,939 (SE, 1149), a difference of -5214 (95% CI, -7542 to -2887; p <
0.0001).

The secondary outcomes of CCI 24hour was analysed according to the time-compliant and
extended time periods and adjusted for pre-transfusion platelet count as a continuous variable
and site. The test for homogeneity of the effect of treatment between sites for the CCI 24hour
was not significant (p = 0.1336) allowing for data to be pooled. The LS mean for time-
compliant CCI 24hour was 6676 (SE, 883) for the PRT-PLTs and 9886 (SE, 915) in the
reference group (difference, -3210; 95% CI, -5160 to -1260). The odds (OR, 0.284; 95% CI,
0.105 to 0.767; p = 0.0130) of achieving a successful response is significantly lower in the
PRT-PLTs arm for the CCI 1hour among time-compliant transfusions but not significantly
lower for the CCI 24hour among time compliant transfusions (OR, 0.481; 95% CI, 0.211 to
1.098; p = 0.0822). Similar results were found when considering transfusions within the
extended time period although the 24-hour CCI result becomes significant in this analysis.
Twenty-eight day treatment period was included in this analysis with the mean number of

21
 days between transfusions was 2.16 (SD, 1.69) for PRT-PLTs and 2.30 (SD, 1.48) for the
reference arm (p = 0.2903). The mean number of platelet transfusions per patient-day during
the treatment period (includes on and off-protocol transfusions) was not significantly
different: PRT-PLTs 0.24 (SD, 0.16) and reference group 0.20 (SD, 0.19; p = 0.2046). None
of the differences observed were significant. RBC requirements were similar in the two
groups.
In the PRT-PLT group 183 RBC units were transfused in the treatment and follow-up
periods: 155 were given in the treatment period with a mean (SD) per patient of 2.8 (1.7). In
the reference group 142 of 160 RBC units were given in the treatment group with a mean per
patient of 2.6 (2.4; p = 0.7257). Overall PLT and RBC utilisation in the two study groups was
not significantly different, suggesting that the lower CCIs with the PRT-PLTs did not
translate into significantly higher blood product use. Therefore the author concluded further
studies are needed to show whether the lower CCI observed with PRT-PLTs is associated
with any change in the risk of bleeding the noninferiority of PRT-PLTs compared to
reference platelets using the surrogate outcome measure of CCI 1hour. Safety data did not
identify any major adverse effects associated with the transfusion of PRT-PLTs. 17 Level 1

Goodrich et al looked at bacterial pathogen reduction in platelets in 2002. Riboflavin (50µM)

was added, and the unit was spiked to 5-6log/mL titers with S. epidermidis, Escherichia Coli,
S. areus, B. cereus, and K. pneumonia. Illumination time was with 6-7J/cm2 of UV light. A
low- titer experiment paralleled the above, spiking the units to 2 log/mL titers and using 7/
Jcm2 of UV light. Significant reduction occurred from 3.1 to 5.5 log/mL reduction. Even the
low titers were 100-1000 times higher than levels expected in platelets at the time of donation.
Thus, the phoactivation of Riboflavin appears to be adequate for pathogen reduction in blood
products. 6 level III

Thionine/UV Light
A narrative review by Bryant BJ et. al. 2007, indicated that thionine (phenothiazine dye) can
be safely used in place of MB with the same type low-pressure sodium UV light leading to a
comparable spectrum of pathogen inactivation. Measurements of platelet function such as
change in morphology, aggregation, and activation as well as biochemical parameters are just
slightly affected by thionine/UV light treatment. There is no change in storage duration after
treatment. Thionine with UV light treatment alone does not inactivate bacteria and leukocytes.
Inactivation of bacteria can be accomplished with an additional step consisting of UV-B
irradiation for approximately 2 to 4 minutes (1.2–2.4 J/cm2). The review claimed that an
added advantage of the extra UV-B irradiation step is more effective for viral inactivation.
However, the Thionine-based pathogen inactivation systems are currently under development
and in clinical testing. 3 Level III

5.4 B. COST EFFECTIVENESS

CADTH Health Technology Assessment report revealed information regarding the economy
evaluations for pathogen reduction technology (PRT). The study by Janssen et. al. 2006,
performed a cost –utility analysis which evaluated the cost- effectiveness of PRT (particular
brand was not specified) compared to an untreated sample diversion pouch. The cost-
effectiveness of these interventions was analysed after the introduction of the diversion pouch

22
during blood collection. The diversion pouch collects the first 20 to 30 ml of blood which
contains the highest level of bacteria. A hypothetical population of 100,000 patients
undergoing platelet transfusions was evaluated. This study took place in the Netherlands. The
price year was 2002, the discounting rate was 4%, and costs were reported in US dollars. The
estimation of quality –adjusted life years (QALYs) was derived using Markov model. The
author reported that the additional cost per QALY gained was $496,674 when using PRT with
the sample diversion pouch compared to sample diversion pouch alone. The results obtained
were most sensitive to the probability of sepsis, probability of death given sepsis, patients’
quality – adjusted life expectancy, and probability of bacterial contamination. Therefore it was
concluded that with the unknown probabilities, the cost effectiveness was uncertain.10 Level 1

Another study by Moeremas et. al. 2006, conducted a cost –utility analysis to evaluate cost
effectiveness of INTERCEPT Blood system. The study compared a hypothetical population
receiving INTERCEPT-treated platelets with a population received control platelets (prepared
conventionally). The study population was people with haematological malignancies, breast
cancer, and patients undergoing coronary artery bypass graft surgery. This study was
conducted in Belgium and the costs were reported in Euros. The price year and the rate of
discounting were not reported. It was indicated that with the assumption of an absence of an
emerging virus, the incremental cost- effectiveness ratio (ICER) ranged from €195,364 to
€3,459,201 per QALY. The author concluded that the implementation of INTERCEPT could
be considered cost-effectiveness. 10 Level 1

Similarly another study was by Postma et al, which was published in 2005. This study was
conducted in Netherlands and the costs were expressed in Euros. The price year was 2003 and
future costs were discounted at a rate of 4%. Two main assumptions were made, the PRT was
100% effective in eliminating pathogens and the PRT- treated platelets were not associated
with AEs. Additional sensitivity analysis was performed taking in account various
assumptions including risk of sepsis. The cost-effectiveness of PRT was €554,000 per life year
gained. The author concluded that PRT was cost- effective. 10 Level 1

There was also a cost-utility analysis comparing the INTERCEPT Blood System with control
single donor platelets. The study was conducted in Japan, by Staginnus and Corash, 2004. The
price year was not stated; however, resource use data was taken from studies that were
published between 2002 and 2004. The cost was presented in Japanese Yen. A one way
sensitivity analysis was performed. The cost per QALY gained of using INTERCEPT-treated
platelets compared with controlled platelet ranged from ¥ 1,076,000 (for hip arthroplasty) to ¥
99,000 (for the treatment of acute lymphocytic leukemia with progenitor cell transplantation).
The author concluded that the treatment of platelets with INTERCEPT Blood System was a
cost- effective treatment strategy compared with treatment strategies in Japan including the use
of nucleic acid testing to detect pathogens. 10 Level 1

23
6.0 . B CONCLUSION
Safety
i) Amotosalen and Mirosal (Riboflavin)
Good evidence was obtained on the safety aspect of Amotosalen (photochemical treatment)
and Mirosal (Riboflavin) used as pathogen inactivation technology for platelets. There was
good patients tolerance for platelets treated with amotosalen. Both Amotosalen and Mirosol
technologies did not affect the quality of platelets concentrates.
Effectiveness
i) Amotosalen and Mirosal (Riboflavin)
Fair evidence was obtained with regards to the effectiveness of Amotosalen
(photochemical treatment) and Mirosal (Riboflavin) pathogen inacativation technology for
platelets
ii) Thionine/UV Light
Low level of evidence on the effectiveness of Thionine/UV Light used as pathogen
inactivation technology for platelets.

Cost effectiveness
i) Amatosalen
Sufficient evidence to show that Amatosalen (photochemical technology) is cost effective
compared with non treated plaletets used for transfusions.
7.0. B RECOMMENDATION
Based on the above review:-
Amotosalen and Mirasol (Riboflavin) pathogen inactivation technologies can be used for
platelet concentrates.
More research is warranted on Thionine /UV Light when used as pathogen inactivation
on platelet concentrates.

RED BLOOD CELL (RBC)

5.1.C SAFETY
S-303
Jorge et. al. 2006, conducted a study which enrolled 42 subjects, in a two-arm, single
blinded RCT. In this study, subjects were assigned to receive either S-303 RBCs or control
RBCs. Subjects received a single transfusion of 10 to 15 mL of either 51Cr-labeled 35-day-
old S-303-treated RBCs (test) or sham-treated (control) 51Cr-labeled 35-day-old RBCs.
Radiolabeling and determination of 24-hour post transfusion recovery for all studies were
performed with established methods Phase 1B design. The second study was a single-arm
open-label trial to investigate whether repeated exposures to S-303-treated autologous RBCs
would lead to immune-mediated destruction of S-303 RBCs. All of the subjects from Phase
1A were invited to participate in the second trial. Twenty-eight subjects were enrolled.
Subjects donated whole blood that was treated with the same S-303 process used in Phase
1A and Phase 1C design. These studies were designed to examine several key aspects of
RBCs function in vivo. The first two trials (1A and 1B) measured post transfusion recovery
of S-303 RBCs after single and repeated exposures to assess viability and immune responses
to potential neoantigens.
24
In the third study (1C), RBCs were prepared with a compound adsorption device (CAD),
stored for 35 days, and median life span after transfusion was determined. Thirty-five days
after donation, aliquots of the autologous study RBCs were removed for radiolabeling and
transfused into subjects following the procedure of Moroff and coworkers. For all three
studies, 24-hour post transfusion recovery was determined by collecting samples at 5, 7.5,
10, 12.5, 15, 20, 30, and 60 minutes after transfusion and 24 hours after transfusion.
In Phase 1C, blood samples were collected from the subjects on the day of transfusion, 24
hours after transfusion, and for 5 weeks after each transfusion to determine circulating life
span based on the methods outlined by the International Committee for Standardization in
Hematology (ICSH). All adverse events were recorded from the start of the first RBC
transfusion through seven days after the last transfusion. The study found that in a Phase 3
trial of chronic transfusion support for patients with thalassaemia and sickle cell anemia,
detected antibodies with specificity to RBC treated with S-303, prepared with the first-
generation process, in two patients after repeated transfusions. Serum samples from these
two patients were nonreactive in an in vitro monocyte phagocytosis assay, previously shown
to correlate with RBC antibody physiologic activity. During the follow-up of patients
transfused with S-303 RBCs for support of cardiovascular surgery, two patients who had not
been exposed to S-303 RBCs were found to have transient low-titer antibodies with
specificity for RBC treated with S-303. Both the test and control treatment groups exhibited
mean post transfusion recovery greater than 75 percent. No evidence of immune responses
to S-303 RBCs after repeated exposures were detected by in vitro crossmatch assays with
autologous S-303 RBC, direct antiglobulin test (DAT) and antibody screen with a
commercial cell pan. RBC life span was calculated by three different methods: a linear
model, an exponential model, and a weighted mean linear-exponential model. RBC
recoveries for test and control RBCs were weakly correlated with the life span
measurements, correlation coefficients of 0.18 and 0.28, respectively. In preclinical studies,
S-303 RBCs prepared with up to 1.0 mmol per L S-303 and 10 mmol per L Glutathione
(GSH) demonstrated no toxicity in a wide range of in vitro and in vivo tests, including the
absence of carcinogenicity with prolonged exposure in p53-deficient mice that are highly
sensitive to carcinogens. It appeared that some subjects may form antibodies to S-303 RBC
or have preformed antibodies cross-reactive with S- 303 RBCs. The mean 24-hour recovery
of 35-day old S-303 RBCs was less than untreated RBCs, but greater than 75 percent. RBCs
treated with S-303 and stored for 35 days exhibited median life span not different from that
of conventional RBCs. 18 Level 1

5.2 C. EFFECTIVENESS
Red Blood Cells (RBCs)
S-303
A randomised, double-blind trial evaluated the clinical efficacy of RBCs treated with
pathogen inactivation with S-303, a synthetic labile alkylating agent. This was a phase III
clinical trial evaluating the efficacy and safety of S-303–treated RBCs compared to
conventional RBCs in the clinical setting of transfusion support for complex cardiac surgical
procedure. A sample size of 100 patients per group was randomised in a one-to-one ratio to
receive either S- 303–treated or control RBC transfusion during surgery and for 6 days
thereafter. The trial was terminated prematurely owing to detection of antibodies to S-303
25
RBCs in a companion trial of S-303–treated RBCs administered in the setting of chronic
transfusion support.

All transfused patients were asked to consent to a follow-up trial for further evaluation for
safety and detection of antibodies to S-303–treated RBCs. Hundred forty eight transfused
patients were equally distributed between the test and control groups in terms of
demographics and indications for surgery. One-hundred forty-one patients (95%) completed
the trial through Day 27; reasons for not completing the trial were death (3 patients), lost to
follow-up (2 patients), and other reasons (2 patients): first-study RBC transfusion received
after Day 6 (1 patient) and issues with transportation to study site (1 patient). Fifty-one
(34.5%) patients who received transfusions participated in the follow-up study: 27 (36.5%)
test patients and 24 (32.4%) control patients. Overall, 41 percent of all RBCs were transfused
during surgery. The mean change in patients’ heamoglobin (Hb) levels from baseline to the
end of study was a decrement of 2.4 g per dL, and the mean change after each transfusion
was an increment of 1.4 g per dL. Thirteen (8.8%) patients, seven in the test group and six in
the control group, received off-study RBC transfusions owing to the difficulty in maintaining
adequate inventory of study units at all sites and the unpredictability of RBC usage in this
setting. The mean number of off-study RBC transfusions in these 13 patients was 1.0. No
significant differences were noted between the test and control groups. The number of
patients transfused with other blood products (PLTs, fresh-frozen plasma [FFP], and
cryoprecipitate), and the mean number of units of other blood products transfused was similar
between treatment groups. Serum samples from 127 patients (64 tests and 63 controls) were
tested for antibodies to S-303–treated RBCs. One control patient had a positive result with S-
303 RBC. The patient’s serum sample was negative on day 32 but had a positive result 535
days after surgery; the reaction was 3+ positive and specific for S-303–treated RBCs in a
PEG assay. A competitive inhibition assay with S-300, a nonreactive S-303 derivative,
confirmed the specificity for S-303 RBC. A second follow-up sample taken on day 838 and
tested with the same protocol in a different reference laboratory was negative for S-303–
treated and conventional RBCs were equivalent with respect to clinical efficacy and safety in
supporting the transfusion needs of cardiac surgery patients. Investigations are under way to
ascertain the significance of S-303 RBC antibodies and to prevent their occurrence.19 Level I

PEN110
Extensive research has been conducted with regards to the pathogen inactivation capacity of
PEN110. These studies were very specific in demonstrating the exact nature of the action of
PEN110 on the virus and on the kinetics of inactivation and demonstrates the activity in the
presence of high and low titers of viruses.

One of the initial studies evaluated the effect of PEN 110 on four enveloped viruses, six non-
enveloped viruses, and cell-associated HIV. The viruses were chosen to include a wide array
of physiological characteristics such as size, structure, biochemical composition, and whether
the virus was cell associated or cell free. Of the enveloped types, Bovine Viral Diarrhea
Virus (BVDV) and pseudorabies virus (PRV) are recognised models for HCV and herpes
virus. Other enveloped viruses included in the study were sinbis (SV) virus and
vesicularstomatitis Indiana virus (VSIV).

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 The non- enveloped viruses included porcine parvovirus (PPV), which is a model for B19
virus. Vesicular exanthema of swine virus (VESV), is a model for hepatitis E and Norwalk
viruses. In these studies, the inactivation process was altered by using 1M sodium
thiosulphate (STS) and 1M 3-morpholinopropanesulphonic acid (MOPS) to chemically stop
the PEN 110 reaction (rather than washing) to ensure that the decreased viral cyctotoxic
effect was due to PEN111 alone and not washing. In addition, red cell units were collected in
three different additive solution storage media CPD/AS-1, CP2D/AS-3, and CPD/As-5. Virus
titers were determined by tissue culture infectious dose 50% (TCID50) assay and plaque-
forming units assay. The kinetics of viral reduction was determined by collecting samples at
3, 6, 12, 18 and 22 hours during incubation with PEN111. Three of the non-enveloped
(VESV, foot and mouth disease virus [FMDV], bluetongue virus [BTV] and two of the
enveloped viruses (SV and VSIV) were reduced to the level of detection of the assay within 3
hours of incubation. These viruses were reduced by factors ranging from 5.4 to 7.5 log10
TCID50 per milliliter. BVDV and PRV took longer 12 hours to be reduced to the level of
detection. Three of the non-envoloped viruses PPV, reo-3 and Adeno-2 took 18 hours to be
reduced to the level of detection. The reduction factors for these viruses were >6.2 log10
TCID50 per milileter, >5.3 and >5.6 log10 PFU per milliliter, respectively. For cellular HIV
the reduction to the level of detection took 6 hours. 20 Level II.

Another series of experiments investigating PEN110 inactivation of HIV. The effect of

PEN110 on the structure of the virus by EM, the ability of virus to enter and the activity after
the treatment of reverse transcriptase using PEN110 were investigated. The demonstration of
the PEN110 treatment effect was estimated via the TCID50 assay. The structure of the virus
did not change and the ability of the virus to infect cells remained intact after PEN110.
PEN110 effectively reduced HIV-1 to the limit of detection for a reduction factor of at least
5.57 log 50 percent tissue culture infectious dose per bulk test. The PEN110-treated virions
maintained their morphology, protein integrity, and functionality However; the viral reverse
transcriptase did not function, consistent with the alteration of nucleic acid by PEN110. The
HIV reduction took place primarily in the first 6 hours and decrease to the level of detection
after 12 hours. 21 Level II

6.0. C. CONCLUSION
RED BLOOD CELLS (RBC)
Safety
i) S 303
Fair level of evidence to show the safety of S303 when using as pathogen inactivation
technology for RBC.
Effectiveness
i) S 303
Fair evidence obtained on the effectiveness of S303 as a pathogen inactivation
technology when used on RBC.
ii) PEN110
Poor evidence for the effectiveness of PEN110 as a pathogen inactivation technology
when used on RBC.
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 Cost effectiveness
No retrievable evidence on the cost effectiveness of S303 and PEN110 pathogen
inactivation technologies when used for RBC.
7.0. REFERENCES
1. Council of Europe Expert Committee in Blood Transfusion Study Group on Pathogen
Inactivation of Labile Blood Components. Pathogen inactivation of labile blood
products. Transfus Med. 2001 Jun; 11(3):149-75.
2. Kaur P, Basu S. Transfusion-transmitted infections: existing and emerging pathogens. J
Postgrad Med. 2005 Apr-Jun; 51(2):146-51.
3. Bryant BJ, Klein HG. Pathogen inactivation: the definitive safeguard for the blood
supply. Arch Pathol Lab Med. 2007 May; 131(5):719-33
4. Chris Powse. Pathogen inactivation of blood components. The author Journal
Compilation 2008 LMS Group Ltd. Transfusion Alternatives in transfusion Medicine.
2008: 10, 139-146.
5. Solheim BG, Seghatchian J. Update on pathogen reduction technology for therapeutic
plasma: an overview. Transfus Apher Sci. 2006 Aug;35(1):83-90
.
6. Pelletier JP, Transue S, Snyder EL. Pathogen inactivation techniques. Best Pract Res
Clin Haematol. 2006;19(1):205-42. Review.
7. Jeffry McCullough . Pathogen inactivation of platelets. Transfusion Alternatives in
Transfusion Medcine 10,1111/j.1778-428X2006.0015
8. Dr. R. Vijayaraghavan. Porphyrins : Dynamic Photosensitizer in Photodynamic
Therapy.http://www.pharmainfo.net/reviews/porphyrins-dynamic-photosensitizer-
photodynamic-therapy) retrieved on: 11-09-2010
9. INHATA Briefs CAHTA, Cathalan Agency For Health Technology Assessment.
Amotosalen (Intercept) for the Inactivation of Pathogens for Transfusion Therapy 2009,
June ,www.gemcet.cat/salut/depsan /units/aatrm/pdf.
10. Stephen K., Doug Coyle, Don Husereau, et al. Octaplas Compared with Fresh Frozen
Plasma to Reduce the Risk of Transmitting Lipid-Enveloped Viruses: An Economic
Analysis and Budget Impact Analysis. Alan Tinmouth, MD, MSc, FRCPC2 Sarah
Normandin, BSc, MLIS1. Canadian Agency for Drugs and Technologies in Health.
HTA Issue 107. CADTH. April 2009.
11. Griffiths E and Padilla A. Guidelines on viral inactivation and removal procedures
intended to assure the viral safety of human blood plasma products. World Health
Organization (WHO) Technical Report, Series No. 924, 2004.
12. Solheim BG, Seghatchian J.) Update on pathogen reduction technology for therapeutic
plasma: an overview. Transfus Apher Sci. 2006 Aug;35(1):83-90.

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13. Riedler GF, Haycox AR, Duggan AK, Dakin HA. Cost-effectiveness of
solvent/detergent-treated fresh-frozen plasma. Vox Sang. 2003 Aug;85(2):88-95.

14. Pereira A. Cost-effectiveness of transfusing virus-inactivated plasma instead of standard

plasma. Transfusion. 1999 May;39(5):479-87.
15. van Rhenen D, Gulliksson H, Cazenave JP, e al. Transfusion of pooled buffy coat
platelet components prepared with photochemical pathogen inactivation treatment: the
euroSPRITE trial. Blood. 2003 Mar 15;101(6):2426-33.
16. Kerkhoffs JL, van Putten WL, Novotny VM, et al. Clinical effectiveness of
leukoreduced, pooled donor platelet concentrates, stored in plasma or additive solution
with and without pathogen reduction. Br J Haematol. 2010 Jul; 150(2):209-17.
17. The Mirasol Clinical Evaluation Study Group. A randomized controlled clinical trial
evaluating the performance and safety of platelets treated with MIRASOL pathogen
reduction technology. Transfusion. 2010 May 18
18. Rios JA, Hambleton J, Viele M, et al. Viability of red cells prepared with S-303
pathogen inactivation treatment. Transfusion. 2006 Oct; 46(10):1778-86.
19. Benjamin RJ, McCullough J, Mintz PD, et al. Therapeutic efficacy and safety of red
blood cells treated with a chemical process (S-303) for pathogen inactivation: a Phase
III clinical trial in cardiac surgery patients. Transfusion. 2005 Nov; 45(11):1739-49.
20. Lazo A, Tassello J, Jayarama V, et al. Broad-spectrum virus reduction inred cell
concentrates using INACTINE PEN110 chemistry. Vox Sang. 2002 Nov;83(4):313-23.
21. Ohagen A, Gibaja V, Aytay S, Horrigan J, Lunderville D, Lazo A. Inactivation of HIV
in blood. Transfusion. 2002 Oct; 42(10):1308-17.

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9. APPENDIX

9.1 Appendix 1

DESIGNATION OF LEVELS OF EVIDENCE

I Evidence obtained from at least one properly designed randomized controlled trial.

II-I Evidence obtained from well-designed controlled trials without randomization.

II-2 Evidence obtained from well-designed cohort or case-control analytic studies, preferably from
more than one centre or research group.

II-3 Evidence obtained from multiple time series with or without the intervention. Dramatic results in
uncontrolled experiments (such as the results of the introduction of penicillin treatment in the
1940s) could also be regarded as this type of evidence.

III Opinions or respected authorities, based on clinical experience; descriptive studies and case
reports; or reports of expert committees.

SOURCE: US/CANADIAN PREVENTIVE SERVICES TASK FORCE (Harris 2001)

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