Quality Control And Pathogen Inactivation of Platelets Ohood Al.Ayyadhi Laboratory Manager Blood Transfusion services Kuwait Central Blood Bank

Pathogen Inactivation Introduction. Why Pathogen Inactivation. Methods. Kuwait Central Blood Bank Results.

Infectious Risks Transfusion of biological fluids = Viruses, Parasites, Bacteria, Prions, ???

Infectious Risks Test vs Risk Known Unknown

Pathogens of highest concern in the Middle East West Nile virus SARS Vaccinia Chikungunya virus Dengue Avian flu virus (H5N1) Borrelia burgdorferi Trypanosoma cruzi Plasmodium falciparum Leishmania Babesia microti HIV HBV HCV HTLV Bacterial Contamination

Kuwait Central Blood Bank

What’s our Scope? The ONLY Central Blood Bank 21 Government Hospitals 20 Private Hospitals 2 Military Hospitals Blood product supplier to Allied armies

What’s our Scope? 69,000 Whole blood donations. 100% Leukocyte Reduced RBC & Plasma. 7,794 Apheresis Platelets (55,000 units).

What’s our Scope? AABB Accredited 1989. CAP Survey 1994. Accredited by the National Quality Program. National Reference Laboratory. Accredited as regional reference center for Arabian countries. AABB Immunohaematology Reference Lab IRL self assessment 2008.

What’s our Scope? Training center for post-graduate hematologist. Training center for pre-graduate medical lab technologist. Training center for regional countries. Therapeutic Apharesis Center. National Antenatal screening program.

Pathogens of highest concern in the Middle East (KCBB) Bacterial Contamination. HBV. HCV. HIV.

Viral Infectious Risks At KCBB

Infectious Risks at KCBB 2011 2010 2009 2008 HIV 68239 62720 60991 61771 Total Tests 4(0.006%) 0 (0.00%) 1 (0.01%) 4 (0.06%) Anti-HIV 2 3 4 NAT-HIV

Infectious Risks at KCBB 2011 2010 2009 2008 HBV 68239 62720 60991 31771 Total Tests 122(0.19%) 163(0.26%) 163(0.27%) 191(0.31%) HBsAg 157 59 175 139 NAT-HBV 5715 (8.76%) 5657(9%) 6187(10%) 6059(9.8%) ANTI-HBC 3950 (6.06%) 3753(5.98%) 3754(60%) 3979(65%) ANTI-HBs

Infectious Risks at KCBB 2011 2010 2009 2008 HCV 68239 62720 60991 61771 Total Tests 110 (0.17%) 163 (0.26%) 208 (0.34%) 317 (0.19%) Anti-HCV 76 53 122 115 NAT-HCV

Infectious Risks at KCBB 2011 2010 2009 2008 HTLV 68239 65218 60991 31771 Total Tests 9 (0.012%) 12 (0.019%) 8 (0.01%) 14 (0.02%) Anti-HTLV

Infectious Risks at KCBB 2011 2010 2009 2008 MALARIA 8207 10830 11319 11218 Total Tests 728 (8.87%) 220 (2.03%) 239 (2.1%) 183 (3%) Anti-Malaria

Bacterial Contamination Risks of Platelets At Kuwait Central Blood Bank

Transmission risk, per unit Comparison of Residual Risks 1:100 Transmission risk, per unit HIV Bacterial Contamination (platelets) 1:1000 1:10 000 HBV HCV Septic Fatalities (platelets) 1:100 000 1:1 000 000 1984 1986 1988 1990 1992 1994 1996 1998 2000 2002 Updated from: Goodnough LT e t al. NEJM 1999;341:126-7

Bacterial Contamination Risks Unit Transfused Risk per Million Units Confirmed Report of Fatality Bacterial Contamination Red Blood Cells 6.0 1.0 Platelet pheresis units 32 7.1 TOTAL, all units 7.4 1.1 Perez P et al. Transfusion 1999;39:2S.

Bacterial Contamination Risks Unit Transfused Risk per Million Units Confirmed Report of Fatality Bacterial Contamination Red Blood Cells 6.0 1.0 Platelet pheresis units 32 7.1 TOTAL, all units 7.4 1.1 1/140,800 Perez P et al. Transfusion 1999;39:2S.

Bacterial Contamination Risks Unit Transfused Risk per Million Units Confirmed Report of Fatality Bacterial Contamination Red Blood Cells 6.0 1.0 Platelet pheresis units 32 7.1 TOTAL, all units 7.4 1.1 1/140,800 1/31,000 Perez P et al. Transfusion 1999;39:2S.

Frequency of Contamination Based on Johns Hopkins’ Data Plt Conc SDP Post-transfusion sepsis 402/million 75/million Fatalities 62/million 14/million Ness PM et al. Transfusion 2001;41:857-61. Recalculation: LJ Dumont.

Frequency of Contamination Based on Johns Hopkins’ Data Plt Conc SDP Post-transfusion sepsis 402/million 75/million Fatalities 62/million 14/million Compare: HIV 0.33/million HCV 1/million Ness PM et al. Transfusion 2001;41:857-61. Recalculation: LJ Dumont.

Bacterial Contamination AABB STD 5.1.5.1 The blood bank or transfusion services shall have methods to limit and detect bacterial contamination in all platelet components.

History of testing in KCBB Date Syphilis 1965 HBVs-Ag 1970 HIV-Ab 1985 Malaria-Ab 1987 HBVc-Ab 1992 HBVs-Ab HCV-Ab HTLV-I&II Ab 1994 HIV-I & II Ab 1997 HIV-Ag Bacterial Detection 2005 NAT- HIV, HCV, HBV 2006 27

Bacterial Risk at KCBB 6,800 Apheresis Platelets (~ 40,000 units). 0.5 – 1 confirmed cases of sepsis per year.

Bacterial Detection March 05 – May 07 26,000 units eBDS Pall Scansystem March 05 – May 07 26,000 units eBDS Pall June 07 – May 08 14,000 units 10% of collection tested by culture as QC

Bacterial Detection Summary results of QC of platelet aphaeresis component (10% collection) Detection System Tested Components Initial Positive False Positive False Negative Scansystem March 05 – May 07 2475 5 (.20%) 1 (.04%) eBDS June 07 – May 08 1292 2 (.15%) 1 (.07%) Total 3767 (22,602) 7 (.19%) 2 (.05%)

Mitigation Delay of component release (2 days) Complexity of procedures Results: False positives (0.2 %) False negatives (0.05%) Safety?: Did not prevent all septic transfusion reactions Did change the risks of bacterial contamination.

Pathogen Inactivation WHY ?

Why is Pathogen Inactivation Important? Reduced risk of bacterially contaminated platelet transfusion Further closing of window period for screened viruses Added protection against untested pathogens (e.g. Chagas) Pro-active protection against emerging pathogens (e.g. Chikungunya, West Nile) Possible reduction in adverse transfusion events Potential to revisit donor deferral strategies and enlarge donor pool Public expectation of ‘ZERO risk’

Pathogen Inactivation Systems

Platelet Pathogen Inactivation Systems Intercept Amotosalen + UV

Platelet Pathogen Inactivation Systems Intercept Mirasol Riboflavin + UV

Platelet Pathogen Inactivation Systems Intercept Mirasol Theraflex UV

Intercept Blood System for Platelets

Mechanism of Action UVA Illumination Amotosalen DNA or RNA of pathogen Intercalation Crosslinking 39

Amatosalen Cross links Both Single and Double Stranded Nucleic Acids Helical Regions Single-stranded DNA or RNA Double-stranded DNA or RNA 40

INTERCEPT Blood System for Platelets Integrated Processing Set 1 Collection 2 Amotosalen 3 Illumination 4 CAD 5 Storage Amotosalen HCl is formulated in saline as a 3 mM solution. The nominal volume used is 15 ml (apheresis) or 17.5 ml (Buffy Coats). When 15 ml of the solution is added to 285 ml of apheresis platelet concentrate, the final volume is 300 ml and the final amotosalen HCl concentration is 150 M. The same concentration is achieved when 17.5 ml of the solution is added to 350 ml Buffy Coat platelets. In vitro studies have shown that the INTERCEPT Blood System platelet process retains its effectiveness with amotosalen HCl concentrations in the range of 120–180 M. UVA Illumination Device 41

Mirasol® Pathogen Reduction Technology System

Mirasol PRT System Overview The Mirasol PRT System uses riboflavin (vitamin B2) and UV light to inactivate pathogens, altering their nucleic acids so they cannot replicate.

Mechanism of Action 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

+ + Mirasol PRT System Concept Platelet or Plasma product Riboflavin (Vitamin B2) solution UV Light Reduction of viruses, bacteria, parasites Inactivation of residual white cell

Theraflex UV System

Theraflex

Theraflex Methylen Blue and UV illumination device used as a technique for Pathogen Reduction

KCBB Clinical Experience with Platelet PI

Pathogen Inactivation (INTERCEPT) Implementation Training, March 2008 Validation, May 2008 100 % Pathogen Inactivation of Platelets replaced 1st May 2008 AABB inspection 15th May 2008

KCBB Results Source Trima Accel version 5.2 / 6.0 Hemonetics (MCS+) 998 CF-E

KCBB Results Collection dose Single dose (3-4 x 1011/unit) Double dose (6-7 x 1011/unit) Baby units (0.5 x 1010/unit)

KCBB Results Source

KCBB Results Collection dose Single dose (3-4 x 1011/unit) Double dose (6-7 x 1011/unit) Baby units (5 x 1010/unit) Double Single

KCBB QC Results Parameters: Volume (Pre & Post) Platelet Count (Pre & Post) Count Loss Culture

KCBB QC Results QC Samples: Total 990 (15.3%) Valid 718 (11.1%)

KCBB QC Results Total: 718 66 % of tested units ≥ 3 x 1011/unit Pre-PI is < 3 x 1011/unit Divided as baby units. Pathogen Inactivation process has no significant effect on the final product.

KCBB QC Average Results Parameter July-August 2008 Pre Platelet Count 3.25 Post Platelet Count 3.05 Loss 0.21 August-September 2008 3.8 3.2 0.6 All Culture NEGATIVE

KCBB QC Average Results All Culture NEGATIVE

leucocytes reduce aphaeresis platelet pathogen inactivation INTERCEPT Pathogen Inactivation Workflow Registration Collection Filtration Process Processing Storage Duration Average time 2hr Average time = 3-6min Average time =6min Average time =6-16hr Time zero 02:00 02:15 02:20 02:30 08:30 Random Donation Filtration Illumination CAD start Final Platelets Haemonitic: no need for resting Trima: Resting time 1hr Print label leucocytes reduce aphaeresis platelet pathogen inactivation End Product: Action Aphaeresis PLT Collection tools Sealer Computer Filtration Stands Clamps Sealers Connection device Computer Scales Intercept set (LV/SV) Incubator Scales Clamps Sealers Computer RAD-SURE (UVA illumination indicator) Equip- ment

Present Testing Test Date Syphilis 1965 HBVs-Ag 1970 HIV-Ab 1985 Malaria-Ab 1987 HBVc-Ab 1992 HBVs-Ab HCV-Ab HTLV-I&II Ab 1994 HIV-I & II Ab 1997 HIV-Ag NAT- HIV, HCV, HBV 2005 Bacterial Detection 2006 61

Present Testing Test Date Syphilis 1965 HBVs-Ag 1970 HIV-Ab 1985 Malaria-Ab 1987 HBVc-Ab 1992 HBVs-Ab HCV-Ab HTLV-I&II Ab 1994 HIV-I & II Ab 1997 HIV-Ag NAT- HIV, HCV, HBV 2006 Bacterial Detection REMOVED 2008 62

Present Testing Processing Date Syphilis 1965 HBVs-Ag 1970 HIV-Ab 1985 Malaria-Ab 1987 HBVc-Ab 1992 HBVs-Ab HCV-Ab HTLV-I&II Ab 1994 HIV-I & II Ab 1997 HIV-Ag NAT- HIV, HCV, HBV 2006 Bacterial Detection REMOVED 2008 Pathogen Inactivation 2008 63

Clinical Effectiveness

Processing effects Platelet count loss. Platelet activation. Frequent platelet transfusion. Compromised CCI.

Processing effects Same day release of Platelets. Simple procedure. No bacterial contamination. Less allo-immunization.

Cost Expensive.

Cost Less expensive than: Bacterial testing.

Cost Effectiveness at KCBB Better platelet products quality on TIME. STOP Bacterial detection tests. STOP Irradiation. STOP Malaria testing.

Platelets Pathogen Inactivation Conclusion PI highly prevalence agent. PI vs. Testing; compromised budgeting. PI in highly endemic areas.

Acknowledgment Dr. Reem Al.Radwan Ghadeer Boland Dr. Nabeel Sanad Suhaila Al.Shatty Samiya Al.Hamdan Badriya Al.Radwan Hanan Sheshtari Ghadeer Ashkanani Maryam Ameer Ghadeer Boland Quality Control Lab Quality Managment Department IT Department Platelets Donation Department

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